Sphingosine 1‐phosphate modulates apoptosis in cytokine treated pancreatic islet B‐cells

Abstract

Pancreatic islet β-cells undergo apoptosis (APO) following culture with cytokines (CTK) in an in vitro model for type 1 diabetes mellitus. The aim was to determine if sphingosine 1-phosphate (S1P) modulates the biochemical pathways that mediate the β-cell APO response. Isolated rat islets or cells were pretreated with S1P (400 nM) for 5 h prior to and during CTK stimulation for 24-48 h. APO was determined by TUNEL staining. CTK (1 ng/ml interleukin-1β and 5 ng/ml interferon-γ ) induced 28±2% APO in islet cells after 48 h. S1P alone did not affect islet cell APO, however, S1P together with CTK reduced APO to 13±3% (P<0.001 vs. CTK) of total cells. Caspase 3 activity in INS-1e cells was also a measure of APO. CTK increased caspase 3 activity after 24 h to 193±33% of control (C) cells without CTK treatment (P<0.001). The presence of S1P or dihydro-S1P (400 nM) with CTK reduced INS-1e cell caspase 3 activity to 127±15% C (P>0.05) and 105±14% C (P>0.05), respectively; the occurrence of APO cells with S1P alone was 94±7% C (P>0.05). Forskolin (50 nM) reduced CTK-induced caspase 3 activity to 121±5% C (P>0.05). Analysis by real-time PCR showed that the mRNA expression for the anti-APO gene Bcl-xL increased by 337±34% C (P<0.001) in CTK-treated islets, and that S1P augmented the expression to 570±63% C (P<0.01); with S1P alone Bcl-xL mRNA levels were 108±15% C. In conclusion, S1P protects the islet β-cell from cytokine-induced APO to a similar extent as forskolin, and increased expression of Bcl-xL and reduced activity of caspase 3 play a role in the antiapoptotic effects. (Supported by Juvenile Diabetes Research Foundation grant 1-2002-613)