Effect Of A-naphthlcetic Acid On The Proliferation And Apoptosis Of Testis Cell

Objective:To investigate the effect of a-naphthlcetic acid on the proliferation and apoptosis of testis cell.Method:1 In vitro experiment:The mouse testis cell suspension were used in this study and maintained in RPMI 1640 medium with 10% calf serum. MTT assay was used to test the influnence of NAA on inhibition of testis proliferation. (1)testis cells were cultured with NAA of different doses. (2)The proliferation activity of testis cells was determined by MTT assay at different time.2 In vivo experiment:Twenty five mice were divided into five groups,5 mice per group. Basal animal food was fed to the mice in the normal control and positive control groups. Five thousand,1000,200mg/kg of a-naphthlcetic acid were added into another three groups. Three days before sacrifice, the mice in positive control group were intraperitoneally injected with cyclophosphamide of 20 mg/kg each day. Four weeks later, all the mice were sacrificed. Effects on mouse body weight and testis/body weight ratio were probed with different doses of NAA. The level of maleic Dialdehyde(MDA)and the activity of GSH-Px in the five experimental groups were detected. The Proliferation of the testis was determined by MTT assay, and the levels of PCNA, Bcl-2 and Caspase-3 protein expression in testis cell were measured by immunohistochemistry method.Result:1 In vitro experiment:The cell proliferation was 0.761±0.006 after normal testis cells were cultured with 50μmol/L NAA for 24h, much higher than that of the control group(0.891±0.004). At the same time, with the increasing dose of NAA, the proliferation activity levels of 50,100,200,400μmol/L NAA groups decreased gradually(P<0.05). At the same dose of NAA, with the increasing time, the proliferation activity levels of 50, 100μmol/L NAA groups decreased gradually(P<0.05).2 In vivo experiment:(1)Proliferative activity:The testis cell proliferation activity of the high dose group and the normal control group were 0.501±0.069 and 0.875±0.082. The difference was significant (P<0.05). The expression level of PCNA protein of the high dose group was 8.9%, and that of the normal control group were 34.9%. The difference was significant (P<0.05).(2) Apoptosis:The expression level of Bcl-2 protein and Caspase-3 of the high dose group were 11.4% and 41.3%, respectively, and those of the normal control group were 30.6% and 9.1%. The differences between two groups were significant (P<0.05).(3) MDA and GSH-Px:The level of MDA and the activity of GSH-Px in the high dose group were 10.41±0.12nmol/ml and 759.01±11.32 NU/ml, and those of the normal control group were 8.41±0.12 nmol/ml and 991.39±10.79 NU/ml. The differences between two groups were significant (P<0.05)(4)Mice body weight:Exposure to NAA for 28 days change mice's weight, the t of the body weight of the high dose group and the normal control group were 14.96±0.83 and 31.48±1.19. The difference was significant (P<0.05).(5) Testis/body weight ratio:Exposure to NAA for 28 days remarkably promoted testis/body weight ratio in high and middle dose group(P<0.05).Conclusion:1 NAA can inhibit the proliferation and reduce the expression of PCNA protein of mice testis cells.2 NAA can decrease the expression level of Bcl-2 protein and increase expression level of Bcl-2 Caspase-3. So NAA can induce the apoptosis of testis cells effectively.3 NAA increased lipid per oxidation products in cells and decreased activities of ant oxidation enzyme.4 Exposure to NAA for can change mice's weight and promote testis/body weight ratio.