枸杞子-丹参对rd10小鼠视网膜Caspase-8、Caspase-12、Caspase-3、αB-crystallin表达的影响

Effects of Lycii Fructus-Salviea Miltiorrhizae Radix et Rhizoma on Expressions of Caspase-8,Caspase-12,Caspase-3 andαB-crystallin in Retinal of rd10 Mice

目的观察枸杞子-丹参对rd10小鼠视网膜Caspase-8、Caspase-12、Caspase-3、α晶状体球蛋白B(αB-crystallin)表达的影响,探讨枸杞子-丹参治疗视网膜色素变性的作用机制。方法40只rd10小鼠随机分为空白组、对照组、枸杞子组、丹参组、杞参组,每组8只,分别予生理盐水、维生素A溶液、枸杞子超微配方颗粒溶液、丹参超微配方颗粒溶液、枸杞子+丹参超微配方颗粒溶液灌胃,另取8只C57小鼠作为野生组,灌胃等体积生理盐水,灌胃剂量20 mL/kg,每日1次,连续28 d。TUNEL染色观察小鼠视网膜形态及感光细胞凋亡情况,RT-qPCR检测小鼠视网膜组织Caspase-8、Caspase-12、Caspase-3、αB-crystallin mRNA表达,免疫组化和免疫荧光检测Caspase-8、Caspase-12、Caspase-3、αB-crystallin蛋白表达。结果野生组小鼠视网膜各层结构排列正常,视网膜外层未见凋亡细胞;空白组小鼠视网膜结构不清晰,视网膜外层可见凋亡细胞;对照组、枸杞子组、丹参组小鼠视网膜结构欠清晰,视网膜色素上皮细胞不连续,视网膜外层可见少量凋亡细胞;杞参组小鼠视网膜结构清晰,视网膜外层凋亡细胞数量少于空白组。与空白组、对照组、枸杞子组和丹参组比较,杞参组小鼠视网膜组织Caspase-8、Caspase-12、Caspase-3 mRNA和蛋白表达降低,差异均有统计学意义(P<0.05);与空白组、对照组和枸杞子组比较,杞参组小鼠视网膜组织αB-crystallin mRNA和蛋白表达明显升高(P<0.05)。结论枸杞子-丹参可抑制rd10小鼠视网膜细胞凋亡,维持视网膜正常结构,从而保护视功能。

Objective To observe the effects of Lycii Fructus-Salviea Miltiorrhizae Radix et Rhizoma on the expressions of Caspase-8,Caspase-12,Caspase-3 andαB-crystallin in the retina of rd10 mice;To explore the mechanism of Lycii Fructus-Salviea Miltiorrhizae Radix et Rhizoma in the treatment of retinitis pigmentosa.Methods Totally 40 rd10 mice were randomly divided into blank group,control group,Lycii Fructus group,Salviea Miltiorrhizae Radix et Rhizoma group,and Lycii Fructus-Salviea Miltiorrhizae Radix et Rhizoma group,with 8 mice in each group.They were given normal saline,vitamin A solution,Lycii Fructus ultrafine formula granule solution,Salviea Miltiorrhizae Radix et Rhizoma ultrafine formula granule solution,and Lycii Fructus-Salviea Miltiorrhizae Radix et Rhizoma ultrafine formula granule solution for gavage,respectively.Another 8 C57 mice were taken as the wild group,and the same volume of normal saline was intragastrically administered.The treatment was given at a dose of 20 mL/kg,once a day for 28 consecutive days.TUNEL staining was used to observe the morphology of the retina and the apoptosis of photoreceptor cells;RT-qPCR was used to detect the expressions of Caspase-8,Caspase-12,Caspase-3,andαB-crystallin m RNA in the retina;Immunohistochemistry and immunofluorescence were used to detect the expressions of Caspase-8,Caspase-12,Caspase-3,andαB-crystallin protein in the retina.Results The structure of the retina in the wild group was normal,and no apoptotic cells were seen in the outer layer of the retina;the structure of the retina in the blank group was unclear,and apoptotic cells could be seen in the outer layer of the retina;the retina structure of the mice in the control group,Lycii Fructus group and Salviea Miltiorrhizae Radix et Rhizoma group were not clear,the retinal pigment epithelial cells were discontinuous,and a small amount of apoptotic cells could be seen in the outer layer of the retina;the retinal structure of the mice in the Lycii Fructus-Salviea Miltiorrhizae Radix et Rhizoma group were clear,and the number of apoptotic cells in the outer layer of the retina was less than that of the blank group.Compared with the blank group,control group,Lycii Fructus group and Salviea Miltiorrhizae Radix et Rhizoma group,the expressions of Caspase-8,Caspase-12,and Caspase-3 mRNA and protein in the retinal tissue of mice in Lycii Fructus-Salviea Miltiorrhizae Radix et Rhizoma group decreased,with statistical significance(P<0.05).Compared with the blank group,control group and Lycii Fructus group,the expression ofαB-crystallin mRNA and protein in the retinal tissue of mice in Lycii Fructus-Salviea Miltiorrhizae Radix et Rhizoma group significantly increased(P<0.05).Conclusion Lycii Fructus-Salviea Miltiorrhizae Radix et Rhizoma can inhibit the apoptosis of retinal cells of rd10 mice,maintain the normal functional structure of the retina,and thereby protect visual function.