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Objective:To investigate overexpression ofαBC will attenuate LPS-induced Pro-inflammatory Response in rat cardiac cells and its related machanisms.Methods:1.H9c2 cells were either untreated or exposed to a mild heat shock(42.5℃for 1h),and recovered at 37℃for 0h,0.5h,1h,6h, 12h,24h.The expression level of Hsp25,αBC,Hsp70,Hsp90 were determined by immunoblot analysis.2.(1)Rat cardiac cell line H9c2 overexpression ofαBC was used in the experiment,normal H9c2 cells served as control.They were stimulated with LPS for 30min、1h and 2h.The unstimulated cells were used as control.At each time point after stimulation,the myoblasts were collected, the protein was extracted,and the activation of IκB was determined.(2) The H9C2 cardiac myoblasts transiently transfected with NFκB-luc、pRL-SV40、PCI or PCI-αBC were challenged with LPS for 2h.And the control groups of unstimulated cells were also established.At each time point,the cells were collected,and the report gene of NF-κB was exanmined.3.Rat cardiac cell line H9c2 overexpression ofαBC was used in the experiment,normal H9c2 cells served as control.They were stimulated with LPS for 30min、1h and 2h.The unstimulated cells were used as control.At each time point after stimulation,the myoblasts were collected, the protein was extracted,and the activation of Akt,and MAPK pathyway were determined.Results:1.H9c2 cells were cultured at 42.5℃for 1h and then recovered at 37℃for different times.Hsp70,Hsp25,αBC were induced a significant increase at 1h,6h,12h,24h for recovery after HSR.While Hsp90 has no obviously change.2.(1)LPS-induced IκB degradation was significant blocked in the cells transfected withαBC gene,compared with the control cells at 0.5h,1h, after LPS stimuation.(2)We also observed that overexpression ofαBC in H9C2 cardiac myoblasts significantly inhibited LPS-induced NFκB activation.3.(1)αBC overexpreesion suppressed MAPK signaling cascade.The phosphorylation levels of ERKs,JNKs,and P38 MAP kinases were obviously increased in control cells,compared with cells overexpressing theαBC gene after LPS stimuation.(2)LPS-induced Akt phosphorilation was augmented byαBC overexpression in H9c2-αBC,in comparison with control groups.(P<0.05)Conclusion:Our findings suggest thatαBC plays an important role in attenuation of LPS-induced Pro-inflammatory Response in rat cardiac cells and the mechanism involves down-regulation of LPS-induced activation of NFκB,MAPK pathway and up-regulation of LPS-induced Akt activation.
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