Cyclosporine A and NAC on the inducible nitric oxide synthase expression and nitric oxide synthesis in rat renal artery cultured cells.
BACKGROUND
The immunosuppressor cyclosporine A (CsA) presents the nephrotoxicity as its major side effect that is mostly attributed to a renal vasoconstriction. This may be due to an excessive generation of vasoconstrictors like reactive oxygen species (ROS), or due to a reduction of vasodilators such as the nitric oxide, which in turn, can be caused by increased amounts of ROS. We evaluated the effect of CsA and the antioxidant N-acetylcysteine (NAC) on inducible nitric oxide synthase (iNOS) mRNA expression and nitric oxide synthesis, in rat renal artery vascular smooth muscle cells (rVSMCs) primary culture.


METHODS
In cells treated during 72 hours with CsA (10 microg/mL), its vehicle (control) (10 microL/mL), Escherichia coli lipopolysaccharide (LPS) (100 microg/mL), CsA + LPS, NAC (6.13 mmol/L), or CsA + NAC, we determined the nitric oxide synthesis (Griess and chemiluminescence methods), iNOS expression [reverse transcription-polymerase chain reaction (RT-PCR)] and cell viability (acridine orange method).


RESULTS
In rVSMCs, LPS increased nitric oxide and iNOS expression; CsA decreased basal and LPS-induced nitric oxide and iNOS expression; NAC increased nitric oxide and blunted the nitric oxide reduction caused by CsA, with no effect on iNOS. CsA reduced cell viability.


CONCLUSIONS
In this study, CsA reduced nitric oxide synthesis in rVSMCs, both through iNOS down-regulation and reduction of cell viability, which could be responsible for the vasoconstrictive effect of the CsA. In the effect of CsA on nitric oxide, probably a role is also played by free radical production, as this effect was blunted by NAC.
Lessio
Camila
C
Nephrology Division, Universidade Federal de São Paulo-Escola Paulista de Medicina, Sao Paulo, Brazil.
de Assunção Silva
Fábio
F
Glória
Maria Aparecida
MA
Di Tommaso
Ana Beatriz Galhardi
AB
Gori Mouro
Margaret
M
Di Marco
Giovana Seno
GS
Schor
Nestor
N
Higa
Elisa Mieko Suemitsu
EM
D000111:Acetylcysteine / Q000494:pharmacology*; D000818:Animals; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D016572:Cyclosporine / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D007166:Immunosuppressive Agents / Q000494:pharmacology*; D049449:Luminescence; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D009569:Nitric Oxide / Q000096:biosynthesis / Q000378:metabolism*; D052247:Nitric Oxide Synthase Type II / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017208:Rats, Wistar; D012077:Renal Artery / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology
D007166:Immunosuppressive Agents; D012333:RNA, Messenger; D009569:Nitric Oxide; D016572:Cyclosporine; D052247:Nitric Oxide Synthase Type II; C496318:Nos2 protein, rat; D000111:Acetylcysteine