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Improved Rodent Model of Myocardial Ischemia and Reperfusion Injury
Published on: March 7, 2022
Activation of ASK1, downstream MAPKK and MAPK isoforms during cardiac ischaemia
Stephen J Harding1, Gareth J Browne, Bryan W Miller
1Department of Biochemistry, Henry Wellcome Building, University of Leicester, Leicester, UK.
Insights
Cardiac ischemia activates p38 mitogen-activated protein kinase (MAPK) via ASK1 and MKK6, not MKK4 or JNK pathways. This reveals specific signaling during heart attacks, aiding therapeutic development.
Area of Science:
- Cardiovascular Research
- Cell Signaling
- Molecular Biology
Background:
- p38 MAPK is activated during cardiac ischemia, but its upstream activators remain unclear.
- Understanding these pathways is crucial for developing targeted therapies for ischemic heart conditions.
Purpose of the Study:
- To investigate the signaling pathways upstream of p38 activation during global cardiac ischemia in an isolated perfused rat heart model.
- To elucidate the specific roles of MAPKKs (MKK3, MKK4, MKK6) and MAPKKKs (ASK1) in p38 activation during ischemia.
Main Methods:
- Utilized an isolated perfused rat heart model subjected to global ischemia.
- Measured the activation of p38 MAPK, JNK pathway, MAPKKs (MKK3, MKK4, MKK6), and MAPKKK (ASK1) using direct assays.
Main Results:
- Ischemia potently activated p38alpha but not the JNK pathway.
- MKK3 and MKK6 were activated, while MKK4 and MKK7 showed no activation, indicating MKK4 is not a key activator in this context.
- Apoptosis Signal-regulating Kinase 1 (ASK1) was activated late during ischemia, preferentially activating MKK6, which then activated p38.
- ASK1 activation during ischemia preferentially targeted the p38 pathway over the JNK pathway.
Conclusions:
- Cardiac ischemia activates p38 MAPK primarily through the ASK1-MKK6 axis, demonstrating pathway specificity.
- The findings highlight distinct activation patterns and physiological roles for MKK3 and MKK6.
- This study provides a clearer understanding of MAPK signaling complexity in the ischemic heart, essential for future therapeutic strategies.
Abstract:
p38 MAPK is activated potently during cardiac ischaemia, although the precise mechanism by which it is activated is unclear. We used the isolated perfused rat heart to investigate the signalling pathways activated upstream of p38 during global cardiac ischaemia. Ischaemia strongly activated p38alpha but not the JNK pathway. The MAPKKs, MKK3, MKK4 and MKK6 have previously been identified as potential upstream activators of p38; however, in the ischaemic perfused heart, we saw activation of MKK3 and MKK6 but not MKK4. MKK3 and MKK6 showed different temporal patterns of activity, indicating distinct modes of activation and physiological function. Consistent with a lack of JNK activation, we saw no activation of MKK4 or MKK7 at any time point during ischaemia. A lack of MKK4 activation indicates, at least in the ischaemic heart, that MKK4 is not a physiologically relevant activator of p38. The MAPKKK, ASK1, was strongly activated late during ischaemia, with a similar time course to that of MKK6 and in ischaemic neonatal cardiac myocytes ASK1 expression preferentially activated MKK6 rather than MKK3. These observations suggest that during ischaemia ASK1 is coupled to p38 activation primarily via MKK6. Potent activation of ASK1 during ischaemia without JNK activation shows that during cardiac ischaemia, ASK1 preferentially activates the p38 pathway. These results demonstrate a specificity of responses seldom seen in previous studies and illustrate the benefits of using direct assays in intact tissues responding to physiologically relevant stimuli to unravel the complexities of MAPK signalling.
