Protective effect and potential molecular mechanism of galangin from Alpinia officinarum Hance on knee osteoarthritis in rats

Osteoarthritis (OA) is the most common form of joint disease with over half of all people older than 65 years demonstrating radiographic changes of osteoarthritis in the knees. Honey is known to contain bioactive compounds that exert chondroprotective effects by counteracting the homeostatic dysregulation of the joint. However, its effect on the radiographic features of osteoarthritis has not been proven. This study was carried out to evaluate the effect of honey on radiographic features of monosodium iodoacetate (MIA)- induced knee osteoarthritis in female Wistar rats. Thirty female Wistar rats were randomly divided into five groups of six animals each. Animals in group one were healthy (control) rats, while animals in groups two to five were subjected to experimental osteoarthritis of the right knee joint induced by a single intra-articular injection of 1mg of MIA. The animals in groups two, three, four, and five were treated with normal saline (1ml/kg b. w.), arthocare (glucosamine/chondroitin sulfate 6.67/8.33mg/kg b. w.), low dose honey (250mg/kg b. w.) and high dose honey (1,000mg/kg b. w.) respectively. All treatments were administered orally once daily using an oral cannula for twenty-one days. All animals were subjected to radiographic assessment of the right knee joint before and after induction of OA, and after treatment. High and low-dose honey reversed the loss of joint space; sclerosis of the tibial plateau, medial, and lateral femoral condyles, when compared to the arthocare-treated and untreated groups. In conclusion, honey improved radiographic features of knee osteoarthritis in a rat model induced by monosodium iodoacetate.

BackgroundOur previous clinical evidence suggested that the direct application of “Sanse powder” the main ingredient of “Yiceng” might represent an alternative treatment for knee osteoarthritis. However, the mechanism underlying its effect is poorly understood. In this study, we investigated the mechanism of the effect of direct “Sanse powder” application for the treatment of knee osteoarthritis (KOA) in rats by using lipidomics.MethodsKOA rats were established by cutting the anterior cruciate ligament, and the cold pain threshold and mechanical withdrawal threshold (MWT) of seven rats from each group were measured before modelling (0 days) and at 7, 14, 21 and 28 days after modelling. Histopathological evaluation of the synovial tissue was performed by haematoxylin and eosin (H&E) staining after modelling for 28 days. Interleukin-1β (IL-1β), pro-interleukin-1β (pro-IL-1β) and tumor necrosis factor-α (TNF-α) proteins in synovial tissue were measured by western blot, and the mRNA expression levels of IL-1β and TNF-α in synovial tissue were measured using Real-time reverse transcription polymerase chain reaction (qRT-PCR), the levels of IL-1β and TNF-α in rat serum were measured by enzyme-linked immunosorbent assay (ELISA), Serum lipid profiles were obtained by using ultra-performance liquid chromatography combined with quadrupole-Exactive Orbitrap mass spectrometry (UPLC-Q-Exactive Orbitrap MS).ResultsThe results confirmed that the direct application of “Sanse powder” had a significant protective effect against KOA in rats. Treatment with “Sanse powder” not only attenuated synovial tissue inflammation but also increased the levels of the cold pain threshold and MWT. In addition, the lipidomics results showed that the levels of diacylglycerol (DAG), triacylglycerols (TAGs), lysophosphatidylcholine (LPC), phosphatidylcholine (PC), fatty acid esters of hydroxy fatty acids (FAHFAs), and phosphatidylethanolamine (PE) were restored almost to control levels following treatment.ConclusionsLipidomics provides a better understanding of the actions of direct application “Sanse powder” therapy for KOA.

ObjectiveTo investigate the correlation between melatonin and osteoarthritis (OA) in rats. To explore the relevant mechanisms in the occurrence and development of osteoarthritis in rats, and to further understand the disease of osteoarthritis.MethodsForty healthy 6‐month‐old male SD rats were randomly divided into two groups: sham and drug intervention groups. Pre‐OA modeling, enzyme‐linked immunosorbent assay was employed to detect the levels of IL‐1β, IL‐6, COX‐2, and melatonin in the serum of the rats in each group. For OA modeling, we administered an injection of papain into the knee cavity of all rats. The levels of IL‐1β, IL‐6, and COX‐2 in the serum of rats in each group were detected 2 weeks after the modeling. Additionally, 2 weeks after the modeling, the rats in the drug intervention group were intraperitoneally injected with melatonin antagonists. The rats in the sham group were intraperitoneally injected with normal saline for 2 weeks. The levels of IL‐1β, IL‐6, and COX‐2 in the serum of each group were measured at the second, third, and fourth weeks after the drug intervention, and the levels of melatonin in the serum were measured at the second week after the drug intervention. Finally, the rats were euthanized by cervical dislocation, and pathological sections were collected from the knee joint to observe the pathological tissue changes under a microscope, and Mankin score was determined. The independent samples t‐test method was used for analysis.ResultsThe imaging examination after the drug intervention showed that the modeling of knee osteoarthritis in rats was successful. In the pathological findings, HE staining showed a legible cartilage structure of each layer, with cartilage proliferation and partial cartilage tearing to the radial layer. The tide line was intact; toluidine blue staining revealed more obvious changes. The differences among the mean values of IL‐6, IL‐1β, and COX‐2 measured in each period were statistically significant (t = 5.50, p < 0.05). The measured mean values of IL‐6, IL‐1β, and COX‐2 revealed statistically significant differences among the groups (t = 2.01, p < 0.05). The intergroup comparison of the Mankin scores in each period showed statistically significant differences.ConclusionMelatonin may inhibit inflammation and associated oxidative stress on the surface of knee cartilage. It may be related to the repair and regeneration of articular surface cartilage during the development of OA in the rat knee joint.

To observe the similarities and differences of effects of moxibustion at "Zusanli" (ST36) on target tissues and macrophages polarization in knee osteoarthritis (KOA) and rheumatoid arthritis (RA) rats, and to summarize its efficacy and characteristics. Thirty rats were equally and randomly divided into control, KOA, RA, KOA+Moxi and RA+Moxi groups. The KOA model and RA model were induced by injection of sodium monoiodoacetate or Freund's complete adjuvant into the rats' knee joints, respectively. Rats of the KOA+Moxi and RA+Moxi groups received moxibustion stimulation at bilateral ST36 for 30 min, once a day for 21 days, beginning from the 7th day on after modeling. The contents of serum interleukin(IL)-1β and IL-10 were detected by ELISA. Histopathological changes (Markin score of the knee cartilage and synovial pathology score) of the knee joints were observed after HE staining. The polarization state of M1 and M2 macrophages in the synovial tissue of the knee joints was assessed by detecting the expression of CD86 and CD206 after immunofluorescence staining. Compared with the control group, the content of serum IL-1β, synovial pathology score, and synovial CD86 expression were significantly increased (P<0.01, P<0.05), while the content of serum IL-10 and synovial CD206 expression markedly decreased (P<0.01) in both KOA and RA groups；the Markin score was increased (P<0.01) in the KOA group. In comparison with the KOA group, the Markin score was obviously decreased (P<0.01), while the content of serum IL-10 and CD206 expression were apparently increased (P<0.01) in the KOA+Moxi group. No significant changes were found in the content of serum IL-1β, synovial pathology score and CD86 expression in the KOA+Moxi group relevant to the KOA group. In comparison with the RA group, the content of serum IL-1β, synovial pathology score, and CD86 expression were considerably decreased (P<0.01) in the RA+Moxi group. No marked differences were found in the serum IL-10 level, Markin score, and CD206 expression between RA+Moxi and RA model groups. The increased Markin score was significantly higher in the KOA group than in the RA group (P<0.01), but the increased synovial pathology score was significantly lower in the KOA group than in the RA group (P<0.01). Correspondingly, the effect of moxibustion at ST36 was significantly better in RA model than in KOA model in reducing serum IL-1β (P<0.05). Moxibustion at ST36 can effectively reduce cartilage injury of knee joint in rats with KOA and reduce synovial injury in rats with RA, which may be related with its effects in lowering IL-1β level in RA model by inhibiting the polarization of M1 macrophages, and up-regulating level of IL-10 in KOA model by promoting the polarization of M2 macrophages. However, the relevant mechanism needs to be further studied.

The role of amentoflavone on cartilage injury in knee osteoarthritis (KOA) rats and the underlying mechanismwere explored. KOA rat and IL-1β-stimulated chondrocyte models were constructed. MTT, colony formation, and ELISA were performed to determine the cytotoxicity, cell proliferation, and inflammatory factors. The role of PTGS2 in IL-1β-stimulated chondrocytes was also confirmed through transfecting PTGS2 overexpression and silencing plasmids. Further, we analyzed how amentoflavone regulated PTGS2 to improve IL-1β-stimulated chondrocytes in vitro. Additionally, we analyzed the expression of PTGS2 after amentoflavone treatment. In vivo, HE and Safranin-O staining were carried out, and the inflammatory response was detected by ELISA and HE staining. In addition, we also analyzed the regulatory effect of amentoflavone on PTGS2 and explored the mechanism effect of PTGS2 in vitro and in vivo. The results indicated that PTGS2 was the downstream molecule of amentoflavone, which was highly expressed in IL-1β-stimulated chondrocytes and KOA rats, and amentoflavone decreased PTGS2 expression. We also confirmed the potential role of amentoflavone on KOA, which was also characterized by the repair of cartilage injury, reduction of inflammatory infiltration, and improvement of functional disability. Consistent with in vivo results, in vitro experiments gave the same conclusions. Amentoflavone reduced PTGS2 expression in IL-1β-stimulated chondrocytes and inhibited inflammation of chondrocytes via PTGS2. Collectively, the results confirmed that this drug was the potential targeted drug for KOA, whose repair effect on cartilage injury was partly related to PTGS2.

Bushen Qiangjin capsule inhibits the Wnt/α-catenin pathway to ameliorate papain-induced knee osteoarthritis in rat.

Objects Osteoarthritis is the most common joint disease and a major cause of functional limitation and pain in adults. This study aims to investigate the effect of wogonoside (WOG) on the progression of knee osteoarthritis (KOA) in model rats. Materials and methods Rats KOA models were established and treated with different doses of WOG (10 mg/kg, 20 mg/kg and 30 mg/kg). The degree of cartilage injury was detected by Mankin scores via HE/Alcian blue staining. The levels of IFN-γ and IL-4 in peripheral blood and synovial fluid and the Th1/Th2 ratio were detected by flow cytometry. The model mice were injected with NF-κB p65 or ERK1/2 inhibitors or activators to further investigate the effect of WOG on KOA. Results WOG significantly improved cartilage tissue damage and reduced the Mankins score. WOG down-regulated the level of IFN-γ while up-regulated the expression of IL-4, which maintained the balance of Th1/Th2 cells. Further studies showed that the expression of NF-κB p65, phosphorylated p65, cytoplasmic ERK1/2 and nuclear ERK1/2 were all inhibited by WOG. The results of reverse verification experiments showed that the activator of NF-κB p65 and ERK1/2 weakened the protective effect of WOG on KOA, and the inhibitor of NF-κB p65ERK1/2 enhanced the protective effect of WOG on KOA. Conclusions WOG inhibited the activation of NF-κB and ERK1/2 to alleviate the articular cartilage injury and Th1/th2 cytokine infiltration in KOA rats.

ObjectiveWe aim to explore whether acupuncture inhibits inflammation and bone destruction in rat model monosodium iodoacetate (MIA)-induced knee osteoarthritis (KOA) by 18F-fluorodeoxyglucose (18F-FDG) small-animal positron emission tomography (PET) and micro-computed tomography (CT) imaging.MethodsKOA was induced in rats by intra-articular injection MIA (2 mg/50 μL) through the right knee of the rats. Forty male Sprague Dawley rats weighing 280 to 340 g (12 weeks old) were randomly divided into four groups including Control group, KOA group, KOA plus manual acupuncture group (KOA+MA), KOA plus sham acupuncture group (KOA+SA). The acupuncture treatment lasted for three weeks (one-day rest after six days of treatment). Paw withdrawal threshold test and open-field test were used to assess mechanical allodynia and locomotor activity respectively for once a week. Hematoxylin and eosin (H&E) staining was used to assess the damage of the cartilage, synovium and infrapatellar fat pad (IFP). 18F-FDG PET was performed to quantify joint inflammation. The influence on the subchondral bone in these rats was confirmed by micro-CT.ResultsMechanical hyperalgesia, joint inflammation, and obvious bone destruction were observed in the KOA group. H&E staining of the knee joint found that manual acupuncture played a protective effect in cartilage, synovium and IFP destruction. However, compared with KOA group, the results in sham acupuncture had no significant difference. After manual acupuncture treatment in KOA rats, inflammation was significantly suppressed shown by 18F-FDG PET imaging. Micro-CT analysis of the knee joint revealed that manual acupuncture protected bone by inhibiting osteophyte development and subchondral bone remodeling.ConclusionThe results of 18F-FDG PET and micro-CT showed that manual acupuncture inhibited inflammation and bone destruction, which provides reliable evidence for the effectiveness of acupuncture in hindering development of KOA, and provides reliable evidence for clinical application of acupuncture.

Acupuncture with near-infrared irradiation using needles loaded with curcumin-polydopamine film attenuates papain-induced knee osteoarthritis in rats.

To compare the effects of moxibustion and scraping of "Yanglingquan" (GB34) and "Xuehai" (SP10) area on changes of bioactive substances in the region of acupoints in rats with knee osteoarthritis (KOA). SD rats were randomly divided into blank, model, moxibustion, scraping, and moxibustion + scraping (combination) groups, with 8 rats in each group. The KOA model was established by injecting 50 μL 0.9% sodium chloride solution into the right knee cavity. Fourteen days after modeling, GB34 and SP10 on the right limb were stimulated by moxibustion (10 min) or scraping (till regional flush) once every other day for 7 times. The mechanical paw withdrawal threshold (PWT) and thermal withdrawal latency (TWL) were tested by Von Frey and hot stabbing instrument, separately. The pathological changes of the right knee joint were observed by HE staining. The serotonin (5-HT) contents of skin tissues in the region of acupoint GB34 and SP10 were detected by ELISA. The expression levels of substance P (SP) and calcitonin gene-related peptide (CGRP) in GB34 and SP10 region skin tissues were detected by Western blot. Compared with the blank group, the PWT and TWL of the rats in the model group were significantly decreased (P<0.001), while the contents of 5-HT and the expression levels of SP and CGRP in GB34 and SP10 region skin tissues were significantly increased (P<0.001, P<0.01). Following intervention and in comparison the with the model group, the TWL and PWT of rats in the three treatment groups were significantly increased (P<0.01), the content of 5-HT and the expression levels of SP and CGRP in GB34 and SP10 region skin tissues were significantly decreased (P<0.01, P<0.001, P<0.05). Except for the expression levels of CGRP, the above indexes of the combination group were significantly superior to those of the moxibustion and scraping groups (P<0.05, P<0.01). Findings of HE staining showed severe damaged cartilage, few chondrocytes on the surface, with subchondral neovascularization in the model group, which was relatively milder in the moxibustion, scraping, and combination groups. Moxibustion and scraping can relieve knee joint pain in KOA rats, which may be associated with its function in down-regulating the expression levels of SP and CGRP, and the content of 5-HT. The therapeutic effect of moxibustion plus scraping is better than that of moxibustion and scraping alone.

To investigate the correlation between osteopontin(OPN) and cartilage oligomeric matrix protein (COMP) levels in synovial fluid of patients with knee osteoarthritis(KOA) and the severity of the disease. A total of 59 patients with KOA admitted to our hospital from February 2018 to May 2020 were selected as the KOA group, including 25 males and 34 females, age ranged 60 to 75 years old with an average of(65.57±1.56) years old, the body mass index(BMI) ranged 21.4 to 30.7(26.12±1.54) kg/m2. After admission, X-ray examination was performed, and Kellgren-Lawrence(K-L) grading system was used to evaluate the X-ray examination results. There were 14 cases in gradeⅡ(K-L2 group), 27 cases in grade Ⅲ(K-L3 group), and 18 cases in grade Ⅳ(K-L4 group). Eighteen patients who underwent arthroscopy for ligament or meniscus disease without cartilage damage were selected as control group, including 7 males and 11 females, age ranged 61 to 78 years old with an average of (64.88±1.60) years old, BMI ranged 22.8 to 29.9(25.89±1.49) kg/m2. Before treatment, synovial fluid samples of subjects were collected, and the OPN and COMP levels of synovial fluid were detected by Elisa. The OPN and COMP levels of synovial fluid in KOA group and control group were compared. The clinical data of KOA patients with different K-L grades were collected, including gender, age and BMI. The biochemical indices of interleukin-1 β(IL-1β), OPN, COMP and matrix metalloproteinase 3(MMP-3) in synovitic fluid were detected by enzyme-linked immunoassay, and the clinical data and biochemical indices of KOA patients with different K-L grades were compared. Logistic regression was used to analyze the factors affecting the K-L classification of KOA patients, and the area under the ROC curve(AUC) was used to predict the severity of KOA. All the 59 patients were followed up for 8 to 27(15.75±3.27) months. The levels of OPN and COMP in synovial fluid in KOA group were significantly higher than those in control group (t=16.991, 17.387, P<0.001). The levels of IL-1β, OPN, COMP and MMP-3 in synovitic fluid were significantly different among those in different K-L grade KOA patients(P<0.001). Compared with the K-L2 group, the levels of IL-1β, OPN, COMP, and MMP-3 in the synovial fluid of K-L3 and K-L4 were increased (P<0.05). Compared with the K-L3 group, the levels of IL-1β, OPN, COMP, and MMP-3 in the K-L4 joint were increased (P<0.05). Multivariate Logistic regression analysis showed that the levels of OPN, COMP and MMP-3 were independent risk factors for K-L grading of KOA patients(OR=6.653, 4.229, 1.579, P<0.001). AUC of OPN in synovial fluid predicting K-L4 KOA was 0.720[95%CI(0.588-0.851)], and the sensitivity was 94.4%. the specificity was 65.9%. The AUC of COMP in synoviac fluid predicting K-L4 KOA was 0.731[95%CI(0.592-0.870)], the sensitivity was 88.9%, the specificity was 63.4%. The AUC of OPN combined with COMP in synoviac fluid predicting K-L4 KOA was 0.839 [95%CI(0.724-0.953)], the sensitivity was 94.4%, and the specificity was 51.2%. The AUC of OPN combined with COMP in synoviac fluid predicting K-L4 grade KOA was greater than that of OPN and COMP alone(Z=4.037, 3.540, P<0.05). The levels of OPN and COMP in synovial fluid increase in patients with KOA, and they increase with the increase of K-L grade. Synovitic fluid OPN and COMP are independent risk factors affecting K-L grade of KOA patients, and they have high AUC, sensitivity and specificity in predicting of K-L4 KOA, and can be used to evaluate the progression of KOA disease.

Quadriceps muscles play a pivotal role in knee osteoarthritis (OA) progression and symptom manifestation, particularly pain. This research investigates the therapeutic effectiveness of muscle enhancement and support therapy (MEST), a recently developed device intended for intramuscular insertion of cog polydioxanone filaments, in quadriceps restoration to alleviate OA pain. Knee OA was induced in Sprague Dawley rats via monoiodoacetate injections. MEST or sham treatment was performed in OA or Naive rat quadriceps. Pain was assessed using paw withdrawal threshold and weight bearing. Quadriceps injury and recovery via MEST were evaluated using biomarkers, tissue morphology, muscle mass, contractile force and hindlimb torque. Satellite cell and macrophage activation, along with their activators, were also assessed. Data were compared at 1- and 3-weeks post-MEST treatment (M-W1 and M-W3). MEST treatment in OA rats caused muscle injury, indicated by elevated serum aspartate transferase and creatinine kinase levels, and local β-actin changes at M-W1. This injury triggered pro-inflammatory macrophage and satellite cell activation, accompanied by heightened interleukin-6 and insulin-like growth factor-1 levels. However, by M-W3, these processes gradually shifted toward inflammation resolution and muscle restoration. This was seen in anti-inflammatory macrophage phenotypes, sustained satellite cell activation and injury markers regressing to baseline. Quadriceps recovery in mass and strength from atrophy correlated with substantial OA pain reduction at M-W3. This study suggests that MEST-induced minor muscle injury triggers macrophage and satellite cell activation, leading to recovery of atrophied quadriceps and pain relief in OA rats.

This study investigated the efficacy of transplantation of allogeneic adipose-derived stem cells (ADMSCs) in rats with knee osteoarthritis (KOA) and the effect on the expression of matrix metalloproteinase 13 (MMP-13) and discoid domain receptor 2 (DDR2). In total, sixty rats were randomly selected. Eleven rats were selected as the blank group. Forty-four rat KOA models were established, and the remaining 5 rats were used for stem cell extraction. The rats were randomly divided into two groups, and the transplantation group was treated with ADMSCs transplantation. The KOA group was intragastrically administered with saline. The expressions of MMP-13 mRNA and DDR2 in rats were detected by RT-qPCR and immunohistochemistry. Correlation analysis was performed in MMP-13 mRNA and DDR2 expression levels in the KOA rats. After treatment, the indexes of Lequesne MG knee joints, MMP-13 mRNA and DDR2 in the transplanted rats were significantly lower than those in the KOA group (P<0.05). In the KOA rats, MMP-13 mRNA and DDR2 was positively correlated (r=0.830, P<0.001). Therefore, the transplantation of ADMSCs has a significant effect on the KOA rats, which can effectively improve the knee joint function of KOA rats and reduce the expression of MMP-13 mRNA and DDR2 in rats, and it is worthy of clinical promotion.

In order to elucidate the therapeutic effect and mechanism of action of Zhengqing Fengtongning Sustained-release Tablets on knee osteoarthritis, this study created a knee osteoarthritis model using 0.2 mL 40 g·L~(-1) papain and randomly divided the rats into the model group, high-dose and low-dose groups of Zhengqing Fengtongning Sustained-release Tablets, and celecoxib group. All groups were given the drug for four weeks, with the diameter of their knee joint being measured during this period. Hematoxylin-eosin staining and Senna solid green staining were utilized to observe the pathology of knee joint tissue in SD rats. The initial therapeutic impact of Zhengqing Fengtongning Sustained-release Tablets on knee osteoarthritis in rats was assessed by monitoring the levels of interleukin-1β(IL-1β) and interleukin-6(IL-6) in the plasma. Using a combination of non-targeted metabolomics and 16S rRNA techniques, researchers determined the variations in endogenous molecules and intestinal flora in rats and identified potential biomarkers. The results showed that Zhengqing Fengtongning Sustained-release Tablets improved the diameter of knee joint swelling, ameliorated the pathological damage of cartilage tissue, and reduced the plasma levels of IL-1β and IL-6 in rats with knee osteoarthritis. Metabolomics analysis identified 22 potential biomarkers associated with the modulatory effects of Zhengqing Fengtongning Sustained-release Tablets, including 5-hydroxytryptamine, corticosterone, methylmalonic acid, and other biomarkers, which were mainly involved in eight metabolic pathways, including tryptophan metabolism, vitamin K metabolism, steroid synthesis, and so on. The results of intestinal flora showed a decrease in the diversity of intestinal flora in the model group, an increase in the diversity of intestinal flora, and an improvement in the microecology of intestinal flora. Significant differences were found in Lachnospiraceae＿NK4A136＿group, Helicobacter, Lactobacillus, Bacteroides, and Parabacteroides. Finally, the results of the combined analysis showed that 22 biomarkers were correlated with five genera. The above results indicate that Zhengqing Fengtongning Sustained-release Tablets can improve the tissue morphology and structure of knee joints, reduce the level of plasma inflammatory factors, regulate the diversity of intestinal flora, and balance the metabolic pathways of steroid synthesis, vitamin K metabolism, and tryptophan metabolism to exert a therapeutic effect on knee osteoarthritis.

This study aimed to investigate the therapeutic effects of ELASEM®Flex and ELASEM®ProFlex, two eggshell membrane (EM) products, on sodium iodoacetate (MIA)-induced osteoarthritis (OA) in rats. An OA model was established by a single intra-articular injection of MIA into the knee joint. After modeling, rats were administered diclofenac sodium, ELASEM®Flex, and ELASEM®ProFlex by gavage daily for 4 consecutive weeks. During the experiment, food intake, water intake, body weight, and plantar mechanical pain threshold (MPT) of rats were measured weekly. Serum levels of TNF-α, COX-2, IL-1β, and CTX-II were assessed at weeks 2 and 4. After 4 weeks, knee joints were harvested for histopathological examination (HE staining and Safranin-O fast green staining). Results indicated that knee joints of OA rats showed significant swelling, which was alleviated to varying degrees in all treatment groups. Both ELASEM®Flex and ELASEM®ProFlex significantly increased the MPT (P < 0.05) and demonstrated sustained analgesic effects. These treatments also significantly reduced the serum levels of IL-1β, COX-2, TNF-α, and CTX-II at weeks 2 and 4 (P < 0.05). Histopathological analysis revealed that both EM preparations markedly alleviated arthritis symptoms, improved cartilage structure, promoted chondrocyte proliferation, enhanced staining of chondrocytes and cartilage matrix, and resulted in significantly lower Mankin's scores compared to the OA model group (P < 0.05). These results indicate that ELASEM®Flex and ELASEM®ProFlex can exert preventive and reparative effects on knee OA in rats by alleviating arthritis pain, inhibiting inflammatory factor expression, reducing type II collagen degradation, and promoting chondrocyte proliferation.