Transforming growth factor beta induces mesangial cell apoptosis through NO- and p53-dependent and -independent pathways.
BACKGROUND
Because transforming growth factor beta (TGF-beta) 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-beta. Morphologic evaluation of MC apoptosis was performed by staining cells with H-33342 and propidium iodide. To confirm the effect of TGF-beta on MC apoptosis, DNA was extracted from control and TGF-beta-treated MCs and run on gel electrophoresis. We evaluated the effect of NG-nitro-L-arginine methyl ester (L-NAME), a nitric oxide (NO) synthase inhibitor, on TGF-beta-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-beta on MCs derived from p53 knockout mice (p53KO-MC) as well as a normogenic strain (N-MC). We also examined the effect of TGF-beta, 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-beta-treated, and SNAP-treated MCs and probed for the expression of p53.


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


CONCLUSIONS
We hypothesize that TGF-beta promotes MC apoptosis through NO generation and p53-dependent and -independent pathways.
Patel
P
P
Molecular Biology and Experimental Pathology Section, Long Island Jewish Medical Center, New York, NY, USA.
Varghese
E
E
Ding
G
G
Fan
S
S
Kapasi
A
A
Reddy
K
K
Franki
N
N
Nahar
N
N
Singhal
P
P
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D002478:Cells, Cultured; D053938:DNA Fragmentation; D004305:Dose-Response Relationship, Drug; D005920:Glomerular Mesangium / Q000187:drug effects*; D051379:Mice; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000502:physiology*; D016212:Transforming Growth Factor beta / Q000494:pharmacology*; D016159:Tumor Suppressor Protein p53 / Q000502:physiology*
D016212:Transforming Growth Factor beta; D016159:Tumor Suppressor Protein p53; D009569:Nitric Oxide; D019331:NG-Nitroarginine Methyl Ester