Molecular biological studies on the roles of NFkappaBETA and NF-IL6 in the experimental glomerulonephritis and diabetic nephropathy.

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To reveal the effcnts of cytokines on the cultured mesangial cells, we examined the interactions between endothelin, IL-1beta, TNF-alpha, and lipopolysaccharide. It was shown that endothelin suppress the induction of NO synthase by these cytokines, which are known to exert their action via NFkappaBETA. We have also made a vector, which has 6 NFkappaBETA binding cite in the upstream of luciferase, and transfected it to mesangial cells. In these transfected cells, angiotensin II induced 6-fold increase in luciferase activity, suggesting that angiotensin II activates NFkappaBETA. From these results, it was suggested that NFkappaBETA plays an important role in the effects of cytokines on mesangial cells, next we made mutated IkappaBETA. This IkappaBETA lacks phosphorilation cite, which is necessary for the atctivation of NFkappaBETA. Therefore, the overexpression of mutated IkappaBETA should abolish the effects of cytokines. In the mesangial cells with the adenovirus-mediated transfection of mutated IkappaBETA, TNF-alpha showed enhanced induction of apoptosis, compared with wild type cells. It was also shown that induction of apoptosis was enhanced with the transfection of mutated IkappaBETA by IL-1 and lipopolysaccharides. From these results, it was shown that NFkappaBETA was a suppressor of apoptosis. Next, we tried to infect the adenovirus to the kidneys for the transfection of mutated IkappaBETA, although the injected adenovirus to the renal artery infected to only proximal tubules. Currently, we are developing a technique for the in vivo transfer of the mutated IkappaBETA to the kidney.