Author + information
- Li Wenhua
To observe the cell proliferation ability, ROS, expressions of SIRT1, FOXO3a in AKI model induced by iodine in HK-2 cells and the effects of astaxanthin on it and its possible mechanisms will be also investigated. Our study will provide a theoretical evidence of astaxanthin on the prevention and treatment of contrast induced acute kidney injury.
HK-2 cells were cultured in DMEM/F12 medium in vitro and then randomly divided into appropriate experimental groups: normal group, dimethyl sulfoxide (DMSO), iohexol group, iohexol + (1.0, 10.0 μmol/L) astaxanthin group, iohexol + SIRT1 inhibitors (Nicotinamide, NA) group and iohexol + siRNA FOXO3a group;when were cultured at 24 hours, cell proliferation ability was tested by CCK-8 method, ROS were detected by flow cytometry, The expression of SIRT1 and FOXO3a were observed using western-blot.
(1) CCK-8 Results: ① with the increase of concentration of astaxanthin HK-2 cells proliferation has not changed, when AST concentrations > 15μmol/L, cell proliferation was significantly reduced,(aP<0.05), the difference was statistically significant. ② when concentration of the iohexol > 200gI/L, the cell proliferation decreased,(aP<0.05), the difference was statistically significant.③nicotinamide concentration> 20mmol/L, the cell proliferation decreased, (aP <0.05), the difference was statistically significant.
(2) Flow cytometry results showed that (after intervention 24 h): compared to the negative control group, the level of ROS in the iohexol group, iohexol + 1.0μmol/L, 10μmol/L astaxanthin group was significantly increased, the differences were statistically significant (aP<0.05); and between iohexol group and iohexol+ (1.0,10.0μmol/L) astaxanthin group, iohexol+1.0μmol/L astaxanthin group and iohexol + 10.0μmol/L astaxanthin group, differences were statistically significant (bP <0.05).
(3) Western blot results are shown, iohexol group than normal group SIRTl protein were significantly increased, FOXO3a protein expression decreased, the differences were statistically significant (aP <0.05); iohexol+(1.0,10.0 μmol/L) astaxanthin group compared to iohexol group SIRTl protein expression in each group were significantly lower, FOXO3a protein increased, the differences were statistically significant (bP <0.05).
Iohexol+NA group than the control group, SIRT1, FOXO3a protein expression decreased significantly (aP <0.05); iohexol + FOXO3a siRNA group than control group, the expression of SIRT1 protein had no significant change (P>0.05), FOXO3a protein decreased expression (aP<0.05), the differences were statistically significant.
astaxanthin inhibits iohexol induced damage in HK-2, and its possible mechanism is related to decrease expression of endothelial cells SIRT1, and on the upstream of FOXO3a plays against damage induced by iohexol.