Doxorubicin Mitochondrial Dysfunction and Cytotoxicity in Human Proximal Tubular Epithelial Cells is Attenuated by Resveratrol (RES)

Abstract

Doxorubicin (DOX, Adriamycin), is a cancer chemotherapy agent used in the treatment regimen for breast, Hodgkin and Non‐Hodgkin lymphomas, small cell lung cancer and acute lymphoid leukemia. Unfortunately, doxorubicin clinical usage is associated with permanent cardiomyopathy and renal impairment. The cellular mechanisms for doxorubicin mediated renal cytotoxicity have yet to be understood. Resveratrol (RES) is a phytoalexin found in many fruits including grapes, blueberries, cranberries as well as chocolate and peanuts. Antioxidant and anticancer properties have been associated with RES in cell culture and in human clinical studies in cancer patients as well as animal models. This study tested the hypothesis that RES protection for DOX renal cytotoxicity, in human noncancerous renal proximal tubular epithelial cells (HK‐2), is mediated by preserving mitochondrial function and attenuating oxidative stress. HK‐2 cells were plated and grown in cell culture equilibrated for 48 h. Cells were next pre‐incubated for 1 h with 0 (DMSO) or 5 or 7.5 uM RES followed by a 24 h co‐incubation with 0–5 uM DOX. All results were obtained from 3 independent experiments. Western analysis probed for protein carbonylation was used as an indicator of oxidative stress. Mitochondrial function was assessed using a Seahorse analyzer system. Prior to examining mitochondrial function, cell number and substrate concentrations were optimized based on the manufacturer's recommendations. Mitochondrial Stress Test was conducted following the treatments described above and each well was normalized by protein. Western blot was conducted on whole cell lysate for SIRT1 and PGC1alpha. Viability was assessed using MTT leakage and tyrpan blue exclusion cell counts. RES did not alter cell viability as indicated by comparable MTT values between DMSO and RES groups (p>0.05). DOX was cytotoxic to HK‐2 cells within 24 h. Pretreatment with RES provided protection from DOX to HK‐2 cells. Mitochondrial respiration assessed as oxygen consumption rate (OCR) was diminished by DOX. DOX impaired mitochondria function within 24 h and RES provided protection. In summary, RES attenuated DOX renal cytotoxicity by maintaining mitochondrial function. Further studies are needed to probe changes in specific mitochondrial proteins mediated by DOX.Support or Funding InformationSupported by NIH Grant P20GM103434 to the West Virginia IDeA Network for Biomedical Research Excellence. M.D. and R.M. supported by WV NASA Undergraduate Research Fellowship.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.