加味桃红四物汤阻断大鼠视网膜静脉阻塞模型JAK2-STAT3通路活化的机制

Mechanism of Modified Taohong Siwu Decoction blocking JAK2-STAT3 pathway activation in rat model of retinal vein occlusion

目的观察加味桃红四物汤对视网膜静脉阻塞(RVO)大鼠模型Janus激酶蛋白2-信号转导及转录因子3(JAK2-STAT3)通路的影响。方法将8周龄90只SD大鼠随机分成空白组(A组)、模型组(B组)、加味桃红四物汤高浓度组(C组)、加味桃红四物汤中浓度组(D组)、加味桃红四物汤低浓度组(E组),每组18只。A组不做处理,其他组大鼠右眼使用激光联合孟加拉玫瑰红的方法建立SD大鼠RVO模型。造模成功后,A组与B组予2 mL无菌注射液灌胃,C组、D组、E组灌胃给予低浓度(10 mg/kg)、中浓度(20 mg/kg)、高浓度(40 mg/kg)的加味桃红四物汤。给药后7、14、21 d用HE染色观察各组大鼠的视网膜结构、厚度和节细胞层(GCL)水肿情况,采用Western blot法检测21 d时视网膜组织中Janus激酶蛋白2(JAK2)、磷酸化Janus激酶蛋白2(P-JAK2)、信号转导及转录因子3(STAT3)、磷酸化信号转导及转录因子3(P-STAT3)的蛋白表达。结果①视网膜结构:7 d时各组视网膜3层结构完整,14 d时各组见散在视网膜静脉血管内血栓部分再通,21 d时各组节细胞数量改变不明显。视网膜厚度:7 d时B组、C组、E组见视网膜神经纤维层的间隙增宽;14 d及21 d时C、D、E组视网膜厚度均变薄。GCL水肿:7 d时各组GCL水肿,14 d时C组、D组、E组GCL水肿减轻,21 d时各组GCL无水肿。②JAK2蛋白表达:与A组比较,B组JAK2蛋白表达降低(P<0.05);与B组比较,C组、D组JAK2蛋白表达升高(P<0.05);与E组比较,C组、D组JAK2蛋白表达较高(P<0.05)。P-JAK2蛋白表达:与A组比较,B组、E组P-JAK2蛋白表达升高(P<0.05);与B组比较,C组、D组、E组P-JAK2蛋白表达降低(P<0.05);与E组比较,C组、D组P-JAK2蛋白表达较低(P<0.05)。STAT3蛋白表达:与A组比较,D组STAT3蛋白表达降低,E组STAT3蛋白表达升高(P<0.05);与B组比较,C组STAT3蛋白表达降低,E组STAT3蛋白表达升高(P<0.05);与E组比较,C组、D组STAT3蛋白表达较低(P<0.05)。P-STAT3蛋白表达:与A组比较,B组、C组P-STAT3蛋白表达升高(P<0.05);与B组比较,C组P-STAT3蛋白表达降低(P<0.05)。结论加味桃红四物汤可改善眼底缺血缺氧,阻断RVO的发展,其机制可能与抑制JAK2-STAT3通路活化相关。

Objective To observe the effect of Modified Taohong Siwu Decoction(MTHSWD)on JAK2-STAT3 pathways in rat model of retinal vein occlusion(RVO).Methods Ninety SD rats aged 8 weeks were randomly divided into blank control group(group A),model control group(group B),MTHSWD high concentration group(group C),MTHSWD medium concentration group(group D),MTHSWD low concentration group(Group E)with 18 rats in each group.Group A was left untreated,and the right eyes of rats in other groups were treated with laser combined with Rose Bengal to establish the RVO model in SD rats.After successful modeling,groups A and B were given 2 mL of sterile injection by gavage,and groups C,D and E were given low(10 mg/kg),medium(20 mg/kg)and high(40 mg/kg)concentrations of MTHSWD by gavage.The retinal structure,thickness and edema of ganglion cell layer(GCL)were observed by HE staining at 7,14 and 21 days after administration.The protein expressions of Janus kinase protein 2(JAK2),Janus kinase protein 2 phosphorylation(P-JAK2),signal transduction and transcription factor 3(STAT3),signal transduction and transcription factor 3 phosphorylation(P-STAT3)in retina were detected by Western blot method.Results①Retinal structure:The structure of three layers of retina in each group was intact on the 7th day,and partial recanalization of thrombus in the retinal vein was found in each group on the 14th day,and there was no significant change in the number of ganglion cells in each group on the 21st day.Retinal thickness:The gap of retinal nerve fiber layer was broadened in group B,C and E on the 7th day,and the retinal thickness became thinner in group C,D and E on the 14th and 21st day.GCL edema:GCL edema in each group on the 7th day and the edema in C,D and E groups on the 14th day alleviated,and GCL had no edema on the 21st day.②Expression of JAK2 protein:Compared with group A,the expression of JAK2 protein in group B was lower(P<0.05),the expression of JAK2 protein in group C and D was higher than that in group B(P<0.05),and the expression of JAK2 protein in group C and D was higher than that in group E(P<0.05).The expression of P-JAK2 protein:Compared with group A,the expression of P-JAK2 in group B and E was higher(P<0.05),the expression of P-JAK2 in group C,D and E was lower than that in group B(P<0.05),and the expression of P-JAK2 in group C and D was lower than that in group E(P<0.05).The expression of STAT3 protein:Compared with group A,the expression of STAT3 protein in group D decreased,and the expression of STAT3 protein increased in group E(P<0.05).Compared with group B,the expression of STAT3 protein decreased in group C and increased in group E(P<0.05).Compared with group E,the expression of STAT3 protein in group C and D was lower(P<0.05).The expression of P-STAT3 protein:Compared with group A,the expression of P-STAT3 protein in group B and C was higher(P<0.05),and the expression of P-STAT3 protein in group C was lower than that in group B(P<0.05).Conclusion Modified Taohong Siwu Decoction can improve fundus ischemia and hypoxia and block the development of RVO,and its mechanism may be related to the inhibition of JAK2-STAT3 pathway activation.