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Mineral Metabolism Laboratory (D.J.B., S.M.), Jerry L. Pettis Memorial Veterans Administration Medical Center, Department of Nutrition (S.R.), School of Public Health, Departments of Medicine, Biochemistry, and Physiology, School of Medicine (D.J.B., S.M), Loma Linda University, Loma Linda, California 92357
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Abstract |
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 Top Abstract I. IntroductionII. Characteristics of the...III. Target Cell Actions...IV. IGF-IGFBP Complexes in...V. Assays for Circulating...VI. Relative Distribution of...VII. Regulation of Serum...VIII. IGFBP Proteases in...IX. Endocrine Functions of...X. ConclusionsReferences |
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I. Introduction |
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TopAbstractI. IntroductionII. Characteristics of the...III. Target Cell Actions...IV. IGF-IGFBP Complexes in...V. Assays for Circulating...VI. Relative Distribution of...VII. Regulation of Serum...VIII. IGFBP Proteases in...IX. Endocrine Functions of...X. ConclusionsReferences |
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In subsequent studies, Zapf et al. (14 ) incubated plasma with radiolabeled IGF and detected the somatomedin activity at 40 kDa. This binding was highly specific for the IGFs with no competition from insulin, suggesting the presence of specific high-affinity binding proteins for the IGFs. Furthermore, Kaufmann et al. (15 ) showed that the half-life of 125I-labeled NSILA was reduced when excess unlabeled NSILA was injected into normal rats, suggesting that NSILA is bound to carrier proteins. The binding of NSILAs to the carrier proteins in serum provided a possible explanation for the absence of insulin-like effects of endogenous NSILA in vivo.
The discovery of the six different binding proteins currently known to
exist did not occur all at the same time (1 2 3 4 5 6 7 16 ). The first three
IGFBPs were purified in the mid-1980s. The association of
[125I]IGF in rat serum initially with a small molecular
binding protein complex and a shift to a larger complex after a few
minutes confirmed the presence of two binding proteins (14 ). The first
was a major binding protein in serum identified to be a GH-regulated
acid-labile 150- to 200-kDa complex eventually designated as
IGF+IGFBP-3+acid labile subunit (ALS) complex. The second was a
GH-independent 50 kDa acid-stable protein (2 17 ). Later it was shown
that this 50-kDa small molecular binding protein consisted of IGFBP-1
and IGFBP-2. By the late 1980s and early 1990s, three additional
binding proteins, IGFBPs 4 through 6, were isolated and characterized.
All six binding proteins share 
35% sequence identity with each
other. Several excellent reviews on the structure, molecular and
cellular aspects, and biological actions of these binding proteins are
available (1 2 3 4 5 6 7 ) and therefore will not be the focus of this review.
The recent development of specific assays for measurement of various
IGFBPs in circulation and in other biological fluids has led to
significant new information on serum regulation of IGFBPs and their
functions. The main focus of this review is to present the data
regarding the characterization of the IGF-IGFBP complexes in serum and
other biological fluids and to evaluate their regulation and functions.
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II. Characteristics of the IGFBPs |
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TopAbstractI. IntroductionII. Characteristics of the...III. Target Cell Actions...IV. IGF-IGFBP Complexes in...V. Assays for Circulating...VI. Relative Distribution of...VII. Regulation of Serum...VIII. IGFBP Proteases in...IX. Endocrine Functions of...X. ConclusionsReferences |
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IGFBP-1, a nonglycosylated protein of 30 kDa, was first isolated from
mid-term amniotic fluid (20 ). This binding protein shares sequence
identity with the placental protein 12 (19 26 ), which is synthesized
by endometrium and decidua. It is present in the amniotic fluid in
concentrations 100–500 times higher than in serum. IGFBP-2, originally
isolated from rat liver (BRL)-3A cell line (27 ), is a nonglycosylated
protein of 31–36 kDa and is found in significant amounts both in serum
and cerebrospinal fluid (1 ). The major form of binding protein present
in human circulation is IGFBP-3; its molecular mass ranges from 38 kDa
to 43 kDa depending on the number of sites glycosylated (28 ). In
circulation, this glycoprotein is associated with an IGF molecule and
an 80-kDa acid-labile subunit (ALS) to form a 150- to 200-kDa complex
(2 29 30 ). This complex consists of IGFBP-3 and IGF-I + IGF-II in an
equimolar ratio, suggesting that most of the IGFBP-3 in serum is likely
to be saturated.
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Although the IGFBPs differ in their structure and binding specificity, it is not clear whether these differences contribute to functional differences among the various IGFBPs. For example, it is not known whether there is any functional significance for glycosylation of the IGFBPs and why some of the IGFBPs bind IGF-II with preferential affinity (39 40 ) compared with IGF-I. Interestingly, none of the IGFBPs bind IGF-I with preferential affinity. In any case, the differences in structure (glycosylation, number of cysteine, RGD sequence), binding affinity, and tissue-specific expression are consistent with the general idea that different IGFBPs have discrete functions. Based on the findings that extracellular fluids of certain tissues (described in Section IV) are enriched with specific IGFBPs and that tissues surrounding the body fluid express the same IGFBPs in high abundance (41 42 ), it is speculated that the IGFBPs may function locally to regulate IGF actions. However, there is no experimental data to demonstrate that this is in fact true.
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III. Target Cell Actions of the IGFBPs |
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TopAbstractI. IntroductionII. Characteristics of the...III. Target Cell Actions...IV. IGF-IGFBP Complexes in...V. Assays for Circulating...VI. Relative Distribution of...VII. Regulation of Serum...VIII. IGFBP Proteases in...IX. Endocrine Functions of...X. ConclusionsReferences |
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A. To modulate IGF actions
Studies in a number of laboratories including ours have shown that
IGFBPs are capable of modulating IGF-induced cell proliferation both in
a positive and negative manner (3 4 43 44 45 ). Several IGFBPs,
including IGFBP-1, IGFBP-2, IGFBP-4, and IGFBP-6, inhibit IGF
action by binding to IGFs and preventing the binding of IGFs to IGF
receptors (3 4 ). In contrast to the phosphorylated IGFBP-1 that
inhibits IGF actions, the nonphosphorylated form of IGFBP-1 potentiates
the effect of IGF-I on DNA synthesis in porcine smooth muscle cells
(4 ). Coincubation of human fibroblasts with IGF and IGFBP-3 showed an
inhibitory effect while preincubation with IGF had growth-potentiating
effect (43 ). It was suggested that binding of IGFBP-3 to the cell
surface reduces its affinity for IGF-I and results in a potentiating
effect (44 ). In contrast to other IGFBPs, IGFBP-5 is stimulatory for a
variety of cell types (45 46 47 48 ).
Thus, the different binding proteins may modulate IGF action differently, and the same binding protein can have an IGF-inhibiting or potentiating role under different conditions. The factors that determine these differences include IGFBP phosphorylation, IGFBP proteolysis, and IGFBP cell surface association, among others. These variables may modulate IGF action in target tissues by altering the binding affinity of the IGFBPs to IGFs.
B. To facilitate storage of IGFs in extracellular matrices
Another important role of IGFBPs may be to help in the storage of
IGFs in the extracellular matrices of certain tissues. In this regard,
Jones et al. (45 ) provided evidence for fixation of IGFs via
IGFBP-5 binding to extracellular matrix proteins. We found evidence
that IGFBP-5 may help fix IGFs in bone since the complex of IGFBP-5 and
IGFs, but not IGFs alone, bind to hydroxyapatite (34 49 ). In terms of
the significance of fixation of IGFs in extracellular matrices such as
bone, it is speculated that the stored IGFs may be released during the
osteoclastic bone resorption phase of bone remodeling to stimulate
nearby osteoblasts during the bone formation phase of remodeling (50 ).
Similarly, IGFs stored in extracellular matrices of soft tissues may
have a role in wound healing.
C. To exert IGF-independent effects
Recent evidence suggests that some of the IGFBPs may mediate their
effects on target cells by an IGF-independent pathway. This concept has
evolved from a number of experimental studies, including the study by
Jones et al., which found that IGFBP-1 stimulated smooth
muscle cell migration by an IGF-independent mechanism involving
integrin receptors (51 ). IGFBP-3 has been shown to inhibit
proliferation of breast and prostate cancer cells by a cellular
signaling pathway independent of IGFs (52 53 ). In addition, Rajah
et al. (54 ) have recently shown that IGFBP-3 induces
apoptosis of the p53-negative prostate cancer cell line, PC3, through a
novel pathway independent of either p53 or the IGF-IGF
receptor-mediated cell survival pathway. Consistent with the idea that
IGFBP-3 may have IGF-independent effects on certain types of cells, two
recent reports have provided evidence for nuclear localization of
IGFBP-3 (55 56 ). The significance of this finding is not clear.
However, this exciting finding may clarify the direct intrinsic actions
of some of the IGFBPs on cells. We and others have found evidence that
IGFBP-5 may promote cell proliferation in osteoblasts, possibly through
putative cell surface-binding sites (46 47 ). Although studies from a
number of laboratories support the possibility that IGFBPs may have
IGF-independent effects in certain cell types, further experimental
evidence is needed to verify this mode of IGFBP action.
Thus, the explosion of IGFBP research during the past several years has provided evidence that IGFBPs may have both IGF-dependent and IGF-independent actions. Based on the complexity of IGFBP functions, it is clear that we cannot fully appreciate the significance of changes in IGFBP levels in serum and local body fluids until we know more about the functions of these IGFBPs.
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IV. IGF-IGFBP Complexes in Biological Fluids |
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TopAbstractI. IntroductionII. Characteristics of the...III. Target Cell Actions...IV. IGF-IGFBP Complexes in...V. Assays for Circulating...VI. Relative Distribution of...VII. Regulation of Serum...VIII. IGFBP Proteases in...IX. Endocrine Functions of...X. ConclusionsReferences |
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Figure 1 shows the relative distribution
of various IGF pools in human serum. In circulation, about 75–80% of
the IGFs are complexed to IGFBP-3 and the acid labile 
-subunit to
form the 150- to 200-kDa complex (57 59 63 ). This is possible because
under normal conditions, the total IGFs and IGFBP-3 in serum are in
equimolar concentrations (63 ). A smaller percentage (20–25%) of the
IGFs are associated with low molecular mass IGFBPs (57 ). Less than 1%
are found in the free form in circulation (64 65 ).
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).
Subsequently, the reconstitution of the 150- to 200-kDa complex was
achieved by using purified acid-labile subunit (ALS or -subunit),
IGFBP-3 (ß-subunit), and IGF-I or IGF-II (
-subunit). The role of
ALS appears to be to increase the molecular mass of the IGF+IGFBP-3
complex so that the access of the circulating IGF to the extracellular
fluid and thus to the various tissues is limited (57 58 ).
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The question of whether ALS can form a binary complex with IGFBP-3 in the absence of the IGF ligand is controversial at this time based on recent reports by Barreca and colleagues (71 72 73 ) and Lee et al. (74 ). Barreca et al. (71 ) demonstrated that incubation of recombinant human IGFBP-3 and ALS resulted in the appearance of a 150- to 200-kDa complex in the absence as well as in the presence of IGF. They also showed that ALS binding to IGFBP-3 increased the affinity of IGFBP-3 to IGF-I, possibly by inducing conformational changes in IGFBP-3. Based on these results, the authors speculate that ALS may play an important role in regulating the affinity of IGFBP-3 to IGF-I, thus regulating the levels of free IGFs. In addition, Yang et al. (75 ) observed that [125I]IGF-II readily bound to the 150-kDa fraction of adult rat serum to sites with a higher affinity for IGF-II than IGF-I. In subsequent studies, Lee and Rechler (76 ) demonstrated two different IGFBP-3 complexes in the 150- to 200-kDa fraction of the adult rat serum, one with similar affinity for IGF-I and -II and the other with greater affinity for IGF-II (77 ). The latter complex is formed from proteolytically nicked IGFBP-3 that is present in the native serum before acidification. The proteolytic cleavage in IGFBP-3 decreases the affinity of the IGFBP-3-ALS for IGF-I and increases the binding of IGF-II. Similar proteolytic nicking of IGFBP-3 occurs during human pregnancy, changing the binding specificity for IGFs (78 ). In contrast to these results, Baxter and co-workers (79 ) demonstrated that proteolyzed IGFBP-3 from maternal serum can bind to IGFs and form a ternary complex with ALS with normal affinity. Thus, they speculated that the altered binding affinity of the IGFBP-3 fragment during Western ligand blotting is an artifact resulting from breakage of a labile peptide bond after prolonged acidification or exposure to SDS.
If human IGFBP-3 must first bind to IGF-I before it can form the ternary complex, then the amount of IGF-I associated with the 150- to 200-kDa complex in rats injected with hIGFBP-3 should be twice that of normal rats. However, Lee et al. (74 ) did not observe an increase in the mobilized IGF and thus concluded that IGFBP-3 and ALS can form a binary complex independent of IGF both in vivo (80 ) and in vitro (71 76 ). Thus the question of whether IGF is required for the formation of a complex between IGFBP-3 and ALS remains controversial. One possibility is that there may be two IGFBP-3 pools, one with a higher binding affinity for ALS after first binding to IGF, and the second, which binds ALS even in the absence of IGF but with lower affinity. With differences in binding affinities, the functional roles of ternary complexes of ALS+IGFBP-3+IGF and binary complex of ALS+IGFBP-3 may also be different. Future studies are required to elucidate the extent to which IGFBP-3 forms a binary complex with ALS and if so, whether these binary complexes play a physiological role in modulating the actions of IGF.
One of the proposed functions of plasma IGFBPs is to increase the
half-life of IGFs in circulation. When IGF-I and IGF-II are injected
into normal rats, they bind to IGFBP-3, increasing their stability and
half-life to 4 h compared to 20 min in hypophysectomized rats
(81 ). Guler et al. (57 ) determined the half-lives of free
and IGFBP-bound [125I]IGF-I and -II after bolus injection
of the tracers in two normal adults. Apparent half-lives of
[125I]IGF-I and -II in each of the three IGF serum pools
(150 to 200 kDa, 50 kDa, and free IGF), calculated from the respective
disappearance rates of the tracer, are shown in Fig. 3. These results demonstrate that the
150- to 200-kDa complex is responsible for the relatively long
half-life of IGFs and that the 50-kDa and the free IGF pool have a
rapid turnover and account for most of the daily IGF production (82 ).
Another important role of IGFBP binding to IGF is in modulating IGF
action. The ternary complex does not permeate the capillary endothelial
barrier, but the smaller IGF-IGFBP complexes can easily do so and
facilitate tissue-specific IGF action (3 4 57 59 ). On the other
hand, the endocrine actions of IGFs bound to IGFBP-3 may be achieved by
specific proteolytic enzymes that dissociate the ALS+IGFBP-3+IGF
complex, and it is suggested that this may increase the bioavailability
of IGFs. An additional mechanism altering IGF bioavailability has been
proposed by Yamamoto and Murphy (83 ). They identified the presence of a
protease in rat serum that cleaves IGF-I into des(1 2 3 )IGF-I. Since
this form dissociates from the binding proteins easily, it may serve to
increase IGF bioavailablity. The role of proteases in modulating IGF
bioavailability is discussed in detail in Sections VIII and
IX.
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B. Milk
Human milk contains IGFBP-1, -2, and -3 (84 ), the functions of
which remain unclear. In addition, IGFBP-4 has been identified in rat,
porcine, and bovine milk (85 86 ). As with serum, IGFBP-3 is the major
binding protein of IGFs in milk. Although maternal serum is the source
of rat milk IGFBP-3 (21 ), the 150- to 200-kDa complex is not
translocated from serum into milk. It is suggested that IGFBP-3 may
enter milk from circulation in the free form or complexed to IGF-I. On
the other hand, IGFBP-2 and -4 are produced locally by the mammary
gland as shown by expression of their respective mRNA in the mammary
tissue (21 ). IGFBP-1 in human milk (87 ) parallels the level of IGF-I in
that immediately after birth, both milk IGF-I and IGFBP-1 decline. The
exact role of IGFBPs in milk remains to be explored, but it is
possible that they protect against the degradation of milk IGF-I or
that they modulate the local mitogenic activity of the IGFs.
C. Urine
IGFBP-1, -2, and -3 have been detected in healthy adult urine by
Western ligand blot analysis (25 ), and the concentration is
approximately 3 orders of magnitude less than that in serum (88 ).
Previously, IGFBP-2 was shown to be the predominant form in dialyzed
adult urine, but when urine is not dialyzed, IGFBP-3 is the major
binding protein (88 ). The reason for this discrepancy is not known. It
appears that urinary IGFBP-3 originates mainly from the kidney and/or
the urinary tract and (89 ), unlike the serum, is not found as a 150- to
200-kDa complex in the urine. Quantification by RIA show that urinary
IGFBP-3 is age dependent, with an increase at approximately age 9–11
yr that corresponds to the pubertal rise in serum IGFBP-3 and is
followed by a decline until it plateaus at approximately age 26 yr
(89 ). Although the antiserum used in the RIA recognizes both intact and
proteolyzed fragments of IGFBP-3, there is no evidence for the presence
of urinary protease in normal individuals from age 4–45 yr (25 89 ).
It is suggested that urinary IGFBPs may have diagnostic utility, but
this has not been established.
D. Cerebrospinal fluid (CSF)
Although several different binding proteins have been identified
in human CSF, IGFBP-2 is the major form present (22 ). The CSF contains
high concentrations of IGF-II (90 ), which binds IGFBP-2 with 10- to-
20-fold greater affinity than IGF-I (91 ). Also purified from the CSF is
IGFBP-6, which is present at slightly lower concentrations in the CSF
than serum (92 ), but also has a preferential affinity for IGF-II over
IGF-I. Both IGFBP-3 and -5 have been identified in the CSF at lower
concentrations than IGFBP-2 and -6. It is suggested that the IGFBPs
found in the CSF may be synthesized locally by glial cells and neurons
and not derived from plasma by crossing the blood-brain barrier (93 ). A
30-kDa IGFBP, corresponding to IGFBP-2, was demonstrated in rat CSF
(94 ). By analogy with other transport proteins synthesized by the
choroid plexus, it is suggested that this IGFBP may facilitate the
secretion of IGF-II to the CSF and modulate its biological action at
distant sites within the brain (94 ). In a number of disease states
related to the central nervous system (CNS) (95 96 ), changes in IGFBP
concentrations have been documented, suggesting a possible diagnostic
utility for the measurement of these binding proteins. Thus IGFBPs are
thought to modulate the biological actions of IGFs (39 94 97 )
including a possible regulatory role in growth and differentiation of
the CNS. However, the mechanisms remain to be investigated.
E. Follicular fluid
IGFBPs 1–4 (41 98 99 100 101 102 103 104 ) were identified by Western ligand
blotting in human follicular fluid, suggesting that the regulation of
IGF action in the ovary is probably under the control of regulatory
binding proteins (98 ). In women with normal menstrual cycles, after
ovulation, progesterone stimulates the endometrium to release IGFBP-1
(105 ), which mediates the cell differentiation effects of IGF-I on the
endometrium. IGFBP-1 is synthesized by granulosa cells and is secreted
into the follicular fluid (101 103 106 107 108 109 ), where the concentration
is 4- to 5 times higher than that found in the serum (110 ). The
preovulatory rise in serum IGFBP-1 is not regulated by insulin or
ingestion of a meal, nor is it associated with diurnal variation (111 ).
This implies that during the preovulatory phase, IGFBP-1 detected in
serum is primarily of follicular origin. IGFBP-1 is thought to inhibit
the biological activity of free IGF on androgen-producing theca cells,
since that might lead to atresia and anovulation (112 ).
Changes in the various IGFBPs during atresia and follicular growth have been reported. The levels of IGFBP-2 and -4 are higher in atretic follicles (99 113 ) compared with healthy developing follicles of serum, suggesting a role for them in inducing atresia. A decrease in proteolytic activity degrading IGFBP-3 and an increase in IGFBP-2, -4, and -5 protease were observed during follicular growth in ovine follicular fluid, and an increase in IGFBP-3 protease and a decrease in IGFBP-4 and -5 protease were observed during atresia (100 101 114 115 ). These observations suggest that changes in intrafollicular IGFBP proteolytic activity could be responsible in part for the changes in IGFBP levels seen during growth and atresia (114 ). Since the expression of various IGFBPs is altered during follicular development and atresia, it is speculated that the changes in IGFBP levels may regulate follicular growth by modulating the local IGF bioavailability.
F. Amniotic fluid
The binding protein isolated from amniotic fluid (AFBP) is a small
molecular mass binding protein that is both heat and acid stable (42 116 ) and was later identified to be the same as IGFBP-1. It is the
major IGFBP in the amniotic fluid and is present in concentrations
100–500 times higher than that found in the serum (117 ). During
pregnancy, a surge in the concentration of amniotic fluid IGFBP-1
reflects its local production in decidual tissues (42 118 ). The
amniotic fluid IGFBP-1 contributes to the increase in serum IGFBP-1
levels during the second trimester of pregnancy (117 ). IGFBP-1 is twice
as high in preterm amniotic fluid as in term amniotic fluid, suggesting
a role for this binding protein in growth and development.
Immunoreactive IGFBP-3 is present in amniotic fluid but at a much lower concentration than that in serum. Western ligand blot analysis of amniotic fluid failed to reveal evidence for the presence of IGFBP-3 in amniotic fluid. This could be due to the presence of IGFBP-3 protease capable of degrading intact IGFBP-3 into fragments that do not bind [125I]IGF tracer. Consistent with this interpretation, incubation of amniotic fluid with radiolabeled IGFBP revealed the presence of a protease(s) specific for IGFBP-3, -4, and -5 (119 ). These proteases alter the binding affinity of the IGFs for their binding proteins and thereby could modulate the bioactivity of IGFs (119 ). The role of IGFBP-6 in the amniotic fluid is not yet known, although the levels present are similar to those seen in serum (92 ).
G. Lymph
The concentration of both IGFs and IGFBPs in lymph are lower than
that found in serum (120 121 ). Using gel filtration chromatography, it
was shown that the IGFBPs present in lymph eluted in the 40- to 50-kDa
size range. The finding that little IGF activity eluted as a 150- to
200-kDa complex from lymph is consistent with the fact that the
IGF+IGFBP-3 complex does not cross the capillary endothelial barrier.
IGFBP-2 is believed to be one of the major binding proteins in the
lymph tissue and may originate from both the serum and surrounding
local tissues (3 ).
H. Seminal fluid
IGFBP-1-like immunoreactivity was detected in human seminal plasma
(122 ), with levels similar to those found in human adult serum. Intact
IGFBP-3 could not be detected in seminal fluid by Western ligand blot
analysis, but Western immunoblot analysis using IGFBP-3 antiserum
revealed the presence of immunoreactive IGFBP-3 fragments (122 ). Since
the amounts and ratio of these binding proteins do not correlate with
those present in the serum, it is likely that the source of these
proteins are specific to the cell population within the local tissues,
such as Sertoli cells. Human seminal plasma also contains intact
IGFBP-2 and IGFBP-4, while IGFBP-3 is present in the fragmented
form (123 124 125 126 ). The prostate-specific antigen (PSA) has proteolytic
activity for not only IGFBP-3, but also for IGFBPs -4 and -5. In
addition to the PSA protease, an IGFBP-5-specific protease has been
identified in seminal plasma (125 126 127 ). Since IGFBP proteolytic
activities in seminal fluid from normal volunteers, vasectomized
patients, or patients with idiopathic azoospermia were not
significantly different, the role of IGFBPs and IGFBP proteases in the
male reproductive system and male infertility remains to be
established.
I. Other biological fluids
The presence of IGFBPs 1 through 4 has been detected in
interstitial fluid obtained from human skin blisters caused by high
negative pressure in healthy volunteers (128 ). The IGFBP-3
concentration was lower than that present in the circulation and was
due to increased IGFBP-3 protease activity. Several IGFBPs have been
identified in vitreous and aqueous humors (129 ), but the predominant
serum carrier protein IGFBP-3 was not detected in these fluids. This
may be due to the presence of increased amounts of IGFBP-3 protease
activity in vitreous and aqueous humors (129 ). Vitreous humor from
diabetics had a higher amount of IGFBP-3 proteolytic fragment compared
with healthy controls, suggesting that the rate of IGFBP-3 proteolysis
is different in vitreous humor of normal and diabetic individuals.
Western ligand blotting and immunoprecipitation of normal synovial
fluid revealed the presence of IGFBPs 1 through 4, with levels higher
in synovial fluid of patients with rheumatoid arthritis (130 ) compared
with controls. These findings suggest that understanding the normal
IGF/IGFBP axis in physiological states and the alterations that occur
in pathological conditions may provide clues to our understanding of
the pathophysiology of different disease states.
Although the enrichment of certain biological fluids with one or more
IGFBPs, together with the increased expression of the same IGFBP in the
local tissues surrounding the body fluid (Table 2), suggests that these IGFBPs may
function locally to regulate IGF actions, more work is required to
understand the specific role of these binding proteins in modulating
IGF action in various biological fluids.
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V. Assays for Circulating Levels of IGFBP |
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TopAbstractI. IntroductionII. Characteristics of the...III. Target Cell Actions...IV. IGF-IGFBP Complexes in...V. Assays for Circulating...VI. Relative Distribution of...VII. Regulation of Serum...VIII. IGFBP Proteases in...IX. Endocrine Functions of...X. ConclusionsReferences |
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B. Western immunoblotting
Antibodies specific for a IGFBP can be used to quantitate IGFBPs
in a conventional Western immunoblotting, after size separation by
SDS-PAGE. Western immunoblotting can be improved by optimizing protein
transfer, antibody binding, and detection systems (e.g.,
chemiluminescence). In general, immunoblot analysis using polyclonal
antiserum usually detects both intact and fragmented forms of the
IGFBPs (25 ). For example, when pregnancy sera were analyzed using the
Western ligand blot technique (25 135 ), there was no evidence of
IGFBP-3, while both immunoblot and RIA detected the presence of
fragmented IGFBP-3 that arose from protease activity. In subsequent
studies, Baxter and co-workers (79 ) demonstrated that the proteolyzed
IGFBP-3 fragment from maternal serum can bind IGF with normal affinity
and that the lack of detection by Western ligand blot analysis is an
analytical artifact resulting from using [125I]IGF-I for
binding. Another example of the usefulness of the immunoblot assay was
shown in diabetics. Patients with untreated insulin-dependent diabetes
mellitus (IDDM) showed lower levels of IGFBP-3 compared with healthy
controls (136 137 ). While the Western ligand blot could only detect
the intact fragment, immunoblot assay was able to show a decrease in
intact IGFBP-3 and also an increase in fragmented IGFBP-3 compared with
controls. This finding led to subsequent investigation of proteolytic
activity in these patients. IGFBP-3 protease activity was found to be
higher in the serum of untreated IDDM patients compared with
age-matched controls (137 ). The usefulness of Western immunoblotting in
the identification of IGFBP fragments has fostered studies on
characterization of IGFBP proteases in a variety of biological fluids.
C. RIA
One of the major problems with Western ligand blot and Western
immunoblot analysis was the lack of precision, which was overcome by
the development of RIA (92 138 139 140 141 142 143 144 ). At present RIAs for IGFBP-1, -2,
and -3 are commercially available. Recent success in purifying IGFBPs
from a variety of sources to homogeneity and recombinant expression of
various IGFBPs has led to the development of specific antibodies
suitable for establishment of RIAs for accurate measurement of the
various IGFBPs. These RIAs have already provided important new
information on the physiological and hormonal regulation of the various
IGFBPs. The RIAs for the various IGFBPs do not require an extraction
procedure as in the case of the IGFs since endogenous IGFs do not
interfere with the assay (138 139 140 141 142 143 144 ). Thus, the various biological
fluids can be assayed directly for IGFBPs. The majority of IGFBP RIAs
thus far developed utilize polyclonal antiserum, which reacts with both
intact and fragment forms of IGFBPs. Although measurement of both
intact and fragmented forms of IGFBPs may provide useful information in
various clinical settings, one of the disadvantages with the use of
polyclonal antisera is that the antisera developed in various
laboratories may recognize different fragments. This could lead to
inconsistent quantitative results using antisera that recognize
dissimilar epitopes for similar biological samples. For example, it is
known that serum from children with end-stage renal disease contains
increased amounts of fragmented forms of various IGFBPs (145 146 ). The
quantitative measurements of various IGFBPs in serum from children with
end-stage renal disease may depend on whether a particular antiserum
used for measurement of a given IGFBP recognizes only selected
fragments or all of the forms of that particular IGFBP.
D. Immunoradiometric assay
The immunoradiometric assay (IRMA) is a noncompetitive assay in
which the IGFBP to be measured is "sandwiched" between two
antibodies. The first antibody, which needs to be specific, is
immobilized to the inside wall of the tubes. The second antibody is
used as a capture antibody (radiolabeled or enzyme conjugated). Since
the two antibodies used for IRMA are typically developed against
amino-terminal and carboxy-terminal ends of the molecule, the advantage
of this assay is that it is often more specific than RIA (147 148 ) and
more likely to measure the intact molecule. The disadvantage with IRMA
is that it may not reflect production rate as well as RIA since the
analyte measured may be degraded during storage or during experimental
conditions, resulting in artifactually lower values than that actually
present. At present, IRMA is commercially available only for IGFBP-3.
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VI. Relative Distribution of IGFBPs in Serum |
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TopAbstractI. IntroductionII. Characteristics of the...III. Target Cell Actions...IV. IGF-IGFBP Complexes in...V. Assays for Circulating...VI. Relative Distribution of...VII. Regulation of Serum...VIII. IGFBP Proteases in...IX. Endocrine Functions of...X. ConclusionsReferences |
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). IGFBP-3 is the
predominant form present in serum with levels more than 10-fold
higher than the other IGFBPs (59 149 ). The concentration of the small
molecular mass binding proteins are found in increasing order
(IGFBP-4 > IGFBP-5 > IGFBP-2 > IGFBP-6 >
IGFBP-1) in human serum (92 138 139 140 141 142 143 144 149 150 ). There is a 50% molar
excess of IGFBPs over IGFs in serum, which implies that a very small
percentage of the IGFs remain in the free form. Since the antisera used
for measurement of various IGFBPs recognize both intact and fragment
forms of IGFBPs, the relative abundance of intact forms of various
IGFBPs in adult human serum is not known at this time.
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) between IGFs and various IGFBPs reveal
that only IGFBP-5, in addition to IGFBP-3, showed positive correlation
with IGF concentration in normal adult human serum (142 ). In contrast,
serum IGFBP-1 and IGFBP-2 levels showed negative correlation while
IGFBP-4 levels did not correlate with IGF concentration. These data
suggest that different mechanisms may regulate the amounts of various
IGFBPs in serum.
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Regarding the MCR, it is known that IGFBPs play a major role in extending the half-life of IGFs in the circulation (see Section IX). In this regard, the half-life of IGF-II in serum may be longer than that of IGF-I since human serum contains IGFBPs with selective affinity for IGF-II over IGF-I (1 3 ). For example, human IGFBP-6 has 50- to 100-fold higher affinity for IGF-II over IGF-I. IGFBP-2 and IGFBP-5 have slightly higher affinity for IGF-II than IGF-I. In addition, Lee and Rechler (76 ) showed that the 150- to 200-kDa protein complexes in the rat serum have higher affinity for IGF-II than IGF-I. They propose that these 150- to 200-kDa complexes in the adult rat serum contain proteolytically nicked IGFBP-3 and ALS that bind to IGF-II preferentially. Based on these data, it is speculated that the presence of IGFBPs with higher affinity for IGF-II over IGF-I could contribute to a greater half-life of IGF-II over IGF-I. This explains some of the observed differences in the greater abundance of IGF-II vs. IGF-I in human serum. However, it appears likely that the differences in IGF-binding affinity of IGFBPs is not the only mechanism that contributes to the greater abundance of IGF-II over IGF-I, since 75% of IGF-II is bound to IGFBP-3 in the form of ternary complex and the intact IGFBP-3 binds IGF-I and IGF-II with similar affinity (3 ).
Indeed, inasmuch as the differences in production rate and IGFBP affinities to IGFs cannot account for the observed differences in the serum levels of IGF-I and IGF-II, it would seem that some aspect of metabolic clearance, such as the degradation rate, is higher for IGF-I than for IGF-II, thereby contributing to the lower level of serum IGF-I compared with IGF-II in adults. Based on the above analysis, it seems reasonable to conclude that three mechanisms may contribute to the greater serum level of IGF-II than IGF-I in humans: 1) greater production rate of IGF-II than IGF-I; 2) the preferential binding of minor IGFBPs for IGF-II as compared with IGF-I; and 3) a lower degradation rate of IGF-II than IGF-I (the latter two mechanisms would lead to a greater MCR for IGF-I than IGF-II).
If we assume that the actions of IGF-I and IGF-II are similar (i.e., both act via the Type I IGF receptor), then the structural differences between IGF-I and IGF-II would serve some other mechanism than functional activity. This raises the possibility that the differential structure of the two IGFs could lead to differences in MCR. Accordingly, we can speculate that serum contains two pools of reserve IGFs — a smaller IGF-I pool, which is rapidly turning over, and a larger IGF-II pool, which is slowly turning over. If so, the differential structure of the IGFs may produce differential three-dimensional structures with the IGFBPs and, therefore, could lead to a lower proteolysis rate of IGF-II than IGF-I. In this regard, recent studies have shown that exogenous addition of IGF-II to cell-free conditioned medium derived from a number of cell types, including human osteoblasts and fibroblasts, increases the rate of IGFBP-4 proteolysis (157 158 159 160 ). Since IGFBP-4 proteolysis is not induced by the addition of insulin, des(1 2 3 )IGF-I, or des(1 2 3 4 5 6 )IGF-II, all of which bind IGFBP-4 with extremely low affinity, it is speculated that the binding of IGF-II to IGFBP-4 may alter the conformation of the protein and enhance the susceptibility of IGFBP-4 to proteolytic degradation (159 ). Although these data are consistent with the possibility that the binding of ligand to binding protein may result in altered proteolysis of the ligand and/or the binding protein due to conformation changes, further studies are needed to establish whether or not there are, in fact, different MCRs for the serum IGF-I pool and the serum IGF-II pool. If this proved to be the case, this would open the possibility that the two reserve serum IGF pools provide a metabolic advantage to maintain overall body economy in the face of dramatic changes in functional demands, such as during growth, pregnancy, and starvation. Regardless of whether or not this concept has merit, the findings that IGF-II circulates in greater abundance than IGF-I in human serum, and that IGF-II is produced by several adult tissues in large amounts, are consistent with an important role for IGF-II in human physiology.
The finding that IGFBP-3 is the most abundant IGFBP present in adult human serum does not necessarily mean that the production rate of IGFBP-3 is more than that of other IGFBPs. In this regard, the higher abundance of IGFBP-3 in serum may be due to the fact that the half-life of IGFBP-3 is considerably longer (15–20 h) since it is bound to the 80- to 85-kDa ALS. In contrast, the half-lives of IGFBP-1 and IGFBP-2 have been estimated to be on the order of 1–2 h (161 ), which suggests that these binding proteins must be produced at a higher rate than that of IGFBP-3 to achieve similar serum levels based on the differences in their half-lives. Thus, it is essential to understand not only the regulation of IGFBP-3 and other IGFBPs in serum, but it is also necessary to know the regulation of different IGFBPs in various extracellular body fluids since the relative levels of these IGFBPs and their corresponding proteases in local body fluids may play a role in regulating the local actions of IGFs depending on the needs of local tissues.
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VII. Regulation of Serum IGFBPs |
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TopAbstractI. IntroductionII. Characteristics of the...III. Target Cell Actions...IV. IGF-IGFBP Complexes in...V. Assays for Circulating...VI. Relative Distribution of...VII. Regulation of Serum...VIII. IGFBP Proteases in...IX. Endocrine Functions of...X. ConclusionsReferences |
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A. Physiological conditions
1. Diurnal variation. Plasma IGFBP-1 values are subject to
diurnal variation with the levels reaching the lowest during the
afternoon and midnight, and highest in the morning (61 ). In
circulation, IGFBP-1 has a free IGF-binding site, suggesting that it is
unsaturated in contrast to GH-dependent IGFBP-3, which is normally
saturated. It appears that the increase in IGFBP-1 during the morning
hours coincides with an increase in IGF-I level, thus reducing
insulin-like activity (20 ). However, this is independent of both IGF
and GH (61 ). In contrast to IGFBP-1, IGFBP-2 and -3 are more stable and
do not exhibit diurnal variation nor are they subject to postprandial
changes (143 144 ). Diurnal variations have not yet been studied for
IGFBPs 4–6.
2. Nutrition. Nutritional regulation of IGFBP-1, -2, and -3 has been discussed briefly in a recent review by Thissen et al. (162 ), but little is known regarding the nutritional regulation of IGFBP-4, -5, and -6 (163 164 ). The metabolic state of an individual is reflected by insulin level, which influences the circulating concentration of IGFBP-1. Insulin-dependent diabetic patients have higher serum IGFBP-1 levels than nondiabetic controls (136 165 ). Further, acute steady state hyperinsulinemia reduces the serum IGFBP-1 concentration to values that are 40–70% lower than baseline values in normal individuals and also in diabetic and insulinoma patients (166 ), suggesting that insulin is involved in the regulation of serum IGFBP-1 levels (167 ). Serum IGFBP-1 levels fluctuate acutely in response to dietary food intake, with a marked increase (3- to 4-fold higher than baseline) after an overnight fast (168 ) or long-term dietary restriction (169 ), and a decline immediately after a meal. This decline in serum IGFBP-1 may be attributed to the direct effect of insulin or insulin-induced changes in glucose transport.
The effect of calorie and protein restriction on the concentrations of the serum IGFBPs is different for adults and children. A 50% calorie reduction for 6 days increased IGFBP-1 levels in healthy adults but not in children. Levels returned to normal after refeeding (170 171 ). The differences in these responses were not due to differences in insulin secretion, since both adults and children had a significant decline in fasting C peptide levels. Although insulin is the major regulator of IGFBP-1 concentration (165 172 173 ), this study showed that IGFBP-1 changes in children may not be linked to changes in insulin secretion.
Long-term dietary deprivation decreases plasma IGF-I and IGFBP-1 and may modify the tissue response to IGF by increasing IGF receptor synthesis (174 ). Another interesting finding is the role of glucagon as a stimulator of plasma IGFBP-1 independent of insulin levels (175 ). This is evident in healthy subjects, patients with GHD, and IDDM patients who have increased levels of IGFBP-1 when glucagon is administered in spite of an increase in plasma glucose and insulin levels. Based on these data, it is speculated that the nutritional regulation of serum IGFBP-1 level is complex and may be dependent on changes in the level of hormones such as insulin and glucagon, in addition to metabolic changes.
The serum IGFBP-2 level is more stable than IGFBP-1 level and is not influenced by postprandial changes (141 ). However, serum IGFBP-2 increased markedly in both adults and children on protein restriction (170 ). Similar observations have been made in patients with anorexia nervosa (171 ), chronic protein-calorie malnutrition (170 ), and in prolonged fasting that lasted more than 1 week (172 ). This increase in serum IGFBP-2 follows a cellular increase in the expression of the IGFBP-2 mRNA in rat liver (174 176 ). Although protein refeeding normalized the serum IGFBP-2 levels of undernourished children, high-protein intake is required to achieve complete normalization (177 ). Thus, nutrition-induced changes in serum IGFBP-2 level appear to be the direct effect of dietary protein on IGFBP-2 expression in liver.
Serum IGFBP-3 levels declined slightly but significantly with calorie restriction in both children and adults (170 ), but protein restriction caused a decrease in IGFBP-3 only in adults. However, this was normalized after protein refeeding (177 ). Although serum IGFBP-3 levels are regulated by IGF-I and GH under normal conditions (140 ), the decrease seen in undernourished children is more likely due to the presence of IGFBP-3-specific protease levels (177 ). The presence of similar proteolytic activity accompanying a low IGFBP-3 level is seen in other catabolic states (135 178 ), pregnancy (135 179 ), and in postsurgical patients (180 ). Age appears to influence changes observed in IGFBPs as a result of dietary modification. The response to calorie restriction among children and adults differs more so than during protein restriction, which may be due, in part, to an overestimation of the energy requirements for children (169 ).
Weight loss or long-term moderate energy restriction does not alter IGFBP-3 (181 ). Serum IGFBP-3 was not influenced by a very low calorie diet (VLCD) consumed by normal and obese subjects, while IGFBP-1 increased markedly in controls on VLCD and not in obese subjects (182 ). The increase in IGFBP-1 is suggested to inhibit the IGF-I feedback regulation of GH secretion, while a similar response is absent in obese individuals. Thus, GH secretion is increased in normal subjects on VLCD, but this response is abolished in obese individuals. The impaired GH secretion in obese subjects resulted in a lowered IGF/IGFBP-3 molar ratio, but this was reversed after weight loss by these subjects (183 ). Thus, changes in serum levels of IGFBPs induced by VLCD appear to be different between normal and obese subjects.
Based on the observations that serum levels of IGFBPs change depending on the nutritional status, two general conclusions can be made: 1) serum IGFBP-1 and IGFBP-2 levels are regulated differently than IGFBP-3 by nutrition, and 2) the decrease in serum IGFBP-3 with corresponding increases in serum IGFBP-1 and IGFBP-2 levels during malnutrition would decrease the half-lives of IGFs but tend to increase the transport of IGFs across the vascular endothelium and thereby could modulate the bioavailability of IGFs to target tissues (see Section IX). Further studies are needed to determine whether nutrition regulates IGF action by altering the ratio of IGFs bound to the 150- to 200-kDa and 50-kDa complexes.
3. Exercise. Exercise increased IGF-I and IGF-II in human adults (184 185 ) with the degree of response influenced by the intensity of exercise (186 ). This increase in IGF level in serum appears to be GH independent, since the increase in serum IGF-I occurred earlier than the increase in serum GH. In addition, GH secretion increased only in high-intensity exercise while IGF increased under both low and high intensities. Circulating IGF-I levels are also influenced differently by different types of exercise. Weight-bearing exercise caused no change in the IGF system (186 187 ), while endurance-type exercise induced significant increases in serum IGF-I level (185 ). Exercise is also accompanied by changes in some of the IGFBPs (increase in IGFBP-1 and IGFBP-3) with or without changes in IGFs, with an overall change in the IGF to IGFBP ratio.
Prolonged exercise increased the need for plasma glucose because of depleted muscle glycogen or increased hepatic glucose output. Prolonged exercise increased serum IGFBP-1 (188 189 ), and this was inversely related to serum insulin and IGF-I levels (188 ). This raised the possibility that serum insulin was the main regulator of IGFBP-1 in circulation during exercise. However, another study (189 ) showed that serum IGFBP-1 increased in response to prolonged exercise even when normal plasma glucose and insulin levels were maintained, suggesting that factors other than insulin levels, e.g., muscle glycogen, may be involved in the regulation of serum IGFBP-1 during exercise.
Increased serum IGFBP-3 levels have been reported in adults after exercise (184 190 ). This increase paralleled an increase in IGFBP-3 proteolytic activity. It is speculated that the proteolysis was induced by activation of calcium-dependent protease (191 ) and might be responsible for the IGF-induced anabolic effect of exercise on muscle tissue. These changes in the IGF system components are observed immediately after exercise and are of short duration (<1 h). It is not known at this time whether the alterations in the IGF system components in circulation play a role in mediating the anabolic effects of physical activity or whether the exercise-associated changes in circulating levels of IGFs and IGFBPs reflect processes that occur in the exercising tissue itself (191 ).
4. Glucocorticoid. Glucocorticoids inhibit somatic growth in humans in part by suppressing GH secretion and IGF activity. When dexamethasone was administered to healthy male volunteers, it suppressed IGFBP-1 and IGFBP-2 levels while increasing IGF-I and IGFBP-3 levels (192 193 ). The mean IGF bioactivity was reduced by 60% over the sampling period (192 ). This decrease in bioactivity could be due to the induction of serum inhibitors, alteration in IGFBP activity, and/or alteration in secretory profiles of GH. Recently it was shown that the negative effects exerted by glucocorticoid on bone formation may be mediated, in part, via changes in endocrine and local action of IGFs (194 ). In this study, the reduction in bone formation after glucocorticoid therapy of chronic obstructive pulmonary disease patients was accompanied by a decrease in stimulatory IGF system components including IGFBP-3. Whether or not other stimulatory and inhibitory IGFBPs are also affected remains to be determined.
B. Development and aging
1. Fetal and neonatal development. It is possible that IGFBPs
play a major role in regulating the mitogenic and
differentiation-promoting effects of IGFs in fetal tissues. IGFBPs 1 to
6 are expressed in the different organ systems of the developing fetus
(12–16 weeks) as shown by Northern blot analyses (195 196 ). Of the
IGFBPs seen in the serum of a human fetus, IGFBPs 1, -2, and -3
originate predominantly from the liver, while only small amounts of
IGFBPs 4, -5, and -6 are expressed in the liver. IGFBP-5 mRNA was
detected in several cell types during early postimplantation stages of
the developing rat, suggesting that IGFBP-5 has a role in the
development of different organ systems (197 ). These binding proteins
either cause inhibitory or stimulatory effects on IGF action, depending
on the amount of IGFs bound to each of the IGFBPs and the pattern of
distribution of these binding proteins in the various fetal tissues.
In order for optimal fetal growth, a constant interaction between the maternal host and the developing embryo/fetus is required. The presence of IGFs, IGFBPs, and fragments of IGFBP-3 in human extraembryonic cavities provide support for maternal-fetal exchange of IGF system components (115 ). It is suggested that the altered affinities of the proteolyzed IGFBP-3 for IGF-II in extraembryonic cavities may play a role in regulating the bioavailability of IGF-II in the chorion and/or the amnion (115 ). However, the role of IGFBP-3 protease in modulating IGF bioavailability remains controversial.
There is also a developmental switch during transition from fetal to neonatal life in the IGFBPs present in circulation. In a fetus of less than 27 weeks of gestation, serum contains IGFBP-1, while cord serum contains mainly IGFBP-3 (198 ). The level of IGFBP-1 is higher in the fetal and cord blood than in adult plasma. In normal weight fetuses, the IGFBP-3 and IGFBP-1 concentration in serum is 15% and 50% of maternal serum levels, respectively (199 ). Low cord serum IGFBP-1 and elevated IGFBP-3 concentrations were reported in large-for-gestational age fetuses at term birth (200 ). Since these infants also had elevated cord serum insulin levels, it was suggested that the changes in IGFBP-1 were mediated by insulin, mainly by directing greater delivery of the IGF/IGFBP complex to the target tissue, resulting in the accelerated growth as seen in large-for-gestational age fetuses. IGFBP-3 levels increase significantly during the last trimester of intrauterine life. This is supported by a study (201 ) that showed an increase in serum IGFBP-3 in preterm infants from birth (3 months preterm) to 2 months past appropriate term age. Thus IGFBP-1 and IGFBP-2 are the predominant binding proteins during fetal life, but they decline during the early neonatal period, with IGFBP-3 becoming the predominant binding protein.
In intrauterine growth-retarded (IUGR) fetuses, there is a marked elevation in cord serum IGFBP-1 and -2 compared with normal fetuses (199 200 202 ). Giudice et al. (200 ) showed that serum IGFBP-3 levels were decreased in IUGR fetuses, but this is in contrast to the results of Lassarre et al. (203 ), who showed that fetal cord serum had higher IGFBP-3 than normal cord serum. Typically, serum IGFBP-3 declines in cord serum due to a specific protease that degrades this protein to increase the amount of IGFs available for stimulation of growth in the target tissues. Thus, in fetal circulation, the increased availability of IGF is due to a molar excess of IGF-I and -II, an increase in IGFBP-2, and a decrease in the ternary IGFBP-3 complex formation. It is therefore speculated that IGFBP-3 protease is less likely to play a significant role in fetal serum in contrast to maternal or neonatal serum (204 ).
During fetal life, not only is the total plasma IGFBP-3 lower than in the adult circulation, but the amounts of IGF-I and IGF-II bound to this binding protein are also low compared with amounts of IGFs bound to IGFBP-3 in adults. IGF-I levels are depressed in infants with IUGR, suggesting that IGFs play a significant role in promoting growth. However, the majority of IGFs are bound to IGFBP-3 as a 50-kDa ALS-independent complex in infant serum, which is capable of crossing the endothelial barrier, thus increasing the bioavailability of the IGFs (203 ). The role of IGFBP-4 in neonatal development has not yet been explored, but given the inhibitory effect of IGFBP-4 on IGF action, it is possible that serum IGFBP-4 levels are higher in children with slow growth compared with normally growing children (205 ). However, future studies need to confirm these speculations.
2. Puberty. Although low at birth, the serum IGFBP-3
concentration rapidly increases during the first years of infancy
(206 207 208 ), reaches a peak at puberty (207 ), and declines during
adulthood (208 ). Girls have higher serum IGFBP-3 levels than boys of
comparable age throughout childhood, and levels peak a year earlier
than boys during puberty (207 208 ). In addition, IGFBP-3 increases
with the increasing stage of pubertal maturation. Both the height and
body mass index of an individual correlates positively with IGFBP-3
levels independent of age, sex, and pubertal stage. The molar ratio
between IGF-I and IGFBP-3 is increased during puberty, suggesting that
more biologically active IGF is available in the free form during the
pubertal growth spurt. Serum levels of IGFBP-2 show marked
age-dependence with high levels at birth and senescence and low levels
during puberty (144 ). Serum IGFBP-1 declines progressively with age,
with the lowest concentration observed during puberty (207 208 ). The
height variability seen among pubertal children correlates with the
concentration of IGF-I and IGFBP-3, with lower values in short stature
children and higher values in tall children (209 ). Also, in patients
with acromegaly and those with high serum GH levels, circulating
IGFBP-3 and IGF-I are increased. Under normal physiological conditions,
most of the serum IGFs are bound to IGFBP-3, with an approximate 1:1
molar ratio of total IGF (IGF-I + IGF-II) and IGFBP-3 (60 63 ).
However, this ratio seems to vary with developmental age, with a
greater increase in circulating IGF-I than IGFBP-3 during puberty
(208 ). Changes in serum concentrations of the various IGFBPs during
puberty (Table 4) have been proposed to
play a role in inducing the growth spurt during puberty.
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3. Pregnancy. Several changes in the IGF system occur in maternal and fetal circulation during pregnancy, suggesting an important role for the IGF system components in producing the anabolic effects on maternal and fetal tissues. Both serum IGFBP-1 and -2 are found in the maternal serum during the initial weeks of gestation. However, serum IGFBP-2 levels are lower toward the latter part of gestation due to the presence of IGFBP-2-specific protease (179 ). Serum IGFBP-1 levels are higher during early gestation and plateaus from week 12 through the end of gestation (118 ).
Serum IGFBP-3 is the major binding protein for the circulating IGFs (211 212 ) in pregnant and nonpregnant women. Binoux et al. (213 ), using competitive binding studies, showed that IGFBP-3 derived from maternal serum had 10 times lower affinity for IGF-I than IGFBP-3 derived from normal human serum and half the affinity for IGF-II. An apparent decline in the acid-stable IGF-binding subunit of IGFBP-3 in the latter part of gestation and the presence of smaller molecular forms of IGFBP-3 in term maternal serum provide strong evidence that specific proteases that cleave the IGF-IGFBP-3 complex (78 135 179 ) are present. The protease detected in the serum of pregnant women is suggested to cause a functional change (78 214 ) resulting in a redistribution of IGF-I. This would increase the proportion of free IGF-I (215 ) at the expense of IGF-I bound to the 150 to 200 kDa complex. Both the intact and fragmented forms of IGFBP-3 have greater affinity for IGF-II than IGF-I (3 ), a role that could be related to the established importance of IGF-II in fetal development.
On the other hand, Baxter and co-workers (67 216 217 ) demonstrated
that IGFBP-3 derived from human maternal serum is not functionally
altered. Baxter et al. (67 ) reported that this proteolyzed
IGFBP-3 circulates in a ternary complex that is normal in molecular
mass and IGF-carrying capacity. IGFBP-3 derived from maternal serum
appears similar in size to the IGFBP-3 from serum of nonpregnant women
dissociated from ternary complex by acid treatment with normal binding
affinity for ALS (217 ). Ternary complex formation measured by
radiolabeled -subunit binding was identical in serum from pregnant
and nonpregnant women after acid treatment of serum that destroys
endogenous -subunit. Based on these data, Baxter and co-workers (79 )
concluded that IGFBP-3 in native maternal serum is functionally normal.
Thus, to date, the functional significance of IGFBP-3 protease in
maternal serum remains controversial.
IGF biological activity is modified in situations in which fetal growth is abnormal, such as in multiple or abnormal pregnancies. Maternal IGFBP-3 increases with gestational age and is higher in multiple pregnancy compared with gestational age-matched twin or single pregnancies (218 ). IGFBP-3 protease activity also increases with increasing fetal number, but is not reversed by embryo reduction. This implies that the protease activity is activated early on in pregnancy and remains irreversible. A similar increase in IGFBP-3 protease activity has been observed in both appropriate-for and small-for-gestational age fetuses with utero placental insufficiency. Based on this evidence, the concept has been advanced that in both multiple pregnancy and under utero placental insufficiency, the increase in protease activity may be an adaptive mechanism to counteract the threat to fetal growth (218 ).
4. Aging. An age-related decline in serum IGF-I level has been reported by several investigators (219 220 221 222 223 ). The decrease in serum IGF-I with advancing age could be explained in part by the decrease in serum GH levels (221 ). Recent studies also demonstrated that serum IGF-II levels declined with age but to a lesser extent compared with IGF-I (222 ). In addition to the IGFs, the serum IGFBP levels were also altered with age. Both the magnitude and direction of change with age were different for the various IGFBPs. Serum levels of IGFBP-1, IGFBP-2, and IGFBP-4 increased with age while those of IGFBP-3 and IGFBP-5 declined with age (138 139 140 141 142 143 144 190 ). Serum IGFBP-1 showed an age-related increase (138 224 ) in both men and women. Fasting IGFBP-1 levels increased with aging (224 ), but there was no correlation between insulin and IGFBP-1 with advancing age. Although insulin correlates inversely with IGFBP-1 (165 ), this relationship ceases to exist in aged individuals. This might be due to the development of age-related insulin resistance among these subjects (225 ). Replacing adrenal steroids suppressed the IGFBP-1 levels while increasing the IGF-I levels and restoring normal mitogenic activities in serum of the elderly (226 ). The age-dependent pattern of serum IGFBP-2 resembles that of IGFBP-1 in that it increased with aging (143 144 ). After puberty there is a steady increase in IGFBP-2 in circulation, and this increase is accelerated at age 60 and above. However, the mechanism that causes the increase in IGFBP-2 is unknown at present.
Serum IGFBP-3 declines with age during adulthood, and this corresponds to an age-related decline in GH secretion (140 150 ). In contrast, serum IGFBP-4 levels increase with age (139 222 ), indicating that different binding proteins are differentially regulated. The increase in serum IGFBP-4 in elderly women was in part due to the age-related increase in PTH, the production of which is stimulated in calcium deficiency states (227 ). A 30% reduction in the serum IGFBP-5 levels in women aged 23–85 yr was observed when compared with IGFBP-5 in the serum of prepubertal girls (142 ). Since serum IGFBP-5 levels show significant positive correlation with age and the treatment of various cell types including osteoblasts with IGFs increased production of IGFBP-5, it is suggested that the decline in IGF production may contribute to the age-related decrease in IGFBP-5 production. Consistent with this interpretation, Nicholas et al. (49 ) reported that the decline in skeletal content of IGFBP-5 showed significant positive correlation with skeletal content of both IGF-I and IGF-II. Whether the age-related decline in skeletal and serum IGFBP-5 reflects an age-related decline in synthesis of this protein by bone and other tissues needs further investigation.
The data in Table 4
demonstrate that multiple deficits in the IGF
system components occur as a consequence of aging. The underproduction
of stimulatory IGF system components and the overproduction of
inhibitory IGF system components could lead to an age-related decrease
in the hormonal as well as local actions of the IGFs, all of which
could contribute to an impairment in the function of various organs
during aging. Further studies are needed to establish the cause and
effect relationship between changes in IGF system components and
impairment in various body functions with age.
C. Hormonal effects: mechanisms
If IGFBPs play an important role in regulating IGF actions, then
the regulation of IGFBPs in various physiological situations would
depend on the hormones involved and the target tissue. Thus, the
changes in serum levels of IGFBPs during normal physiological
conditions such as malnutrition, puberty, pregnancy, and aging may be
influenced in part by corresponding changes in the levels of various
hormones including insulin, GH, sex steroid hormones, glucocorticoids,
and thyroid hormone.
The extent to which changes in the level of a given hormone influences the serum level of a given IGFBP depends on two main factors: 1) the magnitude of change in the production of the corresponding IGFBP in the target tissue(s) and 2) the extent to which the target tissue(s) contribute to the circulating level of the IGFBP in serum. It is thus possible that the changes in the production of one or more IGFBPs in the target tissues in response to hormonal stimuli may be reflected by corresponding changes in serum levels of these IGFBPs. Alternatively, the levels of one or more IGFBPs may change in the local body fluid in response to a hormonal stimuli, and these local changes may not be reflected by changes in corresponding levels of the IGFBPs in serum.
Studies on serum regulation of IGFBPs in response to hormonal stimuli suggest that some hormones may primarily influence the serum level of one IGFBP while other hormones may influence the serum levels of multiple IGFBPs. For example, the level of insulin changes in response to alterations in nutritional status. Accordingly, the serum level of IGFBP-1 changes in response to alterations in insulin level (59 ). The finding that insulin is the major regulator of IGFBP-1 production in liver, one of the target organs for insulin action, suggests a role for insulin in regulating IGFBP-1 function depending on the nutritional status. In contrast to the specific effect of insulin on IGFBP-1, GH appears to influence the serum level of a number of IGFBPs (59 228 ). Since GH mediates somatic growth in a variety of organs and some IGFBPs are more abundantly expressed in certain tissues than in others, it is not surprising that GH influences multiple IGFBPs.
Studies on the mechanisms by which various hormones influence the serum level of known IGFBPs provide evidence for both direct effect of a given hormone on the IGFBP gene in the target tissues and an indirect effect of the hormone secondary to changes in the production of local growth factors. For example, the effect of GH to increase the serum level of IGFBP-3 is thought to be mediated via the following two mechanisms: 1) Based on the findings that GH treatment increases IGFBP-3 expression by a pathway independent of IGFs in liver cells in vitro (229 ), it is proposed that GH may mediate its effects by directly influencing the IGFBP-3 gene in the target tissues; 2) Based on the findings that many of the effects of GH on target tissues are mediated indirectly via IGFs and that changes in the serum levels of IGFBP-3 correlate significantly with changes in serum level of IGFs, it is proposed that GH effects on IGFBP-3 may be secondary to an increase in the production of IGFs (59 228 230 231 ). In addition to these two mechanisms, GH induces ALS production, thus contributing to the prolonged half-life of IGFBP-3 (66 ). The extent to which each of these mechanisms contributes to the GH-induced increase in serum level of IGFBP-3 is not known at this time.
Similar to GH effects, a number of other hormones, including sex steroid hormones and glucocorticoids, may also influence serum level of IGFBPs by both direct and indirect mechanisms. In this regard, studies on regulation of production of IGFBPs using serum-free monolayer cell cultures have provided evidence for the direct effect of estradiol, testosterone, progesterone, and dexamethasone on the expression of various IGFBPs (153 232 ). In addition, it is also known that these hormones influence actions of a variety of locally produced growth factors including IGFs (153 159 232 ). Because many of these locally produced growth factors also influence production of IGFBPs in a variety of target cell types, the changes in the serum level of IGFBPs may be a reflection of not only the direct effect of the given hormone on IGFBP production in the target tissues but also the indirect effect of a given hormone on local growth factor milieu.
Studies on the molecular mechanisms by which hormones influence expression of IGFBPs provide evidence for complex regulation involving both transcriptional and posttranscriptional mechanisms (159 ). Some hormones may regulate IGFBP level by primarily regulating one control mechanism while other hormones may regulate IGFBP level by regulating multiple control mechanisms. In this regard, it is known that the insulin effect on IGFBP-1 production is regulated primarily at the transcriptional level in liver cells (233 ). In contrast, the IGFBP-3 levels in the conditioned medium of various cell types in response to various effectors are regulated by mechanisms involving alterations in both synthesis and degradation (191 ). In addition, there is evidence of the release of cell surface-bound IGFBP-3 by certain growth factors (234 ). Similarly, IGFs and bone morphogenetic proteins influence IGFBP-5 levels in human osteoblasts by both decreased proteolysis and increased gene transcription (235 ). Thus, some hormones appear to modulate the IGFBP level by regulating one control mechanism (e.g., proteolysis) while other hormones appear to modulate IGFBP levels by regulating multiple control mechanisms (e.g., gene transcription, mRNA processing, mRNA stability, proteolysis, release of cell surface-bound IGFBP). It is speculated that the complexity of IGFBP regulation may provide the required flexibility for modulating IGF actions by a multitude of systemic and local effectors in various tissues.
D. Pathological conditions
The following section describes the changes in the IGFBP axis in
serum during various pathological states and how these changes could be
interpreted to explain the corresponding disease states.
1. Diabetes. Serum IGFBP-1 is the only component of the IGF system that is directly regulated by insulin. Several studies (229 236 237 238 ) have shown that in adult and adolescent IDDM patients there is a significant increase in serum IGFBP-1 level. In normal subjects, serum IGFBP-1 levels progressively declined throughout childhood and puberty, but this age dependency did not hold true in diabetic individuals (239 ). Although levels of serum IGFBP-1 in IDDM patients was 7 times higher than in healthy controls before initiation of insulin treatment (240 ), the levels were normalized immediately after insulin infusion. Glucagon is known to be a stimulator of IGFBP-1 production in IDDM patients and is independent of insulin action (175 ). The serum IGFBP-1 in patients with IDDM and severe ketoacidosis was in the phosphorylated form (241 ). This form has 6-fold higher affinity for IGF-I than the nonphosphorylated form in vitro (242 ). Based on these findings, it is speculated that the increase in phosphorylated IGFBP-1 leads to a decrease in free IGF available for stimulating growth in IDDM patients, which may contribute in part to the low lean body mass observed in these patients (241 242 ).
In contrast, in non-insulin-dependent diabetes mellitus (NIDDM) that results in hyperinsulinemia and insulin resistance, conflicting data (165 173 ) have been obtained in regard to serum IGFBP-1 levels (i.e., both increases and decreases in serum IGFBP-1 levels). This discrepancy could be due to the type of treatment provided to the patients and how this in turn affects ß-cell function and insulin resistance. For example, sulfonylurea increases endogenous insulin secretion and thereby decreased IGFBP-1 secretion, while multiple insulin injections inhibit endogenous insulin secretion, resulting in an increase in serum IGFBP-1 (243 ). An increase in fasting IGFBP-1 and -2 levels and a decrease in IGFBP-3 levels were observed in normal and NIDDM patients after IGF-I and insulin infusion (244 ). These changes in the levels of various IGFBPs are likely to alter distribution of IGFs among these binding proteins which, in turn, could modify the bioavailability of IGFs in the target tissues depending on the tissue-specific proteolysis of IGFBPs (see Section VIII).
Serum IGFBP-2 is also increased in diabetes; however, insulin therapy normalized the levels (239 ). This is surprising since it is known that free IGF-I and not insulin is the major regulator of IGFBP-2. Although serum IGFBP-3 is not involved in glucose metabolism, a 30–40% reduction was reported in diabetic patients (237 ) before starting insulin treatment. After treatment, there was an increase in both serum IGF-I and IGFBP-3, with the former showing a slightly greater increase than the latter. This makes more IGF bioavailable and possibly accounts for the improved linear growth seen in children with IDDM receiving insulin treatment (240 ).
In several pathological states, IGFBP-specific protease(s) is increased in the serum, which may lead to an increase in the amount of free IGF-I available for growth stimulation. Intact IGFBP-3 measured by ligand blot analysis of children with untreated IDDM was 50% lower than that of age-matched controls (243 ) while it was 70% lower when measured by immunoblot assay that detected both intact and fragmented IGFBP-3. IGF-I, a known regulator of serum IGFBP-3, was reduced in IDDM patients (245 ) and may be responsible for the decline in IGFBP-3 levels. After insulin treatment, IGFBP-3 levels increased significantly, with an associated decline in serum IGFBP-3 protease (137 ). It is postulated that an increase in IGFBP-3 protease activity during insulin deficiency, as seen in IDDM, helps to partly overcome the catabolic state by increasing the bioavailability of IGF for growth stimulation (137 ).
An increase in protease activity is also observed in gestational diabetes. During the latter part of gestation, there is an alteration in IGFBP-3 levels due to a specific IGFBP-3 protease that increases availability of free IGF-I. Although women with gestational diabetes had a greater increase in free IGF-I compared with nondiabetic pregnant women, this change did not seem to be due to a decrease in serum IGFBP-3 level, but rather to an increase in this protease activity (246 ).
2. Tumors. Extrahepatic non-insulinoma tumor-induced hypoglycemia is accompanied by metabolic abnormalities such as an inhibition of hepatic glucose production, decrease in lipolysis in adipose tissue, and increased consumption of glucose by peripheral tissues (247 ). Although these effects are characteristic of insulin action, plasma insulin levels are low in these patients (248 ). It was evident that the tumors in these patients secreted a bioactive big IGF-II (15–25 kDa) as a result of incomplete processing of pro-IGF-II peptide (249 ). Big IGF-II has an insulin-like action and binds insulin receptor (250 ). The presence of high levels of IGF-II by itself does not address the issue of tumor-related hypoglycemia. However, it was shown that there is a shift in the distribution of IGF-II from the 150- to 200-kDa to the 50-kDa complex in the circulation (251 252 ), since IGFBP-3 and tumor IGF-II failed to complex with ALS (253 ). The increase in 50 kDa complex is mainly due to an increase in the level of IGFBP-2 (251 252 ). Since the 50-kDa IGF complex can cross the vascular endothelial barrier, it seems likely that the bioavailability of IGF-II is increased, resulting in insulin-like actions in target organs.
A proposed mechanism by which big IGF-II secreted by tumors may lead to
hypoglycemia is shown in Fig. 5.
Consistent with this model, Baxter et al. (253 ) showed that
prednisone treatment of a 87-yr-old women with a non-islet cell tumor
suppressed pro-IGF-II formation and corrected the hypoglycemia.
Prednisone treatment caused a redistribution of serum IGFBP-3 from
binary to ternary complex forms. These studies demonstrate the role of
ALS and the ternary ALS+IGFBP-3+IGF complex in glucose regulation.
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b. Endometrial tumors.
IGFs have been shown to stimulate
breast cancer cell proliferation in vitro (257 ). With
respect to breast cancer, tamoxifen therapy suppressed plasma IGF-I
(258 ) and increased serum levels of IGFBP-1 (259 ). This increase in
IGFBP-1 could either result in a decrease in the biologically available
form of IGFs and reduce growth of the tumor or could enhance the
delivery of IGFs to the tissues because of the ability of the
IGF/IGFBP-1 complex to cross the endothelial barrier, making tamoxifen
therapy ineffective in these patients. In women with endometrial
cancer, fasting insulin was found to be higher, and serum IGF-I and
IGFBP-3 levels were lower than in normal controls (260 ). It is not
known at this time whether the changes in the serum levels of various
IGF system components in women with endometrial cancer play a role in
modulating the amount of IGFs available for cancer cell growth.
c. Other tumors.
Several other tumors are also associated with
alterations in IGF and their binding proteins. For example, in one type
of tumor affecting the kidney in children (Wilm’s tumor), IGF-II
levels in serum were lower than in normal controls, but IGFBP-2 was
significantly higher in those with the tumor (261 ). Although it is
known that IGFBP-2 sequesters IGF-II and prevents it from binding
to the IGF receptors, the exact role of IGFBP-2 in tumor development is
not known. Based on the findings that the serum level of IGFBP-2 is
increased in various types of tumors, Zumkeller et al. (261 )
concluded that the serum IGFBP-2 measurements may prove to be useful as
markers for various malignancies.
Elevation of IGFBP-2 in CSF was observed in patients with malignant CNS tumors (96 ). The source of CSF IGFBP-2 was not the serum as there was no correlation between these two measurements. Increased expression of IGFBP-2 mRNA in CNS tumors from one patient suggests the possibility that the tumor itself may be the source of the CSF IGFBP-2. Although it is speculated that the IGFBP-2 in the CSF could be a useful marker for CNS tumors, further studies are required to understand the mechanism that causes the increase in IGFBP-2 and the clinical utility of IGFBP-2 measurement as a diagnostic marker for CNS tumors. Children with highly malignant CNS tumors or CNS leukemia (95 ) have elevated IGFBP-3 in the CSF possibly due to the disruption of the blood-brain barrier and entry of serum IGFBP-3 or to an increase in local production of IGFBP-3 by tumor tissues. Chemotherapy for cranial tumors decreased serum IGF-I and IGFBP-3, although GH levels remained within normal limits (262 ). However, after 12 months, the levels returned to pretreatment values. It is speculated that GH insensitivity seen during chemotherapy might be responsible for the early growth retardation observed in these patients.
3. Laron type dwarfism. Laron type dwarfism (LTD) is an autosomal recessive condition involving a defect in the GH receptor gene. These patients have high circulating levels of GH and low serum IGF-I with impaired receptor function causing growth retardation. In this GH-resistant condition, it appears that the IGFBPs play an important role in modulating the activity of IGFs. Elevated serum IGFBP-1 concentrations were observed in LTD patients, and this may be partly due to low IGF-I levels. After IGF-I administration, the IGFBP-1 levels were down-regulated in LTD patients (263 ). Low levels of serum IGFBP-3 were seen in LTD subjects compared with normal controls, and most of the IGF-II in circulation appeared bound to IGFBP-2 instead of IGFBP-3. Furthermore, an IGFBP-3-specific protease was identified in the serum of LTD patients but not in controls. Based on these data, it is speculated that the decrease in IGF bioactivity in LTD is largely due to variations in IGFBPs and not in IGF-II levels.
Long-term treatment of LTD with IGF-I resulted in a strikingly progressive increase in serum IGFBP-3 levels with continuous treatment (264 265 266 ). This increase in serum IGFBP-3 levels, despite the absence of GH action in LTD, suggests the possibility that IGFBP-3 in humans is regulated directly by IGF-I, independent of GH. An increase in serum levels of other IGFBPs was also detected after IGF therapy. The increase in serum IGFBP-1 levels paralleled the decrease in insulin levels observed with the therapy. Since insulin is the major regulator of IGFBP-1 production, the rise in the latter is most probably due to this observed decrease in insulin (267 ). The serum IGFBP-2 level was also increased after long-term IGF-I therapy of LTD. Similar increases in the serum IGFBP-2 level were also observed in normal human subjects receiving infusions of IGF-I (268 ). The functional significance of the increase in serum levels of various IGFBPs as a result of IGF-I administration in modulating the growth-promoting effects of exogenous IGF-I in LTD is not known at this time.
Two forms of IGF- and ALS-associated IGFBP-3 (44-kDa doublet and 28-kDa band) that are capable of forming the ternary complex have been identified in healthy adult serum (28 ). In GH receptor deficiency (GHRD), the distribution of these two forms differs from that of control serum and explains the alterations in IGF distribution in these patients. Although in adults with GHRD, recombinant IGF-I treatment increases serum IGF-I and decreases serum IGF-II without any change in ALS (269 ), the IGF distribution between the two molecular forms of IGFBP-3 is not corrected (270 ). Thus IGF-I therapy in GHRD patients is expected to have a minimal role in prolonging the half-lives of IGFs since IGF-I therapy in these patients has failed to increase the formation of the 150- to 200-kDa IGF complex. However, a recent study demonstrated the potential beneficial effects of short-term IGF-I therapy in LTD (271 ) although it is not known whether long-term IGF-I treatment would result in continuous improvement in growth velocity in these patients.
4. GHD. In GHD the patients have a low circulating level of GH but normal receptor function. The serum IGFBP-1 levels in GH-deficient patients were higher than in healthy controls (138 272 ), and this could be due to low endogenous insulin production in these subjects (273 274 ). In addition to insulin, the role of other known stimulators of IGFBP-1, such as glucagon and catecholamines, in altering serum IGFBP-1 needs to be explored. As for serum IGFBP-3, both in congenital and idiopathic GHD, the levels were diminished (60 ). Similarly, in patients with hypopituitarism secondary to certain tumors, the low levels of IGFs were accompanied by low serum IGFBP-3.
It is known that the serum levels of IGFBP-3 are dependent on GH and/or IGF-I production. GH and/or IGF effects on IGFBP-3 levels are mediated via an increased synthesis of IGFBP-3 or an alteration in IGFBP-3 breakdown (275 276 277 ). IGF-I can also induce IGFBP-3 level independent of GH activity as shown by an increase in the serum IGFBP-3 level after treatment with recombinant IGF-I (278 ). However, the changes in serum IGFBP-3 level in IGF-I-treated children are much smaller compared with those changes induced by GH treatment, probably due to lack of increase in ALS production during IGF-I treatment (278 279 ). In terms of whether or not changes in IGFBP-3 production play a role in mediating GH-induced growth, it is speculated that the absence or presence of IGFBP-3 plays a role in the growth of pygmy children. It is proposed that the absence of IGFBP-3 in these children accounted for their normal growth during childhood, in spite of the low levels of IGF-I, while during puberty the increase in IGFBP-3 level resulted in the failure of the normal growth spurt (63 ).
In human serum, levels of IGFBP-3 were shown to be low in hypopituitary patients and increased after administration of GH as assessed by immunoanalysis and Western ligand blot analysis. Similarly, circulating levels of human IGF-I and ALS are considered to be directly regulated by GH. Administration of GH in the absence of IGF-I to GH-deficient rats showed that continuous administration of GH, as opposed to twice daily GH injections, is superior in restoring the IGF-IGFBP-3-ALS complex (279 280 ). These data are consistent with the idea that ALS and IGF-I are directly regulated by GH while IGFBP-3 is directly regulated by IGF-I (280 ). Also, the IGF/IGFBP-3 ratio and the induction of all the components of the IGFBP-3 complex were observed with continuous infusion of GH. It is suggested that continuous infusion of GH would up-regulate GH receptors and stimulate production of IGF-I and provide a synergistic effect on growth promotion.
Prepubertal GH-deficient children treated with GH showed an increase in serum IGF-I and IGFBP-2, suggesting that they are the major correlates of pubertal growth (231 ). GH-deficient children treated with GH sustained an increase in circulating levels of IGFBP-5 and IGF-II, suggesting that IGFBP-5 and IGF-II may also play a role in mediating some of the anabolic actions of GH (228 ). However, whether or not this effect is due to a direct influence of GH on IGFBP-5 or due to a GH-induced increase in IGF-I and II on IGFBP-5 (281 282 283 ) is unclear. GHD not only impairs longitudinal growth, but also decreases bone mineral content in humans. GH has been shown to have important effects on skeletal metabolism in vitro and in vivo (123 184 283 284 285 286 ). Thus, in patients with GHD of childhood onset, GH administration is often used to increase the bone mineral content (287 288 ).
In contrast to GH, administration of IGF-I in adults with GHD for 7 consecutive days increased serum levels of IGF-I but decreased serum levels of IGF-II (231 ). Daily injections of IGF-I had no significant effect on serum levels of IGFBP-1, IGFBP-2, and IGFBP-3. In contrast, long-term treatment of 20 GH-deficient patients, aged 22–57 yr, with GH increased serum levels of IGF-I, IGFBP-3, and ALS (289 ). However, the percentage increase in IGF-I was higher than the other two peptides (i.e., IGFBP-3 and ALS) resulting in a higher ratio of IGF-I/ALS and IGF-I/IGFBP-3 after treatment of GH-deficient adults with GH. In addition, the increase in lean body mass during GH treatment showed significant positive correlation with age, thus suggesting that IGF-I can be used as a marker in monitoring changes in lean body mass during GH-replacement therapy.
Determination of spontaneous GH secretion in detecting GHD is a time-consuming procedure, and alternate markers are often evaluated to determine their usefulness as a diagnostic measure in detecting GHD. Blum et al. (140 ) in their earlier work showed that IGFBP-3 was subnormal in GH-deficient patients and that it could serve as an excellent screening parameter for GHD. In a recent study (290 ), a 17.7-kDa form of IGFBP-3 that is GH dependent was detected in the serum and urine of healthy children but was absent in healthy adults and GH-deficient children. Since IGFBP-3 increases in response to GH treatment secondary to an increase in IGF-I, quantification of urinary IGFBP-3 could provide a simple and noninvasive test to detect GHD and monitor the response to GH therapy. The IGFBP-3 in urine is degraded and, therefore, the usefulness of urinary IGFBP-3 as a diagnostic tool may be partly dependent on patient-to-patient variations in this proteolytic cleavage. Further, monitoring the levels of this molecular form of IGFBP-3 as a screening tool for GHD may be considered only after further studies support these findings. Serum levels of IGF-I, IGFBP-3, and ALS were measured after GH treatment in GHD adults (291 ). It was observed that the sensitivity of IGFBP-3 and ALS to the GH dose was lower than that of IGF-I. Thus IGF-I levels in serum are much more useful as a diagnostic tool than serum IGFBP-3 levels for detecting GH excess in adults or in those receiving GH replacement therapy. This is because it is more sensitive than IGFBP-3 and ALS to GH doses in the high range (291 ). Although the value of additional measurement of either IGFBP-3 or ALS levels is limited, these measurements could improve the reliability of diagnostic assessment.
5. Acromegaly. Acromegaly is a disorder caused by hyperfunction of the pituitary gland with normal to high production of GH. Plasma levels of IGF-I and IGFBP-3 were elevated markedly in untreated acromegalic patients compared with age-matched controls (292 293 294 ). Although regulated by GH, the increase in IGFBP-3 was mediated in part via an increase in IGF-I (292 ). Insulin, glucose, and gut peptides have a role in altering the levels of IGFBP-3 (293 294 295 ), but further studies are needed to completely understand the physiological regulation of IGFBP-3 in acromegaly. Acromegalic patients typically demonstrate elevated serum IGFBP-1 and IGFBP-2 (294 ) and an elevated ratio of IGF-I/IGFBP-3, suggesting that the increase in GH level in acromegalic subjects may result in increased availability of free IGF-I (294 ). This ratio, however, does not provide additional diagnostic value over IGF-I measurement in clinical settings, since like IGF-I, the ratio of IGF-I to IGFBP-3 is higher for acromegalic patients compared with healthy controls. In a recent case report, Yoshida et al. (296 ) showed that a nonpulsatile GH secretion and enhanced tissue sensitivity to GH may induce hypersecretion of IGF-I and IGFBP-3 resulting in clinical acromegalic gigantism. The findings that the levels of IGF-I and IGFBP-3 were increased in acromegalic subjects while they were decreased in GH-deficient subjects are consistent with an important role for GH in regulating IGF-I and IGFBP-3 levels in vivo.
6. Osteoporosis. All of the IGFBPs identified thus far (IGFBP-1 through -6) are also produced by human bone cells (31 34 46 153 297 298 299 ). IGFBPs exhibit significant biological effects on bone cells, and the production of the IGFBPs is regulated by a variety of osteoregulatory agents, emphasizing a key role for IGFBPs in regulating osteoblast cell proliferation and differentiation (34 153 297 300 301 302 ). A number of studies demonstrate that circulating levels of IGF-I and bone mass are positively correlated in healthy subjects and that subjects with osteoporosis have low circulating levels of IGF-I (284 303 304 ). Recent studies also showed changes in the serum levels of various IGFBPs in osteoporotic subjects compared with normal individuals (304 305 306 ). Johansson et al. (284 ) reported that serum levels of IGFBP-3 showed significant positive correlation with bone mineral density at multiple sites. Serum IGFBP-3 levels were found to be considerably less in osteoporotic subjects compared with age-matched controls (305 ) and was shown to increase in response to treatment with recombinant human GH (306 ). In addition, serum IGFBP-5 levels, measured by RIA using polyclonal antiserum, were found to be low in osteoporotic subjects (142 ). However, further studies are needed to evaluate whether the low levels of IGFBP-5 in the serum of osteoporotic patients represent that of intact IGFBP-5 or fragments or both.
Using ligand blot analysis, it was shown that serum IGFBP-4 levels are higher in elderly women with hip fractures with elevated PTH levels compared with age-matched controls (227 ). However, this needs to be confirmed using RIA in a larger population. It was also shown that the increased serum IGFBP-4 levels in those who are advanced in age show significant positive correlation with serum PTH levels (139 ). PTH treatment increased IGFBP-4 production in human osteoblasts in vitro (301 ). In addition, serum IGFBP-4 levels are increased during oral 1,25-dihydroxyvitamin D3 therapy in psoriasis patients (307 ). These findings suggest that during calcium deficiency, the increase in serum PTH and 1,25 dihydroxyvitamin D3 may, in addition to stimulating bone resorption, inhibit bone formation by stimulating bone cell production of IGFBP-4 (307 308 309 ). These data demonstrate that patients with senile osteoporosis who have hip fracture and secondary hyperparathyroidism also have a highly significant increase in the ratio of inhibitory to stimulatory IGF system components (222 241 ). These two changes could lead to a decrease in bone formation and a decrease in hip bone density that may increase their propensity toward hip fractures.
7. Renal failure. Patients with chronic renal failure (CRF) demonstrated a higher capacity to bind IGFs compared with age-matched controls (310 311 ). This is contributed by a significant increase in circulating IGFBP-2 and -3 (312 ), with a moderate increase in IGFBP-1 (145 313 ). Serum IGFBP-2 and -3 levels of CRF patients are high, unlike that of serum IGFBP-1, which is reduced due in part to the high insulin levels seen in these patients. The increase in IGFBP-2 seems to be due to an increase in hepatic production and/or lowered rate of renal filtration. This excess IGFBP in CRF patients is believed to act as an inhibitor of IGF action by decreasing the free biologically active IGF (314 315 ). This could explain the growth deficit seen in CRF subjects and the improvement seen in longitudinal growth after GH treatment (146 314 ).
In children with chronic renal insufficiency (CRI), the growth deficit observed is reversed after renal transplants. Growth is not always sustained in these patients because a number of factors, including the treatments they receive, impair growth (316 ). Prednisone treatment of patients with CRI increased serum IGFBP-3 levels (311 312 315 317 318 ) and decreased growth rate. In contrast to IGFBP-3, serum IGFBP-1 levels were higher before prednisone treatment, but returned to normal after treatment, probably as a result of an increase in insulin concentration. It is not known whether the increase in IGFBP-3 and/or decrease in IGFBP-1 after prednisone therapy in CRI patients has any role in mediating the impairment of growth in CRI patients.
Nephrotic syndrome results in damages of the capillary wall of the arteries that supply the glomerulus and clinically leads to the loss of large amounts of protein in the urine, resulting in hypoalbuminemia. It is therefore likely that IGF-I/IGFBP complex is also filtered, affecting the circulating levels of these binding proteins. Serum IGF-I concentration and the binding to the 150-to 200-kDa complex were lower in nephrotic rats than pair-fed controls (317 ), while the serum level of IGFBP-2 was increased due to an increase in liver synthesis. It was shown in this study that the decrease in serum IGF-I was compensated for by an increase in a IGFBP-3 protease, which shifted the binding of IGF-I from the 150- to 200-kDa complex to the low molecular mass complex so that it permeates the vascular compartment and maintains tissue IGF-I availability.
8. Other pathological conditions. Serum IGFBP-1 and GH were elevated in cirrhotic patients compared with control subjects (319 320 ). The authors suggested that this might be due to the insulin resistance seen in cirrhosis as insulin regulates IGFBP-1 production in the healthy state (319 ). On the other hand, a decreased level of IGFBP-3 was observed due to decreases in hepatic production. While the concentration of IGFBP-3 correlated with the severity of the hepatic disease, IGFBP-1 did not show a similar relationship. It is clear that IGFBP-1 and -3 are regulated differently and serve different functions.
Growth and wasting accompany end-stage liver disease in children and accelerated growth is feasible only after liver transplant. Before the transplant, the circulating levels of IGF-I and IGFBP-3 are low (321 ), similar to those observed in acute and chronic renal failure (319 ). The low levels of IGF-I could arise from the state of malnutrition or the presence of liver disease itself. After receiving the liver transplant, these patients demonstrate an increase in both IGF-I and IGFBP-3, with the latter increasing relatively higher than control values. It is believed that the higher IGF-I level in circulation after liver transplantation compared with the pretransplantation level would increase the endocrine action of IGFs for growth stimulation.
Serum IGFBP-1 levels were markedly increased in patients before transplant, and this persisted after transplant. IGFBP-1 thus exerts an inhibitory effect on IGF action. However, unlike in healthy subjects, insulin does not regulate IGFBP-1 levels, as high insulin secretion in these patients does not suppress IGFBP-1 production. A rise in serum IGFBP-2 as a result of dietary restriction in these patients was also observed, but the level continued to be elevated for reasons not understood. A combined increase in serum IGFBP-1 and -3 post-transplant is speculated to play a role in the growth failure or slow catch-up observed in patients after liver transplant surgery.
In severely ill patients, catabolism leads to major changes in IGFBP production and circulation in the serum, which in turn can influence IGF bioavailability. The major binding protein in the serum, IGFBP-3, was greatly decreased in severely ill patients due to the release of substrate-specific IGFBP-3 protease. Similarly, after surgery such as cholecystectomy (180 ) and major heart surgery (322 ), there is an increase in proteolysis of IGFBP-3. Based on these findings, it is speculated that the release of IGFBP-3 protease during surgery is an adaptive mechanism to ensure an adequate supply of IGF to the tissue for counteracting the catabolic state (178 ).
Serum IGFBP-3 decreased in hypothyroid patients and was within normal levels in hyperthyroid patients. In contrast, serum IGFBP-1 was elevated in the hyperthyroid group (323 ) or increased in hypothyroid patients receiving T4 treatment (324 ). The patient is typically in a catabolic state in hyperthyroidism, and this may be due, in part, to elevated IGFBP-1, which suggests a decrease in bioactivity of IGFs in plasma. These conclusions have been questioned since the collection of the plasma from hyperthyroid subjects and the reference pool was not done at the same time of day. This suggests that the changes in IGFBP-1 levels between the healthy controls and hyperparathyroids may have been partly due to diurnal variation.
Based on the above mentioned discussion, it is obvious that the serum
IGFBP levels are altered under various physiological and pathological
conditions. Table 5 summarizes the key
regulators of various IGFBP levels in circulation. Multiple mechanisms
are known to be involved in regulating serum levels of various IGFBPs,
including alterations in the production of IGFBPs in target cell types
and degradation of IGFBPs by specific proteases. The presence of
multiple binding proteins and the complexity of regulation are
consistent with a key role for IGFBPs in regulating the actions of IGFs
in response to a multitude of systemic and local effectors.
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VIII. IGFBP Proteases in Circulation |
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TopAbstractI. IntroductionII. Characteristics of the...III. Target Cell Actions...IV. IGF-IGFBP Complexes in...V. Assays for Circulating...VI. Relative Distribution of...VII. Regulation of Serum...VIII. IGFBP Proteases in...IX. Endocrine Functions of...X. ConclusionsReferences |
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A. Proteases under normal conditions
IGFBP-3 proteolysis is known to occur under normal conditions (25 191 325 326 327 ) in a variety of biological fluids such as peritoneal,
follicular, amniotic, seminal, and cerebrospinal fluids. In addition,
amniotic fluid contains a IGFBP-1-specific protease and cerebrospinal
fluid contains a IGFBP-2 protease (91 ). In addition to tissue fluids, a
variety of cell types in culture secrete proteases capable of degrading
IGFBPs. Some of these proteases are relatively specific to a given
IGFBP while others are relatively nonspecific. For example, bone cells
in culture produce matrix metalloproteases and plasmin, which degrade
multiple IGFBPs as well as other proteins (328 ). In addition, human
bone cells as well as fibroblasts in culture produce proteases that are
relatively specific to IGFBP-5 (281 282 329 ). Endogenous IGF-II and
tumor necrosis factor-ß stimulate IGFBP-4 protease (282 330 331 ),
and IGF-II inhibits IGFBP-5 protease (282 ). The finding that IGFs
regulate IGFBP levels by regulating proteolysis in a variety of cell
types raises the interesting possibility that IGFs function to regulate
IGFBPs, as well as vice versa. IGFBP-2-, -4-, and
-5-specific proteases have been detected in smooth muscle cells (332 )
with IGFBP-4 protease capable of increasing the bioavailable IGF-I.
IGFBPs 1–4 were found in human skin interstitial fluid (128 ) with
IGFBP-3 in the modified form due to an increased IGFBP-3 protease
activity (128 333 ). Although IGF concentration in the interstitial
fluid is lower than in serum, it is more available due to proteolytic
activity for IGFBPs (128 ). Synovial fluid from both normal individuals
and patients with rheumatoid arthritis (130 ) also contain IGFBP-3
protease activity, which acts to regulate IGF bioavailability. IGFBP-4
levels in estrogen-dominant follicular fluid were lower than levels
found in androgen-dominant follicular fluid because of the presence of
a metalloserine IGFBP-4 protease activity in the former. It is
speculated that the presence of IGFBP-4 protease in estrogen-dominant
follicular fluid would decrease the inhibitory IGFBP-4 level and
increase bioavailability of IGF. It is also speculated that the
available IGFs stimulate estradiol production along with gonadotropins
in the growing follicular tissues (41 ). Significant proteolysis of
IGFBP-3 was shown in the reproductive tract of men, and the intact
IGFBP-3 levels were inversely related to the PSA concentration (334 ).
Thus, proteases have been postulated to aid in modulating IGF action in
a tissue-specific manner.
B. Pregnancy-associated proteases
One of the first identified IGFBP proteases was a
pregnancy-associated IGFBP-3-specific protease that is active from the
second trimester of pregnancy in humans (135 179 ). This is specific to
the reproductive tissues and helps to meet the in utero
fetal growth demand. The potential role of IGFBP-3 protease in
regulating the bioavailability of IGFs has been discussed earlier. In
addition to IGFBP-3 protease, a IGFBP-5 protease that degrades IGFBP-3,
-4, and -5 was detected in maternal serum and amniotic fluid (119 ). The
protease activity is also increased in relation to fetal demands as
seen by a higher serum protease activity in mothers with multiple
fetuses or with fetuses affected by in utero placental
insufficiency (203 ) and in GH-deficient pregnant rats (275 ). The
regulation of these proteases is now being investigated.
C. Proteases under catabolic and disease states
Another common condition that increases serum protease activity is
a general catabolic state. For example, IGFBP-3 specific protease was
detected in patients with severe illnesses and in a catabolic negative
nitrogen balance (178 180 ). Protease activity is more significant
during fasting and declines after parenteral nutrition (322 325 ),
suggesting that there is a specific response to catabolism. Another
example of a catabolic state occurs in patients with acute and chronic
renal failure. These patients showed an increase in urinary
IGFBP-3-specific protease, which was responsible for the complete
absence of intact IGFBP-3 in the urine (276 ). Significant proteolytic
activity of IGFBP-3 protease was observed in acute diseases of
childhood such as Kawasaki disease, bacterial meningitis, and
mycoplasma pneumonia (277 ). A high amount of IGFBP-3 protease activity
was detected in the CSF of children with CNS tumors (95 ). It is
speculated that IGFBP-3 proteolysis plays a role in overcoming
catabolic conditions by increasing the availability of IGF-I.
Insulin has an important role in regulating the activity of IGFBP-3-specific protease as shown in studies that demonstrated an elevated serum IGFBP-3 protease activity in untreated NIDDM (335 ) and IDDM (137 ) patients. This appears to be regulated by insulin levels, as shown by a decline in the protease activity after insulin therapy. Therefore, the catabolic state induced by insulin deficiency is counteracted to some extent by an increase in the IGFBP protease activity.
The observations that IGFBP protease activity was detected in various
biological fluids during pathological and physiological situations
suggest a role for protease in conditions of increased growth such as
in pregnancy, and also when a person is in a catabolic state indicated
by an illness, infection, cancer, or diabetes. The increase in protease
activity during these situations helps to increase the dissociation of
IGF from the 150- to 200-kDa complex so that more free IGF is available
for promoting growth. The finding that a number of IGFBP proteases have
been identified in a variety of biological fluids (Table 6) and the finding that the rate of IGFBP
proteolysis can be regulated by a variety of systemic and local
effectors (327 336 337 338 339 340 341 342 ) raise interesting possibilities for the
involvement of IGFBP protease in regulating the endocrine and local
actions of IGFs (see Figs. 6 and 7).
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IX. Endocrine Functions of IGFBPs in Serum |
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TopAbstractI. IntroductionII. Characteristics of the...III. Target Cell Actions...IV. IGF-IGFBP Complexes in...V. Assays for Circulating...VI. Relative Distribution of...VII. Regulation of Serum...VIII. IGFBP Proteases in...IX. Endocrine Functions of...X. ConclusionsReferences |
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A. To prevent insulin-like effects
In adult humans, the mean IGF concentration is about 800
µg/liter, with IGF-I and -II contributing about 200 µg/liter and
600 µg/liter, respectively. This amount is 1000-fold higher than mean
insulin concentration. Despite the fact that insulin-like activity of
IGFs is only 5% of insulin, the IGFs could in theory contribute 50
times more insulin-like activity than insulin alone due to their
abundance (6 58 59 149 343 ). However, this does not occur because
the activity of IGFs is largely neutralized as a consequence of IGFBP
binding. Thus, without the IGFBPs, the effect of IGFs would overwhelm
the effect of insulin alone, and because IGF expression is not tightly
controlled by blood glucose levels, blood glucose homeostasis would not
be possible.
B. To increase the half-lives of IGFs
IGFBPs are known to facilitate IGF endocrine action by increasing
the half-life of the IGFs in circulation. Both IGF-I and IGF-II, when
injected into hypophysectomized rats, disappeared in approximately
20–30 min, while in normal rats the half-life was extended to about
15–20 h (81 ). This difference is due to the fact that the majority of
IGFs circulate in serum as a 150- to 200-kDa ALS+IGFBP-3+IGF ternary
complex as described previously. The stabilization of IGFs thus depends
mainly on IGFBP-3, with other IGFBPs having minimal roles in this
function. IGFBP-1 and -2, for example, had shorter half-lives in serum
(161 ) and thus seem unlikely to play any role in stabilizing the IGFs
in the circulation. Thus, by increasing the half-life of IGFs and
protecting them from rapid degradation, IGFBP-3 alters the ratio of
free IGF to bound IGF, which in turn regulates the metabolic actions of
the IGFs. Consistent with the idea that IGFBPs may increase the
endocrine actions of IGFs by increasing their half-lives, several
reports document that systemic administration of an equimolar dose of
IGFBP-3+IGF-I complex is more effective than IGF-I alone in stimulating
bone formation in ovariectomized rats (344 345 ).
C. To control the transport of IGFs from the vascular space
The transportation of serum IGFs from the vasculature into the
extracellular fluid is necessary in order for IGFs to elicit a
growth-stimulating response. For this to be accomplished, the IGFBPs
bound to the IGFs in circulation must be either degraded or should be
able to cross the endothelial barrier (Fig. 6). The majority of IGFs exist as a
ALS-IGFBP-3-IGF complex and, as such, other IGFBPs, including
IGFBP-1 and IGFBP-2, remain unsaturated in plasma since they are
not the predominant binding proteins (59 ) for IGFs. However, their
ability to cross the endothelial barriers intact (346 ) makes them
important IGF transporters to target tissues. Unlike the complex with
IGFBP-1 and -2, the IGFBP-3-IGF-ALS complex, which contains more than
75% of the circulating IGFs, does not migrate from the vascular
compartment. The ALS+IGFBP-3+IGF complex serves as a reservoir for IGF
release, which is determined by 1) a drop in free IGF or 2) the action
of certain proteases. Proteolytic cleavage can reduce the affinity of
the IGFBP-3 for IGFs by 20–30 times (135 ) and facilitates
equilibration with the smaller molecular mass binding protein complex
in serum (Fig. 6). Because the affinity of IGFs to IGFBP-3 is reduced
upon binding to glycosaminoglycans, it is speculated that
glycosaminoglycans present on the surface of endothelial cells may
control the efflux of the IGFs out of the vascular compartment (346 ).
Although this is an attractive theory, further experimental data are
required to support this function for extracellular matrix proteins
produced by vascular endothelial cells.
The shifting of IGF from the 150- to 200-kDa complex to a 50-kDa IGF
complex does not necessarily increase IGF bioavailability in target
tissues since many of the IGFBPs (e.g., IGFBP-1, IGFBP-2,
IGFBP-4, and IGFBP-6) inhibit IGF actions by preventing the binding of
IGFs to their receptors. Thus, the inhibitory IGFBP in the 50-kDa
IGFBP+IGF complex must be degraded in the local tissue to release the
IGF from this circulating complex in a bioavailable form (Fig. 7). Based on the above mentioned
discussion, two sequential mechanisms are essential to increase the
availability of free IGFs to local tissues: First, release of IGFs from
the 150- to 200-kDa complex by disruption of ALS+IGFBP-3+IGF complex
(e.g., proteolysis of IGFBP-3); second, release of IGFs from
a 50-kDa inhibitory IGFBP+IGF complex by disruption of this complex
(e.g., proteolysis of inhibitory IGFBP-4). Consistent with
the idea that locally produced IGFBP proteases may play a role in
regulating IGF action, a number of studies have shown that the activity
of the IGFBP proteases in biological fluids may be regulated based on
the physiological and/or pathological conditions. In addition, in
vitro studies have shown evidence that a number of cell types and
cultures produce proteases capable of degrading one or more IGFBPs,
which are regulated by both local and hormonal effectors (100 124 157 282 329 331 347 348 ).
Thus the 150- to 200-kDa and 50-kDa IGFBP complexes may have a specific role either to increase half-life, prevent hypoglycemia, or to facilitate transport of IGF out of the vascular compartment. These functions ultimately are responsible for increasing the bioavailability of free IGF for mitogenic/differentiation activity in the target tissues. Several mechanisms have been suggested to cause an increase in the bioavailability of free IGFs in the target tissues, such as 1) increasing the ratio of IGFs/inhibitory IGFBPs; 2) increasing the ratio of stimulatory IGFBPs/inhibitory IGFBPs; 3) increasing or decreasing the rate of proteolysis of inhibitory or stimulatory IGFBPs, respectively; and 4) increasing IGF receptor abundance in the target tissue (149 ). Essentially, factors that can alter the effective concentration of IGFBPs can, in turn, regulate the availability of free IGF.
Since IGFs present in the 50-kDa complex can exist as complexes of both stimulatory and inhibitory binding proteins, it is essential that the regulation of these two types of binding proteins occur in a reciprocal manner to produce a significant biological effect. Several physiological and pathological conditions, including age, development, nutrition, exercise, surgery, and pregnancy, have been shown to affect the ratio of IGF to IGFBP (78 162 178 179 180 214 231 273 ). For example, the relative ratio of inhibitory IGFBP-4 to stimulatory IGFBP-5 increases with advancing age, a change that would tend to decrease the endocrine actions of IGFs (222 ). It is anticipated that future research studies would provide experimental data to demonstrate a role for IGFBPs and their proteases in regulating the endocrine IGF actions depending on the needs of the target tissues.
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X. Conclusions |
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TopAbstractI. IntroductionII. Characteristics of the...III. Target Cell Actions...IV. IGF-IGFBP Complexes in...V. Assays for Circulating...VI. Relative Distribution of...VII. Regulation of Serum...VIII. IGFBP Proteases in...IX. Endocrine Functions of...X. ConclusionsReferences |
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Footnotes |
|---|
1 Supported by funds from NIH (AR-31062), Veteran’s
Administration, and Department of Nutrition, School of Public
Health, Loma Linda University. 
2 Oh et al. (349 ) have recently shown
that purified recombinant human mac25 protein (termed IGFBP-7), which
shares 20–25% identity with IGFBPs, binds IGFs but with several fold
lower affinity than that of IGFBP-3.
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References |
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TopAbstractI. IntroductionII. Characteristics of the...III. Target Cell Actions...IV. IGF-IGFBP Complexes in...V. Assays for Circulating...VI. Relative Distribution of...VII. Regulation of Serum...VIII. IGFBP Proteases in...IX. Endocrine Functions of...X. ConclusionsReferences |
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K. Nosho, H. Yamamoto, H. Taniguchi, Y. Adachi, Y. Yoshida, Y. Arimura, T. Endo, Y. Hinoda, and K. Imai Interplay of Insulin-Like Growth Factor-II, Insulin-Like Growth Factor-I, Insulin-Like Growth Factor-I Receptor, COX-2, and Matrix Metalloproteinase-7, Play Key Roles in the Early Stage of Colorectal Carcinogenesis Clin. Cancer Res., December 1, 2004; 10(23): 7950 - 7957. [Abstract] [Full Text] [PDF]
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M. Dehoux, R. Van Beneden, N. Pasko, P. Lause, J. Verniers, L. Underwood, J.-M. Ketelslegers, and J.-P. Thissen Role of the Insulin-Like Growth Factor I Decline in the Induction of Atrogin-1/MAFbx during Fasting and Diabetes Endocrinology, November 1, 2004; 145(11): 4806 - 4812. [Abstract] [Full Text] [PDF]
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K.-C. Leung, G. Johannsson, G. M. Leong, and K. K. Y. Ho Estrogen Regulation of Growth Hormone Action Endocr. Rev., October 1, 2004; 25(5): 693 - 721. [Abstract] [Full Text] [PDF]
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P. A. Pennisi, J. J. Kopchick, S. Thorgeirsson, D. LeRoith, and S. Yakar Role of Growth Hormone (GH) in Liver Regeneration Endocrinology, October 1, 2004; 145(10): 4748 - 4755. [Abstract] [Full Text] [PDF]
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E D Watson, S-E Bae, R Thomassen, S R M Thomson, K Woad, and D G Armstrong Insulin-like growth factors-I and -II and insulin-like growth factor-binding protein-2 in dominant equine follicles during spring transition and the ovulatory season Reproduction, September 1, 2004; 128(3): 321 - 329. [Abstract] [Full Text] [PDF]
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J. L. King and C. Guidry Insulin-Like Growth Factor Binding Proteins Modulate Muller Cell Responses to Insulin-Like Growth Factors Invest. Ophthalmol. Vis. Sci., August 1, 2004; 45(8): 2848 - 2855. [Abstract] [Full Text] [PDF]
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 A. De Luca, M. De Falco, V. Fedele, L. Cobellis, A. Mastrogiacomo, V. Laforgia, I. L. Tuduce, M. Campioni, D. Giraldi, M. G. Paggi, et al. The Serine Protease HtrA1 Is Upregulated in the Human Placenta During Pregnancy J. Histochem. Cytochem., July 1, 2004; 52(7): 885 - 892. [Abstract] [Full Text] [PDF]
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C. F. Singer, M. Mogg, W. Koestler, M. Pacher, E. Marton, E. Kubista, and M. Schreiber Insulin-Like Growth Factor (IGF)-I and IGF-II Serum Concentrations in Patients with Benign and Malignant Breast Lesions: Free IGF-II Is Correlated with Breast Cancer Size Clin. Cancer Res., June 15, 2004; 10(12): 4003 - 4009. [Abstract] [Full Text] [PDF]
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D. Vanderschueren, L. Vandenput, S. Boonen, M. K. Lindberg, R. Bouillon, and C. Ohlsson Androgens and Bone Endocr. Rev., June 1, 2004; 25(3): 389 - 425. [Abstract] [Full Text] [PDF]
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C. H. Lang, R. A. Frost, E. Svanberg, and T. C. Vary IGF-I/IGFBP-3 ameliorates alterations in protein synthesis, eIF4E availability, and myostatin in alcohol-fed rats Am J Physiol Endocrinol Metab, June 1, 2004; 286(6): E916 - E926. [Abstract] [Full Text] [PDF]
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C. Y. Bowers, R. Granda, S. Mohan, J. Kuipers, D. Baylink, and J. D. Veldhuis Sustained Elevation of Pulsatile Growth Hormone (GH) Secretion and Insulin-Like Growth Factor I (IGF-I), IGF-Binding Protein-3 (IGFBP-3), and IGFBP-5 Concentrations during 30-Day Continuous Subcutaneous Infusion of GH-Releasing Peptide-2 in Older Men and Women J. Clin. Endocrinol. Metab., May 1, 2004; 89(5): 2290 - 2300. [Abstract] [Full Text] [PDF]
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L. J. Spicer Proteolytic Degradation of Insulin-Like Growth Factor Binding Proteins by Ovarian Follicles: A Control Mechanism for Selection of Dominant Follicles Biol Reprod, May 1, 2004; 70(5): 1223 - 1230. [Abstract] [Full Text] [PDF]
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J. Ayuk, R. N. Clayton, G. Holder, M. C. Sheppard, P. M. Stewart, and A. S. Bates Growth Hormone and Pituitary Radiotherapy, But Not Serum Insulin-Like Growth Factor-I Concentrations, Predict Excess Mortality in Patients with Acromegaly J. Clin. Endocrinol. Metab., April 1, 2004; 89(4): 1613 - 1617. [Abstract] [Full Text] [PDF]
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K. Wolk, S. C. Larsson, B. Vessby, A. Wolk, and K. Brismar Metabolic, Anthropometric, and Nutritional Factors as Predictors of Circulating Insulin-Like Growth Factor Binding Protein-1 Levels in Middle-Aged and Elderly Men J. Clin. Endocrinol. Metab., April 1, 2004; 89(4): 1879 - 1884. [Abstract] [Full Text] [PDF]
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Z. T. Resch, B.-K. Chen, L. K. Bale, C. Oxvig, M. T. Overgaard, and C. A. Conover Pregnancy-Associated Plasma Protein A Gene Expression as a Target of Inflammatory Cytokines Endocrinology, March 1, 2004; 145(3): 1124 - 1129. [Abstract] [Full Text] [PDF]
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S. Baron-Hay, F. Boyle, A. Ferrier, and C. Scott Elevated Serum Insulin-Like Growth Factor Binding Protein-2 as a Prognostic Marker in Patients with Ovarian Cancer Clin. Cancer Res., March 1, 2004; 10(5): 1796 - 1806. [Abstract] [Full Text] [PDF]
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S. R. Ravelich, B. H. Breier, S. Reddy, J. A. Keelan, D. N. Wells, A. J. Peterson, and R. S.F. Lee Insulin-Like Growth Factor-I and Binding Proteins 1, 2, and 3 in Bovine Nuclear Transfer Pregnancies Biol Reprod, February 1, 2004; 70(2): 430 - 438. [Abstract] [Full Text] [PDF]
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S.'i. Miyamoto, K. Yano, S. Sugimoto, G. Ishii, T. Hasebe, Y. Endoh, K. Kodama, M. Goya, T. Chiba, and A. Ochiai Matrix Metalloproteinase-7 Facilitates Insulin-Like Growth Factor Bioavailability through Its Proteinase Activity on Insulin-Like Growth Factor Binding Protein 3 Cancer Res., January 15, 2004; 64(2): 665 - 671. [Abstract] [Full Text] [PDF]
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F. Haddad and G. R. Adams Inhibition of MAP/ERK kinase prevents IGF-I-induced hypertrophy in rat muscles J Appl Physiol, January 1, 2004; 96(1): 203 - 210. [Abstract] [Full Text] [PDF]
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D. Nemet, P. H. Connolly, A. M. Pontello-Pescatello, C. Rose-Gottron, J. K. Larson, P. Galassetti, and D. M. Cooper Negative energy balance plays a major role in the IGF-I response to exercise training J Appl Physiol, January 1, 2004; 96(1): 276 - 282. [Abstract] [Full Text] [PDF]
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S. R. Edmondson, S. P. Thumiger, G. A. Werther, and C. J. Wraight Epidermal Homeostasis: The Role of the Growth Hormone and Insulin-Like Growth Factor Systems Endocr. Rev., December 1, 2003; 24(6): 737 - 764. [Abstract] [Full Text] [PDF]
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Y. Matsumoto, Y. Oda, M. Uryu, and Y. Hayakawa Insect Cytokine Growth-blocking Peptide Triggers a Termination System of Cellular Immunity by Inducing Its Binding Protein J. Biol. Chem., October 3, 2003; 278(40): 38579 - 38585. [Abstract] [Full Text] [PDF]
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J. A. Clowes, H. C. Allen, D. M. Prentis, R. Eastell, and A. Blumsohn Octreotide Abolishes the Acute Decrease in Bone Turnover in Response to Oral Glucose J. Clin. Endocrinol. Metab., October 1, 2003; 88(10): 4867 - 4873. [Abstract] [Full Text] [PDF]
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X. Wu, S.-T. Pang, L. Sahlin, A. Blanck, G. Norstedt, and A. Flores-Morales Gene Expression Profiling of the Effects of Castration and Estrogen Treatment in the Rat Uterus Biol Reprod, October 1, 2003; 69(4): 1308 - 1317. [Abstract] [Full Text] [PDF]
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B. C. Nindl, J. W. Castellani, A. J. Young, J. F. Patton, M. J. Khosravi, A. Diamandi, and S. J. Montain Differential responses of IGF-I molecular complexes to military operational field training J Appl Physiol, September 1, 2003; 95(3): 1083 - 1089. [Abstract] [Full Text] [PDF]
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B.-K. Chen, K. M. Leiferman, M. R. Pittelkow, M. T. Overgaard, C. Oxvig, and C. A. Conover Localization and Regulation of Pregnancy-Associated Plasma Protein A Expression in Healing Human Skin J. Clin. Endocrinol. Metab., September 1, 2003; 88(9): 4465 - 4471. [Abstract] [Full Text] [PDF]
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C. H. Lang, T. C. Vary, and R. A. Frost Acute in Vivo Elevation of Insulin-Like Growth Factor (IGF) Binding Protein-1 Decreases Plasma Free IGF-I and Muscle Protein Synthesis Endocrinology, September 1, 2003; 144(9): 3922 - 3933. [Abstract] [Full Text] [PDF]
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 J A Mohammed, A Y. Mok, A Parbtani, and D G Matsell Increased expression of insulin-like growth factors in progressive glomerulonephritis of the MRL lpr mouse Lupus, August 1, 2003; 12(8): 584 - 590. [Abstract] [PDF]
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