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Accelerated apoptosis characterizes cyclosporine-associated interstitial fibrosis
S E Thomas1, T F Andoh, R H Pichler
1Department of Medicine, University of Washington, Seattle, USA.
Abstract:
Recently we developed a model of cyclosporine nephropathy in rats characterized by tubulointerstitial (TI) injury, macrophage infiltration, and progressive interstitial fibrosis [1, 2]. To determine if the TI injury accompanying cyclosporine A (CsA) nephropathy was associated with accelerated apoptosis and ischemia, we treated rats for five weeks with CsA with or without losartan (to block angiotensin II type 1 receptor), or hydralazine/furosemide (H/F) (protocol #1). In protocol #2, rats received CsA with or without L-NAME (to block nitric oxide) or L-arginine (to provide a precursor to nitric oxide formation). Cyclosporine A treated rats had increased apoptosis of tubular and interstitial cells documented by PAS, propidium iodide staining, TUNEL assay, and electron microscopy compared to vehicle treated controls. Macrophages containing apoptotic cells could be confirmed by TUNEL/ED-1 doublestaining and colocalized in areas of TI injury. Animals treated with CsA + losartan had a statistically significant decrease in apoptosis (TUNEL + cells/mm2) when compared to CsA treated animals (6.0 vs. 19.9, P < or = 0.0001). The decrease in apoptosis in the CsA + H/F group was not statistically significant. Animals treated with CsA + L-NAME had a statistically significant increase in apoptosis compared to the CsA treated animals (12.3 vs. 6.4, P = 0.001). L-arginine administration with CsA resulted in a decrease in tubulointerstitial apoptosis versus CsA treated animals, however, this did not reach statistical significance. The addition of L-arginine did result in a significant reduction in interstitial fibrosis (P < 0.0001). Regression analysis revealed a significant correlation between apoptosis and interstitial fibrosis in both protocols. (CsA vs. CsA + losartan r = 0.63, P < 0.0001; CsA vs. CsA + L-NAME r = 0.83, P < 0.0001). We conclude that CsA nephropathy is associated with a marked increase in apoptosis of tubular and interstitial cells. Cyclosporine A induced apoptosis is partially mediated by angiotensin II and nitric oxide inhibition, suggesting a role for renal ischemia in this process, and CsA induced apoptosis correlates with interstitial fibrosis.
Insights
Cyclosporine A (CsA) causes kidney injury by increasing cell death (apoptosis) in rats. Blocking angiotensin II or nitric oxide partially reduces this apoptosis, which is linked to fibrosis.
Area of Science:
- Nephrology
- Cell Biology
- Pathology
Background:
- Cyclosporine A (CsA) induces tubulointerstitial (TI) injury, macrophage infiltration, and fibrosis in a rat model.
- The role of apoptosis and ischemia in CsA-induced TI injury requires further investigation.
Purpose of the Study:
- To investigate the association between CsA nephropathy, apoptosis, and ischemia.
- To evaluate the effects of losartan, hydralazine/furosemide, L-NAME, and L-arginine on CsA-induced apoptosis and fibrosis.
Main Methods:
- Rats were treated with CsA alone or in combination with losartan, hydralazine/furosemide, L-NAME, or L-arginine.
- Apoptosis was quantified using PAS, propidium iodide staining, TUNEL assay, and electron microscopy.
- Interstitial fibrosis was assessed, and correlations between apoptosis and fibrosis were analyzed.
Main Results:
- CsA treatment significantly increased tubular and interstitial cell apoptosis compared to controls.
- Losartan significantly decreased CsA-induced apoptosis; L-NAME significantly increased it.
- L-arginine reduced interstitial fibrosis but not apoptosis significantly.
- A strong positive correlation was observed between apoptosis and interstitial fibrosis.
Conclusions:
- CsA nephropathy is characterized by increased tubular and interstitial cell apoptosis.
- Angiotensin II and nitric oxide pathways are involved in CsA-induced apoptosis, suggesting a role for renal ischemia.
- CsA-induced apoptosis is significantly correlated with the development of interstitial fibrosis.