女贞子提取物干预大鼠膝骨关节炎的实验研究

目的观察女贞子提取物对膝骨关节炎(KOA)大鼠关节疼痛的影响,研究其对KOA大鼠软骨的保护作用。方法32只SD大鼠随机分为正常组、模型组、女贞子灌胃组和女贞子注射组。模型组、女贞子灌胃组和女贞子注射组均采用碘乙酸关节腔注射造模法构建大鼠KOA模型,女贞子灌胃组大鼠进行女贞子提取物灌胃处理,女贞子注射组大鼠予以关节腔内注射女贞子提取物。治疗持续4周,于最终给药后1周测定行为学数据;取各组大鼠膝关节软骨组织,进行番红O染色和Mankin′s评分,免疫组化检测Col2的表达。取大鼠原代软骨细胞,以不同浓度的女贞子提取物进行干预,采用MTT法测定女贞子提取物对软骨细胞增殖的影响,选取最佳浓度。RT-PCR测定女贞子提取物对软骨细胞中OA相关基因表达的调节作用,并进行统计分析。结果疼痛模型数据显示,模型组的压痛阈值与正常组相比显著下降(P<0.01);女贞子注射组与模型组比较有显著提高(P<0.01);女贞子灌胃组与模型组比较有提高(P>0.05)。组织病理学结果显示,模型组的Mankin′s评分高于空白组(P<0.01);女贞子注射组Mankin′s评分较模型组显著下降(P<0.01);女贞子灌胃组与模型组比较有下降(P>0.05)。免疫组化分析显示女贞子注射组和女贞子灌胃组软骨中Col2表达增加。女贞子提取物可促进大鼠原代软骨细胞的增殖,RT-PCR分析结果显示与TNF-α处理组相比,女贞子组显著下调Col10、MMP13的表达,上调Col2和Aggrecan的表达。结论女贞子提取物关节腔注射对软骨细胞的保护效果优于灌胃给药;女贞子提取物对大鼠KOA疼痛模型有干预作用,其机制与抑制软骨细胞肥大和分解代谢有关。

Experimental Study on the Effect of Ligustrum Lucidum Extract on Knee Osteoarthritis in Rats

Objective:To observe the effect of Ligustrum lucidum extract on joint pain and cartilage in KOA rats,and to study the protective effect of fructus ligustri extract on cartilage in KOA rats.Methods:32 SD rats randomly divided into four groups:the normal group,the model group,Ligustrum lucidum extract lavage group and Ligustrum lucidum extract injection group.The model group,the lavage group and the injection group adopted iodoacetate articular cavity injection mold method to build KOA rat model.The treatment lasted 4 weeks and behavioral data were measured one week after final administration.After treatment,the knee cartilage tissues of each group were taken,stained with saffron-o and scored by Mankin's.The expression of Col2 was detected by immunohistochemistry.Primary rat chondrocytes were taken,and different concentrations of Ligustrum lucidum extract were used for intervention.MTT method was used to determine the effect of Ligustrum lucidum extract on chondrocytes proliferation,and the optimal concentration was selected.RT-PCR was used to determine the effect of Ligustrum lucidum extract on the specific expression of OA in chondrocytes.Results:The data of pain model showed that the tenderness threshold of the model group was significantly lower than that of the normal group,with significant difference(P<0.01);compared with the model group,the injection group was significantly improved,with significant difference(P<0.01);compared with the model group,the lavage group increased,however,without significant difference(P>0.05).The results of histopathology showed that the Mankin's score of the model group was higher than that of the blank group,with significant difference(P<0.01);compared with the model group,the injection group was significantly decreased,with significant difference(P<0.01);compared with the model group,the lavage group decreased,however,without significant difference(P>0.05).Immunohistochemical analysis showed that the expression of Col2 in cartilage of the injection group and the lavage group increased.Ligustrum lucidum extract could promote the proliferation of primary chondrocytes in rats.RT-PCR analysis showed that the expression of Col10 and MMP13 was down-regulated,Col2 and Aggrecan up-regulated in Ligustrum lucidum extract groups,compared with that in TNF-αgroup.Conclusion:The effect of intraarticular injection of Ligustrum lucidum extract on chondrocytes is better than that of intragastric administration.It can intervene the pain model of knee osteoarthritis in rats.The mechanism is related to the inhibition of chondrocyte hypertrophy and catabolism.