Effect of 2‐mercaptoethanol on glutathione levels, cystine uptake and insulin secretion in insulin‐secreting cells

Abstract

The role of glutathione (GSH) in the differentiated state of insulin‐secreting cells was studied using 2‐mercaptoethanol as a means of varying intracellular GSH levels. 2‐Mercaptoethanol (50 μM) caused a marked increase of GSH in two rat insulinoma cell lines, RINm5F and INS‐1, the latter being dependent on the presence of 2‐mercaptoethanol for survival in tissue culture. The effect of 2‐mercaptoethanol on GSH was shared by other thiol compounds. Since in other cell types 2‐mercaptoethanol is thought to act on cystine transport, thereby increasing the supply of cysteine for GSH synthesis, we have studied [35S]cystine‐uptake in INS‐1 cells. At equimolar concentrations to cystine, 2‐mercaptoethanol caused stimulation of [35S]cystine‐uptake. The effect persisted in the absence of extracellular Na+, probably suggesting the involvement of the Xc− carrier system. INS‐1 cells with a high GSH level, cultured 48 h with 2‐mercaptoethanol, displayed a lower cystine uptake than control cells with a low GSH content. The effect of variations of the GSH levels on short‐term insulin release was studied. No alteration of glyceraldehyde‐induced or KCl‐induced insulin release in RINm5F cells was detected. In contrast, both in islets and in INS‐1 cells, a high GSH level was associated with a slighly lower insulin release. In INS‐1 cells the effect was more marked at low glucose concentrations, resulting in an improved stimulation of insulin secretion. On the other hand, in islets, a decrease in the incremental insulin release evoked by glucose was seen. As in other cell types, oxidized glutathione (GSSG) was less than 5% of total GSH, and in INS‐1 cells no change in the GSH/GSSG ratio was detected during glucose‐induced or 3‐isobutyl‐1‐methylxanthine‐induced insulin release. In conclusion, 2‐mercaptoethanol‐dependent INS‐1 cells, as well as RINm5F cells and islets of Langerhans, display a low capacity in maintaining intracellular levels of GSH in tissue culture without extracellular thiol supplementation; 2‐mercaptoethanol possibly acts by promoting cyst(e)ine transport; changes in GSH levels caused a moderate effect on the differentiated function of insulin‐secreting cells. Copyright © 1992, Wiley Blackwell. All rights reserved