Inhibition of nitric oxide synthase ameliorates cellular injury in sickle cell mouse kidneys

Background. In previous studies of transgenic sickle cell mice, increased r enal expression of inducible nitric oxide syn thase (iNOS) and endothelial cell isoform of NOS (EcNOS) was found by Western blot and immunohistochemis try. In addition, putative evidence of peroxynitrite (ONOO-) formation was found in the form of positive immunostaining and immunoblot for nitrotyrosi ne. Apoptosis was also detected by DNA strand breakage and TUNEL assay. The present study was carried out to examine the role of NO/ONOO- in mediating renal tubular cell apoptosis in sickle cell mouse kidneys. Methods. Mercaptoethylguanidine (MEG), a compound that selectively inhibits iNOS and also is a scavenger of ONOO-, was administered intraperitoneally over a five-day period to control and beta(s) mice. Immunohistochemistry of iNOS and nitrotyrosine, DNA electrophoresis, ApoTACS assay for apoptosis, and Western blot of poly(ADP-ribose) polymerase (PARP) were carried out. Results. MEG administration virtually eliminated renal im munostaining of i NOS and nitrotyrosine and prevented DNA strand breakage. In addition, Weste rn blot analysis of PARP, a nuclear DNA-reparative enzyme activated in resp onse to DNA strand breakage, was found to be cleavaged in hypoxic beta(s) m ice, but was partially protected in MEG-treated beta(s) hypoxic mice. Final ly, apoptosis was markedly reduced by MEG in beta(s) hypoxic mice. Conclusions. These observations provide evidence that NO and/or ONOO- are r esponsible for initiating cell damage, which leads to apoptosis in sickle c ell mouse kidneys.