Transition from Cyclosporine-Induced Renal Dysfunction to Nephrotoxicity in an in Vivo Rat Model

Abstract

Cyclosporin A (CsA), a calcineurin inhibitor, remain the cornerstone of immunosuppressive regimens, regardless of nephrotoxicity, which depends on the duration of drug exposure. The mechanisms and biomarkers underlying the transition from CsA-induced renal dysfunction to nephrotoxicity deserve better elucidation, and would help clinical decisions. This study aimed to clarify these issues, using a rat model of short- and long-term CsA (5 mg/kg bw/day) treatments (3 and 9 weeks, respectively). Renal function was assessed on serum and urine; kidney tissue was used for histopathological characterization and gene and/or protein expression of markers of proliferation, fibrosis and inflammation. In the short-term, creatinine and blood urea nitrogen (BUN) levels increased and clearances decreased, accompanied by glomerular filtration rate (GFR) reduction, but without kidney lesions; at that stage, CsA exposure induced proliferating cell nuclear antigen (PCNA), transforming growth factor beta 1 (TGF-β1), factor nuclear kappa B (NF-κβ) and Tumor Protein P53 (TP53) kidney mRNA up-regulation. In the long-term treatment, renal dysfunction data was accompanied by glomerular and tubulointerstitial lesions, with remarkable kidney mRNA up-regulation of the mammalian target of rapamycin (mTOR) and the antigen identified by monoclonal antibody Ki-67 (Mki67), accompanied by mTOR protein overexpression. Transition from CsA-induced renal dysfunction to nephrotoxicity is accompanied by modification of molecular mechanisms and biomarkers, being mTOR one of the key players for kidney lesion evolution, thus suggesting, by mean of molecular evidences, that early CsA replacement by mTOR inhibitors is indeed the better therapeutic choice to prevent chronic allograft nephropathy.

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“…Plasma levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT), urea and creatinine were used as indicators of hepatotoxicity [38] and nephrotoxicity [39], respectively. These assays were performed using a diagnostic kit from Labtest (Lagoa Santa, Minas Gerais, Brazil) in samples collected in the 14 th day after surgery after behavioral evaluation.…”

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“…23,24 The CsA rats given 10 or 20 ng/kg 2AMD did not gain weight or lost up to 8% of their weight, respectively, suggesting a worsening of CsA pathology with hypercalcemia. However, when 2AMD or 2MD was administered at 2.5 or 5 ng/kg, rat weight increased by 20%, indicating a trend in amelioration of the deleterious effects caused by CsA.…”

“…Sections from rats that received CsA plus vehicle exhibited the typical areas of tubular atrophy and increased fibrosis apparent as basophilia, likely due to mesenchymal cell proliferation and inflammatory cell infiltration typical for CsA-treated kidneys. 23,24 Mesenchymal cells are responsible for the production of excess collagens. 2 Vehicle-treated kidney sections were mostly acidophilic, with intact tubules and no apparent areas of interstitial cell growth.…”

“…Cyclosporine is an immunosuppressant given to reduce organ rejection but that eventually promotes renal vasoconstriction, ending in epithelial cell apoptosis and interstitial fibrosis. 23,24 Neither 2AMD nor 2MD has been tested previously for renoprotective or antifibrotic effects in models of kidney disease. We found that at 5 ng/kg both 2AMD and 2MD decreased kidney fibrosis and ameliorated nephrotoxic effects of CsA without raising serum calcium levels.…”

“…In addition, we report the effects of higher 2AMD doses to be avoided due to toxicity in the CsA rat model. 24 hours before blood collection and organ retrieval. Animals were placed under anesthesia, blood (aorta) was collected, and kidneys were removed, with subsequent death by pneumothorax puncture.…”