蒙医针刺疗法治疗干眼症模型大鼠的实验研究

Experimental Study on Mongolian Medicine Acupuncture Therapy in the Treatment of Dry Eye Model Rats

目的:探讨蒙医针刺疗法通过调控角膜炎症相关基因治疗干眼症模型大鼠角膜机制。方法:选取SPF级SD大鼠60只,使用随机数表法分为正常组、模型组、玻璃酸钠滴眼液组、蒙医针刺疗法组,每组各15只。除正常组以外的其余3组建立阿托品诱导的干眼症大鼠模型。大鼠造模成功后,模型组不做处理;玻璃酸钠滴眼液组给予玻璃酸钠滴眼,3次/d,每次1滴,连续点眼4周;蒙医针刺疗法组取赫依穴、希拉穴、肝穴行针刺,1次/d,每次留针15 min,共针刺4周。每组分别于实验前和造模成功后2周、4周时末次给药/针刺2 h后,检测大鼠泪液分泌量和泪膜破裂时间;HE染色观察角膜形态变化;采用RT-PCR法检测角膜组织中IL-1βmRNA、IL-6 mRNA、TNF-a mRNA的表达情况。结果:HE染色结果示蒙医针刺疗法组上皮细胞增生与基底部细胞排列紊乱,明显优于模型组。针刺后2周,与模型组比较,蒙医针刺疗法组大鼠泪膜破裂时间、泪液分泌量显著增加(P<0.01),角膜组织中IL-1βmRNA、IL-6 mRNA、TNF-a mRNA的表达明显降低(P<0.05);与蒙医针刺疗法组比较,玻璃酸钠滴眼液组大鼠泪膜破裂时间、泪液分泌量明显降低(P<0.01),角膜组织中IL-1βmRNA、TNF-a mRNA的表达明显增加(P<0.05)。针刺后4周,与模型组比较,蒙医针刺疗法组大鼠泪膜破裂时间、泪液分泌量明显增加(P<0.01),角膜组织中IL-1βmRNA、IL-6 mRNA、TNF-αmRNA的表达明显降低(P<0.01);与蒙医针刺疗法组比较,玻璃酸钠滴眼液组大鼠泪膜破裂时间、泪液分泌量明显降低(P<0.01),角膜组织中IL-1βmRNA、IL-6 mRNA、TNF-αmRNA的表达明显增加(P<0.01);与正常组比较,蒙医针刺疗法组大鼠泪液分泌量、泪膜破裂时间及IL-1βmRNA、IL-6 mRNA、TNF-αmRNA的表达无明显改变(P>0.05);蒙医针刺疗法组4周与蒙医针刺疗法组2周比较,泪液分泌量、泪膜破裂时间明显增加,IL-1βmRNA、IL-6 mRNA、TNF-αmRNA的表达明显减少(P<0.01);玻璃酸钠滴眼液组4周与玻璃酸钠滴眼液组2周比较,泪液分泌量、泪膜破裂时间无明显增加,IL-1βmRNA、IL-6 mRNA、TNF-αmRNA的表达无明显改变(P>0.05)。结论:蒙医针刺疗法可以增加干眼症模型大鼠的泪液分泌量,延长泪膜破裂时间,降低角结膜炎症反应,这可能是通过调控IL-1βmRNA、IL-6 mRNA、TNF-αmRNA等炎症相关基因实现的。

Objective:To explore the corneal mechanism of Mongolian medicine acupuncture therapy in the treatment of dry eye model rats by regulating keratitis-related genes.Methods:A total of 60 SPF SD rats were randomly divided into normal group,model group,sodium hyaluronate eye drops group and Mongolian medicine acupuncture group,with 15 rats in each group.Atropine-induced dry eye rat model was established in the other three groups except the normal group.After the rats were modeled,the model group was not treated.The sodium hyaluronate eye drops group was given sodium hyaluronate eye drops,3 times/d,1 drop each time,for 4 weeks.The Mongolian medicine acupuncture therapy group took Heyi point,Xila point and liver point for acupuncture,once a day,15 minutes each time,for 4 weeks.The tear secretion and tear film rupture time of rats in each group were detected before the experiment and 2 weeks and 4 weeks after the last administration/acupuncture 2 h.HE staining was used to observe the changes of corneal morphology.The expression of IL-1βmRNA,IL-6 mRNA and TNF-a mRNA in corneal tissue was detected by RT-PCR.Results:HE staining results showed that the epithelial cell proliferation and basal cell arrangement disorder in the Mongolian medicine acupuncture group were significantly better than those in the model group.Two weeks after acupuncture,compared with the model group,the tear film rupture time and tear secretion of the Mongolian medicine acupuncture group were significantly increased(P<0.01),and the expression of IL-1βmRNA,IL-6 mRNA and TNF-a mRNA in corneal tissue was significantly decreased(P<0.05).Compared with the Mongolian medicine acupuncture group,the tear break-up time and tear secretion of the sodium hyaluronate eye drops group were significantly decreased(P<0.01),and the expression of IL-1βmRNA and TNF-a mRNA in the corneal tissue was significantly increased(P<0.05).At 4 weeks after acupuncture,compared with the model group,the tear film rupture time and tear secretion of the Mongolian medicine acupuncture group were significantly increased(P<0.01),and the expression of IL-1βmRNA,IL-6 mRNA and TNF-αmRNA in the corneal tissue was significantly decreased(P<0.01).Compared with the Mongolian medicine acupuncture group,the tear film rupture time and tear secretion of the rats in the sodium hyaluronate eye drops group were significantly decreased(P<0.01),and the expression of IL-1βmRNA,IL-6 mRNA and TNF-αmRNA in the corneal tissue was significantly increased(P<0.01).There was no significant difference in the expression of SIT,BUT,IL-1βmRNA,IL-6 mRNA and TNF-αmRNA between the Mongolian medicine acupuncture group and the normal group(P>0.05).Compared with the Mongolian medicine acupuncture group at 2 weeks,the SIT and BUT were significantly increased,and the expression of IL-1βmRNA,IL-6 mRNA and TNF-αmRNA was significantly decreased in the Mongolian medicine acupuncture group at 4 weeks(P<0.01).There was no significant difference in the expression levels of SIT,BUT,IL-1βmRNA,IL-6 mRNA and TNF-αmRNA between the sodium hyaluronate eye drops group at 4 weeks and the sodium hyaluronate eye drops group at 2 weeks(P>0.05).Conclusion:Mongolian medicine acupuncture therapy can increase the tear secretion of dry eye model rats,prolong the tear film rupture time,and reduce the inflammatory response of the cornea and conjunctiva,which may be achieved by regulating inflammation-related genes such as IL-1βmRNA,IL-6 mRNA,and TNF-αmRNA.