Novel growth factors have often been given names which relate to their original site of discovery or their first known target cell or biological action. Subsequent studies have frequently shown such nomenclature to be misleading as more is learned about the factor concerned. Epidermal growth factor (EGF) and fibroblast growth factor (FGF) which in fact act as mitogens for a great variety of cell types and transforming growth factor-β (TGFβ) which is better recognized as a growth inhibitor are all typical examples. Moreover, in addition to their growth regulatory roles, peptide growth factors also serve to modulate cell differentiation and function. The recently characterized hepatocyte growth factor (HGF) joins this illustrious family of cytokines which have proven rather more versatile than their original descriptive title suggests.

ABSTRACT

We have investigated the ability of glucose, human GH and human placental lactogen (hPL) to alter the content and release of somatomedin C/insulin-like growth factor I (SM-C/IGF-I), and the biosynthesis, content and release of insulin from cultured human fetal pancreas. Fetal pancreatic explants obtained from glands following prostaglandin-induced abortion between 12 and 21 weeks of gestation were maintained in free-floating culture for 3–5 days before the experiments. The explants were then cultured for 3 days in medium containing either 2·7 or 16·7 mmol glucose/l with or without GH (4·5 or 45·5 nmol/l) or hPL (4·6 or 46·5 nmol/l). Serum-free medium from the final 24 h of culture was collected and SM-C/IGF-I and insulin were measured radioimmunologically in both conditioned medium and tissue explants extracted with acid ethanol. Insulin biosynthesis, determined by immunoprecipitation of [³H]leucine incorporated into insulin, was not significantly altered by any experimental variable. Incubation in the presence of 16·7 mmol glucose/l caused an increase of insulin release from explants, but had no consistent action on insulin content, compared with medium containing 2·7 mmol glucose/l. The pancreatic content and release of SM-C/IGF-I were independent of these glucose concentrations. Neither GH nor hPL altered insulin or SM-C/IGF-I content or release in the presence of the lower glucose concentration. At the higher glucose concentration, 45·5 nmol GH/1 did not alter insulin release but caused a significant increase in SM-C/IGF-I content. When present with 16·7 mmol glucose/l hPL (46·5 nmol/l) promoted significant increases in both insulin content and release (59 and 47% respectively) and significantly increased pancreatic SM-C/IGF-I content and release (114and 117% respectively).

The results suggest that, during tissue culture, glucose enhances the insulin output of human fetal pancreatic explants obtained in the second trimester. The ability of hPL to modulate B-cell function raises the possibility that this peptide may contribute to pancreatic development during fetal life and to total fetal body growth by increasing insulin production.

J. Endocr. (1987) 113, 297–303

Two isozymes of the enzyme 11 beta-hydroxysteroid dehydrogenase (11 beta-HSD) are responsible for the interconversion of the active glucocorticoid, cortisol in man, (corticosterone in the rodent), to the inactive 11-keto metabolite, cortisone (11-dehydrocorticosterone). We have examined the regulation of type 1 11 beta-HSD (11 beta-HSD1) using primary cultures of rat and human hepatocytes, both of which express only 11 beta-HSD1. Only 11 oxo-reductase activity could be demonstrated in cultured hepatocytes (apparent Km for cortisone 382 +/- 43 nM in human hepatocytes, apparent Km for 11-dehydrocorticosterone 14.6 +/- 1.5 microM in rat hepatocytes). There exists a marked discrepancy between 11 beta-HSD oxo-reductase activity and 11 beta-HSD1 mRNA levels in cultured human hepatocytes and human liver. Thus oxo-reductase specific activity is much higher in the cultured hepatocytes (7.2 +/- 0.01 nmoles cortisol/mg/h vs 0.89 +/- 0.06 for whole liver homogenates) whilst the converse is true for steady state 11 beta-HSD1 mRNA levels (0.78 +/- 0.02 vs 1.94 +/- 0.07 in whole liver, 11 beta-HSD1/18S expressed as arbitrary units). Carbenoxolone has a significant inhibitory effect on 11 oxo-reductase activity in both rat and human hepatocytes. However, there is clear species-specific regulation of 11 oxo-reductase activity by thyroid hormone (tri-iodothyronine (T3)), which increases 11 oxo-reductase activity in rat hepatocytes but has no effect on activity in human hepatocytes, and progesterone which inhibits activity in human hepatocytes, but has no effect on activity in rat hepatocytes. Neither T3 nor progesterone altered 11 beta-HSD1 mRNA levels. A series of growth factors (hepatocyte growth factor, epidermal growth factor, basic fibroblast growth factor, transforming growth factor beta 1) were without effect on 11 oxo-reductase activity in cultured rat hepatocytes. In contrast to homogenates of human liver, cultured hepatocytes express only 11 beta-HSD oxo-reductase activity. This is inhibited by carbenoxolone and shows species-specific regulation by T3 and progesterone. Growth factors do not appear to regulate activity or expression of 11 beta-HSD1. The discrepant enzyme activity data and 11 beta-HSD1 mRNA expression in hepatocytes and whole liver could reflect unstable 11 beta-HSD1 oxo-reductase activity or, alternatively, an additional 11 beta-HSD oxo-reductase isoform in cultured hepatocytes.

ABSTRACT

The presence of insulin-like growth factors (IGFs) in blood is regulated by their association with specific IGF-binding proteins (IGFBPs). In turn, the level of IGFBPs in the blood is likely to depend on a dynamic equilibrium between peptide production and clearance to extravascular tissues or organ-specific degradation. Since circulating IGFBPs may largely derive from liver we have employed partial hepatectomy in the rat to study the clearance rate of endogenous IGFBPs from blood once a major site of production is removed. Adult male rats were partially hepatectomized and serum and the remaining liver removed between 30 min and 7 days after surgery. Ligand blot analysis revealed two major species of IGFBP, of 28–30 kDa and 40–44 kDa in sera from control rats or sham-operated rats respectively. The larger species corresponded in size to rat IGFBP-3, but the smaller form was not recognized by antisera against rat IGFBP-1, bovine IGFBP-2 or human IGFBP-5 following Western immunoblot. Following hepatectomy, the levels of both IGFBP forms in the serum declined within 30 min and were barely detectable after 3 h or 6 h. They began to increase again in serum 24 h following surgery. The reduction in IGFBPs following hepatectomy was not primarily due to degradation by specific proteases in serum. Circulating levels of insulin were increased fivefold 3 h after hepatectomy but subsequently returned to control values. The rise in insulin was accompanied by a significant (P < 0·05) reduction in circulating IGF-I after 3 h which persisted at 24 h. Glucose levels in serum showed a transient but non-significant reduction between 90 min and 6 h after hepatectomy. Total RNA was extracted from remnant liver and subjected to Northern blot hybridization with ³²P-labelled cDNAs encoding rat IGFBP-1, -2 or -3. Messenger RNA encoding IGFBP-1 was barely detectable in liver from control or sham-operated animals, but increased within 30 min of partial hepatectomy and peaked at 3 h. It subsequently declined and was again barely detectable after 24 h. No expression of IGFBP-2 or -3 mRNAs was found by Northern blot analysis in the liver of control animals or following partial hepatectomy. These results suggest that both IGF-I and IGFBPs in rat serum decreased rapidly following partial hepatectomy, and that this was due largely to the rapid clearance of the peptide and its binding proteins once the major source of production was removed. A rapid induction of IGFBP-1 in the remaining liver may be unrelated to the circulating IGFBPs since immunoreactive IGFBP-1 was not detected in rat serum.

Journal of Endocrinology (1993) 137, 271–280

ABSTRACT

Insulin-like growth factors (IGFs) are bound to specific binding proteins in extracellular fluids in vivo and when released by cells in vitro. One class of binding protein (IGF-BP), a peptide of 26 kDa purified from amniotic fluid, has been shown to modulate IGF bioactivity on isolated human fibroblasts. We have determined the factors that control release of IGF-BP from monolayers of human fetal fibroblasts using a radioimmunoassay, and have compared this with the effects of these factors on the release of IGF-I and -II. Separation of cell-conditioned cultured medium on SDS-PAGE, and subsequent immunoblotting with antibody against IGF-BP showed that fibroblasts released a single species of immunoreactive protein of an estimated molecular weight of 30 kDa. This was not the predominant binding protein released by cells since major bands of approximately 42 kDa and 39 kDa were visualized following separation by SDS-PAGE and ligand blotting with ¹²⁵I-labelled IGF-I. The 30 kDa IGF-BP was released in parallel with radioimmunoassayable IGF-I and -II over 48 h. However, a significant inverse correlation was found between the release of IGF-BP, IGF-I, IGF-II and cell density. The exposure of fibroblasts to 1·3 nmol/l or greater of IGF-I or -II caused a significant release of IGF-BP. Maximum release was seen in sparse cultures with little or no release from confluent cultures. IGF-I and -II were approximately equipotent with a fourfold increase in IGF-BP release at 19·7 nmol/l. Insulin caused a release of IGF-BP and IGF-I and -II from fibroblasts at supraphysiological concentrations (16·7 nmol/l) which again was maximal on sparse cell cultures. Increases in IGF-BP, IGF-I and -II release were also found in the presence of human placental lactogen (23·3 nmol/l), but human GH, epidermal growth factor, fibroblast growth factor and platelet-derived growth factor were without effect. The results show that human fetal fibroblasts released an IGF-BP immunologically similar to that seen in amniotic fluid together with IGF-I and -II, that IGF-BP release was enhanced by exogenous addition of IGF peptides, and that the release of all three peptides was a property of sparsely plated, rapidly growing cells. These findings strengthen the hypothesis that the cellular expression of IGF-binding proteins may represent an important level of control in IGF physiology.

Journal of Endocrinology (1989) 122, 87–98

A reciprocal relationship between the endocrine and immune system has been demonstrated under pathophysiological conditions. However, few studies have assessed the relationship between thyroid hormones and immune function in apparently healthy individuals. Therefore, to clarify our understanding of normal physiological endocrine–immune interactions this study aimed to examine the interrelationships between thyroid hormones and immunity in healthy individuals. Total triiodothyronine (T₃), total thyroxine (T₄) and markers of immune status were assessed in 93 free-living and apparently healthy individuals aged 55–70 years. T₃ and T₄ concentrations were determined by commercially available kits. Immune status was assessed using flow cytometry and biochemical markers. Statistical analysis was performed by partial correlation, controlling for age. Thyroid hormone concentration was positively associated with markers of inflammation (P≤0.05), natural killer-like T cells (P≤0.001), expression of interleukin-6 (IL6) by activated monocytes (P≤0.05); percentage expression of memory T-lymphocytes (P≤0.01), memory T-helper lymphocytes (P≤0.05) and memory T-cytotoxic lymphocytes (P≤0.05), and higher IL2 receptor density on CD3+T-lymphocytes (P≤0.05). Thyroid hormone concentration was inversely associated with early lymphocyte apoptosis (P≤0.05) and the ratio of naïve- to memory T-cytotoxic lymphocytes (P≤0.05). The current study provides preliminary evidence of a role for T₃ and T₄, within normal physiological ranges, in the maintenance of lymphocyte subpopulations, and in mediating the inflammatory response. In conclusion, these findings highlight the potential implications of altered thyroid function in older individuals and the importance of future research examining thyroid–immune interactions.