Mechanism Study of Calcineurin Inhibitor Cyclosporine A Induced Renal Fibrosis

Fibrosis is the mechanism by which there is an excess production of a fibrous material, such as collagen and fibronectin, within the extracellular matrix. When there is a surplus amount of fibrous material, the extra cellular matrix will become mesh-like. In regard to kidney fibrosis, fibrous accumulation causes tubulointerstitial damage which triggers a loss of overall kidney functions. Cyclosporine A (CsA) is an immunosuppressive drug used to prevent organ rejection, in transplant patients. CsA has several side effects which include: viral infection, chronic renal insufficiency, end-stage renal disease, cancer and new-onset diabetes. We are focused on its nephrotoxicity properties. CsA immunosuppressive properties are accomplished by inhibiting the activation of T-cells. Our previous investigation confirmed the critical role Matrix Metalloproteases (MMP) play in CsA induced renal fibrosis. This study will examine the expression levels of specific per-fibrotic proteins, when kidney epithelial (HEK 293) and fibroblast cells are treated with 100ng/ml of CsA. The pre-fibrotic proteins are as follows: Cyclophilin A (CyPA), Fibronectin, E-Cadherin, E-Selectin, TGF-Beta, MMP-2 and MMP-9. To evaluate protein expression levels, cells are treated with 100ng/ml of CsA, in a six well plate, for 24 and 48 hours. The protein lysates are collected using TNESV lysis buffer and a Western Blot are used to observe the protein expression. We expect all protein levels to increase when cells are treated with CsA. There is a significant difference in the expression levels of MMP-9 and MMP-2, when cells are treated with CsA. There is an increase in CyPA, Fibronectin and TGF-Beta expression, when cells are treated with CsA, but there is no significance. In conclusion, our data suggest that Cyclophilin A (CyPA), Fibronectin, E-Cadherin, E-Selectin, TGF-Beta, MMP-2 and MMP-9 play a key role in CsA induced Renal Fibrosis. Further research is needed to fully understand the effects of long-term treatment with the immune-suppressive drug CsA