Expression of several growth factors and their receptor genes in human colon carcinomas

We have examined the expression of mRNAs for epidermal growth factor (EGF), transforming growth factor-alpha (TGF-α), EGF receptor (EGFR), PDGF-A chain (PDGFA), PDGF-B chain (PDGFB) and PDGF receptor (PDGFR) genes in seven human colorectal carcinoma cell lines and 18 human colorectal carcinomas. In surgically resected specimens of the 18 colorectal tumors, TGF-α, EGFR, PDGFA, PDGFB and PDGFR mRNAs were detected at various levels in 15 (83%), 9 (50%), 18 (100%), 8 (44%) and 12 (67%), respectively. They were also detected in normal tissues. Interestingly, EGF mRNA was detected in only five (28%) of the tumors, but not in normal mucosa. Expression of EGF was also confirmed immunohistochemically in tumor cells. Of the five tumors expressing EGF, four expressed EGFR mRNA and showed a tendency to invade veins and lymphatics. All the colorectal carcinoma cell lines expressed TGF-α mRNA, and five cell lines expressed EGFR mRNA simultaneously. Production of TGF-α protein by DLD-1 and CoLo320DM cells was confirmed by TGF-α specific monoclonal antibody binding assay. The spontaneous³H-thymidine uptake by DLD-1 was suppressed by an anti-TGF-α monoclonal antibody. PDGFA and PDGFB mRNA were also expressed in four cell lines, but PDGFR and EGF mRNA was not detected. These results suggest that human colorectal carcinomas express multi-loops of growth factors and that TGF-α produced by tumor cells functions as an autocrine growth factor in human colonic carcinoma.     

Secretion of a new growth factor, smooth muscle cell derived growth factor, distinct from platelet derived growth factor by cultured rabbit aortic smooth muscle cells

In attempts to determine the mechanism of proliferation of arterial smooth muscle cells (SMC) in intimal atheromatous lesions, autocrine secretion of growth factors by SMC has recently received much attention. Here we report a new growth factor named smooth muscle cell derived growth factor (SDGF). Cultured rabbit medial SMC secreted SDGF for 1 week during their incubation in serum-free media only after at least 4 passages. SDGF differed from platelet derived growth factor (PDGF) physicochemically, immunologically, and biologically. The properties of SDGF also seemed different from those of other known growth factors that stimulate the proliferation of mesenchymal cells.     

Distribution of EGF and its receptor in growing red deer antler

Autografts of the osteogenic part of early antler buds placed elsewhere on the skull have been shown by others to give rise to an antler at the site of grafting. This antler becomes covered in velvet skin, is shed at the end of the growing season and will regrow the following year. Thus, it can be concluded that the nature of antler velvet skin is primarily determined by the underlying osteogenic antler tissue to which it is attached. We hypothesise that a paracrine mechanism operates here and is central to communication between the antler osseous compartment and the integument. A signalling system comprising epidermal growth factor (EGF) and its receptor (EGFR) is known to be expressed in osteogenic cells and to play an important role in skin development and growth. This system may therefore play a significant role in determining the nature and speed of growth of velvet skin via paracrine signalling from osteogenic tissue. We have used bright-field microscope immunohistochemistry to determine the distribution of EGF and its receptor in developing red deer antler osseous compartment and integument. EGF was localized throughout the epidermis and epidermal appendages, in cells of the mesenchyme, in chondrocytes, and in cells of the osteoblastic lineage, including osteoprogenitor cells, osteoblasts and osteocytes. There was strong evidence supporting nuclear and nucleolar staining in sebaceous glands and in keratinocytes. The EGFR was similarly expressed in mesenchyme, chondrocytes and osteoblasts. In skin, the distribution of the EGFR was more localized, being expressed strongly in the deeper cells of the epidermis but not in superficial layers, and was absent from nuclei of cells of the epidermis and its appendages. We conclude that this signalling system is widely distributed in growing antler in a manner which suggests it is predominantly autocrine. No clear-cut evidence for paracrine signalling pathways for this system in either integument or osseous compartments was found. The pattern of distribution of the EGFR in the integument was similar to that seen by others in adult human skin. By contrast, in developing antler osseocartilage, the patterns of distribution were similar to those seen in rodent fetal bone. We conclude that antler consists of rapidly growing fetal osseocartilage overlayed by mature velvet.     

Autocrine anti-apoptotic and proliferative effects of insulin in pancreatic beta-cells

Insulin and glucose inhibited apoptosis in the MIN6 insulin-secreting cell line. The protective effect of 25 mM glucose was prevented by an anti-insulin antibody and this antibody-induced increase in apoptosis was reversed by the presence of excess insulin. Glucose stimulated MIN6 cell proliferation and this was inhibited by blockade of insulin secretion, by an anti-insulin antibody and by phosphatidylinositol-3 kinase (PI-3K) inhibition. Furthermore, MIN6 cell proliferation was stimulated by depolarising concentrations of KCl and by insulin itself. These data indicate that insulin secreted by β-cells in response to elevated glucose exerts autocrine effects to protect against apoptosis and stimulate proliferation, and suggest that the insulin signalling cascade, through the PI-3K pathway, may be an effective means of maintaining β-cell mass in diabetes.     

G proteins and autocrine signaling differentially regulate gonadotropin subunit expression in pituitary gonadotrope

Gonadotropin-releasing hormone (GnRH) acts at gonadotropes to direct the synthesis of the gonadotropins, follicle-stimulating hormone (FSH), and luteinizing hormone (LH). The frequency of GnRH pulses determines the pattern of gonadotropin synthesis. Several hypotheses for how the gonadotrope decodes GnRH frequency to regulate gonadotropin subunit genes differentially have been proposed. However, key regulators and underlying mechanisms remain uncertain. We investigated the role of individual G proteins by perturbations using siRNA or bacterial toxins. In LβT2 gonadotrope cells, FSHβ gene induction depended predominantly on Gα(q/11), whereas LHβ expression depended on Gα(s). Specifically reducing Gα(s) signaling also disinhibited FSHβ expression, suggesting the presence of a Gα(s)-dependent signal that suppressed FSH biosynthesis. The presence of secreted factors influencing FSHβ expression levels was tested by studying the effects of conditioned media from Gα(s) knockdown and cholera toxin-treated cells on FSHβ expression. These studies and related Transwell culture experiments implicate Gα(s)-dependent secreted factors in regulating both FSHβ and LHβ gene expression. siRNA studies identify inhibinα as a Gα(s)-dependent GnRH-induced autocrine regulatory factor that contributes to feedback suppression of FSHβ expression. These results uncover differential regulation of the gonadotropin genes by Gα(q/11) and by Gα(s) and implicate autocrine and gonadotrope-gonadotrope paracrine regulatory loops in the differential induction of gonadotropin genes.     

自分泌运动因子受体在大鼠胃肠的定位

目的 研究自分泌运动因子受体在大鼠胃肠的表达及分布特点。方法应用免疫组织化学方法染色。结果在大鼠胃壁内有自分泌运动因子受体的表达,免疫反应产物主要定位于胃底腺,阳性细胞胞体大,胞核呈阴性,小肠和大肠组织均呈阴性反应。结论正常大鼠胃底腺有自分泌运动因子受体的表达,提示可能参与胃腺的多种生理活动。     

An in situ evidence for autocrine function of NO in the vasculature

The concept of endothelium derived relaxing factor (EDRF) implies that nitric oxide (NO) generated by NO synthase in the endothelium diffuses to the underlying vascular smooth muscle cells (VSMC) modulating thereby vascular tone. VSMC were regarded as passive recipients of NO from endothelial cells. However, this paradigm of a paracrine function of NO became currently subject to considerable debate. To address this issue, we examined the localization of enzymes engaged in l-arginine-NO-cGMP signaling in the rat blood vessels. Employing multiple immunocytochemical labeling complemented with signal amplification, electron microscopy, Western blotting, and RT-PCR, we found that NO synthase was differentially expressed in blood vessels depending on the blood vessel type. Moreover, the expression pattern of NO synthase in VSMC showed striking parallels with arginase and soluble guanylyl cyclase. Our findings challenge the commonly accepted view that the expression of NO synthase is restricted to vascular endothelial cells and lends further support to an alternative mechanism, by which constitutive local NOS expression in VSMC may modulate vascular functions in an endothelium-independent manner. Moreover, the co-expression of enzymes engaged in l-arginine-NO-cGMP signaling (NO synthase, arginase, and soluble guanylyl cyclase) in VSMC is indicative of an autocrine fashion of NO signaling in the vasculature in addition to the paracrine role of NO generated in the endothelium.