Cytoprotective and anti-apoptotic role of Terminalia arjuna on oxalate injured renal epithelial cells

Abstract

Urolithiasis is one of the painful multifactorial disorders caused by metabolic abnormalities influencing the composition of body fluids and urine. The bark of Terminalia arjuna (T. arjuna), very well known in Ayurveda for the treatment of cardiovascular diseases, possesses antioxidant and diuretic activity. The present study was undertaken to investigate the antiurolithiatic efficacy of aqueous extract of bark of T. arjuna on oxalate-induced injury to renal tubular epithelial cells. Madin-Darby canine kidney (MDCK) cells were exposed to 2 mM oxalate for 48 h to evaluate the protective effect of T. arjuna aqueous extract on cell viability, CaOx crystal adherence and apoptotic changes caused by oxalate. The results confirmed that oxalate injured MDCK cells were protected by T. arjuna extract. On treatment with a range concentrations, the cell viability increased in a concentration dependent manner. Moreover, the extract prevented the interaction of the calcium oxalate (CaOx) crystals with the cell surface and reduced the number of apoptotic cells. The current data suggests that T. arjuna bark confers a cytoprotective role and based on our results it could be a potential candidate from natural plant sources against urolithiasis.

Keywords: Calcium oxalate; Madin–Darby canine kidney cells; Phytotherapy; Renal epithelial cells; Terminalia arjuna; Urolithiasis.

Fig. 1

Effect of aqueous extract of T. arjuna on MDCK cell viability assessed by MTT assay. Data are mean ± SD of three independent observations. ns not significant. ****p < 0.0001 versus untreated control; and ^## p < 0.01, ^### p < 0.0005 versus oxalate injury control

Fig. 2

Evaluation of T. arjuna extract on CaOx crystal adherence in oxalate induced injury to MDCK cells, visualized under polarization and phase contrast at magnification ×20 and scale bar 100 microns

Fig. 3

Modulation of apoptosis in oxalate induced injury to MDCK cells by T. arjuna extract, visualized under fluorescence microscopy at magnification ×20 and scale bar 100 microns. Nuclear staining was performed with Hoechst 33258 (blue signal). Inset shows enlargement of areas indicated by white arrows. (Color figure online)

Fig. 4

Flow cytometry evaluation of apoptosis in oxalate induced injury to MDCK cells, visualized by Annexin V/PI staining

Fig. 5

Flow cytometry assessment of apoptosis in oxalate induced injury to MDCK cells, visualized by Anti-Active Caspase-3 antibody staining