Protective effect of ginsenoside Rk1, a major rare saponin from black ginseng, on cisplatin-induced nephrotoxicity in HEK-293 cells

Abstract

Cisplatin, as one of the most effective chemotherapeutic agents, its clinical use is limited by serious side effect of nephrotoxicity. Cisplatin-induced nephrotoxicity is closely related to apoptosis induction and activation of caspase. The present study aimed to explore the potential protective effect of ginsenoside Rk1 (Rk1), a rare ginsenoside generated during steaming ginseng, on cisplatin-induced nephrotoxicity and the underlying mechanisms in human embryonic kidney 293 (HEK-293) cells. Our results showed that the reduced cell viability induced by cisplatin could significantly recover by Rk1. Furthermore, glutathione (GSH) as an oxidative index, was elevated and the lipid peroxidation product malondialdehyde (MDA) was significantly decreased after Rk1 treatment compared to the cisplatin group. Additionally, Rk1 can also decrease the ROS fluorescence expression and increase the protein levels of nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) compared to the cisplatin group, which suggested a suppression of oxidative response. More importantly, the cisplatin-induced elevated protein levels of Bax, cleaved caspase-3, cleaved caspase-9, and decreased protein level of Bcl-2 were reversed after treatment with Rk1. Our results elucidated the possible protective mechanism of Rk1 for the first time, which may involve in its anti-oxidation and anti-apoptosis effects.

Keywords: HEK-293; anti-apoptosis; cisplatin; ginsenoside Rk1; nephrotoxicity.

FIGURE 1

A, Chemical structures of ginsenoside Rk1. B, Effect of ginsenoside Rk1 on cells viability. C, Effect of cisplatin on cells viability. D, Protective effect against cisplatin‐induced HEK‐293 cell injury. All data were expressed as the mean ± SD, n = 6. **P < .01, *P < .05 vs normal group; ^## P < .01, ^# P < .05 vs cisplatin group

FIGURE 2

Effect of ginsenoside Rk1 on cells morphological changes. HEK‐293 cells treated with ginsenoside Rk1 (10, 20, and 30 μM) and cisplatin (20 μM)

FIGURE 3

Effects of different concentrations of ginsenoside Rk1 against cisplatin‐induced oxidative stress. The levels of A, GSH and B, MDA in cisplatin‐induced HEK‐293 cell injury. C, Improvement effect of ginsenoside Rk1 with different concentrations (10, 20, and 30 μM) on ROS generation in cisplatin‐induced HEK‐293 cells. D, Relative fluorescence intensity. E, Effects of ginsenoside Rk1 on the protein expressions of Nrf2 and HO‐1. F, Quantification of relative protein contents were performed by densitometric analysis. **P < .01, *P < .05 vs normal group; ^## P < .01, ^# P < .05 vs cisplatin group

FIGURE 4

Ginsenoside Rk1 inhibits cisplatin‐induced HEK‐293 cell apoptosis. The nuclear morphology was evaluated using fluorescence microscopy after A, Hoechst 33258 staining, arrows show necrotic and injured nephrocyte. B, The percentage of apoptosis. **P < .01 vs normal group; ^## P < .01, ^# P < .05 vs cisplatin group

FIGURE 5

A, Effects of ginsenoside Rk1 on the protein expressions of Bcl‐2, Bax, cleaved caspase‐3, caspase‐3, cleaved caspase‐9 and caspase‐9. B, Quantification of relative protein contents were performed by densitometric analysis. **P < .01, *P < .05 vs normal group; ^## P < .01, ^# P < .05 vs cisplatin group