The various response to Jo two observed in ILK KO and con trol mice may be attributable in component to reduced hepa tic expression of Fas receptor, since the basal levels of Fas as determined by Western blotting was lower within the livers on the ILK KO liver. The expres sion was also reduce inside the hepatocytes isolated from ILK KO mice when compared with WT mice. As a result, it can be probably that ILK regulates the expression of Fas receptor. Similarly, TUNEL assay on the liver sections demon strated additional abundant apoptotic nuclei in handle mice than in ILK KO mice. Activation of capase3 7 was also higher inside the handle mice than ILK KO mice at 6 and 12 h just after Jo two administration. Moreover, expression of cleaved caspase 3 and PARP have been also greater in the handle than the ILK KO mice at each 6 and 12 h just after a sublethal dose of Jo two.
Mechanism of protection of ILK KO mice against Jo 2 induced hepatic failure We looked at the protein expression of many anti apop totic proteins involved in Fas induced apoptosis. Bcl two loved ones proteins inhibit apoptosis induced by range of sti muli, including Fas mediated apoptosis. We assessed the expression in the antiapoptotic protein Bcl xL and Bcl two by FTY720 solubility Western blotting at 0, 6 and 12 h soon after the injection of sublethal dose of anti Fas antibody. Bcl xL and Bcl 2 proteins levels had been decreased inside the liver of control mice treated with Jo2, having said that, in ILK KO mice Bcl xl and Bcl two protein levels have been main tain in response to a sublethal dose of Jo 2. The ILK KO mice also had greater expression of Bcl 2 at basal levels.
We also looked at the protein expression of Bcl 2 related death promoter soon after Jo two administration. Dephosphorylated Terrible forms a heterodimer with Bcl 2 and Bcl xl, inactivating them, and hence allowing Fas triggered apoptosis to take spot. Poor phosphorylation is hence Hesperadin anti apoptotic, and Bad dephosphorylation is pro apoptotic. In the handle mice the Terrible levels didn’t alter before and following Jo 2 administration but there was an induction of Poor just after Jo two administration inside the ILK KO mice. The expression of p Negative which is antiapoptotic was larger in the ILK KO mice after JO 2 administration as com pared to the manage mice. The basal level of p Undesirable was also greater in the ILK KO mice as compared to the con trols. Expression of p Negative in control was barely detectable at basal levels. To understand the molecular events underlying the resistance of ILK KO mice to Jo 2 induced apoptosis, we examined the activation of many survival pathways known to be involved in cytoprotection against Fas induced apoptosis. We investigated phosphorylation of Akt, Erk1 two, and NF B activation that are known to become involved in cytoprotection against Fas induced apop tosis.