Exosomes derived from miR-146a-overexpressing fibroblast-like synoviocytes in cartilage degradation and macrophage M1 polarization: a novel protective agent for osteoarthritis?

Introduction: Pathological changes in the articular cartilage (AC) and synovium are major manifestations of osteoarthritis (OA) and are strongly associated with pain and functional limitations. Exosome-derived microRNAs (miRNAs) are crucial regulatory factors in intercellular communication and can influence the progression of OA by participating in the degradation of chondrocytes and the phenotypic transformation in the polarization of synovial macrophages. However, the specific relationships and pathways of action of exosomal miRNAs in the pathological progression of OA in both cartilage and synovium remain unclear. Methods: This study evaluates the effects of fibroblast-like synoviocyte (FLS)-derived exosomes (FLS-Exos), influenced by miR-146a, on AC degradation and synovial macrophage polarization. We investigated the targeted relationship between miR-146a and TRAF6, both in vivo and in vitro, along with the involvement of the NF-κB signaling pathway. Results: The expression of miR-146a in the synovial exosomes of OA rats was significantly higher than in healthy rats. In vitro, the upregulation of miR-146a reduced chondrocyte apoptosis, whereas its downregulation had the opposite effect. In vivo, exosomes derived from miR-146a-overexpressing FLSs (miR-146a-FLS-Exos) reduced AC injury and chondrocyte apoptosis in OA. Furthermore, synovial proliferation was reduced, and the polarization of synovial macrophages shifted from M1 to M2. Mechanistically, the expression of TRAF6 was inhibited by targeting miR-146a, thereby modulating the Toll-like receptor 4/TRAF6/NF-κB pathway in the innate immune response. Discussion: These findings suggest that miR-146a, mediated through FLS-Exos, may alleviate OA progression by modulating cartilage degradation and macrophage polarization, implicating the NF-κB pathway in the innate immune response. These insights highlight the therapeutic potential of miR-146a as a protective agent in OA, underscoring the importance of exosomal miRNAs in the pathogenesis and potential treatment of the disease.

Extracellular Vesicle-Mediated miR-150-3p Delivery in Joint Homeostasis: A Potential Treatment for Osteoarthritis?

BACKGROUND: The disruption of joint homeostasis is a critical event during the process of joint injury in osteoarthritis (OA). As regulatory molecules, microRNAs (miRNAs) can be released from secretory cells and delivered to recipient cells through extracellular vesicles (EVs), thereby playing an important role in regulating joint homeostasis. We hypothesized that the fibroblast-like synoviocytes (FLSs) in healthy joints could release EVs enriched in miRNAs that can maintain joint homeostasis by regulating the signal transduction pathways in the joints, whereby the articular cartilage (AC) is protected from degeneration, and OA progression is delayed. METHODS: Via high-throughput sequencing and qPCR, we found that miR-150-3p was enriched in the circulating EVs in healthy rats. Next, we established an in vitro cell model in which chondrocytes were cultured with (i) FLSs transfected with miR-150-3p mimics or (ii) EVs released by FLSs (FLS-EVs) inside the healthy synovial membrane (SM). The transportation mechanism from FLSs to chondrocytes was studied using the EV inhibitor GW4869, and the FLSs were transfected with a miR-150-3p mimic or inhibitor. To assess the therapeutic effect of miR-150-3p-carrying EVs (EVs-150) in vivo, healthy FLS-derived EVs (H-FLS-EVs) were injected into the tail vein of rats with OA at various stages of the pathogenesis and evaluated for the progression of OA. RESULTS: The chondrocytes could uptake fluorescent-labeled miR-150-3p mimics and FLS-EVs, and GW4869 suppressed this uptake. The overexpression of miR-150-3p could significantly reduce the concentrations of pro-inflammatory cytokines in the cell culture medium and the expression of the miR-150-3p target T cell receptor-interacting molecule 14 (Trim14), as well as the innate immune-related factors, including nuclear factor kappa B (NF-κB) and interferon-ß (IFN-ß). Similarly to the in vitro findings, the miR-150-3p level in the serum EVs was significantly upregulated among the EV-treated rats. In the AC of the OA rat model injected with H-FLS-EVs, the joint degeneration was suppressed, and Type II collagen (COLII) and aggrecan (ACAN) were significantly upregulated, whereas the innate immune-related factors Trim14, NF-κB, and IFN-ß were downregulated compared with the levels in the untreated OA rats. Notably, the suppression of joint degeneration was more significant when H-FLS-EVs were administered at the early stages of OA rather than the late stages. CONCLUSION: H-FLS-EVs protect chondrocyte function and maintain joint homeostasis by modulating the innate immune response by suppressing the Trim14/NF-κB/IFNß axis. These effects are achieved through the EV-mediated transport of miR-150-3p from the FLSs to the chondrocytes. Our findings show that EV-mediated miR-150-3p can be used to suppress OA, thus providing a novel therapeutic strategy. Additionally, the EV-mediated miR-150-3p transport may also serve as a potential biomarker in the diagnosis, treatment, and prognosis of OA.

[Correlation of genes expression on MyD 88-dependent signaling pathway in progression of knee osteoarthritis].

OBJECTIVE: To investigate expression features and correlation of genes expression on MyD88-dependent signaling pathway in synovial membrane (SM) of progression of knee osteoarthritis (OA). METHODS: Sixty Wistar rats were randomly divided into 6 groups, including blank group (N), false surgical group, model groups[2 weeks (2W), 4 weeks (4W), 8 weeks (8W) and 12 weeks (12W)], with 10 rats in each group. The models were established by using Hulth method. Control group was experienced no surgery, while false surgical group was only opened joint cavity and sutured. The SM samples was collected according to the time designed above. The relative expression quantity of MyD88, TLR4 and NF-κB was detected by Real-time PCR after the extraction of the total RNA and reverse transcription. The correlation analysis was obtained by SPSS. RESULTS: There was no significant difference in each gene mRNA expression between false surgical and blank group(P> 0.05), while enhanced expression was found in the model groups(P<0.05). The correlation index among MyD88, TLR4 and NF-κB was 0.91 and 0.86 respectively, and had significant difference among them. CONCLUSIONS: Positively relative among MyD88, TLR4 and NF-κB played main role in TLR4/NF-κB signal passway, and could predicate the expression of other genes in the passway. It also could further provide the basis for clarify the pathologic mechanism of knee OA.

Histomorphology and innate immunity during the progression of osteoarthritis: Does synovitis affect cartilage degradation?

Osteoarthritis (OA) is a common chronic degenerative disease that affects all joints. At present, the pathological processes and mechanisms of OA are still unclear. Innate immunity, a key player in damage to the structure of the joint and the mechanism by which the host attempts to repair OA, affects all pathological stages of the disease. In the present study, our aim was to assess changes in innate immunity during the pathological processes of OA in articular cartilage (AC) and the synovial membrane (SM), which are the major structures in joints, and to systematically examine the histological changes in AC and SM in mild, moderate and severe cases of OA, in order to further speculate about the manner in which the interactions of AC and SM are facilitated by innate immunity. Histological methods (including HE and Safranin O-fast green staining), immunofluorescent double staining, TUNEL stain, and Western blots were used to assess the morphological changes within AC and SM tissues in healthy and mild, moderate, or severe OA rats. Our results showed that the damage to AC and SM within the joints progressively worsened in different degrees during the course of the disease, and that the innate immune system was closely involved in the AC and SM during each stage of OA. These findings also confirmed that SM may affect the pathological changes in AC through the innate immune system, and therefore affect the progress of OA.

[Study on the clinical effect of the massage method of micro-regulating with vertical cross pressing lying on one side in treating cervicogenic headache].

OBJECTIVE: To evaluate the clinical effects of the massage method of micro-regulating with vertical cross pressing lying on one side in treating cervicogenic headache (CEH). METHODS: Total 136 patients with CEH were collected in the study from August 2012 to April 2014. They were divided randomly into two groups according to random digits table. Sixty-nine patients accepted the treatment of micro-regulating with vertical cross pressing lying on one side (pressing micro-regulating group), including 29 males and 40 females with an average age of (50.55 ± 11.38) years old; 67 patients received the treatment of traditional massage (traditional massage group), including 28 males and 39 females with an average age of (51.20 ± 11.90) years old. Clinical effect was observed according to the standard of curative effect of State Administration of Traditional Chinese Medicine; the function of cervical vertebra and all body status were evaluated according to NDI score. VAS score, frequency and time of headache were recorded and compared before and after treatment. RESULTS: No adverse reactions were found after treatment, all patients were followed up from 1 to 6 months with an average of 3.1 months. In pressing micro-regulating group, 25 cases got fully recover, 26 excellence, 14 effectiveness and 4 inefficiency; and in traditional massage group, the results were 12,21,22, 12;clinical effect of pressing micro-regulating group was better than that of traditional massage group (P<0.01). NDI score in pressing micro-regulating group decreased from preoperative 13.48 ± 4.83 to postoperative 6.23 ± 3.76; in traditional massage group also decreased from preoperative 13.82 ± 5.78 to postoperative 8.25 ± 4.75; the improvement of the pressing micro-regulating group was obviously better than that of traditional massage group (P < 0.01). VAS score in pressing micro-regulating group decreased from preoperative 4.75 ± 0.97 to postoperative 1.88 ± 1.78; and in traditional massage group decreased from pre-operative 4.78 ± 0.98 to postoperative 2.84 ± 1.94; pressing micro-regulating group was more notable than that of traditional massage group (P < 0.01). The frequency per week,the pain time in pressing micro-regulating group decreased from preoperative (5.38 ± 1.96) times and (6.87 ± 3.67) hours to postoperative (1.71 ± 2.04) times and (0.97 ± 1.74) hours,respectively,in traditional massage group the above parameters decreased from preoperative (5.22 ± 1.81) times and (6.90 ± 3.79) hours to postoperative (2.81 ± 2.42) times and (1.83 ± 2.21) hours;pressing micr-regulating group was more notable than that of traditional massage group (P < 0.01 or P < 0.05). CONCLUSION: Using the tuina method of micro-regulating with vertical cross pressing lying on one side to treat CEH can improve function of cervical vertebra and all body status, lessen the intensity, frequency, duration time of pain, and had advantage of higher security, simple operation, and evident effect.

[Effect of Acupuncture Stimulation of Different Acupoint Groups on Levels of Stress Hormones and Serum Brain-derived Neurotrophic Factor in Depression Rats].

OBJECTIVE: To screen the effective acupoints for anti-depression in depression rats and to explore the mechanisms of acupuncture for relieving depression. METHODS: Fifty rats were randomly divided into normal control, model, "Yin-tang" (EX-HN 3)-"Baihui" (GV 20, 2-acupoints), EX-HN 3-GV 20-"Fengchi" (GB 20)-"Shenshu" (BL 23, 4-acupoints) and medication groups, with 10 rats in each group. The depression model was established by unpredictable chronic mild stress (UCMS) for 28 days. For rats of the 2-acupoints and 4-acupoints groups, EX-HN 3 and GV 20, and EX-HN 3, GV 20, GB 20 and BL 23 were punctured with filiform needles respectively before performing mild stress every time. The acupuncture needles were retained for 30 min during each intervention and the treatment was conducted once daily for 28 days. The rats of the medication group were treated by intragastric administration of Fluoxetine (0.18 mg/kg) once a day for 28 days. The rats' anxiety-like behavior (rearing and crossing times) was detected by open-field test. The contents of adrenocorticotropic hormone (ACTH) in the pituitary, 5-hydroxytryptamine (5-HT) in the hippocampus, the cortisol (CORT) in the adrenal gland, and the brain derived neurotrophic factor (BDNF) in the serum were examined by ELISA. RESULTS: Compared to the normal control group, the numbers of both rearing and crossing motions in the model group were significantly decreased (P<0.01, P<0.05), while in comparison with the model group, the rearing and crossing numbers of rats in the 2-acupoints and 4-acupoints and medication groups were significantly increased (P<0.01). ELISA showed that after modeling, the content of adrenal CORT was significantly increased (P<0.01), and those of hippocampal 5-HT and serum BDNF were obviously down-regulated in the model group (P<0.01). After the treatment, the adrenal CORT levels in the three intervention groups were notably down-regulated, and hippocampal 5-HT and serum BONE evidently up-regulated in these 3 intervention groups (P<0.05, P<0.01). No marked changes were found in the pituitary ACTH contents of the model and 3 intervention groups (P>0.05), and no significant differences were shown among the three intervention groups in the levels of the aforementioned 6 indexes (P>0.05). CONCLUSION: Acupuncture intervention can effectively improve the unprompted activates of the depression rats, which may be related to its effects in up-regulating hippocampal 5-HT and serum BONE levels, and in down-regulating adrenal CORT content.