Autocrine stimulation of AR4-2J rat pancreatic tumor cell growth by glycine-extended gastrin

Abstract

Glycine-extended gastrin (gastrin-Gly) stimulates proliferation of AR4-2) pancreatic tumor cell line through a specific receptor, different from the gastrin-cholecystokinin B receptor. Our purpose was to determine whether AR4-2) cells produced gastrin-Gly and then whether the peptide was involved in an autocrine loop. First, proliferation of AR4-2) cells in serum-free medium was inhibited by a gastrin anti-sense oligodeoxynucleotide phosphorothioate and by antibodies specific for gastrinGly. In contrast, antibodies specific for a-amidated gastrin were without effect. By using RT-PCR, we have shown that AR4-2J cells expressed gastrin mRNA. The presence of gastrin-Gly, but not a-amidated gastrin, in serum-free media was detected by radioimmunoanalysis. Gel chromatography revealed that the predominant molecular forms secreted were glycine-extended gastrin-34 and gastrin-17. Furthermore, epidermal growth factor (EGF), a stimulator of gastrin gene transcription, modulates gastrin-Gly secretion by AR4-2J. These data together suggest that gastrin-Gly is an autocrine growth factor for AR4-2) cells and that it participates with EGF in the regulation of AR4-2)-cell proliferation.o 1996 Wiley-Liss, Inc.The major physiological role of the gastrointestinal hormone, gastrin, is to stimulate gastric-acid secretion. Gastrin can also promote growth effects on normal gastric mucosa, on colorectal tumors in vivo (Winsett et al., 1986), and on colonic (Kuzyk et al., 1986), broncogenic (Sethi and Rozengurt, 1992) and pancreatic cell lines (Seva et al., 1990). Bioactivity of gastrin and several other regulatory peptides requires Cterminal a-amidation using glycine as amide donor (Hiked and Rehfeld, 1986). Carboxyamidation is the final step in the maturation to a bioactive gastrin, which is able to bind its CCK-B gastrin (RCCK-B) receptor with high affinity.Progastrin is expressed in several normal tissues, such as pituitary gland, cerebellum, antrum, vagal neurons and duodenum (Rehfeld, 1986(Rehfeld, , 1991Marino et al., 1994). The tissue concentration of progastrin and processing intermediates is in the same range as that of the mature a-amidated peptide (Sugano et al., 1987). It has been demonstrated that the immediate precursor of amidated gastrin, glycine-extended gastrin (gastrin-Gly), possesses specific biological functions. Thus, like a-amidated gastrin, gastrin-Gly stimulates proliferation of a tumor-derived pancreatic cell line (AR4-2J), through a specific receptor, different from the CCK-B receptor (Seva et al., 1994). Moreover, colorectal (Singh et al., 1994), gastric (Van Solinge and Rehfeld, 1992) and broncogenic tumors (Rehfeld et aL, 1989) contain glycine-extended gastrins, and occasionally at concentrations higher than that of amidated gastrin.A possible mechanism by which tumor cells show unrestricted growth is a response to the growth factors that they produce. An autocrine role for a "gastrin-like" peptide in human colonic tumor cells has already been suggested (Hoosein et al., 1990). More recently, immunologica...

Pathways of processing of the gastrin precursor in rat antral mucosa.

“…There are, however, substantial amounts of G17-Gly generated in rat antrum; the data suggest that this material should be regarded as an end product in its own right. This conclusion is interesting in the context of recent reports that -Gly-extended gastrins might have characteristic biological activities, notably stimulation of cell growth and of proton pump gene expression, independent of those of the amidated gastrins (12)(13)(14).…”

“…However, recent reports indicate that gastrins extended at the COOH terminus by glycine also have biological actions, notably growth. promoting effects on the AR4-2J cell line and stimulation of H+/K+ ATPase a-subunit gene expression, that are distinct from those of the amidated gastrins and appear to be mediated by novel receptors (12)(13)(14). Peptides terminating in gastrins with COOH-terminal glycine (-Gly)' are converted to COOHterminally amidated peptides by the enzyme, peptidyl a-amidating mono-oxygenase (PAM) (15).…”

Gastrin Enhances the Angiogenic Potential of Endothelial Cells via Modulation of Heparin-Binding Epidermal-Like Growth Factor

“…Two gastrin peptides were compared; gastrin-17 (G17), the form involved in endocrine pathways and GlyG17, a form secreted by tumor cells themselves and ascribed a role as an autocrine factor (20). It was confirmed that both G17 and GlyG17 increased tubule formation of human endothelial cells grown on an extracellular matrix and enhanced HB-EGF expression and shedding.…”

The Biological and Therapeutic Importance of Gastrin Gene Expression in Pancreatic Adenocarcinomas

“…In terms of specific gastrin proteins secreted by tumor cells, Goetze et al (7), using a specific radioimmunoassay, confirmed amidated gastrin expression in 14 of 19 carcinomas, whereas a second study detected mainly precursor gastrin forms, progastrin and glycineextended gastrin (8). Glycine-extended gastrin peptides were also shown to be secreted by the rat pancreatic adenocarcinoma cell line AR42J (14).…”