Transforming Growth Factor Beta Induces Mesangial Cell Apoptosis through NO- and p53-Dependent and -Independent Pathways

Abstract

Background

Because transforming growth factor beta (TGF-β) has been shown to have a bimodal effect on mesangial cell (MC) proliferation, we studied its effect on MC apoptosis.

Methods

Cultured mouse MCs were used to evaluate the effect of TGF-β. Morphologic evaluation of MC apoptosis was performed by staining cells with H-33342 and propidium iodide. To confirm the effect of TGF-β on MC apoptosis, DNA was extracted from control and TGF-β-treated MCs and run on gel electrophoresis. We evaluated the effect of N^G-nitro-L-arginine methyl ester (L-NAME), a nitric oxide (NO) synthase inhibitor, on TGF-β-induced MC apoptosis to determine the role of NO and studied the effect of sodium nitroprusside (SNP) and SNAP (S-nitroso-N-acetyl-penicillamine) on MC apoptosis to confirm the effect of NO. We examined the role of p53 by studying the effect of TGF-β on MCs derived from p53 knockout mice (p53KO-MC) as well as a normogenic strain (N-MC). We also examined the effect of TGF-β, SNP, and SNAP on apoptosis of p53 mutant (MDAMB-231) and wild-type p53 (MCF-7) breast cancer cell lines. In addition, Western blots were generated from control, TGF-β-treated, and SNAP-treated MCs and probed for the expression of p53.

Results

TGF-β promoted MC apoptosis. Moreover, TGF-β-treated MCs displayed integer multiples of 180 base pairs (ladder pattern). L-NAME inhibited TGF-β-induced MC apoptosis. Furthermore, SNP and SNAP, NO donors, promoted MC apoptosis. TGF-β also enhanced the MC expression of p53. TGF-β induced only a moderate degree of apoptosis in MCs derived from p53KO-MC when compared with N-MCs. Similarly, the TGF-β-induced apoptosis of MDAMB-231 was of a moderate degree when compared with MCF-7 cells.

Conclusions

We hypothesize that TGF-β promotes MC apoptosis through NO generation and p53-dependent and -independent pathways.