Bavachin ameliorates cisplatin-induced nephrotoxicity by enhancing mitochondrial β-oxidation and lipid metabolism through MFN2

Figures

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  Fig. 1. Bavachin ameliorates renal tubular injury in cisplatin-induced AKI mice. A, B serum creatinine and blood urea nitrogen levels after 72 h of cisplatin administration. C Serum cholesterol levels in each group of mice. D Representative images of hematoxylin and eosin staining. Scale bar = 100 μm. E Tubular damage score. F Representative image of immunohistochemical staining of KIM-1 in renal tissues from each group. Scale bar = 100 μm. G Relative immunostaining density of KIM-1. H Representative western blotting analysis of KIM-1 and NGAL protein levels in mouse kidneys. I Relative band intensity of KIM-1 and NGAL. n = 6 animals per group. All data were expressed as the mean ± SD (ns, no significant. ****p < 0.0001)

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  Fig. 2. Bavachin attenuates cisplatin-induced renal oxidative stress, lipid accumulation, and cell apoptosis. A Representative image of DHE staining. Scale bar = 50 μm. B Representative images of immunofluorescence and immunohistochemical staining of ADRP in renal tissues from each group. Scale bar = 100 μm. C Representative images of Oil red O staining. Scale bar = 50 μm. D Relative mean fluorescence density of DHE. E Relative mean fluorescence density of ADRP. F Relative immunostaining density of ADRP. Scale bar = 100 μm. G Oil Red O area. H Representative western blotting analysis of BAX and BCL-2 protein levels in mouse kidneys. I, J Relative band intensity of BAX and BCL-2. n = 6 animals per group. All data were expressed as the mean ± SD (*p < 0.05, ****p < 0.0001, ns: no significant)

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  Fig. 3. Bavachin improves mitochondrial fatty acid β-oxidation by targeting MFN2. A Molecular docking of MFN2 and bavachin. B Representative western blotting analysis of MFN2 and CPT1α protein levels in mouse kidneys. C Relative band intensity of MFN2 and CPT1α. D Representative images of immunohistochemical staining of MFN2 in renal tissues from each group. Scale bar = 100 μm. E Relative immunostaining density of MFN2. F Representative images of immunofluorescence staining of CPT1α in renal tissues from each group. G Relative immunofluorescence density of CPT1α. Scale bar = 100 μm. n = 6 animals per group. All data were expressed as the mean ± SD (*p < 0.05, ****p < 0.0001, ns: no significant)

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  Fig. 4. Bavachin attenuates cisplatin-induced oxidative stress, lipid accumulation, and apoptosis in HK-2 cells. A Observation of the representative morphology and nucleus of HK-2 cells under a microscope. Scale bar = 100 μm. B Representative images of DCFH-DA staining. Scale bar = 20 μm. C Representative images of Oil Red O staining. Scale bar = 20 μm. D Relative immunofluorescence density of DCFH-DA. E Oil Red O area. F Representative western blotting image of MFN2 and CPT1α in cisplatin-treated HK-2 cells with or without different concentrations of bavachin. G Relative band intensity of MFN2 and CPT1α in cisplatin-treated HK-2 cells with or without different concentrations of bavachin. H Representative western blotting analysis of BAX and BCL-2 protein levels in cisplatin-treated HK-2 cells with or without different concentrations of bavachin. I Relative band intensity of BAX and BCL-2 in cisplatin-treated HK-2 cells with or without different concentrations of bavachin. n = 3. All data were expressed as the mean ± SD (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, ns: no significant)

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  Fig. 5. Bavachin reduces ROS generation and lipid deposition in cisplatin-induced HK-2 cells via an MFN2-dependent mechanism. A Verification of the knockdown effect of MFN2 siRNA. B Relative band intensity of MFN2. C Western blot analysis revealed the expression of MFN2, CPT1α, BAX, and BCL-2 in cisplatin-induced HK-2 cells with or without bavachin after siRNA-mediated MFN2 knockdown. D-E Relative band intensity of MFN2, CPT1α, BAX, and BCL-2. F Representative images of Oil Red O staining. Scale bar = 50 μm. G Oil Red O area. H Representative image of DCFH-DA staining. Scale bar = 20 μm. I Relative immunofluorescence density of DCFH-DA. n = 3. All data were expressed as the mean ± SD. (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, ns: no significant)