中医正骨2025，Vol.37Issue(2)：4-12,9.
肾髓安康饮干预小鼠膝骨关节炎的效果和安全性及作用机制研究
Efficacy,safety,and mechanism of Shensui Ankang Yin(肾髓安康饮)against knee osteoarthritis in mice:an experimental study
摘要
Abstract
Objective:To observe the outcomes and safety of Shensui Ankang Yin(肾髓安康饮,SSAKY)against knee osteoarthritis(KOA)in mice,and to explore its mechanism.Methods:Thirty C57BL/6J mice were selected and randomized into sham-operated group,model group,and SSAKY group.The mice in model group and SSAKY group were subjected to destabilization of the medial meniscus(DMM)on the right hind limbs to build KOA models,while the ones in sham-operated group were merely incised the skin at the corre-sponding site and then sutured after exposing the knee joints.From day 2 after the modeling surgery,the mice in SSAKY group were inter-vened by intragastric administration with SSAKY concentrate(the crude drug concentration was 1.6 g/mL),once a day,0.5 mL at a time for consecutive 8 weeks;while the ones in model group and sham-operated group with the same dosage of normal saline.After 8-week inter-vention,the mechanical and thermal pain thresholds of the mice were measured by Up-down method and hot plate test,respectively,and the gait of the mice was analyzed using the DigiGait gait analysis system.After that,the mice were executed,and the knee joints were harvested from their right hind limbs,and scanned by Micro-CT scanner to measure the bone volume fraction(BVF)of the subchondral bone in tibial plateau.Furthermore,the knee joint cartilage tissue sections were prepared and stained with alcian blue-hematoxylin/orange G(ABH/OG)to observe the degeneration of knee joint cartilage,and then evaluate by using Osteoarthritis Research Society International(OARSI)scoring criteria,apart from that,the expression of type Ⅱ collagen and matrix metalloproteinase(MMP)-13 in the knee joint cartilages was detected by employing immunohistochemical staining.In addition,the tissue sections of the subchondral bone of mice knee joints and the right dorsal root ganglion(DRG)of L4 were prepared for detecting the expression of nerve growth factor(NGF)by immunofluorescence staining;besides,the heart,liver,spleen,lung and kidney of mice were harvested to make tissue sections for observing the histopathological changes via HE staining.Results:①The pain threshold.The 50％paw withdrawal threshold(PWT)was lower,and the reaction time of the right hind paw was shorter in model group compared to sham-operated group(P=0.000,P=0.000);while,the 50％PWT was higher,and the reaction time of the right hind paw was longer in SSAKY group compared to model group(P=0.000,P=0.000).②The gait.The model group ex-hibited longer swing phase duration,shorter stance phase duration,smaller stride length and paw contact area in the right hind limbs com-pared to the sham-operated group(P=0.001,P=0.000,P=0.000,P=0.000);while,the SSAKY group showed shorter swing phase du-ration,longer stance phase duration,greater stride length and paw contact area compared to the model group(P=0.001,P=0.000,P=0.000,P=0.000).③Micro-CT analysis on the knee joints.The obviously abnormal subchondral bone remodeling was observed in the tibial plateau in mice of model group,while that was significantly improved in SSAKY group compared to model group.The BVF of subchondral bone in tibial plateau was greater in model group compared to sham-operated group,while it was smaller in SSAKY group compared to model group(P=0.000,P=0.000).④The pathological observation of knee joint cartilage tissues.The changes,manifesting as severely worn car-tilage with thinned and lost cartilage layer and fissures extending deep into the subchondral bone,were observed in the knee joints of mice in model group;while,compared with that of model group,the knee joint cartilage injury in mice of SSAKY group was significantly im-proved,only presented as partially damaged cartilage surface and a relatively intact matrix structure.The OARSI score was higher in model group compared to sham-operated group,while it was lower in SSAKY group compared to model group(P=0.000,P=0.000).⑤The ex-pression of type Ⅱ collagen and MMP-13 in the knee joint cartilage.The positive expression area ratio of type Ⅱ collagen in knee joint carti-lage was smaller,while that of MMP-13 was greater in model group compared to sham-operated group(P=0.000,P=0.000);The positive expression area ratio of type Ⅱ collagen in knee joint cartilage increased,while that of MMP-13 decreased in SSAKY group compared to mo-del group(P=0.000,P=0.000).⑥The expression of NGF in the subchondral bone of knee joint and the right DRG of L4.The positive expression area ratio of NGF in the subchondral bone of knee joint and the right DRG of L4 was greater in model group compared to sham-operated group(P=0.000,P=0.000),while it was smaller in SSAKY group compared to model group(P=0.000,P=0.000).⑦The pathological observation of visceral tissues in mice.The pathological changes failed to be observed in the heart,liver,spleen,lung,or kidney in the mice of the 3 groups.Conclusion:SSAKY can effectively alleviate the pain sensitization and cartilage degeneration in KOA mice with high safety.It may work by inhibiting the expression of NGF and regulating the cartilage extracellular matrix metabolism.关键词
骨关节炎,膝/肾髓安康饮/疼痛/敏化/软骨细胞/神经生长因子/小鼠Key words
osteoarthritis,knee/Shensui Ankang Yin/pain/sensitization/chondrocytes/nerve growth factor/mice引用本文复制引用
李雪枫,王玉杭,邹凯奥,温经渊,陈文哲,袁文华,肖鲁伟,金红婷..肾髓安康饮干预小鼠膝骨关节炎的效果和安全性及作用机制研究[J].中医正骨,2025,37(2):4-12,9.基金项目
国家自然科学基金项目(82274280) （82274280）
浙江省基础公益研究计划项目(LR23H270001) （LR23H270001）
