Expression Of Protein And MRNA Associated With Pi3k-AKT1 And Their Significance In Rats Liver Tissues With Chronic Fluorosis

Objective To investigate the expression of protein and m RNA associated with PI3 K signaling pathway, and the effect of LY294002 in rat hepatocytes with chronic fluorosis. Methods 48 SD rats were randomly divided into 4 groups according to the body weight, 12 in each group, half male and half female. The control group was provided with the solid feed in which the fluorine concentration was 1.5mg/kg. The fluorosis group was provided with the feed made from the maize containing fluorine at a concentration of 17mg/kg in the endemic fluorosis areas. Both the blocking group and the blocking control group were treated in the same way as the fluorosis group, and in the three weeks before the end of the experiment, the blocking group received the caudal vein injection with 10mg/kg blocking agent LY294002 once every other day. The blocking control group was given injection of blocking agent diluent PBS in the same way. Dental fluorosis was observed in experimental rats. The fluoride contents of urinary and skeletal were determined by the ion selective electrode method. Levels of aspartate transaminase(AST), alanine transaminase(ALT), total protein(TP) and albumin(Alb) in the serum were determined by using autobiochemical machine. Histological changes in liver tissue were observed with Hematoxylin & Eeosin(H&E) staining under light microscopy. The protein expression of PI3 K and Akt1 in hepatocytes of experimental animals was determined by immunohistochemistry(IHC) and Western blotting(Wb), respectively. The m RNA expression of PI3 K and Akt1 in hepatocytes was determined by Real-time quantitative PCR(RT-q PCR). The numbers of liver cell apoptosis and [Ca2+] were counted by Flow Cytometry. Results 1. Fluoride contents of the urine and bone were increased in the fluorosis rats compared to those in the control group(P < 0.05).2. ALT and AST activities were increased while TP and ALB activities were decreased in the fluorosis group and the blocking control group, compared to the control group. ALT and AST activities were decreased while TP and ALB activities were increased in the fluorosis group and the blocking control group, compared to the blocking group. 3. The expression of the m RNA and protein of PI3 K and Akt1 was significantly increased in hepatocytes in the fluorosis rats, and the expression of the m RNA and protein of PI3 K and Akt1 blocking group was lower than that of the fluorosis group(P<0.05). 4. Flow cytometry analysis showed that there were more hepatic apoptosis cells in the fluorosis group than those in the control group and the blocking group; there were slightly more apoptosis cells in the blocking group than that in the control group, no obvious difference has been found between the fluorosis group and the blocking control group. 5. The increase of the concentration of calciumion fluorescence intensity has been obviously found in the fluorosis group and the blocking control group, no obvious difference has been found between the control group and the blocking group. Conclusion PI3K-Akt signaling pathway can be activated by fluoride in liver tissue while LY294002 may inhibit this pathway. PI3K-Akt signaling pathway may be one of the signaling pathways which is involved in the pathogenesis of liver injury caused by fluorosis.