Mechanisms of somatostatin action in RINm5F cells in culture: Preliminary evidence for possible altered G protein function

Abstract

Octreotide (SMS), a somatostatin analogue, is an established antigrowth peptide, but it does not effectively inhibit the growth of insulinoma cells. In order to study the mechanisms that underlie this apparent lack of an antiproliferative effect on insulinoma tumor cells we established the rat insulinoma cell line, RINm5F, in culture. Cells in culture were tested by incubation in media with and without SMS. To study tritiated [3H]-thymidine incorporation into extracted DNA (TTID), 2 [mu]Ci/well of 3H was added for 24 hr, and cells were harvested and assayed for TTID (cpm/[mu]g DNA). Insulin (IRI) and intracellular cAMP (cAMPi) were measured by RIA. To study the effects of SMS on insulin secretion, conditioned media were sampled after 24 hr. To study the effects of cAMP1, conditioned medium was used to extract cAMPi, following incubation with SMS for 15 min. Increasing concentrations of SMS had no significant effect on TTID in the presence of 1% FBS. Trypan blue exclusion tests showed>90% viable cells throughout all stages of these experiments. There were no significant differences in cell numbers and protein content in the presence of SMS. There was a significant decrease in the secretion of insulin and intracellular cAMP levels in response to 50 nM SMS. However, SMS significantly inhibited TTID in RINm5F cells following a 4-hr pretreatment with pertussis toxin (PT) (23553 +/- 1747 vs 20635 [cpm/[mu]g DNA] +/- 1983 [SEM], P < 0.01). We conclude that the inhibition of insulin secretion by SMS is associated with an attenuation of CAMP formation. However, the effects of SMS on cell proliferation appear to be distinct and more complex. The unexpected ability of PT to induce the inhibition of TTID by SMS suggests that SMS mediates, at least some of, its effects on cell proliferation in RINm5F cells via a possibly altered G protein or related signal transducing mechanism. Further study of the signal transduction system in RINm5F cells may elucidate the mechanisms of tumor growth in insulinoma.