This study examined the expression of receptors of the bombesin (BBS) family in human gastric-cancer cell lines. Of 5 cell lines screened, only one, St42, demonstrated specific binding sites for 125I-Tyr4-BBS, which have been further characterized. This binding was saturable, and temperature- and time-dependent. Scatchard analysis of displacement data performed at 37 degrees C revealed 2 binding sites: a high-affinity, low-capacity site (KD = 0.13 nM, Bmax = 1500 sites/cell) and a lower-affinity, higher-capacity site (KD = 11 nM, Bmax = 35,000 sites/cell); the latter was lost when internalization of peptide was prevented, suggesting that it may be an artefact. Displacement assays with gastrin-releasing peptide (GRP) and neuromedin B (NMB) revealed that the receptor was of the GRP-preferring sub-type (GRP IC50 = 0.35 nM; NMB IC50 = 112 nM). Co-valent cross-linking of 125I-Tyr4-BBS to the receptor demonstrated the presence of a single band corresponding to a molecular weight of 37 to 44 kDa on SDS-PAGE, similar to that of the cloned GRP receptor protein core. G-protein linkage of this receptor was demonstrated by selective inhibition of 125I-Tyr4-BBS binding by guanosine nucleotides. The binding of BBS to the receptor resulted in a rise in intracellular calcium. Three of four structurally distinct BBS antagonists bound to the receptor with high affinity, but [DPhe12, Leu14]-bombesin did not cause any displacement of 125I-Tyr4-BBS even at 10 mM. The functional significance of GRP receptors on human gastric-cancer cells is as yet unknown, but further studies may determine whether such receptors have importance in the therapy of gastric cancer.
This study characterises the somatostatin binding site in human gastrointestinal cancer and mucosa in terms of cationic specificity and relative affinity for three somatostatin analogues. Competitive displacement assays were performed on plasma membranes from human gastric and colonic tissues using radiolabelied somatostatin-14 as ligand. Comparison was made with the somatostatin binding site in rat cerebral cortex. In gastrointestinal tissue, magnesium decreased and sodium increased specific binding. By contrast, in rat cerebral cortex, the converse cationic effect was seen. These changes resulted from alterations in receptor density, with no change in receptor affinity. Displacement studies were then performed with somatostatin-14 and somatostatin analogues RC-160, somatuline, and octreotide. RC-160 and somatuline displaced radiolabel from binding sites in gastric and colonic cancer and mucosa with 10-fold lower affinity than the native peptide. Octreotide did not displace radioligand in gastric or colonic cancer at any concentration tested. By contrast, in rat cortex, although all three analogues displaced with a lower affinity than the native peptide, there was no difference between analogues. These data suggest a distinct somatostatin receptor subtype in gastrointestinal tissues.
Summary Somatostatin is a regulatory peptide implicated in the control of cellular proliferation in epithelial tissues and this regulation may occur directly via membrane bound receptor activation. The aim of this study was to investigate somatostatin binding in human gastrointestinal cancer and normal mucosa. Plasma membranes were prepared from specimens of tumour and normal mucosa from 51 patients undergoing surgical resection for malignancy (28 gastric, 23 (Horowitz et al., 1975;Higgins et al., 1941;Crile, 1957;Osei et al., 1985). There is, however, no currently defined role for endocrine therapy in non-endocrine gastrointestinal cancer. Surgical intervention is the most effective treatment for this condition and, even then, can only afford a 'cure' in early disease. Advanced tumours are associated with a poor prognosis, marginally improved by chemotherapeutic regimens. Any benefit from systemic therapy is usually abrogated by systemic toxicity (Rake et al., 1979;Engstrom et al., 1985;Bleiberg, 1990). For this reason a non-toxic, effective, treatment modality would be of value to improve the outlook for those patients with advanced disease.There is increasing evidence that tumours arising from the gastrointestinal tract are, at least in part, hormone dependent. Numerous hormones have now been implicated in the pathogenesis and development of gastrointestinal malignancy, including gastrin, epidermal growth factor, enteropancreatic hormones and oestrogenic steroids (Sirinek et al., 1985;Watson et al., 1988;Li et al., 1980;Howatson & Carter, 1985;McMichael et al., 1980). These findings, together with the observation that transformed gut epithelial cells may retain functional hormone receptors , have suggested a role for hormonal manipulation in these malignancies.Somatostatin and its analogues are good candidates for use as endocrine agents in the treatment of gastrointestinal cancer. The native peptide is widely distributed in the body and, amongst its many inhibitory actions, has a putative role as an anti-proliferative agent in both normal (Lehy et al., 1979) and malignant tissue. The mechanisms involved in this anti-proliferative action have not, as yet, been confirmed. Both the native peptide and its analogues act at the somatotrophs of the anterior pituitary suppressing growth hormone production and release (Adrian et al., 1981). This may influence the proliferation of target tissues directly or may result in a substantial reduction in local growth factor release (Kirkegaard et al., 1984 (Reubi et al., 1987a;Reubi et al., 1987b;Reubi et al., 1990;Ikuyama et al., 1985). Somatostatin (1981). In brief, frozen tumour and mucosal samples were mechanically pulverised and homogenised on ice in homogenising buffer (sucrose 250 mM, KCI 25 mM and MgCl2 10 mM in Tris-HCI 50 mM; pH 7.4) at 10,000 r.p.m. in short bursts for 2 min using an Ultraturrax T25 homogeniser (Scientific
Interleukin-10 gene expression correlates with the fall in monocyte HLA-DR antigen expression in patients undergoing major abdominal surgery and may account for the immunosuppression associated with surgical injury.
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