Sijunzi Decoction Reduces Photoreceptor Cells Apoptosis Associated With Experimental Retinal Reattachment In Rats With Spleen-Qi Deficiency Syndrome By Regulating Mitochondrial Pathway

Objective:To observe whether Sijunzi Decoction(SJZD)protects photoreceptor cells apoptosis associated with experimental retinal reattachment in rats with Spleen-Qi Deficiency Syndrome(SQDS)by regulating mitochondrial pathway.Methods:Fifty-two adult Spregue-Dawley rats were randomly divided into control group(Group A),retinal reattachment group(Group B),retinal reattachment with SQDS group(Group C)and SJZD treatment group(Group D),remnant four rats were used as mitochondrial transmembrane potential positive control.The SQDS model was established at first by dietary disorder and Spleen-Qi consumption medicine in Group C and D.After 6 weeks,the model of RD was prepared by sub-retinal injecting0.1%sodium hyaluronate to rats in Group B,C,and D.After the injection,group D were treated with SJZD by gavage.Animals were sacrificed randomly at 8w and 9w respectively.Food intake,body weight,and mental states of rats were recorded and the contents of D-xylose in serum,Na~+-K~+-ATP activity of erythrocyte membrane were measured at the beginning of the experiment,6w,8w and 9w respectively.The thickness of the outer nuclear layer(ONL)and the inner nuclear layer(INL)was measured in HE staining paraffin sections of eyeball.Apoptotic index(AI)in outer nuclear layer was measured by TUNEL staining.The localization of AIF and Cyt C was detected by immune-fluorescence staining.Mitochondrial transmembrane potential(ΔΨm)of retinal tissue was measured by JC-1 fluorescence probe.The expression of Bax、Bcl-2、Caspase-3 in retinal tissue and AIF in retinal nucleus were analyzed by Western Blot at 8w and 9w respectively.Results:At the beginning of the experiment,food intake,body weight,the serum D-xylose level and Na+-K+-ATP activity of erythrocyte membrane were no difference in all groups.At 6w,body weight,food intake,the serum D-xylose level and Na+-K+-ATP activity of erythrocyte membrane were significantly lower in Group C and D,as compared with Group A and B(P<0.05).In Group C,the above symptoms lasted to 9 weeks.Rats’symptoms were improved in Group D after taking SJZD.food intake and Na~+-K~+-ATP activity of erythrocyte membrane in Group D significantly increased(P<0.05)at 9w.At 8w,compared with Group A,the thickness ratio of ONL and INL was decreased significantly in Group B,Group C and Group D and it was no difference between Group B,Group C and Group D.At 9w,the thickness ratio of ONL and INL in Group D was significantly increased than group C.At 8w,AI in outer nuclear layer in Group C and Group D were increased than Group B(P<0.05).At 9w,AI of outer nuclear layer in Group D was decreased than Group C.At 8w,ΔΨm was no difference between Group C and Group D(P>0.05)as it was decreased in Group C than Group B(P<0.05).At9w,ΔΨm in Group D was increased than Group C(P<0.05).At 8w and 9w,the expression of Bax were no difference between Group A and Group B(P>0.05)as,it increased in Group C than Group A(P<0.05).At 9w,the expression of Bax in Group D was decreased significantly than Group C(P<0.05).The expression of Bcl-2 was no difference in all groups at 8w and 9w(P>0.05).At 8w and 9w,the expression of Caspase-3 in Group B and Group C were increased significantly than Group A(P<0.05)and it increased significantly in Group C than Group B(P<0.05).At 9w,the expression of Caspase-3 in Group D was decreased than Group C.(P<0.05).Immunofluorescence results showed that AIF and Cyt C in group B,Group C and Group D were diffuse than Group A.A small amuont of AIF in Group B was expressed in nucleus in outer nuclear layer.The positive expressions of AIF and Cyt C in Group C were mainly found in outer nuclear layer and they were scattered in outer nuclear layer in Group D.Conclusions:SJZD played neuroprotective roles in experimental retinal reattachment with SQDS by regulating mitochondrial pathway to protec photoreceptor cells apoptosis.