摘要
目的观察当归拈痛汤对膝关节骨性关节炎(KOA)大鼠软骨损伤的影响,并分析其作用机制。方法50只SD雄性大鼠通过随机数表法分为对照组(n=10)与造模组(n=40),通过木瓜蛋白酶关节腔内注射法建立KOA大鼠模型。将造模成功大鼠随机分为模型组(n=10),当归拈痛汤组(低、中、高剂量,n=10)。当归拈痛汤组分别按照低剂量6.5 g/kg、中剂量13.0 g/kg、高剂量26.0 g/kg的药量进行灌胃,对照组与模型组给予等量生理盐水灌胃,连续给药1个月。采用游标卡尺测量大鼠双侧膝关节直径;通过酶联免疫吸附法(ELISA)检测大鼠血清肿瘤坏死因子(TNF-α)、白细胞介素-1β(IL-1β)水平;采用甲苯胺蓝染色观察各组大鼠膝关节软组织病理变化;通过实时荧光定量PCR(Real time-PCR)与免疫印迹法(Western blot)检测各组大鼠蛋白激酶R样内质网激酶(PERK)、免疫球蛋白结合蛋白(Bip)、半胱胺酸天冬氨酸蛋白酶-9(Caspase-9)mRNA与蛋白表达水平。结果给药期间对照组大鼠未见膝关节肿胀,与对照组比较,模型组大鼠双侧膝关节直径显著增加,采用中、高剂量当归拈痛汤治疗后大鼠膝关节直径显著缩短(P<0.05);病理观察结果显示模型组大鼠膝关节软骨局部裂隙缺损,采用中、高剂量当归拈痛汤治疗后大鼠膝关节软骨局部裂隙缺损症状显著改善;与对照组比较,模型组大鼠血清TNF-α、IL-1β含量,PERK、Bip、Caspase-9 mRNA与蛋白表达水平均显著提高,采用当归拈痛汤治疗可以下调大鼠血清TNF-α、IL-1β含量,PERK、Bip、Caspase-9 mRNA与蛋白表达水平,中、高剂量当归拈痛汤组大鼠以上各项指标与模型组比较均有统计学意义(P<0.05)。结论当归拈痛汤可以有效减轻KOA大鼠膝关节肿胀程度,降低血清炎性细胞因子含量,减轻软骨损伤,其机制可能与调控PERK/Bip通路,下调PERK、Bip、Caspase-9 mRNA与蛋白表达水平有关。
Objective To observe the effect of Danggui Niantong Decoction(当归拈痛汤)on cartilage injury in rats with knee osteoarthritis and analyze its mechanism.Methods 50 male SD rats were randomly divided into control group(n=10)and model group(n=40)by random number table method.Knee osteoarthritis rat model was established by intra-articular injection of papain.The rats were randomly divided into model group(n=10)and Danggui Niantong Decoction group(low,medium and high dose,n=10).Danggui Niantong Decoction group was given low dose of 6.5 g/kg,middle dose of 13.0 g/kg and high dose of 26.0 g/kg,respectively.The control group and model group were given the same amount of normal saline by gavage for 1 month.Vernier caliper was used to measure the diameter of bilateral knee joint in rats;the serum levels of tumor necrosis factor(TNF-α)and interleukin-1β(IL-1β)were detected by enzymelinked immunosorbent assay(ELISA);the pathological changes of knee joint soft tissue in each group were observed by toluidine blue staining;and the real-time quantitative PCR(real-time PCR)was used to detect the pathological changes of knee joint,the mRNA and protein expression levels of protein kinase R-like endoplasmic reticulum kinase(PERK),immunoglobulin binding protein(Bip)and Caspase-9 were detected by time-PCR and Western blot.Results During the administration period,no swelling of knee joint was observed in the control group.Compared with the control group,the diameters of bilateral knee joint in the model group were significantly increased,and the diameters of knee joint of the rats in the model group were significantly reduced after the treatment with medium and high dosage of Danggui Niantong Decoction(P<0.05);the pathological observation showed that the cartilage of knee joint in the model group was local the results showed that the symptoms of local fissure defect of knee joint cartilage in rats were significantly improved after treatment with medium and high dose Danggui Niantong Decoction;compared with the control group,serum TNF-α,IL-1βcontent,PERK,Bip,Caspase-9 mRNA and protein expression levels of model group rats were significantly increased,Danggui Niantong Decoction treatment could reduce serum TNF-α,IL-1βcontent,PERK,Bip,Caspase-9 levels.Compared with the model group,the expression of mRNA and protein in the medium and high dose Danggui Niantong Decoction groups were significantly different(P<0.05).Conclusion Danggui Niantong Decoction can effectively reduce the degree of knee swelling in KOA rats,reduce the content of serum inflammatory cytokines,and reduce cartilage damage.The mechanism may be related to the regulation of PERK/Bip pathway and down-regulation of PERK,Bip,Caspase-9 mRNA and protein expression levels.
作者
张有为
李小宏
李凯
ZHANG Youwei;LI Xiaohong;LI Kai(Baoji Central Hospital,Baoji 721008,Shaanxi,China)
出处
《辽宁中医药大学学报》
CAS
2023年第5期34-38,F0003,共6页
Journal of Liaoning University of Traditional Chinese Medicine
基金
宝鸡市科技计划项目(16SFGG3-20)。
关键词
当归拈痛汤
膝关节骨性关节炎
软骨损伤
蛋白激酶R样内质网激酶
免疫球蛋白结合蛋白
Danggui Niantong Decoction(当归拈痛汤)
knee osteoarthritis
cartilage injury
protein kinase R-like endoplasmic reticulum kinase
immunoglobulin binding protein