[1]郑传睿 王蔡未 周莉 童培建 肖鲁伟 单乐天△.血小板裂解液联合脂肪干细胞干预大鼠膝骨关节炎的实验研究[J].中国中医骨伤科杂志,2019,27(08):9-14.
　ZHENG Chuanrui WANG Caiwei ZHOU Li TONG Peijian XIAO Luwei SHAN Letian.Experimental Study on Therapeutic Effect of Platelet Lysate Combined with Adipose-Derived Stem Cells on the Rat Models of Osteoarthritis[J].Chinese Journal of Traditional Medical Traumatology & Orthopedics,2019,27(08):9-14.
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血小板裂解液联合脂肪干细胞干预大鼠膝骨关节炎的实验研究()
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《中国中医骨伤科杂志》[ISSN:1005-0205/CN:42-1340/R]

卷:
第27卷
期数:
2019年08期
页码:
9-14
栏目:
实验研究
出版日期:
2019-07-30

文章信息/Info

Title:
Experimental Study on Therapeutic Effect of Platelet Lysate Combined with Adipose-Derived Stem Cells on the Rat Models of Osteoarthritis
文章编号:
1005-0205(2019)08-0009-06
作者:
郑传睿1 王蔡未1 周莉1 童培建1 肖鲁伟1 单乐天1△
1浙江中医药大学(杭州,310053)
Author(s):
ZHENG Chuanrui1 WANG Caiwei1 ZHOU Li1 TONG Peijian1 XIAO Luwei1 SHAN Letian1△
1Zhejiang Chinese Medical University,Hangzhou 310053,China.
关键词:
骨关节炎;  血小板裂解液;  脂肪干细胞;  软骨;  动物实验
Keywords:
osteoarthritis;  platelet lysate;  adipose-derived stem cells;  cartilage;  animal experimentation
分类号:
R-33
文献标志码:
A
摘要:
目的:观察血小板裂解液(PL)联合脂肪干细胞(ADSCs)对膝骨关节炎(KOA)大鼠关节疼痛和软骨修复的作用。方法:从SD大鼠腹部抽取脂肪制备培养ADSCs,SD大鼠外周血中分离制备PL,以碘乙酸(MIA)法建立大鼠KOA疼痛模型,分别以低、中、高浓度PL,低、中、高浓度ADSCs以及低、中、高浓度PL联合ADSCs对KOA模型大鼠进行双侧关节腔注射干预,每周注射1次,定期观察生理行为及评价疼痛指标,注射4次后取膝关节进行组织病理学观察及Mankin's评分。结果:压痛结果表明:造模后第4周,模型组压痛阈值较正常组显著下降,差异有统计学意义(P<0.01); 低、中、高浓度PL组,高浓度ADSCs组,以及低、中、高浓度联合组压痛阈值分别与模型组比较有显著提高,差异有统计学意义(P<0.01)。热痛结果表明:造模后第4周,模型组的热痛阈值较正常组显著下降,差异有统计学意义(P<0.01); 低、中、高浓度PL组,中、高浓度ADSCs组,以及低、中、高浓度联合组热痛阈值分别与模型组比较有显著提高,差异有统计学意义(P<0.01)。膝关节软骨病理学观察结果:大鼠膝关节软骨Mankin's评分比较,差异有统计学意义(P<0.01); 模型组的Mankin's评分高于空白组,差异有统计学意义(P<0.01); 低、中、高浓度PL组,低、中、高浓度ADSCs组,低、中、高浓度联合组Mankin's评分较模型组显著下降,差异有统计学意义(P<0.01); PL高浓度组的Mankin's评分低于PL低、中浓度组,差异有统计学意义(P<0.01); ADSCs高浓度组的Mankin's评分低于ADSCs低、中浓度组,差异有统计学意义(P<0.01); 联合高浓度组Mankin's评分低于联合低、中浓度组,差异有统计学意义(P<0.01); 联合高浓度组Mankin's评分低于PL高浓度组和ADSCs高浓度组(P<0.01)。结论:PL联合ADSCs可提高 KOA 模型大鼠疼痛阈值,修复软骨损伤,且其作用效果优于PL和ADSCs单独使用,PL和ADSCs在治疗膝骨关节炎上有一定的协同作用,可用于临床治疗。
Abstract:
Objective: To observe the effect of platelet lysate(PL)combined with adipose-derived stem cells(ADSCs)on cartilage repair in rats with knee osteoarthritis(KOA).Methods: The ADSCs with different concentration(1×106/mL,1×107/mL,1×108/mL)were obtained by the collagenase digestion method from fats in 4 SD rats and PL was isolated from their peripheral blood.Another 88 SD rats were seleted and were randomly divided into blank group,KOA model group,PL low-concentration group,PL middle-concentration group,PL high-concentration group,ADSCs low-concentration group,ADSCs middle-concentration group,ADSCs high-concentration group, PL with ADSCs low-concentration group,PL with ADSCs middle-concentration group and PL with ADSCs high-concentration group,8 cases in each group.The KOA models were created in KOA model group,PL low-concentration group,PL middle-concentration group,PL high-concentration group,ADSCs low-concentration group,ADS-Cs middle-concentration group,ADSCs high-concentration group,PL with ADSCs low-concentration group,PL with ADSCs middle-concentration group and PL with ADSCs high-concentration group,by intra-articular injecting iodoacetic acid into bilateral knee joint.One week later,the rats in PL low-concentration group,PL middle-concentration group,PL high-concentration group,ADSCs low-concentration group,ADSCs middle-concentration group,ADSCs high-concentration group,PL with ADSCs low-concentration group,PL with ADSCs middle-concentration group and PL with ADSCs high-concentration group,were administrated with intra-articular injection of PL low-concentration group,PL middle-concentration group,ADSCs low-concentration group,ADSCs middle-concentration group,ADSCs high-concentration group,PL high-concentration group,PL with ADSCs low-concentration group,PL with ADSCs middle-concentration group and PL with ADSCs high-concentration group,with dose of 50 μL in bilateral knee joint,while the rats in the other two groups were administrated with intra-articular injection with the same dose of normal saline,once a week for consecutive 4 weeks.After the end of drug intervention,pressure-pain threshold, paw withdrawal latency(PWL),pathological observation and Mankin's scoring of knee joint were performed on all of the rats. Results: The results of tenderness showed that the tenderness threshold of the model group was significantly lower than that of the normal group(P<0.01).The tenderness threshold of the the low,middle and high concentration PL group,high concentration ADSCs group,the low,middle and high concentration PL with ADSCs group,was significantly higher than that of the model group(P<0.01).The results of thermal pain showed that the model group was significantly lower than that of the normal group(P<0.01).The thermal pain of the PL low-concentration group,PL middle-concentration group,PL high-concentration group,ADSCs middle-concentration group,ADSCs high-concentration group,PL with ADSCs low-concentration group,PL with ADSCs middle-concentration group and PL with ADSCs high-concentration group was significantly higher than that of the model group.(P<0.01).There was statistical difference in Mankin's scores of knee articular cartilage(P<0.01).The pathological results showed that the Mankin's score of the model group was significantly higher than the normal group(P<0.01).the Mankin's score of the PL low-concentration group,PL middle-concentration group,PL high-concentration group,ADSCs low-concentration group,ADSCs middle-concentration group,ADSCs high-concentration group, PL with ADSCs low-concentration group,PL with ADSCs middle-concentration group and PL with ADSCs high-concentration group was significantly lower than the model group(P<0.01),the PL high-concentration group lower than PL low-concentration group and PL middle-concentration group(P<0.01),the ADSCs high-concentration group lower than ADSCs low-concentration group and ADSCs middle-concentration group(P<0.01),the PL with ADSCs high-concentration group lower than PL with ADSCs low-concentration group and PL with ADSCs middle-concentration group(P<0.01),the PL with ADSCs high-concentration group lower than PL high-concentration group and ADSCs high-concentration group(P<0.01).Conclusion: Both the PL,ADSCs and PL with ADSCs can improve joint pain in KOA rats and repair cartilage damage,and PL with ADSCs surpasses PL or ADSCs.This technique can be used for adjuvant treatment of clinical KOA.

参考文献/References:

[1] CLOUET J,VINATIER C,MERCERON C,et al.From osteoarthritis treatments to future regenerative therapies for cartilage[J].Drug Discov Today,2009,14(19/20):913-925. [2] ABRAMSON S B,ATTUR M.Developments in the scientific understanding of osteoarthritis[J].Arthritis Res Ther,2009,11(3):227-235. [3] HILDNER F,EDER M J,HOFER K,et al.Human platelet lysate successfully promotes proliferation and subsequent chondrogenic differentiation of adipose-derived stem cells:a comparison with articular chondrocytes[J].J Tissue Eng Regen Med,2015,9(7):808-818. [4] PEREIRA R C,SCARANARI M,BENELLI R,et al.Dual effect of platelet lysate on human articular cartilage:a maintenance of chondrogenic potential and a transient proinflammatory activity followed by an inflammation resolution[J].Tissue Eng Part A,2013,19(11/12):1476-1488. [5] ZUK P A,ZHU M,MIZUNO H,et al.Multilineage cells from human adipose tissue:implications for cell-based therapies[J].Tissue EngJ,2001,7(2):211-228. [6] TER HUURNE M,SCHELBERGEN R,BLATTES R,et al.Antiinflammatory and chondroprotective effects of intraarticular injection of adipose-derived stem cells in experimental osteoarthritis[J].Arthritis Rheum,2012,64(11):3604-3613. [7] SOFFER E,OUHAYOUN J P,DOSQUET C,et al.Effects of platelet lysates on select bone cell functions[J].Clin Oral Implants Res,2004,15(5):581-588. [8] 贺端端,郭向阳.cAMP-PKA 信号通路介导大鼠骨癌痛的产生和维持[J].中国疼痛医学杂志,2015,21(1):10-14. [9] WANG J,DONG F F,LI X F,et al.Effects of Yiqi Huayu prescription on knee cartilage degenemtion in HIF-1 gene knockout mice[J].China J Orthop Trauma,2013,26(6):486-492. [10] CLOUET J,VINATIER C,MERCERON C,et al.From osteoarthritis treatments to future regenerative therapies for cartilage[J].Drug Discov Today,2009,14(19-20):913-925. [11] SATO M,YAMATO M,HAMAHASHI K,et al.Articular cartilage regeneration using cell sheet technology[J].Anat Rec(Hoboken),2014,297(1):36-43. [12] PIRVU T N,SCHROEDER J E,PEROGLIO M,et al.Platelet-rich plasma induces annulus fibrosus cell proliferation and matrix production[J].Eur Spine J,2014,23(4):745-753. [13] 张希,刘萍.骨性关节炎所致慢性持续性疼痛的发病机制[J].中国骨质疏松杂志,2017,23(2):248-255. [14] 刘,邓廉夫.骨关节炎相关细胞因子及蛋白的研究进展[J].现代生物医学进展,2013,13(1):187-190. [15] 鞠昌军,姜红江,高广凌,等.体外人工膝支具辅助血小板裂解液治疗膝关节骨性关节炎的临床疗效分析[J].中华老年骨科与康复电子杂志,2018,4(4):224-230. [16] SELLBERG F,BERGLUND E,RONAGHI M,et al.Composition of growth factors and cytokines in lysates obtained from fresh versus stored patho-gen-inactivated platelet units[J].Transfus Apher Sci,2016,55(3):333-337. [17] 凌晓宇,周康,姚黎佳,等.血小板裂解液对膝骨关节炎模型大鼠疼痛和软骨损伤的影响及作用机制研究[J].中医正骨,2017,29(11):8-14. [18] 杨雨润,TIAN Hua.自体脂肪原性间充质干细胞修复兔软骨缺损的实验研究[J].中华医学杂志,2008,88(31):2214-2218. [19] 刘兆杰,胡永成,张银光,等.血管内皮生长因子转染后脂肪组织来源干细胞蛋白分泌表达及成骨郭兴德检测[J].中华医学杂志,2013,93(23):1845-1849. [20] 戎露江,夏永法,钱涛,等.富含血小板血浆联合骨髓间充质干细胞治疗骨性关节炎的WOMAC评分分析[J].浙江创伤外科,2017,22(2):294-295.

备注/Memo

备注/Memo:
基金项目:国家自然科学基金资助项目(81774331)通信作者 E-mail:letian.shan@zcmu.edu.cn(收稿日期:2019-03-12)
更新日期/Last Update: 1900-01-01