摘要
目的探讨Kartogenin(KGN)联合脂肪干细胞(adipose-derived stem cells,ADSCs)对兔前交叉韧带(anterior cruciate ligament,ACL)重建术后腱骨愈合的影响。方法取原代ADSCs传代培养,将第3代细胞与10μmol/L KGN溶液培养72 h后,取KGN-ADSCs上清液与纤维蛋白胶以1∶1比例混合备用;另取第3代ADSCs与纤维蛋白胶混合作为对照。取成年新西兰大白兔80只,随机分为4组,分别为生理盐水组(A组)、ADSCs组(B组)、KGN-ADSCs组(C组)、假手术组(D组)。A~C组制备ACL重建模型后,分别于腱骨界面和肌腱间隙中注射生理盐水、ADSCs与纤维蛋白胶混合液、KGN-ADSCs上清液与纤维蛋白胶混合液;D组仅暴露ACL,不作其他处理。术后观察动物一般情况,于6、12周取腱骨界面组织及ACL标本,通过HE染色、c-Jun氨基末端激酶(c-Jun N-terminal kinase,JNK)免疫组织化学染色、TUNEL细胞凋亡检测观察腱骨愈合情况,计数成纤维细胞,测算JNK蛋白阳性表达率及细胞凋亡指数(apotosis index,AI);同时行抗拉力强度测试,测量最大载荷及最大拉伸距离,观察生物力学性能。结果术后28只动物因切口感染或死亡排除研究,最终A~D组分别12、12、12、16只纳入研究。术后A、B组腱骨界面愈合欠佳,C组愈合良好;6、12周时C组成纤维细胞计数、JNK蛋白表达率均高于A、B、D组,差异均有统计学意义(P<0.05);与6周比较,C组术后12周成纤维细胞计数减少、AI及JNK蛋白表达率下降,差异均有统计学意义(P<0.05)。生物力学测试示,术后6、12周,C组最大载荷高于A、B组、但低于D组,而最大拉伸距离结果相反,但组间差异均有统计学意义(P<0.05)。结论兔ACL重建后局部注射KGN-ADSCs可促进腱骨愈合,增强腱骨界面机械强度和抗拉力性。
Objective To investigate the effect of Kartogenin(KGN)combined with adipose-derived stem cells(ADSCs)on tendon-bone healing after anterior cruciate ligament(ACL)reconstruction in rabbits.Methods After the primary ADSCs were cultured by passaging,the 3rd generation cells were cultured with 10μmol/L KGN solution for 72 hours.The supernatant of KGN-ADSCs was harvested and mixed with fibrin glue at a ratio of 1∶1;the 3rd generation ADSCs were mixed with fibrin glue as a control.Eighty adult New Zealand white rabbits were taken and randomly divided into 4 groups:saline group(group A),ADSCs group(group B),KGN-ADSCs group(group C),and sham-operated group(group D).After the ACL reconstruction model was prepared in groups A-C,the saline,the mixture of ADSCs and fibrin glue,and the mixture of supernatant of KGN-ADSCs and fibrin glue were injected into the tendon-bone interface and tendon gap,respectively.ACL was only exposed without other treatment in group D.The general conditions of the animals were observed after operation.At 6 and 12 weeks,the tendon-bone interface tissues and ACL specimens were taken and the tendon-bone healing was observed by HE staining,c-Jun N-terminal kinase(JNK)immunohistochemical staining,and TUNEL apoptosis assay.The fibroblasts were counted,and the positive expression rate of JNK protein and apoptosis index(AI)were measured.At the same time point,the tensile strength test was performed to measure the maximum load and the maximum tensile distance to observe the biomechanical properties.Results Twenty-eight rabbits were excluded from the study due to incision infection or death,and finally 12,12,12,and 16 rabbits in groups AD were included in the study,respectively.After operation,the tendon-bone interface of groups A and B healed poorly,while group C healed well.At 6 and 12 weeks,the number of fibroblasts and positive expression rate of JNK protein in group C were significantly higher than those of groups A,B,and D(P<0.05).Compared with 6 weeks,the number of fibroblasts gradually decreased and the positive expression rate of JNK protein and AI decreased in group C at 12 weeks after operation,with significant differences(P<0.05).Biomechanical tests showed that the maximum loads at 6 and 12 weeks after operation in group C were higher than in groups A and B,but lower than those in group D,while the maximum tensile distance results were opposite,but the differences between groups were significant(P<0.05).Conclusion After ACL reconstruction,local injection of a mixture of KGN-ADSCs and fibrin glue can promote the tendon-bone healing and enhance the mechanical strength and tensile resistance of the tendon-bone interface.
作者
陈刚
郑乾
刘蒙飞
何海洋
巨啸晨
姜侃
CHEN Gang;ZHENG Qian;LIU Mengfei;HE Haiyang;JU Xiaochen;JIANG Kan(Department of Joint Surgery,the Sixth Affiliated Hospital of Xinjiang Medical University,Urumqi Xinjiang,830000,P.R.China)
出处
《中国修复重建外科杂志》
CAS
CSCD
北大核心
2023年第8期1002-1010,共9页
Chinese Journal of Reparative and Reconstructive Surgery
基金
新疆维吾尔自治区卫生健康青年医学科技人才专项科研项目(WY-202219)。
关键词
Kartogenin
脂肪干细胞
前交叉韧带重建
腱骨愈合
兔
Kartogenin
adipose-derived stem cells
anterior cruciate ligament reconstruction
tendon-bone healing
rabbit
作者简介
通信作者:巨啸晨,Email:31841528@qq.com。
