Apoptotic Effect Of Organophosphorus Insecticide Chlorpyrifos On Mouse Retina In Vivo Via Oxidative Stress And Protection Of Combination Of Vitamin C And E

Organophosphorus insecticide chlorpyrifos is widely used in agricultural production and public health practice,however,its widespread use has brought high risks of severe environmental pollution and health problems,including acute and chronic poisoning in human being.Retina is one of the target organs affected by chlorpyrifos,but the mechanisms of its effects on retina are not well elucidated. Previous studies focused on inhibition of acetylcholinesterase activity by chlorpyrifos, and our studies discussed chlorpyrifos-induced cell apoptosis in retinal cells by the pathway of oxidative stress and and the protection of combination of vitamins C and E.Experimental mice were randomly divided into four groups(n=20 in each group): control group(Control),vitamin C plus vitamin E-treated group(VC+VE), chlorpyrifos-treated group(CPF),and chlorpyrifos plus vitamin C plus vitamin E-treated group(CPF+VC+VE).Mice of VC+VE and CPF+VC+VE groups were pretreated with 250 mg/kg VC and 150 mg/kg VE by hypodermic injecting once daily for 6 consecutive days.The other groups were injected hypodermically with the same volume of physiological saline solution at the same time.Then mice of CPF and CPF+VC+VE groups were treated with 63 mg/kg CPF via oral garage,single treatment,which is about 1/4 of 50%of the lethal dose,and the other groups received vehicle treatment.Twenty-four hours after chlorpyrifos treatment,all the mice were anaesthetized and sacrificed,then retinas were separated.Some retinas in each group were used for electron microscopic observation,and laser scanning confocal microscope(LSCM) analysis of[Ca²⁺]_I respectively,and some retinas were isolated into single cells to perform comet assay,acridine orange/ethidium bromide staining and detection of apoptosis by flow cytometer respectively.The remaining retinas in each group were mixed and homogenized to measure malondialdehyde(MDA)content,superoxide dismutase(SOD)activity,catalase(CAT)activity,glutathione peroxidase(GPX) activity,acetylcholinesterase(ACHE)activity,and protein content.Our results showed that cell apoptosis,lipid peroxidation and DNA damage were increased,and activities of antioxidant enzymes superoxide dismutase,catalase and glutathione peroxidase were decreased in retina of chlorpyrifos-administrated mice.Pretreatment of combination of antioxidants vitamin C and vitamin E significantly attenuated these cytotoxic effects of chlorpyrifos,demonstrating oxidative stress was involved in chlorpyrifos-induced cell apoptosis in mouse retina.Moreover,chlorpyrifos treatment inhibited acetylcholinesterase activity and promoted[Ca²⁺]_i level in mouse retinal cells,which were also attenuated by combination of vitamins C and E.In conclusion,our results demonstrated:(1)organophosphorus insecticide chlorpyrifos-induced cell apoptosis in mouse retina via oxidative stress,involved in AChE activity inhibition and[Ca²⁺]_i level increase.Our results may contribute to an understanding of the toxicological mechanisms of chlorpyrifos and other organophosphorous insecticides.(2)Combination of exogenous antioxidants vitamins C and E provided appropriate protection against chlorpyrifos-induced retinal damage. Our results give clue to antioxidant therapy,besides traditional regimens concerning AChE,or combination of both therapies in organophosphorus insecticides-induced ocular damage.