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...

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