摘要
目的探索富血小板血浆(PRP)与软骨细胞力学信号转导通路的协同机制,并将该机制应用于PRP治疗大鼠软骨损伤的修复治疗。方法体外实验部分,采用酶消法获取大鼠软骨细胞,给与PRP处理和(或)流体剪切力刺激(摇床每日3 h,40 r/min),分组为对照组、PRP组、力学+PRP组、先力学后PRP组,通过实时定量反转录聚合酶链反应(RT-qPCR)法检测Ⅱ型胶原、蛋白聚糖及整合素(Integrinβ2)的mRNA表达。体内实验部分,大鼠经木瓜蛋白酶注射后建立膝关节炎/软骨损伤模型,分为对照组(制动+注射生理盐水)、PRP组(制动+注射PRP),运动+PRP组(运动+注射PRP),运动后PRP组(先运动+后注射PRP),4周后取材切片进行苏木精-伊红(HE)、番红O染色评估软骨修复程度,并通过PCR测定Ⅱ型胶原、蛋白聚糖的mRNA表达。两组间比较行One way-ANOVA及LSD-t检验。结果体外实验正常培养的大鼠软骨细胞,PRP及力学+PRP均促进了Ⅱ型胶原及蛋白聚糖转录增多,但差异无统计学意义(3.085±1.166比2.153±0.705/0.818±0.116比1.045±0.266,t=0.561、1.550,P>0.05),然而先力学刺激后给予PRP处理则Ⅱ型胶原及蛋白聚糖转录增多最明显,显著高于PRP组及力学+PRP组(倍数为5.308±0.978及2.845±1.100,t=4.439、5.000、3.695、5.245,P<0.01)。这一机制可能与力学反应性Integrinβ2表达升高有关。该体外实验趋势与大鼠体内实验结果相符合。大鼠对照组软骨损伤明显且表面缺损,PRP组及运动+PRP组软骨损伤有所恢复但表面仍有不平整。大鼠给与先主动运动后PRP治疗方案后,软骨损伤恢复最佳,Ⅱ型胶原纤维及蛋白聚糖mRNA含量最高(倍数分别为46.833±7.400及15.177±3.288,t=6.161、2.683、5.109、3.545,P<0.05)。结论PRP通过整合素β2受体,与软骨细胞的力学反应性信号转导存在协同效应,PRP治疗宜结合运动后PRP的组合方式,提高软骨修复治疗的疗效。
Objective To explore the synergistic mechanism between platelet-rich plasma(PRP)and chondrocyte mechanotransduction,which was applied to improve the repair effects of PRP to cartilage injury in rats.Methods In vitro,rat chondrocytes were obtained by enzymatic digestion,given PRP treatment and(or)fluid shear force stimulation(shaking 3 h per day,40 r/min),divided into control group,PRP group,pro-mechanical+PRP group.The expression of typeⅡcollagen,aggrecan and integrinβ2 mRNA was detected by real-time quantitative reverse transcriptase-polymerase chain reaction(RT-qPCR).In vivo,rats were injected with papain to establish knee arthritis/cartilage injury model,and the animals were divided into control group(external fixation+injection of normal saline),PRP group(external fixation+injection of PRP),exercise+PRP group(exercise+injection of PRP),and pro-exercise PRP group(pro-exercise+PRP injection later).The slices were taken after 4 weeks for hematoxylin and eosin(HE)and safranin O staining to evaluate the degree of cartilage repair,and RT-qPCR was used to determine the mRNA expression of typeⅡcollagen and aggrecan.One way-ANOVA and LSD-t test were performed for comparison between the two groups.Results In normally cultured rat chondrocytes in vitro,PRP and mechanical+PRP all promoted the transcription of typeⅡcollagen and proteoglycan,but there was no statistically significant difference(3.085±1.166 vs.2.153±0.705/0.818±0.116 vs.1.045±0.266,t=0.561,1.550,P>0.05).TypeⅡcollagen and proteoglycan transcription increased most significantly,significantly higher than those in the PRP group and mechanical+PRP group(fold changes of 5.308±0.978 and 2.845±1.100,t=4.439,5.000,3.695,5.245,P<0.01).This mechanism may be related to the increased expression of mechanically responsive Integrinβ2.The in vitro experimental trend was consistent with the rat in vivo experimental results.In the rat control group,the cartilage damage was obvious and the surface was defective.In the PRP group and the exercise+PRP group,the c
作者
谢莹
付苏
杨乾坤
严旭
刘欣
董树岭
陈李影慧
尚春风
郑赛磊
刘宏建
Xie Ying;Fu Su;Yang Qiankun;Yan Xu;Liu Xin;Dong Shuling;Chen Liyinghui;Shang Chunfeng;Zheng Sailei;Liu Hongjian(Department of Blood Transfusion,the First Affiliated Hospital of Zhengzhou University,Zhengzhou 450052,China;Department of Orthopedics,the First Affiliated Hospital of Zhengzhou University,Zhengzhou 450052,China)
出处
《中华实验外科杂志》
CAS
北大核心
2023年第9期1795-1798,共4页
Chinese Journal of Experimental Surgery
基金
中国博士后科学面上基金(2020M682359)。
