An interleukin-6-neutralizing antibody prevents cyclosporine-induced nephrotoxicity in mice.
BACKGROUND
Chronic use of cyclosporine A (CyA) induces nephrotoxicity primarily due to endothelial dysfunction. In our previous studies, potential mechanisms were identified in vitro and implicated nicotinamide adenine dinucleotide phosphate (NADPH) oxidase and interleukin-6 (IL-6) as key components in causing endothelial dysfunction. In this study, we tested the hypothesis that NADPH oxidase activity and IL-6 are key components in renal damage in an in vivo model.


METHODS
Male mice C57B/6 mice from Jackson Laboratory (Bar Harbor, ME) at 6-8 wks were subjected to a low-salt diet throughout the trial. After 1 week on a low-salt diet, the mice were injected daily with treatments in 50 muL vehicle composed of 75% cremaphor (Sigma, St. Louis, MO) and ethanol for 5 wks. A vehicle-alone group was also set aside. Mice were weighed and 25 mg/kg/day cyclosporine (Novartis Pharma, St. Louis, MO) was injected daily. Apocynin (Calbiochem, Gibbstown, NJ) 20 mg/kg were injected either alone or concomitantly with CyA. Another group of mice were administered IL-6 antibody (Cat no. MAB406; R&D Systems, Minneapolis, MN) at 2 mug/day along with CyA. The kidneys were removed en bloc immediately and submitted in formalin for paraffin sections. Trichrome stains were performed. Slides were blinded and 10 photographs of cortical areas per treatment group were taken, which covered an estimate of 10% surface area in random fashion. Areas of renal damage, which were determined by tubular necrosis, were identified and quantified by amount of necrosis per photograph. Each photograph was divided into 10 blocks, and the number of blocks that contained necrotic tubules per photo was recorded.


RESULTS
The two control mice (low salt only) had no damage. The four vehicle mice had trace amounts of tubular necrosis. CyA treatment group demonstrated the highest amount of damage (29/70; 41%). CyA with apocynin, a specific NADPH oxidase inhibitor, was found to have 36% (22/60) damage, whereas the CyA with IL-6 antibody only was observed to have 15% (6/40) damage. Comparing imaging analysis, there was no difference between mice treated with CyA alone and with CyA with apocynin. However, the amount of damage in mice treated with CyA and IL-6 antibody was found to be significantly lower than both CyA and CyA with apocynin.


CONCLUSIONS
CyA action as a calcineurin inhibitor has allowed prolongation of kidney transplants, but its chronic use has led to devastating consequences such as allograft nephropathy. Previously, we have identified potential mechanisms of CyA-induced endothelial dysfunction in vitro. The current study identifies increased IL-6 expression as a mechanism by which CyA induces renal damage and that the use of an IL-6-neutralizing antibody may be useful in reducing CyA-induced renal damage.
Department of Surgery, SUNY Upstate Medical University, St. Syracuse, New York 13210, USA.
Mark
M
LaSpina
Sudipta
S
Tripathi
Louis A
LA
Gatto
David
D
Bruch
Kristopher G
KG
Maier
Dilip S
DS
Kittur
D000098:Acetophenones / Q000494:pharmacology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology*; D016572:Cyclosporine / Q000633:toxicity*; D004039:Diet, Sodium-Restricted; D004195:Disease Models, Animal; D007166:Immunosuppressive Agents / Q000633:toxicity*; D015850:Interleukin-6 / Q000097:blood / Q000276:immunology*; D007668:Kidney / Q000187:drug effects* / Q000378:metabolism; D007674:Kidney Diseases / Q000139:chemically induced / Q000378:metabolism / Q000517:prevention & control*; D007684:Kidney Tubules / Q000187:drug effects / Q000473:pathology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000378:metabolism; D009336:Necrosis
D000098:Acetophenones; D000911:Antibodies, Monoclonal; D007166:Immunosuppressive Agents; D015850:Interleukin-6; D016572:Cyclosporine; C056165:acetovanillone; D019255:NADPH Oxidases