Kevin M. Kelley

Insulin-like growth factor-binding proteins (IGFBPs) and their regulatory dynamics.

The IGFBPs are a family of homologous proteins that have co-evolved with the IGFs and that confer upon the IGF regulatory system both functional and tissue specificity. IGFBPs are not merely carrier proteins for IGFs, but hold a central position in IGF ligand-receptor interactions through influences on both the bioavailability and distribution of IGFs in the extracellular environment. In addition, IGFBPs appear to have intrinsic biological activity independent of IGFs. The current status of research on IGFBPs is reviewed herein. Following a brief introduction to the entire IGF/IGFBP system, separate sections for each of the six cloned mammalian IGFBPs, the most extensive for IGFBP3, cover selected topics that emphasize the dynamics of IGFBPs--that is, their regulation in cells, their functionally important post-translational modifications, and their interactions in the cellular microenvironment--and how these dynamics influence physiological function.

Stimulation of coho salmon growth by insulin-like growth factor I ☆

The effect of insulin-like growth factor I on growth rate of coho salmon (Oncorhynchus kisutch) was examined. Juvenile coho salmon received implants of osmotic minipumps containing recombinant bovine insulin-like growth factor I (rbIGF-I) or saline for a period of 3 to 4 weeks. High doses of rbIGF-I (greater than 0.13 microgram.g-1.d-1) resulted in hypoglycemia and death. In 2-year-old coho salmon, 0.09 microgram.g-1.d-1 rbIGF-I administered for 25 days doubled linear growth rate and increased growth rate in weight by 40%. In rapidly growing, 1-year-old coho salmon, growth rate was not altered by rbIGF-I at 0.01 or 0.05 micrograms.g-1.d-1 for 31 days. In ration-limited fish exhibiting slow growth in the control group, rbIGF-I (0.02 microgram.g-1.d-1) increased linear growth rate by up to threefold and growth rate in weight by up to fourfold. The results indicate that exogenous treatment with mammalian IGF-I can stimulate coho salmon growth under some conditions, and that endogenous IGF-I may be an important factor in regulating growth of teleosts.

Serum insulin-like growth factor binding proteins (IGFBPs) as markers for anabolic/catabolic condition in fishes

In fishes as well as in all vertebrates in which it has been assessed, physiological shifts toward catabolism (e.g. such as during food deprivation) are consistently associated with elevations in the serum levels of at least one (often two in fishes) IGFBP in the < or =31-kDa size range. In mammals, 30-kDa IGFBP-1 is strongly up-regulated under catabolic circumstances, and it plays an important physiological role by sequestering IGF peptides to inhibit energy-expensive growth until conditions are more favorable (e.g. with resumed feeding). Similarly in fishes, it has been found that when the < or =31-kDa IGFBPs are elevated in serum, somatic growth is inhibited, suggesting a similar growth-inhibitory role of these proteins to that of mammalian IGFBP-1. Three different experimentally-induced catabolic states in fishes are compared in this paper: fasting; insulin-dependent diabetes mellitus (IDDM); and stress. A strong relationship between elevated serum cortisol concentrations and the presence of IGFBPs in each case is noted, and the utility of serum IGFBP measurement to serve as an effective indicator (marker) of catabolic condition in fishes is discussed.

Evidence for thyroid endocrine disruption in wild fish in San Francisco Bay, California, USA. Relationships to contaminant exposures

It is well documented that many coastal and estuarine environments adjacent to developed and industrialized urban centers, such as the San Francisco Bay Area, are significantly contaminated by anthropogenic chemicals. However, it is not well understood to what extent existing contaminants, many with continuing inflows into the environment, may impact exposed wildlife. This study provided an initial characterization of thyroid endocrine-related effects and their relationship to accumulated contaminants in two indigenous fish species sampled from different San Franicsco Bay Area study sites. Plasma concentrations of thyroxine (T4) were significantly reduced in fish sampled from highly impacted locations such as Oakland Inner Harbor and San Leandro Bay as compared with fish from other locations representing relatively lower human impact, including Bodega Bay, Redwood City and a remote site on Santa Catalina Island. Triiodothyronine (T3) levels also varied significantly by location, with differing T3/T4 ratios in fish from some locations suggestive of altered peripheral deiodinase activity. The changes in thyroid endocrine parameters were significantly correlated with hepatic concentrations of certain environmental contaminants. A large number of polychlorinated biphenyl (PCB) congeners, both co-planar (dioxin-like) and non-co-planar, exhibited significant inverse correlations with T4 levels in the fish, while in contrast, T3 and T3/T4 ratio were positively correlated with PCB exposures. The positive correlation between T3/T4 ratio and PCBs supports the hypothesis that environmental PCBs may alter T4 deiodination or turnover, actions of PCBs reported in laboratory experiments. Some relationships between chlorinated pesticides including DDT and chlordanes, but fewer relationships with PAHs, were also observed. Together, these findings indicate that the thyroid endocrine system is exhibiting alterations associated with different aquatic environments in the San Francisco Bay Area, which are significantly related to current-day exposures of the fish to contaminant chemicals such as PCBs.

Analysis of Endocrine Disruption in Southern California Coastal Fish Using an Aquatic Multispecies Microarray

Background Endocrine disruptors include plasticizers, pesticides, detergents, and pharmaceuticals. Turbot and other flatfish are used to characterize the presence of chemicals in the marine environment. Unfortunately, there are relatively few genes of turbot and other flatfish in GenBank, which limits the use of molecular tools such as microarrays and quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR) to study disruption of endocrine responses in sentinel fish captured by regulatory agencies. Objectives We fabricated a multigene cross-species microarray as a diagnostic tool to screen the effects of environmental chemicals in fish, for which there is minimal genomic information. The array included genes that are involved in the actions of adrenal and sex steroids, thyroid hormone, and xenobiotic responses. This microarray will provide a sensitive tool for screening for the presence of chemicals with adverse effects on endocrine responses in coastal fish species. Methods We used a custom multispecies microarray to study gene expression in wild hornyhead turbot (Pleuronichthys verticalis) collected from polluted and clean coastal waters and in laboratory male zebrafish (Danio rerio) after exposure to estradiol and 4-nonylphenol. We measured gene-specific expression in turbot liver by qRT-PCR and correlated it to microarray data. Results Microarray and qRT-PCR analyses of livers from turbot collected from polluted areas revealed altered gene expression profiles compared with those from nonaffected areas. Conclusions The agreement between the array data and qRT-PCR analyses validates this multispecies microarray. The microarray measurement of gene expression in zebrafish, which are phylogenetically distant from turbot, indicates that this multispecies microarray will be useful for measuring endocrine responses in other fish.

Down-regulation in the insulin-like growth factor (IGF) axis during hibernation in the golden-mantled ground squirrel, Spermophilus lateralis: IGF-I and the IGF-binding proteins (IGFBPs).

The golden-mantled ground squirrel, Spermophilus lateralis, undergoes a profound winter hibernation that represents, among other changes, a prolonged period of starvation. In addition to dramatic metabolic and other physiological adaptations during hibernation which serve to reduce fuel energy expenditure, we have hypothesized that there may also be significant changes in the endocrine axis that regulates energetically-expensive somatic growth. As compared with euthermic, non-hibernating controls, hibernating S. lateralis were found to have 75%-reduced serum concentrations of insulin-like growth factor-I (IGF-I; from approximately 625 to approximately 150 ng/ml in both females and males, P < 0.05). While IGFBP-3 was the predominant IGFBP in serum of the euthermic controls, its levels were reduced to a similar degree in serum from the hibernating animals. IGFBP-4 was present at relatively low levels in the euthermic controls, and was reduced to undetectable levels in hibernating animals. Surprisingly, there was no IGFBP detectable in the 30 kDa range in either euthermic or hibernating S. lateralis, suggesting that IGFBP-1 does not play a role in hibernation-related changes in the IGF axis. In accordance with these endocrine changes, when serum from hibernating S. lateralis was added to cartilage explant cultures (at a 5% v/v concentration), it exhibited no ability to alter (35)S-proteoglycan synthetic rate, whereas serum from the euthermic squirrels significantly stimulated synthetic activity by 2-fold. These results suggest that part of hibernation adaptation in S. lateralis includes down-regulation in the growth-regulatory IGF axis. J. Exp. Zool. 289:66-73, 2001.

Novel effect of vasoactive intestinal polypeptide and peptide histidine isoleucine: Inhibition of in vitro secretion of prolactin in the tilapia, Oreochromis mossambicus

The effects of vasoactive intestinal polypeptide (VIP) and peptide histidine isoleucine (PHI) on the in vitro secretion of two prolactins (PRL) from the rostral pars distalis (RPD) and of growth hormone (GH) from the proximal pars distalis (PPD) of the pituitary of the tilapia (Oreochromis mossambicus) were studied. RPDs were incubated for 20 hr in hypoosmotic (280-300 mOsm) or hyperosmotic (340-350 mOsm) Krebs-Ringer bicarbonate medium with added peptide concentrations of 0 (control), 0.3, 3.0, 30, and 300 nM; similarly, PPDs were incubated with the same peptide concentrations in isoosmotic (325 mOsm) medium supplemented with cortisol. PRL and GH in the tissue and secreted into the medium were measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by soft laser densitometry of the protein band(s). Neither VIP nor PHI has a detectable effect on the secretion of GH. Secretion of the two PRLs is significantly inhibited by VIP and PHI in both hyperosmotic and hypoosmotic medium. In hyperosmotic medium, 300 nM VIP inhibits secretion of both PRLs by 47%, whereas in hypoosmotic medium, 300 nM VIP inhibits their secretion by 27%. PHI inhibits their secretion by ca. 65% in hyperosmotic medium and by 40% in hypoosmotic medium. There is preliminary immunocytochemical evidence for some VIP-like immunoreactivity (IR), but no conclusive indication of PHI-like IR, in the hypothalamo-hypophysial area. The inhibitory actions of VIP and PHI on PRL secretion in tilapia are in contrast to the known stimulatory actions of VIP and PHI on PRL secretion in tetrapods.

Molecular Analysis of Endocrine Disruption in Hornyhead Turbot at Wastewater Outfalls in Southern California Using a Second Generation Multi-Species Microarray

Sentinel fish hornyhead turbot ( Pleuronichthys verticalis ) captured near wastewater outfalls are used for monitoring exposure to industrial and agricultural chemicals of ~ 20 million people living in coastal Southern California. Although analyses of hormones in blood and organ morphology and histology are useful for assessing contaminant exposure, there is a need for quantitative and sensitive molecular measurements, since contaminants of emerging concern are known to produce subtle effects. We developed a second generation multi-species microarray with expanded content and sensitivity to investigate endocrine disruption in turbot captured near wastewater outfalls in San Diego, Orange County and Los Angeles California. Analysis of expression of genes involved in hormone [e.g., estrogen, androgen, thyroid] responses and xenobiotic metabolism in turbot livers was correlated with a series of phenotypic end points. Molecular analyses of turbot livers uncovered altered expression of vitellogenin and zona pellucida protein, indicating exposure to one or more estrogenic chemicals, as well as, alterations in cytochrome P450 (CYP) 1A, CYP3A and glutathione S-transferase-α indicating induction of the detoxification response. Molecular responses indicative of exposure to endocrine disruptors were observed in field-caught hornyhead turbot captured in Southern California demonstrating the utility of molecular methods for monitoring environmental chemicals in wastewater outfalls. Moreover, this approach can be adapted to monitor other sites for contaminants of emerging concern in other fish species for which there are few available gene sequences.

Evaluation of sources and loading of pesticides to the Sacramento River, California, USA, during a storm event of winter 2005

A monitoring study was conducted in the tributaries and main stem of the Sacramento River, California, USA, during the storm event of January 26 to February 1, 2005. The purpose of the study was to evaluate the sources and loading of pesticides in the Sacramento River watershed during the winter storm season. A total of 26 pesticides or pesticide degradates were analyzed, among which five pesticides and one triazine degradate were detected. Diuron, diazinon, and simazine were found in all streams with a total load of 110.4, 15.4, and 15.7 kg, respectively, in the Sacramento River over the single storm event. Bromacil, hexazinone, and the triazine degradate diaminochlorotriazine were only detected in two smaller drainage canals with a load ranged from 0.25 to 7 kg. The major source of pesticides detected in the main stem Sacramento River was from the most upstream subbasin, the Sacramento River above Colusa, where detected pesticides either exceeded or were close to those at the main outlet of the Sacramento River at Alamar Marina. The higher precipitation in this subbasin was partly responsible for the greater contribution of pesticides observed. Diazinon was the only pesticide with concentrations above water quality criteria, indicating that additional mitigation measures may be needed to reduce its movement to surface water.

An Insulinlike Growth Factor I-Resistant State in Cartilage of Diabetic Rats Is Ameliorated by Hypophysectomy: Possible Role of Metabolism

This study investigated the effect of IDDM on cartilage anabolic activity in rats. Rats were injected with STZ to induce IDDM, were hypophysectomized, or were injected with STZ and hypophysectomized. After 14 days, control (intact and sham-Hx) and Hx rats were normoglycemic, whereas the rats with IDDM exhibited hyperglycemia and glycosuria. The HxDb rats, however, had normal blood glucose levels and no glycosuria. Both growth, serum levels of IGF-I, and basal cartilage 35SO4 incorporation measured in vitro were decreased in the Hx, IDDM, and HxDb groups. IGF-I added in vitro significantly stimulated 35SO4 incorporation by cartilage explants from control and Hx animals, whereas explants from the animals with IDDM were unresponsive. Explants from the HxDb rats, however, were stimulated by IGF-I in a dose-related manner. Because Hx corrected the glycemic status of the IDDM rats and restored cartilage responsiveness to IGF-I, a second set of experiments was undertaken to further investigate the relationship between cellular metabolism and anabolic activity in cartilage. Cartilage explants from rats fasted for 48 h showed significantly decreased basal 35SO4 incorporation, which was as low as that in explants from rats with severe IDDM. Whereas explants from the IDDM rats were completely unresponsive, those from the fasted rats (and fed rats) were significantly stimulated by the added IGF-I. However, incubation in the presence of 2-D-G, which causes intracellular glucopenia, or in the absence of glucose, completely blocked the anabolic response to IGF-I in otherwise responsive tissues. In conclusion, an important component of diabetic growth inhibition appears to be tissue resistance to the anabolic action of IGF-I, a condition that is correctable by Hx and that may be a result of metabolic impairment at the tissue level.