Abstract
Hepatoprotection mediated by free radical scavenging molecules such as dimethyl sulfoxide (Me(2)SO) arose the question as to whether this effect involved one or several anti-apoptotic signals. Here, using primary cultures of rat hepatocytes and in vivo thioacetamide-induced liver failure, we showed that Me(2)SO failed to prevent any cleavage of initiator caspase-8 and -9 but constantly inhibited procaspase-3 maturation and apoptosis execution, pointing to an efficient inhibition of cleaved initiator caspase activities. Evidence was recently provided that apoptosis might require both caspase and ASK1/JNK-p38 activities. We demonstrated that this kinase pathway was strongly inhibited in the presence of Me(2)SO whereas overexpression of ASK1 was able to restore caspase-3 activity and apoptosis. Interestingly, we also found that GST M1/2 and GST Alpha1/2 dropped under apoptotic conditions; furthermore transfection of GST M1, A1, or P1 to cells overexpressing ASK1, abolished caspase-3 activity and restored viability. This role of GSTs was further assessed by showing that their high expression level was tightly associated with inhibition of ASK1 activity in Me(2)SO-protected hepatocytes. Together, these results demonstrate that Me(2)SO-mediated hepatoprotection involves a dual inhibition of cleaved initiator caspase and ASK1/JNK-p38 activities. Furthermore, in highlighting the control of apoptosis by GSTs, these data provide new insights for analyzing the complex mechanisms of hepatoprotection.
Highlights
Control of apoptosis is essential to development and homeostasis in mammals
Appearance of cleaved forms of caspases was concomitant with the proteolytic cleavage of the endogenous substrate poly-ADP-ribose polymerase (PARP)
As in untreated cells, cleaved caspase-3 and -9 and PARP were found in TGF1treated cells but no cleaved caspase-8 was detected (Fig. 1C)
Summary
Control of apoptosis is essential to development and homeostasis in mammals. Progression of the apoptotic signal can be inhibited at different levels depending on death inducers and cell types. Addition of Me2SO to Basal Hepatocyte Culture Constantly Prevents Maturation of Executioner Caspase-3 and Blocks Late Stage of Apoptosis—To determine the mechanism involved in the inhibition of apoptosis by Me2SO, 2% Me2SO was added to the basal medium 24 h after plating according to Isom et al [35]; cell morphology, DNA degradation, and expression and activation of the three caspases were studied (Fig. 2).
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