Abstract: BACKGROUND: Preliminary studies have found that Rongjin Niantong Fang can effectively inhibit the degradation of cartilage matrix. Long non-coding RNA GAS5 can act as a competitive endogenous RNA that adsorbs miR-21-5p to regulate the expression of matrix metalloproteins, thereby influencing the metabolism of articular cartilage matrix. However, whether Rongjin Niantong Fang regulates cartilage matrix metabolism through long non-coding RNA GAS5/miR-21 and plays a role in preventing and treating osteoarthritis needs to be further studied.
OBJECTIVE: To explore the mechanism by which Rongjin Niantong Fang treats knee osteoarthritis by long non-coding RNA GAS5/miR-21 pathway regulating cartilage matrix synthesis and catabolism. 
METHODS: Sixty male SPF-grade Sprague-Dawley rats aged 8 weeks were randomized into four groups (n=15 per group): blank group, model group, treatment group and control group. Knee osteoarthritis models were established using the modified Hulth method in the latter three groups. The blank group and the model group were intragastrically administered with normal saline, the treatment group was intragastrically administered with water extract of Rongjin Niantong Fang, and control group was intragastrically administered with glucosamine hydrochloride capsules, once a day for 12 weeks. After treatment, microstructure and morphological changes of cartilage were observed by hematoxylin-eosin staining and safranin O-fast green staining. The mRNA expression of long non-coding RNA GAS5, miR-21, and matrix metabolic factors in cartilage was detected by real-time PCR. The protein expression of matrix metabolic factors in cartilage was detected by western blot. 
RESULTS AND CONCLUSION: In the model group, the four layers of the cartilage were disordered and arranged irregularly, the tide line and adhesion line were irregular or disappeared, chondrocytes proliferated and arranged disorderly, the subchondral bone proliferated, and a large amount of collagens and proteoglycans were lost. In the treatment and control groups, the four layers of the cartilage and tide line were recognizable, chondrocytes arranged regularly, the subchondral bone structure was relatively intact, and part of collagens and proteoglycans were lost. Compared with the blank group, the expression levels of long non-coding RNA GAS5, matrix metalloproteinases 3, 9, 13, and a disintegrin and metalloproteinase with thrombospondin motif-5 mRNAs and proteins in cartilage were significantly increased in the model group (P < 0.01 or P < 0.05), while the expression levels of miR-21, tissue inhibitors of metalloproteinase 3, type II collagen, and Aggrecan proteins and mRNAs in cartilage were significantly decreased in the model group (P < 0.01 or P < 0.05). Compared with the model group, the expression trends of these genes and proteins in cartilage were reversely changed in the treatment and control groups (P < 0.01 or P < 0.05). To conclude, Rongjin Niantong Fang can reduce the loss of collagen and proteoglycan in cartilage matrix, delay the degradation of cartilage matrix, and reduce the damage of cartilage morphology and structure by down-regulating the expression of long non-coding RNA GAS5, matrix metalloproteinases 3, 9, 13, and a disintegrin and metalloproteinase with thrombospondin motif-5, and up-regulating the expression of miR-21, tissue inhibitors of metalloproteinase 3, type II collagen, and Aggrecan in cartilage of knee osteoarthritis rats.

Key words: Rongjin Niantong Fang, knee osteoarthritis, long non-coding RNA GAS5, miR-21, cartilage matrix metabolism