【作者】 王茂林；

【导师】 陈世益； 李云霞；

【作者基本信息】 复旦大学 ， 运动医学， 2009， 硕士

【摘要】 骨骼肌钝挫伤是运动医学领域中常见的一类创伤,发生率10%～55%不等,损伤后骨骼肌愈合时间长且多为疤痕愈合,严重影响运动功能,是目前运动医学领域研究的热点与难点。研究表明骨骼肌损伤后可立即再生,但骨骼肌再生能力有限,结构及功能恢复常常不完全,愈合缓慢,并伴有一定程度的瘢痕化,易再次发生损伤。研究发现骨骼肌损伤后的早期,损伤局部的微血管首先遭到破坏,出现细胞膜损害,内皮细胞肿胀,微血管渗透性增加,周围血管出血,甚至无法控制的血栓形成,从而导致膜磷脂退变,自由基的聚集以及炎性反应诱导的氧化应激,进一步加剧周围正常骨骼肌组织的损害。国内外研究表明,钝挫伤导致的微循环紊乱和继发的组织损害在损伤后24h时最为严重。局部微循环障碍、损伤细胞渗出物氧化均可导致组织活性氧簇(ROS)的过度生成,ROS可进一步损害脂质膜从而加重骨骼肌细胞的损伤,甚至坏死,而抗氧化酶(超氧化物歧化酶、谷胱甘肽过氧化物酶、过氧化氢酶)可对抗氧化应激损伤,保护骨骼肌细胞。因此如何早期改善局部微循环,减少自由基的生成,提高抗氧化酶的活性,已成为骨骼肌损伤修复的研究热点之一。我们先前的研究已经证明黄芪丹参注射液具有促进骨骼肌肌球蛋白重链表达、缩短肌肉愈合时程以及提高愈合质量的作用,但其促进愈合的机制目前尚不清楚。本研究目的旨在通过观察黄芪丹参注射液对骨骼肌钝挫伤后氧化应激及早期微循环的影响,探讨黄芪丹参注射液在促进骨骼肌愈合过程中是否具有抗氧化应激、改善微循环、保护损伤骨骼肌的作用,从而为其进一步的临床应用提供实验依据本实验第一部分研究黄芪丹参注射液对大鼠急性骨骼肌钝挫伤后氧化应激的影响。通过测定骨骼肌钝挫伤以后组织中脂质过氧化产物丙二醛,抗氧化酶(超氧化物歧化酶、谷胱甘肽过氧化物酶、过氧化氢酶)的活性来探讨黄芪丹参注射液是否是通过清除自由基,提高抗氧化酶活性从而促进了骨骼肌损伤后的愈合本实验第二部分研究黄芪丹参注射液对大鼠急性骨骼肌钝挫伤后微循环的影响。通过测定骨骼肌钝挫伤后大鼠全身血流动力学的改变,骨骼肌的湿、干重比值,以及损伤局部微动脉的管径变化,毛细血管流速变化,观察给予黄芪丹参注射液后是否具有改善微循环,减轻水肿从而促进损伤骨骼肌的愈合。第一部分黄芪丹参注射液对大鼠急性骨骼肌钝挫伤后氧化应激的影响目的:观察黄芪丹参注射液对大鼠骨骼肌钝挫伤后修复过程中氧化应激的影响方法:72只雄性SD大鼠随机分为中药治疗组、生理盐水组、自然愈合组,每组24只。采用打击装置造成右侧腓肠肌中段钝挫伤模型,分别于损伤局部注射黄芪丹参注射液、生理盐水、不作处理。另6只做空白对照(未经任何处理)。于损伤后1、4、7和14天取腓肠肌测定丙二醛(MDA)含量、过氧化氢酶(CAT)活性、超氧化物歧化酶(SOD)活性、谷胱甘肽过氧化物酶(GSH-px)活性。结果:与生理盐水组和自然愈合组相比,中药组腓肠肌SOD、GSH-px的活性在1、4、7天明显升高(P＜0.05),而CAT活性则无明显变化(P＞0.05)。中药组腓肠肌MDA含量在伤后4、7天明显低于生理盐水组和自然愈合组(P＜0.05)。结论:黄芪丹参注射液可明显减少氧化产物MDA含量,减轻骨骼肌钝挫伤愈合过程中的氧化性损伤,并显著提高骨骼肌还原酶SOD和GSH-px的活性。第二部分黄芪丹参注射液对大鼠急性骨骼肌钝挫伤后微循环的影响目的:观察黄芪丹参注射液对大鼠骨骼肌钝挫伤后修复过程中局部微循环的影响。方法:78只雄性SD大鼠随机分为中药治疗组、生理盐水组、自然愈合组和空白对照组4组。前3组采用打击装置造成右侧腓肠肌中段钝挫伤模型,分别于损伤局部注射黄芪丹参注射液、等量生理盐水以及不注射药物。于损伤后2h、24h、72h、120h在体观察腓肠肌的微循环情况。空白对照组为正常大鼠,未经任何处理,同时进行微循环的观察作为对照结果:与生理盐水组和自然愈合组相比,中药治疗组微动脉直径在第24h、72h明显扩张(P＜0.05),且毛细血管血流量在24h时明显增加(P＜0.05)。与空白对照组相比,各组的湿/干重比(W/D)在2h、24h时明显升高(P＜0.05),但中药治疗组升高程度明显低于生理盐水组和自然愈合组(P＜0.05)。各组间血流动力学无明显差异(P＞0.05)结论:骨骼肌钝挫伤后早期给予黄芪丹参注射液可明显改善损伤骨骼肌的局部微循环情况,并减轻组织水肿程度,从而达到促进骨骼肌损伤愈合的作用更多还原

【Abstract】 Skeletal muscle injury presents the most common injury in sports medicine,whose incidence varies from 10%-55%.Skeletal muscle injury presents a challenging problem in sports medicine,as injured muscles heal very slowly and often with scar recovery, which severe influence motor function.It has been observed that injured muscles can initiate regeneration promptly,but the healing process is often inefficient and hindered by the formation of scar tissue,which may contribute to the tendency for muscle injury to recur.It has been reported that skeletal muscle contusion at early can direct trauma to microvessels results in membrane damage,endothelial cell swelling,perivascular hemorrhage,and uncontrolled clot formation.leading to additional local ischemia,in turn,induces further degradation of membrane phospholipids,accumulation of free radicals,and inflammation-induced oxidative stress may aggravate evolving tissue damage,previous experimental studies have shown that maximum posttraumatic microcirculatory disturbances and secondary tissue damage evolve over time and are not observed before 24 hours after contusion.The presence of local microcirculation damage and inflammation reactive in muscle may lead to overproduction of reactive oxygen species.These species may damage lipid membranes and produce cell necrosis, but antioxidant enzymes such as superoxide dismutase,catalase and glutathione peroxidase could reduce oxidative stress and protect skeletal muscle cell.Thus,early restoration local microcirculations,reduce free radicals,elevate antioxidant enzymes activities are becoming a hot research.Our previous experimental studies have shown that Chinese medicine HuangqiDanshen could promote skeletal muscle expression of myosin heavy chain,short the healing process of injured muscle and enhance regeneration of injured muscle.However the detailed mechanism underlying beneficial effects of Huangqidanshen on skeletal muscle contusion remain to be established. Therefore,the aims of this experimental study are to asses the effects of HuangqiDanshen on oxidative stress and microcirculation after acute skeletal muscle contusion.In the part one of our study,we observe the effect of Chinese medicine HuangqiDanshen on oxidative stress in rat skeletal muscles after acute contusion. The present study aimed to investigate the effect of HuangqiDanshen on parameters of oxidative stress,namely lipid peroxidation product malondialdehyde(MDA) and the activities of antioxidant enzymes,catalase(CAT),superoxide dismutase(SOD) and glutathione peroxidase(GSH-px),in skeletal muscle after contusion.Thus,to establish whether use of HuangqiDanshen could scavenge free radical,elevate antioxidant enzymes activities in the process of skeletal muscle after contusion.In the part two of our study,we observe the effect of Chinese medicine HuangqiDanshen on microcirculation in rat skeletal muscles after acute contusion. This study quantitatively analyzed the change of blood hemodynamics,the edema index of skeletal muscle,the changes of arteriolar internal diameter,microvascular flow rate.Therefore,to observe whether administration HuangqiDanshen could amelioration microcirculation injury,attenuate tissue edema,and promote the healing progress of the contused skeletal muscle. Part One Effects of Chinese Medicine HuangqiDanshen on Oxidative Stress in Rat Skeletal Muscles after Acute ContusionObjective To investigate the effects of Chinese medicine HuangqiDanshen on oxidative stress in rat skeletal muscles after acute contusion..Methods Seventy two male Sprague-Dawley rats were randomly divided into three groups(24 rats each group):(1) Chinese medicine HuangqiDanshen group;(2) physiological saline group;(3) natural healing group.The muscle contusion was produced at the gastrocnemius by specific device;Local injection by Chinese medicine HuangqiDanshen at injured site;group physiological saline treated with the injection of physiological saline,and group natural healing for control.The remaining six rats were used as normal control.The muscles were sampled at the day 1,4,7 and 14 after injury respectively,and then investigated the parameters of oxidative stress,namely MDA and the activities of antioxidant enzymes(CAT,SOD and GSH-px) in gastrocnemius after contusion..Result Compared with the physiological saline group and natural healing group,SOD and GSH-px activities in Chinese medicine group were significantly increase after contusion(P＜0.05).However,the CAT activity had no change(P＞0.05).The concentrations of MDA in Chinese medicine group were lower than that of physiological saline group and natural healing group at days 4,7 after injury(P＜0.05)Conclusion The Chinese medicine HuangqiDanshen could significantly reduce the content of oxidative product MDA,protect the skeletal muscles in the healing process from oxidative injury and markedly increase the reducing enzyme SOD,GSH-px activities of contused skeletal muscle. Part Two Effects of Chinese Medicine HuangqiDanshen on Microcirculation in Rat Skeletal Muscles after Acute ContusionObjective To investigate the effects of Chinese medicine HuangqiDanshen on microcirculation in rat skeletal muscles after acute contusion.Methods Seventy eight male Sprague-Dawley rats were randomly divided into four groups:(1) Chinese medicine treatment group(A);(2) physiological saline group(B); (3) natural healing group(C);(4) control group(D).The acute skeletal muscle contusion model was produced at the right gastrocnemius by specific device.Local injection at the injury site with Chinese medicine HuangqiDanshen in group A;local injection with physiological saline in group B;The group C without intervention at injured site,and the blank group D as control.Microcirculations of gastrocnemius in vivo were observation at 2h,24h,72h and 120h respectively after contusion.Rats that were not injured served as control.Result Compared with the group B and group C,the internal diameters of arteriole in group A were significantly dilation at 24h and 72h,the blood flows of capillary were markedly increase at 24h after contusion(P＜0.05).Compared with group D,the Wet/ Dry Weight Ratios of each group were significantly increase at 2h and 24h,but edema in group A was less pronounced compared with group B or group C.The hemodynamics,however,have no significantly changed in each group(P＞0.05).Conclusions Administration Chinese medicine HuangqiDanshen at early could amelioration microcirculation injury,attenuate tissue edema,and promote the healing process of the contused skeletal muscles.更多还原