We describe the use of four monoclonal antibodies (MAbs) to the rabbit liver growth hormone (GH) receptor and one raised against purified rat liver GH receptor to characterize liver receptor subtypes which differ in their hormone-binding regions. The anti-(rat liver GH receptor) MAb both inhibited and precipitated rat and rabbit GH receptors, but only one-half of 125I-oGH (ovine GH) binding to liver microsomes could be inhibited by excess antibody. Conversely, only one-half of 125I-anti-(rat GH receptor) MAb binding was inhibited by excess oGH and Scatchard plots for this MAb exhibited two components. Although only 50% of 125I-oGH binding to membranes was inhibited by this MAb, all solubilized receptor could be immunoprecipitated. We postulate two epitopes for the anti-(rat GH receptor) MAb, one located at the hormone-binding site (inhibitory site) and one elsewhere (immunoprecipitating site). A second, rabbit-specific antibody (MAb 7) inhibited 85% of hormone binding but only 30% of 125I-anti-(rat GH receptor) MAb binding to rabbit liver microsomes. A combination of this MAb with the anti-(rat GH receptor) MAb totally inhibited 125I-oGH binding. MAb 7 alone totally inhibited 125I-rat GH binding to rabbit liver microsomes, as it did with 125I-oGH binding to purified receptor. On the basis of these results and others we postulate three types of GH receptor in rabbit liver membranes and ascribe approximate extents of 125I-oGH binding to each. A cytosolic 'GH receptor' which is not poly(ethylene glycol)-precipitable is shown to share five epitopes with 'type 2' microsomal receptors. Purified plasma membrane and endoplasmic reticulum fractions derived from a rabbit liver microsomal preparation have identical antigenic characteristics with respect to the GH-binding region, indicating that the heterogeneity we describe is not related to receptor processing. Of the three types of GH receptor in the plasma membrane of the rabbit (and possibly rat) we postulate that one (type 1) corresponds to the GH receptor involved in stimulating growth and possesses all of the epitopes studied here. A second (type 2) appears to be identical with the cytosolic 'GH receptor' and lacks the epitope for the anti-(rat GH receptor) MAb in the hormone binding site region. A third (type 3) does not possess the epitope for the inhibitory anti-(rabbit GH receptor) MAb, appears not to bind rat GH and is lost during purification. The availability of type-specific MAbs will facilitate assignment of specific functions to liver receptor subtypes which mediate the multiple functions of GH.
GH receptors have not hitherto been demonstrated or characterized on osteoblasts. In this study we report the characterization of functional GH receptors on the clonal rat osteoblast-like cell line UMR 106.06. The receptors have a typical somatogenic specificity, with high affinity for human GH, 10-fold lower affinity for rat GH, and very poor affinity for rat PRL. The affinity for rat GH is 1.2 +/- 0.4 x 10(9) M-1, and there are approximately 9000 receptors per cell. GH binding increased over several hours when incubations were carried out in serum free minimal essential medium, but binding reduced rapidly when incubations were carried out in Tris-NaCl-Mg++, HEPES, or bicarbonate buffer, suggesting a critical dependence of receptor expression on nutritional factors. Rat GH stimulated proliferation of UMR 106.06 cells in a dose-dependent fashion with a maximum 43 +/- 2% stimulation above control and half-maximal effect at a final hormone concentration of 15 +/- 3 ng/ml. A proliferative response was not observed at low cell density, suggestive of a requirement for a threshold concentration of autocrine mediators or density-dependent receptor expression. A monoclonal antibody (MAb 263) which blocks GH binding to a subset (type 1) of rat hepatic GH receptors did not block binding to osteoblast GH receptors, and did not block the proliferative response. Thus, the proliferative response appears to be mediated by a class of GH receptors not blocked by MAb 263 and possibly related to the type 2 hepatic GH receptors. RNA was extracted from UMR 106.06 cells and a second osteoblast-like cell line UMR 201. Hybridization to a 32P-labeled complementary DNA probe to the rabbit hepatic growth hormone receptor revealed two major labelled bands (3.5 and 1.2 kilobases) and one minor band (2.4 kilobases) in both cell types. In summary, GH receptors are present in clonal osteoblast-like cells, and the receptors mediate a proliferative response to GH. The UMR 106.06 cells provide a valuable model system for studying the mechanism of GH action.
Summary Prognostic information is essential for the evaluation, judgement and optimal treatment of patients with squamous cell cancers (SCCs)
Acromegaly is characterized by coarsening of facial features, acanthosis nigricans, hypertrichosis and oily skin. To determine the site through which GH exerts these effects, we have used immunohistochemistry to localize the GH receptor/binding protein (BP) in rat, rabbit and human skin. Three monoclonal antibodies (MAb 1, 43, 263) were immunoreactive in identical locations, whereas no immunoreactivity was evident when control monoclonal antibodies (MAb 50.8 and MAb 7 (rat] were used. Skin from neonatal and adult animals was used to determine whether GH receptor/BP expression was developmentally regulated. Immunoreactivity of the GH receptor/BP in the three species was consistently localized in the stratum basale and stratum spinosum. Intermittent staining was observed in the stratum granulosum. Scattered basal epidermal cells often displayed more intense immunoreactivity. This distribution was observed at all maturational stages examined. Intense GH receptor/BP immunoreactivity was observed in all histological layers of the lower one-third of hair follicles and in hair matrix cells of the dermal papillae. Immunoreactivity was also detected in the outer epithelial root sheath of the upper two-thirds of hair follicles, in sebaceous glands and in fibroblasts of the connective tissue sheath surrounding the follicle. GH receptor/BP immunoreactivity was also present in the secretory duct and myoepithelial cells of human eccrine sweat glands. Fibroblasts, Schwann cells of peripheral nerve fascicles, skeletal muscle cells and adipocytes of the dermis were also immunoreactive as were medial smooth muscle and endothelial cells of arteries. These results provide evidence that GH acts locally on the epidermis and epidermal appendages concordant with our recent localization of GH receptor/BP to epithelial cell types of the gastrointestinal and reproductive systems.
Growth factors play an important role in the regulation of cell growth, division and differentiation. In this study the distribution and regulation of insulin-like growth factor-I (IGF-I) in the continuously erupting rat incisor was determined by immunohistochemistry. Results were evaluated both visually and with a computer-based image analysis system. The distribution and intensity of IGF-I immunoreactivity varied with developmental stage of the rat incisor. Strong IGF-I immunoreactivity was observed in differentiating odontoblasts and ameloblasts. The most intense immunoreactivity was observed in secretory ameloblasts, secretory odontoblasts and in maturation ameloblasts. Staining was weak or absent in post-secretory ameloblasts but persisted in post-secretory odontoblasts. Weak to moderate immunoreactivity was also seen in cells of the stratum intermedium and in the reduced enamel epithelium. Surrounding alveolar bone showed strong IGF-I immunoreactivity in osteoblasts and in the stratum basale and stratum spinosum of the adjacent labial gingival epithelium. In order to assess the role of GH in IGF-I expression, GH (65 micrograms/100 g bw) was administered for six days to dwarf GH deficient rats, producing a significant increase in body weight (P < 0.01). Measurements at different stages of odontogenesis showed that the staining intensity of secretory ameloblasts (P < 0.01) and maturation ameloblasts (P < 0.001) was significantly different between untreated and treated animals. These results indicate that IGF-I is present in cell populations of the enamel organ of the rat incisor found previously to exhibit growth hormone receptors, and that expression of IGF is GH dependent.(ABSTRACT TRUNCATED AT 250 WORDS)
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
customersupport@researchsolutions.com
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
This site is protected by reCAPTCHA and the Google Privacy Policy and Terms of Service apply.
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.