USP2 Promotes the Proliferation and Inflammation of Fibroblast-Like Synovial Cells in Rheumatoid Arthritis Through Deubiquitination of TRAF2.

Rheumatoid arthritis (RA) is a prevalent inflammatory disorder affecting about 1% of the global population. The ubiquitin-specific protease 2 (USP2) is known to have a substantial influence on the regulation of several cellular processes. Both in vivo (using rats with collagen-induced arthritis, CIA) and in vitro (using human fibroblast-like synoviocytes, HFLS-RA) models of RA were used to examine the role of USP2 in RA. The proliferation of HFLS-RA cells was assessed using the cell counting kit 8 test and EdU staining. The technique used for the assessment of gene expression was quantitative real-time PCR. Protein expression was quantified using Western blot (WB) analysis, while the quantities of inflammatory factors and matrix metalloproteinases were assessed using an ELISA test. The co-immunoprecipitation and ubiquitination tests investigated the relationships between proteins and the underlying molecular pathways. The results of this study demonstrate an upregulation of USP2 expression in both vivo and vitro models of RA. In addition, our findings indicate that the overexpression of USP2 notably exacerbates both proliferation and inflammation. The consistent downregulation of USP2 resulted in a reduction in the secretion of inflammatory cytokines and a suppression of cellular proliferation. Furthermore, it was shown that USP2 interacts with tumor necrosis factor receptor-associated factor 2 (TRAF2) and facilitates the removal of ubiquitination chains from TRAF2, enhancing its stability. Our findings propose that USP2 functions as a favorable modulator of proliferation and inflammatory reactions in HFLS-RA, thereby indicating its potential as a therapeutic target for the treatment of RA.

Summary of Outpatient Visits for Characteristic Nursing in Traditional Chinese Medicine.

The aim of this study was to investigate the practical outcomes of traditional Chinese medicine specialty nursing clinics in the clinical setting. Outpatient services have become increasingly popular for seeking medical care. Establishing traditional Chinese medicine specialty nursing clinics can meet the medical needs of the general public, and provide patients with convenient and efficient medical services. This study employed a retrospective cross-sectional observational design to analyze the medical service status of all patients who attended the clinic since its opening. Five qualified traditional Chinese medicine nursing experts identified and implemented 5 categories of traditional Chinese medicine characteristic nursing techniques, including cupping, moxibustion, needle acupuncture, and massage. Nurses and patients evaluated the treatment outcomes for various diseases. Since the establishment of the nursing outpatient department 2 years ago, there have been over 7046 visits, with a satisfaction rate of 97.1%. Currently, 5 nursing experts are nurturing a total of 11 graduate students, conducting 5 free clinics in the nursing outpatient department, and organizing 3 visits by overseas experts. The traditional Chinese medicine specialty nursing outpatient service effectively meets the diverse medical needs of patients, alleviates the outpatient pressure on hospitals, enhances the specialized development of nurses, increases the prominence of traditional Chinese medicine specialty nursing techniques, and promotes traditional Chinese medicine culture.

Regulatory effect of zinc finger protein A20 on rheumatoid arthritis through NLRP3/Caspase-1 signaling axis mediating pyroptosis of HFLS- RA cells.

As a chronic inflammatory autoimmune disease, rheumatoid arthritis (RA) causes significant destruction to joints and cartilage. So far, from RA patients, the synovial cells and subsynovial tissues reflected the positive expression of IL-18, IL-1ß, Caspase-1 and NLRP3, with the synovial tissues of those patients also expressing the zinc finger protein A20 at a significantly lower level compared with osteoarthritis (OA) ones. Thus, the inhibition of the NLRP3/caspase-1 signaling pathway can effectively down-regulate the expression of IL-1ß, but when NLRP3 inflammasomes are activated, they can also shear GSDMD and induce pyroptosis. These suggest that the Gasdermin family of proteins, downstream of the NLRP3 inflammasome, could be involved in pyroptosis. Previous studies have shown that A20 contributes largely as an anti-inflammatory factor in many inflammatory diseases, but it remains unclear whether zinc finger protein A20, as an inhibitor of NLRP3 inflammasomes, can play a protective role against RA by inhibiting NLRP3 inflammasome-mediated pyroptosis. Therefore, this study aimed to verify the effects of zinc finger protein A20 on NLRP3/ Caspase-1-mediated pyroptosis in rheumatoid arthritis synovial fibroblasts (HFLS-RA) cells through cell experiments and clinical bidirectional verification, aim to understand the regulatory mechanism of A20 on RA. The results of clinical trials showed that NLRP3, Caspase-1, IL-1ß and IL-18 were positively scattered in RA synovial cells and subsynovial tissue. The expression level of the zinc finger protein A20 in RA synovial tissues was significantly lower than that in OA synovial tissue and was negative, while zinc finger protein A20 was strongly positive in OA synovial tissue. In addition, HFLS-RA cells with siRNA-interfering zinc finger protein A20 were constructed at the cellular level, with the results also confirming that zinc finger protein A20 can play a protective role against RA by inhibiting NLRP3 inflammasome-mediated pyroptosis. In conclusion, this study is of great significance for understanding the role of the NLRP3-caspase-1-IL-1ß/ pyroptosis signaling pathway in the occurrence and development of RA. It is expected that the results will provide a theoretical basis for the immune regulation of innate immunity in the occurrence and development of RA, while providing a new therapeutic target for the clinical treatment of RA.

Diagnostic accuracy of multiparametric ultrasound for peripheral nerve schwannoma.

BACKGROUND: Ultrasound (US) diagnostic techniques have the advantages of low cost, convenient operation, and high availability. PURPOSE: To explore the diagnostic accuracy of multiparametric US in evaluating signs of peripheral schwannoma. MATERIAL AND METHODS: This retrospective case-control study included patients with soft-tissue masses on the limbs (divided into the schwannoma and non-schwannoma groups) between January 2017 and November 2020. US features were compared between the two groups, and receiver operating characteristics analysis was used to evaluate the diagnostic efficacy of these features. RESULTS: A total of 165 patients were included in this study; of them, 63 (38.2%) were diagnosed with schwannoma. Regular morphology (95.2% vs. 39.2%), cystic degeneration (71.4% vs. 27.5%), target sign on elastography (82.5% vs. 0), and polar blood supply sign (87.3% vs. 14.7%) were more common in schwannomas than in non-schwannoma lesions (all P < 0.001). Combining the four signs for diagnosis of schwannomas, the sensitivity, specificity, and accuracy were 95.24%, 96.08%, and 95.76%, respectively, with an area under the curve (AUC) of 0.987 (95% confidence interval = 0.955-0.998). Entering and exiting nerve sign was observed in 87.3% of schwannomas and in 3.0% of non-schwannoma lesions (P < 0.001), while split-fat sign was similar between the two groups (9.5% vs. 2.0%; P = 0.068). CONCLUSION: Polar blood supply sign and target sign on elastography are specific US signs in peripheral schwannomas. The combination of two-dimensional imaging, color flow imaging, and elastography can achieve an excellent diagnostic accuracy in schwannomas.

Roles of CD3, CD4 and CD8 in synovial lymphocytes of rheumatoid arthritis.

In this study, the immunohistochemical EnVision method was applied to detect CD3, CD4 and CD8 in synovial tissues of 40 patients with rheumatoid arthritis (RA) and 10 patients with osteoarthritis (OA). In 92.5% (37/40) RA cases, lymphocytes were focally aggregated, and even germinal centers appeared, forming lymphoid follicle-like structures. The expression of CD3, CD4, and CD8 were high in synovial tissue of RA group, but low in OA group. The number of CD3, CD4+, and CD8+ lymphocytes in OA group were significantly lower than that in RA group (p < 0.05); CD4+lymphocytes in RA accounted for the majority, and mostly were focally distributed. The number of CD8+lymphocytes in the synovial tissue were small, and were mostly scattered. The number of CD4+lymphocytes were significantly higher than CD8+lymphocytes (p<0.05). Compared with the OA group, the number of CD4+T and CD8+T lymphocytes in RA group were higher, and the ratio of CD4/CD8 was higher in RA group (p < 0.05). In conclusion, the CD3, CD4 and CD8 with high level may promote the occurrence and development of RA. The ratio of CD4+/CD8+ may be used as a reference index for the diagnosis and prognosis of RA.

IGF2BP1 Bolsters the Chondrocytes Ferroptosis of Osteoarthritis by Targeting m6A/MMP3 Axis.

Introduction: Chondrocyte degeneration and senescence are characteristics of osteoarthritis (OA) and other joint degenerative diseases, and ferroptosis has been observed to regulate the development of OA. However, the role of the N6-methyladenosine (m6A) modification in OA ferroptosis remains unclear. Methods: This study performed series of assays to investigate the function of the m6A reader IGF2BP1 in OA ferroptosis, including m6A quantitative analysis, Iron (Fe2+) release analysis, Malondialdehyde (MDA) measurement, lipid peroxidation (ROS) detection and Glutathione (GSH) measurement. The molecular interaction and mechanism analysis was performed by Luciferase reporter assay, mRNA stability analysis and RNA immunoprecipitation (RIP) assay. Results: These results indicate that IGF2BP1 is upregulated in IL-1ß-induced chondrocytes. Functionally, IGF2BP1 silencing represses ferroptosis, including iron (Fe2+) accumulation, malondialdehyde, and reactive oxygen species (ROS). Mechanistically, among the potential downstream targets, matrix metalloproteinase-3 (MMP3) was observed to harbor a significant m6A modified site in the 3'-UTR. IGF2BP1 combines with MMP3 through the binding of m6A sites, thereby enhancing MMP3 mRNA stability. Discussion: In conclusion, our findings revealed the functions and mechanisms of m6A regulator IGF2BP1 in OA chondrocyte's ferroptosis, providing a novel target for OA treatment.

Tumor necrosis serves as an important pathological characteristic of stage I-II colon cancer.

BACKGROUND: The long-term prognosis of colon cancer patients remains little changed with relatively high mortality and morbidity. Since the most widely used prognostic parameter TNM staging system is less satisfactory in predicting prognosis in early-stage cancers, numerous clinicopathological factors, including tumor necrosis, have been proposed for prognosis stratification, but substantial evidences are still lacking for early-stage colon cancer. MATERIALS AND METHODS: In the retrospective study, a total of eligible 173 stage I-II colon cancer patients, who received tumor radical resection and lymphadenectomy in the local hospital between January 1, 2010, and December 31, 2018, were enrolled for analyzing the prognostic role of tumor necrosis. The primary endpoints included 5-year overall survival (OS) and progression-free survival (PFS). RESULTS: The median follow-up of enrolled early-stage colon cancer patients was 58.3 months. The 2-year and 5-year OS rates were 88.3% and 68.2%, respectively, and the 2-year and 5-year PFS rates were 85.6% and 62.7%, respectively. Seventy-eight patients (45.1%) were diagnosed with tumor necrosis by pathological examination. Demographic analysis revealed a significant association of tumor necrosis with larger tumor size and a marginal association with vascular invasion. Kaplan-Meier survival curves demonstrated that tumor necrosis was associated with worse OS (log-rank P = 0.003) and PFS (log-rank P = 0.002). The independent unfavorable prognostic effect of tumor necrosis was further validated in univariate and multivariate Cox regression analysis (hazard ratio = 1.91 (1.52-2.40), P = 0.004). CONCLUSIONS: The current study confirmed the independent prognostic role of tumor necrosis from pathological review in early-stage colon cancer patients. This pathological criterion promises to help in identifying high-risk subgroup from early-stage colon cancer patients, who may benefit from strict follow-up and adjuvant therapy.

Authentication and validation of key genes in the treatment of atopic dermatitis with Runfuzhiyang powder: combined RNA-seq, bioinformatics analysis, and experimental research.

Background: Atopic dermatitis (AD) is inflammatory disease. So far, therapeutic mechanism of Runfuzhiyang powder on AD remains to be studied. This study aimed to mine key biomarkers to explore potential molecular mechanism for AD incidence and Runfuzhiyang powder treatment. Methods: The control group, AD group, treat group (AD mice treated with Runfuzhiyang powder were utilized for studying. Differentially expressed AD-related genes were acquired by intersecting of key module genes related to control group, AD group and treatment group which were screened by WGCNA and AD-related differentially expressed genes (DEGs). KEGG and GO analyses were further carried out. Next, LASSO regression analysis was utilized to screen feature genes. The ROC curves were applied to validate the diagnostic ability of feature genes to obtain AD-related biomarkers. Then protein-protein interaction (PPI) network, immune infiltration analysis and single-gene gene set enrichment analysis (GSEA) were presented. Finally, TF-mRNA-lncRNA and drug-gene networks of biomarkers were constructed. Results: 4 AD-related biomarkers (Ddit4, Sbf2, Senp8 and Zfp777) were identified in AD groups compared with control group and treat group by LASSO regression analysis. The ROC curves revealed that four biomarkers had good distinguishing ability between AD group and control group, as well as AD group and treatment group. Next, GSEA revealed that pathways of E2F targets, KRAS signaling up and inflammatory response were associated with 4 biomarkers. Then, we found that Ddit4, Sbf2 and Zfp777 were significantly positively correlated with M0 Macrophage, and were significantly negatively relevant to Resting NK. Senp8 was the opposite. Finally, a TF-mRNA-lncRNA network including 200 nodes and 592 edges was generated, and 20 drugs targeting SENP8 were predicted. Conclusion: 4 AD-related and Runfuzhiyang powder treatment-related biomarkers (Ddit4, Sbf2, Senp8 and Zfp777) were identified, which could provide a new idea for targeted treatment and diagnosis of AD.

Effects of combining acupuncture with exercise training on relieving dyspnea and improving exercise tolerance in chronic obstructive pulmonary disease patients: a protocol for a single-blind, randomized, sham-controlled trial.

BACKGROUND: Exercise training is beneficial for patients with chronic obstructive pulmonary disease (COPD). However, the effect of exercise training is limited by patients' impaired exercise capacity, exertional dyspnea and other respiratory problems. Acupuncture, as a reliable and safe therapy, is effective in reducing dyspnea, relieving respiratory muscle fatigue, and improving exercise capacity of COPD patients. However, it is not known whether the combination of exercise training and acupuncture reduces dyspnea and improves quality of life of COPD patients or induces more pronounced effects in dyspnea and exercise tolerance. This trial aims to determine whether acupuncture enhances the effect of exercise training in COPD patients compared to sham acupuncture. METHODS: In this single-blind, randomized, sham-controlled trial, 70 COPD patients will be enrolled and randomly assigned (1:1) to the following 2 groups: (I) real acupuncture and exercise training group; and (II) sham acupuncture and exercise training group. For acupoint selection, CV 4, CV 12, CV 17, ST 40, ST 16 and ST 25 will be used for all patients. For sham acupuncture group, Streitberger placebo needles will be used. A single-blind method will be adopted in this trial. Data collectors and statisticians will be blinded in this trial, only the acupuncturists will know the group allocation. The intervention will be conducted 3 times a week for 8 weeks, totaling 24 treatments. Patients will be evaluated at the baseline, after 14 treatments during the 5th week, after 24 treatments during the 8th week, and at a 5-month follow-up period. The primary outcomes will be assessed by a modified British medical research council questionnaire (mMRC), COPD assessment test (CAT), and 6-minute walk test (6MWT). The secondary outcomes will be measured by changes in variables from the lung function test, cardiopulmonary exercise testing, and blood gas analysis. Two-independent sample t-tests will be used to compare differences in the changes in all outcome measures after the intervention between two groups. Safety evaluation will be performed at each treatment visit and assessment by recording adverse events (AE) in the AE Report Form. DISCUSSION: This study will help to determine whether acupuncture increases the benefits obtained from exercise training in COPD patients. TRIAL REGISTRATION: Chinese Clinical Trial Registry: ChiCTR1900028627. Registered on December 29, 2019.

Effect of resveratrol on abnormal bone remodeling and angiogenesis of subchondral bone in osteoarthritis.

PURPOSE: The effect of resveratrol on subchondral bone in osteoarthritis was explored by constructing a mouse model of osteoarthritis and giving resveratrol as intervention. METHODS: The degree of proteoglycan loss in articular cartilage was assessed by safranine fast green staining. The expressions of Lubricin and Aggrecan, COLX, and MMP-13, the co-expression of CD31 and Endomucin, and the expression of angiogenesis-related factors were determined by immunohistochemistry. TRAP stain and immunostaining were used to assess abnormal subchondral bone resorption and bone formation. Angiography was employed to analyze the effect of resveratrol on the proliferation of subchondral bone vessels. RESULTS: Resveratrol inhibited cartilage thickening and the increase of COLX and MMP-13 expression, delayed the loss of proteoglycan, Lubricin, and Aggrecan, and inhibited osteoclast differentiation by up-regulating osteoprotegerin (OPG) and down-regulating the expression of RANKL. Angiography showed that resveratrol can reduce the abnormally elevated number and volume of blood vessels in the subchondral bone. Immunostaining showed that resveratrol inhibited CD31hiEmcnhi angiogenesis and high expression of VEGFA and Angiopoietin-1. CONCLUSION: Resveratrol inhibits osteoclast differentiation and reduces active bone resorption by regulating the OPG/RANKL/RANK pathway, and inhibits the abnormal proliferation of CD31hiEmcnhi blood vessels by downregulating the expression of VEGFA and Angiopoiein-1, thereby eliminating the pathologic coupling mechanism of osteogenesis and vascularization, and delaying the progression of osteoarthritis.

Biochanin A Regulates Cholesterol Metabolism Further Delays the Progression of Nonalcoholic Fatty Liver Disease.

PURPOSE: To discover the possible target of biochanin A (BCA) in the lipid metabolism pathway and further explore its mechanism to nonalcoholic fatty liver disease (NAFLD). METHODS: We adopted a high-fat and high-glucose diet for 12 weeks to build the NAFLD rat model, which was then treated with different proportions of BCA for 4 weeks. General condition, body weight, Lee index, and liver index were then evaluated. Furthermore, blood lipid level and insulin resistance (IR) were detected. Moreover, hematoxylin and eosin and oil red O staining were used to observe the pathological changes in the liver. Finally, Western blotting was used to detect the protein expression levels of CYP7A1, HMGCR, LDLR, PPAR-α, PPAR-γ, and SREBP-1c in the liver. RESULTS: The vital signs of rats in each group were stable. The treatment with BCA effectively reduced Lee index and liver index (F = 104.781, P < 0.05); however, the weight was not effected in each group. Additionally, BCA effectively reduced the related lipid metabolism indexes of NAFLD, such as total cholesterol (TC), triglyceride (TG), low-density lipoprotein (LDL), blood glucose, insulin, IR (F =12.463 (TC), 6.909 [TG], and 15.3 effected 75 [LDL], P < 0.05), and increased HDL (F = 11.580, P < 0.05). We observed that BCA could significantly improve steatosis and inflammatory cell infiltration in liver slices. Furthermore, BCA significantly increased the CYP7A1, LDLR, and PPAR-α protein expression in the liver and downregulated the HMGCR, SREBP-1c, and PPAR-γ protein expression. CONCLUSION: BCA could delay the liver damage of NAFLD induced by a high-fat diet, regulate the blood lipid level, and improve the expression of lipid metabolism-related genes in rats.

A multicenter, randomized controlled trial of massage in children with pediatric cerebral palsy: Efficacy of pediatric massage for children with spastic cerebral palsy.

BACKGROUND: Cerebral palsy is 1 of the diseases critically affecting the health of children. The spasmodic type is the most common, characterized by the increased muscular tension. It often leads to lifelong disability, bringing a heavy economic burden to families and society. As a key treatment in traditional Chinese medicine, pediatric massage has a significant clinical effect on cerebral palsy in children; however, high-quality randomized controlled studies are lacking. The main objective of this study was to evaluate the efficacy of pediatric massage for children with spastic cerebral palsy. METHODS/DESIGN: The study will be a multicenter, single-blinded, randomized-controlled pilot trial. During the period from June 2019 to December 2020, 182 children with spastic cerebral palsy will be randomly divided into experimental and control groups in a 1:1 ratio. The experimental group will undergo the modified selective spinal massage method combined with the basic rehabilitation treatment, while only the basic rehabilitation treatment would be performed for the control group. The intervention period of the study will last 12 weeks, 5 days weekly on weekdays. The primary outcomes include a modified Ashworth scale assessment and gross motor function test. The secondary outcomes include the 4-diagnostic scale of Chinese medicine and children's intelligence. The observation index will be measured during the complete 12 weeks duration after the treatment of the child, that is, before treatment, after 4 weeks of treatment, after 8 weeks, and after 12 weeks of treatment. DISCUSSION: This study aims to evaluate the efficacy of pediatric massage on children with spastic cerebral palsy; if the outcome is positive, it can provide a reference for the further promotion and application of pediatric massage in the treatment of spastic cerebral palsy. TRIAL REGISTRATION: Chinese ClinicalTrials.gov, ID: ChiCTR1900021666. Acupuncture-Moxibustion Clinical Trial Registry, AMCTR: (AMCTR-IPR-19000260) Registered on 04 March 2019.

Factor analysis on symptoms and signs of chronic low-back pain based on Traditional Chinese Medicine theory.

OBJECTIVES: The objective of this study was to use factor analysis to explore patterns of symptoms and signs from patients with chronic low-back pain (CLBP) based on the Traditional Chinese Medicine (TCM) theory. DESIGN, SETTINGS, AND SUBJECTS: A cross-sectional study was carried out among 513 patients with CLBP in four hospitals affiliated with Yunnan University of Traditional Chinese Medicine, China. OUTCOME MEASURES: Outcome measures comprised 31 symptoms and signs on a six-level self-report questionnaire. RESULTS: Four (4) factors were extracted. They were eventually interpreted as (1) "Qi and/or Blood Stagnation," which includes eight items such as piercing pain; activity limited by feeling of local heaviness, lumbar and flank stiffness with bending limitation and purple tongue, etc.; (2) "Cold/Damp," which has seven items (for example, Cold/Damp pain, pallid face and greasy coating, etc.); (3) a part of "Kidney Deficiency," which includes two items: "dull pain and recurrent vague pain"; (4) "Warmth/Heat," which is related to three items (namely, purple tongue, yellow tongue coating, and burning pain). The four factors accounted for 12.7%, 8.2%, 8.2%, and 7.8% of the total variance, respectively. There are seven items with uniqueness over 0.8. CONCLUSIONS: Four (4) TCM major groups of pathophysiology could explain 37% of variance of 31 symptoms and signs of the CLBP patients. Thirteen (13) items are not groupable.