关节腔注射脂肪间充质干细胞对碘乙酸钠诱导大鼠急性软骨损伤的抗炎及组织学修复作用

doi:10.3877/cma.j.issn.2095-1221.2026.05.002

中华细胞与干细胞杂志(电子版) ›› 2026, Vol. 16 ›› Issue (05) : 266 -277. doi: 10.3877/cma.j.issn.2095-1221.2026.05.002

论著

关节腔注射脂肪间充质干细胞对碘乙酸钠诱导大鼠急性软骨损伤的抗炎及组织学修复作用

李文芳¹, 董孟炜², 杨万彭², 巴特³^,⁴, 南楠³^,⁴, 刘杨²^,³^,⁴, 郝慧琴²^,³^,⁴^,^†(

)

¹030619　晋中，山西中医药大学第一临床学院
²030619　晋中，山西中医药大学基础医学院
³030619　晋中，山西中医药大学国家中医药管理局中医药基因表达调节实验室
⁴030619　晋中，山西省中医药交叉创新工程研究中心

收稿日期:2026-03-23 出版日期:2026-10-01
通信作者: 郝慧琴
基金资助:
国家中医药管理局高水平中医药重点学科建设项目(zyyzdxk-2023022); 山西省优秀研究生导师团队建设项目(2024TD33); 风湿免疫病中西医结合基础研究学科建设项目(2026XK29); 山西省重点实验室建设项目(zyyyjs2024021); 山西省中医药科技创新工程项目(2024KJZY006); 山西省教育厅科技创新项目(2024L269)

Anti-inflammatory and histological repair effects of intra-articular injection of adipose-derived mesenchymal stem cells on acute cartilage injury induced by sodium iodoacetate in rats

Wenfang Li¹, Mengwei Dong², Wanpeng Yang², Te Ba³^,⁴, Nan Nan³^,⁴, Yang Liu²^,³^,⁴, Huiqin Hao²^,³^,⁴^,^†()

¹First Clinical College, Administration of Traditional Chinese Medicine, Shanxi University of Chinese Medicine, Jinzhong 030619, China
²School of Basic Medicine, Administration of Traditional Chinese Medicine, Shanxi University of Chinese Medicine, Jinzhong 030619, China
³Laboratory of Gene Regulation by Traditional Chinese medicine, Administration of Traditional Chinese Medicine, Shanxi University of Chinese Medicine, Jinzhong 030619, China
⁴Engineering Research Center for Cross-Innovation in Chinese Traditional Medicine of Shanxi Province, Jinzhong 030619, China

Received:2026-03-23 Published:2026-10-01
Corresponding author: Huiqin Hao

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引用本文：

李文芳, 董孟炜, 杨万彭, 巴特, 南楠, 刘杨, 郝慧琴. 关节腔注射脂肪间充质干细胞对碘乙酸钠诱导大鼠急性软骨损伤的抗炎及组织学修复作用[J/OL]. 中华细胞与干细胞杂志(电子版), 2026, 16(05): 266-277.

Wenfang Li, Mengwei Dong, Wanpeng Yang, Te Ba, Nan Nan, Yang Liu, Huiqin Hao. Anti-inflammatory and histological repair effects of intra-articular injection of adipose-derived mesenchymal stem cells on acute cartilage injury induced by sodium iodoacetate in rats[J/OL]. Chinese Journal of Cell and Stem Cell(Electronic Edition), 2026, 16(05): 266-277.

目的

探讨脂肪间充质干细胞（ADSCs）对膝骨关节炎（KOA）大鼠软骨损伤及炎症反应的影响及作用持续时间。

方法

将45只雄性SD大鼠随机分为空白组、模型组、治疗组。制备KOA模型14 d后，治疗组大鼠在关节腔注射1 × 10⁶个ADSCs/50 μL，空白组和KOA模型组大鼠注射等量生理盐水，每周1次，连续注射4次，观察4周、8周及12周各指标变化，各时间点设立独立的空白组、模型组及治疗组（n = 5）。监测并记录各组大鼠的机械痛阈值和热痛阈值，取材后通过micro-CT、HE染色、番红O-固绿染色和阿利新蓝染色观察软骨损伤退变程度；免疫组化检测Ⅱ型胶原（COL2A1）、聚集蛋白聚糖（ACAN）的蛋白表达；ELISA法检测关节腔冲洗液中的炎症因子IL-1β、TNF-α的表达水平。采用双因素析因设计方差分析分析分组与观察时间的主效应，两两比较采用Bonferroni事后检验。

结果

各组机械痛阈值、骨体积分数、骨小梁分离度、Mankin评分、OARSI评分、COL2A1及IL-1β的分组主效应具有统计学意义（P 均 < 0.05），时间主效应无统计学意义（P 均 > 0.05）；热痛阈值、骨小梁厚度、骨小梁数目、ACAN、TNF-α的分组主效应与时间主效应均有统计学意义（P 均 < 0.05）。与空白组相比，模型组大鼠机械痛阈值、热痛阈值、骨体积分数、骨小梁厚度及骨小梁数目[4周：（0.92 ± 0.02）比（1.17 ± 0.05）mm^-1，8周：（0.81 ± 0.07）比（1.17 ± 0.04）mm^-1，12周：（0.75 ± 0.03）比（1.16 ± 0.05） mm^-1]均降低，骨小梁分离度、Mankin评分及OARSI评分增加，COL2A1、ACAN蛋白的阳性细胞表达率降低；IL-1β[4 周：（53.87 ± 2.32）比（29.71 ± 3.26）pg/mL，8周：（56.31 ± 5.85）比（27.70 ± 2.69） pg/mL、（58.59 ± 3.36）比（25.34 ± 3.23）pg/mL]和TNF-α表达水平[4周：（142.52 ± 14.33）比（71.52 ± 6.68） pg/mL，8周：（153.94 ± 7.36）比（75.45 ± 2.78）pg/mL、（160.60 ± 10.93）比（75.67 ± 3.20） pg/mL]均升高（P 均 < 0.05）。与模型组相比，治疗组机械痛阈值、热痛阈值升高，骨体积分数、骨小梁厚度及骨小梁数目[4周：（1.09 ± 0.04）比（0.92 ± 0.02）mm^-1，8周：（1.02 ± 0.15）比（0.81 ± 0.07）mm^-1，12周：（1.00 ± 0.05）比（0.75 ± 0.03）mm^-1]均增加，骨小梁分离度、Mankin评分及OARSI评分降低，COL2A1、ACAN蛋白的阳性细胞表达率升高，IL-1β[4 周：（37.15 ± 7.16）比（53.87 ± 2.32） pg/mL，8周：（41.56 ± 7.24）比（56.31 ± 5.85）pg/mL、12周：（42.39 ± 3.41）比（58.59 ± 3.36） pg/mL]和TNF-α表达水平[4周：（84.49 ± 7.12）比（142.52 ± 14.33）pg/mL，8周：（88.22 ± 3.25）比（153.94 ± 7.36） pg/mL、12周：（90.31 ± 5.29）比（160.60 ± 10.93）pg/mL]降低（P 均 < 0.05）。与4周模型组相比，12周模型组的骨小梁厚度增加，骨小梁数目降低，ACAN蛋白的阳性细胞表达率降低，TNF-α的表达水平显著升高（P 均 < 0.05）。

结论

ADSCs可以缓解KOA的疼痛症状，减轻关节炎症，可能通过促进软骨生成修复软骨损伤，且疗效具有一定的时效性，可为临床干细胞治疗KOA的时效特征提供实验依据。

关键词: 脂肪间充质干细胞, 膝骨关节炎, 炎症, 软骨损伤, 远期疗效

Objective

To explore the effects and duration of adipose-derived mesenchymal stem cells (ADSCs) on cartilage injury and inflammatory response in knee osteoarthritis (KOA) rats.

Methods

Forty-five male SD rats were divided into the blank group, model group and treatment group randomly. Fourteen days after the KOA model establishment, the rats in the treatment group were injected with 1×10⁶ ADSCs/50 μL into the articular cavity while rats in the blank group and the KOA model group were injected with the same volume of normal saline once a week for four consecutive injections. The rats were observed at 4 weeks, 8 weeks and 12 weeks (n = 5) after the first injection. Independent blank group, model group and treatment group were set up at each observation time point. The mechanical pain threshold and thermal pain threshold of rats in each group were monitored and recorded. The degree of cartilage degeneration was observed by micro CT, HE-staining, Safranine O-fast green staining and Alcian blue staining. The protein expressions of collagen Ⅱ (COL2A1) and aggrecan (ACAN) were detected by immunohistochemistry. The expression levels of inflammatory factors IL-1β and TNF-α in articular cavity lavage fluid were detected by ELISA. Two-way factorial design ANOVA was used to analyze the main effect of group and observation time for all measurement data, and Bonferroni post-test was used for pairwise comparison.

Results

The main effects of group in mechanical pain threshold, bone volume fraction, trabecular bone separation, Mankin score, OARSI score, COL2A1 and IL-1β in each group were statistically significant (all P < 0.05), while there was no significant difference in the main effects of time (all P > 0.05); The main effects of group and time were significant in terms of thermal pain threshold, trabecular thickness, trabecular number, ACAN and TNF-α (all P < 0.05). Compared with the blank group, the mechanical pain threshold, the thermal pain threshold, bone volume fraction, trabecular thickness and trabecular number [4 weeks: (0.92 ± 0.02) vs (1.17 ± 0.05) mm^-1, 8 weeks: (0.81 ± 0.07) vs (1.17 ± 0.04) mm^-1, 12 weeks: (0.75 ± 0.03) vs (1.16 ± 0.05) mm^-1] as well as positive cell expression rate of COL2A1 and ACAN protein were significantly decreased in the model group, while the bone trabecular separation, Mankin score, OARSI score and the expression levels of IL-1β [4 weeks: (53.87 ± 2.32) vs (29.71 ± 3.26) pg/mL, 8 weeks: (56.31 ± 5.85) vs (27.70 ± 2.69) pg/mL, (58.59 ± 3.36) vs (25.34 ± 3.23) pg/mL] and TNF-α [4 weeks: (142.52 ± 14.33) vs (71.52 ± 6.68) pg/mL, 8 weeks: (153.94 ± 7.36) vs (75.45 ± 2.78) pg/mL, (160.60 ± 10.93) vs (75.67 ± 3.20) pg/mL] were significantly increased (all P < 0.05). Compared with the model group, the mechanical pain threshold, the thermal pain threshold, the bone volume fraction, trabecular thickness, and trabecular number [4 weeks: (1.09 ± 0.04) vs (0.92 ± 0.02) mm^-1, 8 weeks: (1.02 ± 0.15) vs (0.81 ± 0.07) mm^-1, 12 weeks: (1.00 ± 0.05) vs (0.75 ± 0.03) mm^-1] and positive cell expression rate in the COL2A1 and ACAN protein was significantly increased in the treatment group, while the bone trabecular separation, Mankin score, OARSI score and the expression level of IL-1β [4 weeks: (37.15 ± 7.16) vs (53.87 ± 2.32) pg/mL, 8 weeks: (41.56 ± 7.24) vs (56.31 ± 5.85) pg/mL, 12 weeks: (42.39 ± 3.41) vs (58.59 ± 3.36) pg/mL] and TNF-α [4 weeks: (84.49 ± 7.12) vs (142.52 ± 14.33) pg/mL, 8 weeks: (88.22 ± 3.25) vs (153.94 ± 7.36) pg/mL, 12 weeks: (90.31 ± 5.29) vs (160.60 ± 10.93) pg/mL] were significantly decreased (all P < 0.05). Compared with the 4-week model group, the trabecular thickness and the expression level of TNF-α in the 12-week model group were significantly increased, while the number of trabeculae and the positive cell expression rate of ACAN protein were significantly decreased (all P < 0.05).

Conclusions

ADSCs can relieve the pain symptoms of KOA, reduce joint inflammation, and may repair cartilage damage by promoting chondrogenesis, which has a certain timeliness, providing experimental basis for certain timeliness characteristics of clinical stem cell therapy for KOA.

Key words: Adipose-derived mesenchymal stem cells, Knee osteoarthritis, Inflammation, Cartilage injury, Long-term efficacy

图1 膝骨关节炎大鼠造模方法

图2 ADSCs的形态与增殖曲线注：ADSCs为脂肪间充质干细胞；a图为倒置生物显微镜下的ADSCs形态观察（×100）；b图为ADSCs的增殖曲线，10%、2.5%为培养基中胎牛血清的含量，2.5 : 1、1 : 1为培养基中胰岛素样生长因子-1体积与青链霉素体积的比值，n = 5

图3 流式细胞术对ADSCs特异性表面标志物分析结果注：ADSCs为脂肪间充质干细胞；流式结果显示ADSCs高表达CD73、CD90和CD105，低表达CD34和CD45

图4 倒置生物显微镜下观察ADSCs成脂/成骨/成软骨三系诱导分化及相应染色鉴定（×200）注：a图为ADSCs成脂分化鉴定，油红O染色显示脂滴阳性；b图为ADSCs成骨分化鉴定，茜素红染色显示钙盐沉积阳性；c图为ADSCs成软骨分化鉴定，阿利新蓝染色显示酸性黏多糖阳性

图5 ADSCs对KOA大鼠机械痛、热痛阈值的影响注：^*P < 0.05，^**P < 0.01，^***P < 0.001，n = 5

图6 ADSCs对KOA软骨立体光学解剖结构的影响注：随着时间的进展，模型组大鼠膝关节面逐渐粗糙并伴炎性增生，治疗组炎性增生有所减少，软骨表面附着新生的透明软骨

图7 ADSCs对KOA大鼠膝关节micro-CT骨微结构及骨参数的影响注：a图为大鼠膝关节2D、3D冠状面和矢状面，随着时间进展，模型组骨侵蚀加重，骨赘增多，治疗组骨侵蚀有所减轻；b ~ e图为各组大鼠膝关节骨参数：骨体积分数、骨小梁厚度、骨小梁分离度、骨小梁数目，^*P < 0.05，^**P < 0.01，^***P < 0.001，n = 3

图8 ADSCs对KOA大鼠膝关节的病理学结果及评分的影响注：a图为切片扫描仪观察大鼠膝关节HE染色、番红O-固绿染色、阿利新蓝染色，随着时间进展，模型组糖胺聚糖与胶原蛋白丢失愈发明显，骨损伤严重，治疗组糖胺聚糖与胶原蛋白表达有所恢复，骨损伤有所改善（×100）；b、c图为各组大鼠Mankin评分和OARSI评分比较，^*P < 0.05，^**P < 0.01，^***P < 0.001，n = 5

图9 ADSCs对KOA大鼠软骨组织中COL2A1、ACAN蛋白表达的影响注：a图为大鼠膝关节软骨组织中COL2A1、ACAN蛋白的免疫组化染色结果，随着时间进展，模型组的COL2A1、ACAN蛋白表达减少，治疗组COL2A1、ACAN蛋白表达有所改善（×200）；b、c图为各组大鼠软骨组织中COL2A1、ACAN蛋白的阳性细胞表达率比较。^***P < 0.001，n = 3

表1 ADSCs对KOA大鼠关节腔滑液炎症因子表达的影响（

± s）

分组	样本量	IL-1β （ pg/mL）			TNF-α （ pg/mL）
分组	样本量	4周	8周	12周	4周	8周	12周
空白组	5	29.71 ± 3.26	27.70 ± 2.69	25.34 ± 3.23	71.52 ± 6.68	75.45 ± 2.78	75.67 ± 3.20
模型组	5	53.87 ± 2.32^a	56.31 ± 5.85^a	58.59 ± 3.36^a	142.52 ± 14.33^ac	153.94 ± 7.36^a	160.60 ± 10.93^a
治疗组	5	37.15 ± 7.16^b	41.56 ± 7.24^b	42.39 ± 3.41^b	84.49 ± 7.12^b	88.22 ± 3.25^b	90.31 ± 5.29^b

表1 ADSCs对KOA大鼠关节腔滑液炎症因子表达的影响（

± s）

分组	样本量	IL-1β （ pg/mL）			TNF-α （ pg/mL）
分组	样本量	4周	8周	12周	4周	8周	12周
空白组	5	29.71 ± 3.26	27.70 ± 2.69	25.34 ± 3.23	71.52 ± 6.68	75.45 ± 2.78	75.67 ± 3.20
模型组	5	53.87 ± 2.32^a	56.31 ± 5.85^a	58.59 ± 3.36^a	142.52 ± 14.33^ac	153.94 ± 7.36^a	160.60 ± 10.93^a
治疗组	5	37.15 ± 7.16^b	41.56 ± 7.24^b	42.39 ± 3.41^b	84.49 ± 7.12^b	88.22 ± 3.25^b	90.31 ± 5.29^b

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Anti-inflammatory and histological repair effects of intra-articular injection of adipose-derived mesenchymal stem cells on acute cartilage injury induced by sodium iodoacetate in rats

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Anti-inflammatory and histological repair effects of intra-articular injection of adipose-derived mesenchymal stem cells on acute cartilage injury induced by sodium iodoacetate in rats