Roles of MAPKs in cHSP60-induced eNOS downregulation in HCAECs. (A) MAPK phosphorylation was determined by Bio-Plex luminex immunoassay. HCAECs were treated with cHSP60 (2 µg/mL) for different times (5, 10, 20, 30, 45, 60, and 90 min). (B) The eNOS mRNA levels were determined by real-time PCR for cHSP60 and MAPK chemical inhibitor treatment groups. HCAECs were treated with cHSP60 (2 µg/mL) in the presence or absence of inhibitors of p38, JNK, or ERK for 24 h. (C) The eNOS mRNA levels were determined by real-time PCR for cHSP60 and MAPK dominant-negative mutant (DN) adenovirus (AD) transfection groups. HCAECs were infected with AD-GFP (control), AD-p38-DN, AD-JNK-DN, or AD-ERK2-DN for 24 h and then treated with cHSP60 (2 µg/mL) for additional 24 h. *P < 0.05, n = 3, t-test.