Supplemental Data

Supplemental Data

Files in this Data Supplement:

• Figure S1 - Adenoviral overexpression of SAPK and MKP in MIN6 cells and primary islet cells MIN6 cells (A) and dispersed mouse pancreatic islet cells (B) were infected with adenoviruses integrating LacZ, p38-Flag, JNK-Falg, MKP-1-Flag, or MKP-7-Flag for 12 h. Following infection, cells were cultured in fresh normal media for 24 h. Fixed cells were subjected to immunofluorescence analysis using anti-p38, anti-JNK, and anti-Flag antibodies.
• Figure S2 - Reactive oxygen species (ROS) were generated by LG culture in MIN6 cells MIN6 cells were placed in 25 mM glucose (HG) or 2 mM glucose (LG) for up to 24 h. Intracellular ROS generation was monitored by fluorescence of CM-H₂DCF-DA as in Fig. 1A.
• Figure S3 - Phosphorylation of p38 MAPK is promoted in the LG-cultured MIN6 cells MIN6 cells were incubated in Krebs-Ringer buffer containing 2 mM glucose for 2 h, then exposed to 2 mM glucose (LG) or 25 mM glucose (HG) for an indicated time up to 30 min. The cells were lysed and equal amounts of lysates were subjected to immunoblot analysis with antibodies to the total protein and the phosphorylated form of three MAPKs: ERK, p38, and JNK. Experiments were repeated three times with reproducible results.
• Figure S4 - Effect of long-term LG or HG culture on the expression of MKPs A. MIN6 cells were cultured in LG or HG for 24 h. Total cell RNAs were prepared and subjected to RT-PCR analysis for Wip-1, VHR, MKP-1, MKP-3, MKP-7, and MKP-5. B. MIN6 cells were cultured in LG for an indicated time up to 36 h. Cell lysates were analyzed by immunoblotting with antibodies to MKP-1, MKP-3, or MKP-7. Experiments were repeated three times with reproducible results.