Proteomics analysis of lung tissue reveals protein makers for the lung injury of adjuvant arthritis rats.

Lung injury is one of the common extra­articular lesions in rheumatoid arthritis (RA). Due to its insidious onset and no obvious clinical symptoms, it can be easily dismissed in the early stage of diagnosis, which is one of the reasons that leads to a decline of the quality of life and subsequent death of patients with RA. However, its pathogenesis is still unclear and there is a lack of effective therapeutic targets. In the present study, tandem mass tag­labeled proteomics was used to research the lung tissue proteins in RA model (adjuvant arthritis, AA) rats that had secondary lung injury. The aim of the present study was to identify the differentially expressed proteins related to RA­lung injury, determine their potential role in the pathogenesis of RA­lung injury and provide potential targets for clinical treatment. Lung tissue samples were collected from AA­lung injury and normal rats. The differentially expressed proteins (DEPs) were identified by tandem mass spectrometry. Bioinformatic analysis was used to assess the biological processes and signaling pathways associated with these DEPs. A total of 310 DEPs were found, of which 244 were upregulated and 66 were downregulated. KEGG anlysis showed that 'fatty acid degradation', 'fatty acid metabolism', 'fatty acid elongation', 'complement and coagulation cascades', 'peroxisome proliferator­activated receptor signaling pathway' and 'hypoxia­inducible factor signaling pathway' were significantly upregulated in the lung tissues of AA­lung injury. Immunofluorescence staining confirmed the increased expression of clusterin, serine protease inhibitors and complement 1qc in lung tissue of rats with AA lung injury. In the present study, the results revealed the significance of certain DEPs (for example, C9, C1qc and Clu) in the occurrence and development of RA­lung injury and provided support through experiments to identify potential biomarkers for the early diagnosis and prevention of RA­lung injury.

Network analysis and transcriptome profiling in peripheral blood mononuclear cells of patients with rheumatoid arthritis.

The present study aimed to investigate the differential expression of long non-coding RNAs (lncRNAs) in rheumatoid arthritis (RA). High-throughput gene sequencing technology was used to detect the expression of lncRNA and mRNA in three patients with RA (RA group) and normal controls (NC group). A Bioinformatics analysis was used to assess the effects of differentially expressed mRNAs on signaling pathways and biological functions. The selected dysregulated lncRNAs were verified by reverse transcription-quantitative (RT-q)PCR in the peripheral blood mononuclear cells (PBMCs) of patients with RA and age- and sex-matched controls. A correlation analysis was used to analyze the relationship between lncRNAs and clinical indexes. From the lncRNA sequencing data, significantly differentially expressed lncRNAs between the RA and NC groups were identified by a fold change ≥2 and P<0.05. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analysis suggested that the differentially expressed mRNAs were mainly involved in organelle composition, intracellular regulation, signaling pathways, cancer, virus and inflammation. A total of four of these lncRNAs were confirmed by RT-qPCR to be significantly differentially expressed (LINC00304, MIR503HG, LINC01504 and FAM95B1). Through the correlation analysis, it was confirmed that there was a strong correlation between these lncRNAs and clinical laboratory indicators and indexes such as course of disease, arthrocele and joint tenderness. Overall, the present results suggested that the expression levels of LINC00304, MIR503HG, LINC01504 and FAM95B1 in PBMCs from patients with RA may serve as potential biomarkers for RA diagnosis, influencing the occurrence and progress of RA.

lncRNA expression profiles related to apoptosis and autophagy in peripheral blood mononuclear cells of patients with rheumatoid arthritis.

Long noncoding RNAs (lncRNAs) are >200-bp molecules that do not generally code for proteins. Human lncRNAs have well-characterized roles in gene expression regulation, particularly with regard to protein-coding genes, and their dysregulation has been linked to disease. Here, we set out to investigate changes in the expression of lncRNAs related to apoptosis and autophagy in the peripheral blood mononuclear cells (PBMCs) of rheumatoid arthritis (RA). In addition, we aimed to correlate lncRNA expression profiles with clinical indexes and self-perception of patients (SPP). To this end, we employed RNA sequencing of lncRNAs in PBMCs from three patients with RA and three healthy controls. We used bioinformatics to screen several dysregulated lncRNAs related to apoptosis and autophagy. To validate key lncRNA candidates, we performed quantitative reverse transcriptase-PCR on 20 patients with RA and 20 healthy controls. We found the expression of seven lncRNAs (MAPKAPK5-AS1, ENST00000619282, C5orf17, LINC01189, LINC01006, DSCR9 and MIR22HG) was significantly altered in PBMCs of patients with RA. Receiver operating characteristic curve analysis suggested that MIR22HG [area under the curve (AUC) = 0.846, P = 0.000], DSCR9 (AUC = 0.783, P = 0.005), LINC01189 (AUC = 0.677, P = 0.034), MAPKAPK5-AS1 (AUC = 0.644, P = 0.025) and ENST00000619282 (AUC = 0.636, P = 0.043) are potential biomarkers of RA. Spearman's correlation analysis revealed selected lncRNAs correlated with clinical indexes and SPP. Therefore, we highlight that some lncRNAs related to apoptosis and autophagy may serve as potential biomarkers for diagnosis and monitoring of RA progression, which also correlate with several clinical indexes and SPP.

RNA-seq reveals the circular RNA and miRNA expression profile of peripheral blood mononuclear cells in patients with rheumatoid arthritis.

OBJECTIVE: Circular RNAs (circRNAs) are a significant class of molecules involved in a wide range of diverse biological functions that are abnormally expressed in many types of diseases. The present study aimed to determine the circRNAs specifically expressed in peripheral blood mononuclear cells (PBMCs) from rheumatoid arthritis (RA) patients to identify their possible molecular mechanisms. METHODS: To identify the circRNAs specifically expressed in RA, we started by sequencing the of PBMCs circRNA and microRNAs (miRNAs) from a RA group (n = 3) and a control group (n = 3). We constructed a network of differentially expressed circRNAs and miRNAs. Then, we selected differentially expressed circRNAs in PBMCs from 10 RA patients relative to 10 age- and sex-matched controls using real-time quantitative reverse transcription-polymerase chain reaction (RT-qPCR). Spearman's correlation test was used to evaluate the correlation of circRNAs with biochemical measurements. RESULTS: A total of 165 circRNAs and 63 miRNAs were differently expressed between RA patients and healthy people according to RNA-seq, including 109 circRNAs that were significantly up-regulated and 56 circRNAs that were down-regulated among the RA patients. RT-qPCR validation demonstrated that the expression levels of hsa_circ_0001200, hsa_circ_0001566, hsa_circ_0003972, and hsa_circ_0008360 were consistent with the results from the sequencing analysis. Then, we found that there were significant correlations between the circRNAs and disease severity. CONCLUSION: Generally, these results suggest that expression of hsa_circ_0001200, hsa_circ_0001566, hsa_circ_0003972, and hsa_circ_0008360 in PBMCs from RA patients may serve as potential biomarkers for the diagnosis of RA, and these circRNAs may influence the occurrence and development of RA.

Triptolide protects against TGF-ß1-induced pulmonary fibrosis by regulating FAK/calpain signaling.

The present study aimed to investigate the mechanism of anti-proliferative, anti-inflammatory and anti-fibrotic effects of triptolide (TPL) on activated lung fibroblasts by regulating the focal adhesion kinase (FAK) and calpain signaling pathways. The HFL-1 human foetal lung fibroblast cell line was cultured in vitro and treated with 50 ng/ml transforming growth factor (TGF)-ß1 for 48 h to establish the model of pulmonary fibrosis. Subsequently, the cells were divided into five groups, including a control, model, TPL, FAK inhibitor and calpeptin group. Subsequently, the proliferation of lung fibroblasts was detected using the Cell Counting Kit-8 assay. The concentration of interleukin (IL)-6 in the cell culture supernatant was examined by ELISA and the mRNA expression levels of collagen type I (ColI)α and ColIII in lung fibroblasts were quantified by reverse transcription-quantitative PCR. The protein levels of FAK, phosphorylated (p)-FAK, calpain 1 and calpain 2 were detected by western blot analysis. TGF-ß1 induced the proliferation of lung fibroblasts, whereas TPL inhibited this proliferation in a dose-dependent manner. TPL also decreased the TGF-ß1-induced production of IL-6 and reduced the upregulation of ColIα, ColIII, FAK, p-FAK, and inhibited the decrease of calpain 1 and calpain 2 induced by TGF-ß1. In addition, the FAK inhibitor acted synergistically with TPL to decrease TGF-ß1-induced production of IL-6 and attenuate TGF-ß1-induced synthesis of ColIα and ColIII, while calpeptin had an antagonistic effect on the function of TPL. Furthermore, treatment with the FAK inhibitor and TPL markedly decreased the protein levels of FAK and p-FAK, and increased the protein expression of calpain 1 and calpain 2 in lung fibroblasts stimulated by TGF-ß1 to a greater extent than TPL alone, while calpeptin had an antagonistic effect on the action of TPL. In conclusion, the present study indicated that TPL protected against TGF-ß1-induced proliferation, inflammation and fibrosis by regulating the FAK and calpain signaling pathways.

[Effects of Triptolide on the Autophagy in Synovial, Spleen and Thymus of Rats with Adjuvant Arthritis].

OBJECTIVE: To determine the effect of triptolide (TP) on the expression of ATG /LC3-Ⅱ Beclin1 in synovial, spleen, and thymusof rats with adjuvant arthritis (AA). METHODS: Rats were divided for four groups: normal control (NC), model control (MC), leflunomide (LEF) treatment, and triptolide (TP)treatment, with 12 rats in each group.The AA model was established through Freund's complete adjuvant (0.1 mL each) injection into the right foot plantar skin to introduce inflammation and 10 days of tail root injection of 0.05 mL Freund's complete adjuvant for immunity strengthening. Drug administration started 13 days after induction of inflammation. Rats in the NC and MC groups were given normal saline (1 mL/100 g) once a day for 30 days, compared with 5 mg/kg of oral LEF for the rats in the LEF group and 50 µg/kg of oral TP for the rats in the TP group. Paw swelling (E), joint arthritis index(AI) and joint pathological changes of the rats were recorded. The serum expressions of cytokines B lymphocyte stimulating factor (BAFF), interleukin (IL)-1, tumor necrosis factor (TNF) alpha, IL-15,and IL-10 were detected by ELISA. The expressions of Atg5, Atg7, and Atg12 mRNA in synovial, spleen, and thymus of the rats were detected by RT-PCR.The expressions of LC3-Ⅱ and Beclin1 in synovial, spleen, and thymus of the rats were detected by Western blot assay. RESULTS: The AA model rats had lower serum BAFF, IL-1, TNF alpha, IL-15, and IL-10; lower Atg5and Atg12 mRNA in synovial; lower Atg5 mRNA, Atg7, and Atg12 mRNA in spleen; higher Atg12 mRNA in thymus; and lower LC3-Ⅱ and Beclin1 in synovial, spleen and thymus(P<0.05 or 0.01). TP treatment led to reduced paw swelling and arthritis index; declined Atg7 and Atg12 mRNA in synovial; declined Atg5, Atg7 mRNA and Atg12 mRNA in spleen; decreased Atg5 and Atg7mRNA in thymus; increased Atg12 mRNA in thymus; and increased LC3-Ⅱ and Beclin1 in synovial, spleen and thymus (P<0.05 or 0.01). Compared with rats treated with LEF, TP treated rats had lower TNF-α and BAFF and higher E and IL-15 (P<0.05 or 0.01); as well as decreased expressions of Atg7 mRNA (synovial) and Atg5, Atg7 mRNA (thymus), and increased expressions of Atg12 mRNA (thymus) and Atg5, Atg7, Atg12 mRNA (spleen). CONCLUSION: TP regulates autophagy in synovial, thymus and spleen of AA rats, and improves theirjointinflammatory response.

[Xinfeng Capsule reduces hypercoagulative state in patients with Sjogren's syndrome by inhibiting miR-155/SOCS1/NF-κB signaling pathway].

Objective To explore the relationship between Xinfeng Capsule (XFC) improving the hypercoagulative state in patients with Sjogren's syndrome (SS) and miR-155/suppressor of cytokine signaling 1 (SOCS1)/nuclear factor κB (NF-κB) signaling pathway. Methods Sixty-six SS patients were randomly divided into XFC-treated group and hydroxychloroquine (HCQ)-treated control group (n=33 per group), which were respectively treated with XFC and HCQ. In addition, 20 healthy volunteers were enrolled as a normal control group. The levels of prothrombin time (PT), activated partial thromboplastin time (APTT), fibrinogen (FIG), thrombin time (TT) and D-dimer (D-D) were detected using automatic coagulation analyzer. Interleukin-1ß (IL-1ß), IL-4, IL-10, tumor necrosis factor-α (TNF-α), P50, P65, inhibitor of NF-κB α (IκBα) were tested using ELISA. Meanwhile, the mRNA expressions of p50, p65 and IκBα were determined using quantitative real-time PCR, and the level of microRNA-155 (miR-155) was examined by one-step fluorescence quantitative PCR. The protein levels of P50, P65 and SOCS1 were detected using Western blotting. Erythrocyte sedimentation rate (ESR) was evaluated by Westergren method. Hypersensitive C-reactive protein (hs-CRP) was detected using automatic biochemical analyzer. Results Compared with the normal control group, the levels of D-D and FIB significantly increased in SS group; simultaneously, the serum levels of miR-155, IL-1ß, TNF-α, P50, P65, IκBα, hs-CRP, ESR were significantly elevated in SS patients, while IL-4 and IL-10 were significantly reduced. Spearman correlation analysis showed that the coagulation parameters were remarkably correlated with cytokines, NF-κB and activity indexes. In the two treated groups, coagulation parameters and related indexes were demonstrated having some improvement, especially in the XFC group, which had a much higher efficiency, and better outcomes in reducing the levels of FIB, D-D, miR-155, TNF-α, IL-1ß, P50, P65, ESR and hs-CRP, as well as increasing the expressions of SOCS1, IL-4 and IL-10. Conclusion XFC can significantly alleviate the hypercoagulative state of patients with SS, and the mechanisms may be related to the inhibition of miR-155/SOCS1/NF-κB signaling pathway.

[Xinfeng capsule improves hypercoagulative state by inhibiting miR-155/NF-κB signaling pathway in patients with active ankylosing spondylitis].

Objective To observe the effect of Xinfeng capsule (XFC) on miR-155, nuclear factor kappa B (NF-κB) signal pathway, and indexes related to hypercoagulative state in patients with active ankylosing spondylitis (AS), and investigate the possible mechanism. Methods Fifty-six cases in active AS were randomly divided into XFC group and sulfasalazine (SASP) group. All cases in the XFC group took three capsules three times daily for twelve consecutive weeks. The ones in the SASP group took four tablets two times daily for twelve consecutive weeks. The expression of miR-155 was detected by real-time PCR. The mRNA expressions of nuclear factor κB activator 1(Act1), NF-κB inhibitor-alpha (IκBα), inhibitor of kappa-B kinase beta (IKKß), NF-κB p65, and NF-κB p50 were tested by reverse transcription PCR (RT-PCR). Meanwhile, the protein expressions of NF-κB P65 and NF-κB P50 were determined by Western blotting. Tumor necrosis factor-alpha (TNF-α), interleukin (IL)-4, IL-10, IL-17, thromboxane B2 (TXB2), 6-ketone-prostaglandin F1 (6-keto-PGF1), platelet granular membrane protein 140 (GMP140), platelet activation factor (PAF), and plasminogen activator inhibitor-2 (PAI-2) were determined by ELISA. Clinical efficacy was evaluated in the two groups. Results Compared with the SASP group, 50% Bath ankylosing spondylitis disease activity index (BASDAI50) was significantly higher in the XFC group. Compared with the SASP group after treatment, platelet (PLT), fibrinogen (FBG) and D-D dimer (D-D), TXB2, GMP140, PAF, PAI-2, IL-17, erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), visual analog scale (VAS), BASDAI, BASFI, and BAS-G were reduced more obviously in the XFC group after treatment; meanwhile, 6-keto-PGF1, IL-4, and IL-10 significantly increased. Compared with the SASP group after treatment, the expressions of IKKß mRNA, IκBα mRNA, NF-κB p65 mRNA, NF-κB p50 mRNA, NF-κB P65 protein, NF-κB P50 protein, and miR-155 were lower in the XFC group after treatment. Conclusion XFC could effectively improve hypercoagulative state in active AS patients. The potential mechanism may be associated with the inhibition of miR-155 and NF-κB signal pathway.

[Hypercoagulable state is associated with NF-kappa B activation and increased inflammatory factors in patients with rheumatoid arthritis].

OBJECTIVE: To investigate the mechanism of hypercoagulable state based on nuclear factor κB (NF-κB) pathway in patients with rheumatoid arthritis (RA). METHODS: Thirty-five RA patients were enrolled as well as 20 healthy volunteers as a control group. Interleukin-10 (IL-10), IL-6, IL-4, IL-17, NF-κB activator 1 (Act1), p50, p65, IκBα, platelet activating factor (PAF), PAF-acetylhydrolase (PAF-AH) and anti-cyclic citrullinated peptide (CCP) were detected using ELISA. The number of platelet (PLT) was detected using Sysmex XT-2000i automated hematology analyzer. The levels of D-dimer (D-D), fibrinogen (FBG), thrombin time (TT), prothrombin time (PT), and partial thromboplastin time (APTT) were detected using Sysmex CA-1500 automatic coagulation analyzer. Erythrocyte sedimentation rate (ESR) was detected using Westergren method. C-reactive protein and rheumatoid factor (RF) were detected using Hitachi 7060 automatic biochemical analyzer. Meanwhile, the mRNA expressions of Act1, p65, p50, IκBα and IκB kinase α (IKKα) were detected using semi-quantitative reverse transcription PCR. The expressions of p65, p50 and IκBα proteins were examined using Western blotting. The correlations of the above indexes were analyzed by Spearman correlation test. RESULTS: Compared with the normal group, the levels of DD, FBG, PLT significantly increased in the peripheral blood of RA patients, TT decreased, while APTT and PT were not significantly changed. IL-4, IL-10 and PAF-AH were significantly reduced in the sera of RA patients, while IL-6, IL-17, Act1, p50, p65, IκBα, IKKα and PAF were significantly elevated. Spearman correlation analysis showed that coagulant and fibrinolytic indexes were significantly correlated with cytokines, NF-κB, activity indexes and clinical symptoms and signs. CONCLUSION: The hypercoagulable state is common in the peripheral blood of RA patients, and it is closely related to inflammatory factors, activity indexes and abnormal activation of NF-κB.

[Xinfeng Capsule Improved Blood Stasis State of Rheumatoid Arthritis Patients Based on Actl/NF-KB Signaling Pathway: Mechanism and Effects].

Objective To observe the mechanism of Xingfeng Capsule (XFC) for improving blood stasis state in rheumatoid arthritis (RA) patients based on Actl/NF-κB signaling pathway. Methods Totally 76 RA patients were equally assigned to two groups by random digit table, the XFC group (XFC, 3 pills each time, three times per day) and the Leflunomide group (LEF, 10 mg each time, once per day). All patients were intervened for 3 successive months. Clinical efficacy of symptoms of Chinese medicine (CM) was assessed. Serum levels of interleukin-10 (IL-10) , IL-17, IL-6, NF-κB activator 1 (Actl), p50, p65, platelet activating factor ( PAF) , platelet activating factor acetyl hydrolase ( PAF-AH) were detected using ELISA. Symptoms of blood stasis syndrome (BSS) were also assessed. mRNA expressions of Act1, p50, and p65 were detected using fluorescent quantitative PCR. Protein expressions of p50 and p65 were detected using Western blot. Correlation analyses were performed in RA patients' peripheral blood coagulation indicators, total score of BSS, and IL-1 0, IL-6, IL-17, Act1 , p50, p65 using Spearman. Results The total effective rate was 89. 5% (34t38) in the XFC group, with no statistical difference as compared with that of the LEF group [94. 7% (36)38), P >0. 05]. Compared with before treatment in the same group, serum levels of D-dimer (DD) , fibrinogen (FBG) , platelet (PLT) , PAF, IL-17, and IL-6 all decreased, mRNA expressions and serum levels of Act1, p50, and p65 were lowered, protein expres- sions of p50 and p65 decreased, scores for each symptoms in BSS all decreased, serum levels of PAF- AH and IL-10 increased in the two groups after treatment (P <0. 05, P <0. 01). Compared with the LEF group, serum levels of DD, FBG, PLT, IL-17, and IL-6 decreased, mRNA expressions and serum levels of Act1 and p65 were lowered, protein expression of p65 decreased, scores for joint prickling pain, tongue proper, subcutaneous ecchymosis, and BSS total score all decreased in the XFC group (P < 0. 05, P <0. 01). Peripheral blood DD was positively correlated with IL-17, IL-6, Act1, and and p65, but negatively correlated with IL-10 (P <0. 05, P <0. 01). FBG was positively correlated with IL-6 (P <0. 05). PLT was positively correlated with IL-17 (P <0. 05). BSS total score was positively correlated with IL-6, Act1, and p65, but negatively correlated with IL-10 (P <0. 05, P <0. 01). PAF was positively correlated with IL-17, IL-6, Act1 , and p65 (P <0. 05, P <0. 01), while PAF-AH was negatively correlated with p50 (P <0. 05). Conclusion The pathogenesis of BSS in RA patients and the effects of XFC on blood stasis state might be closely correlated to the Act1/NF-KB signaling pathway.

[Efficay Observation of Xinfeng Capsule for Treating 58 Cases of Sjogren's Syndrome].

Objective To observe the clinical effect of Xinfeng Capsule (XFC) in treating Sjögren's syndrome (SS) and its effect on coagulation parameters, peripheral blood cytokines, and NF- kappa B signaling pathway protein. Methods Totally 58 SS patients were assigned to the treatment group and the control group according to random digit table, 29 cases in each group. Patients in the treat- ment group took XFC, 3 pills each time, 3 times per day. Those in the control group took Hydroxychloro- quine ( HCQ) Sulfate, 0. 1 g per tablet, 2 tablets each time, twice per day. Three months consisted of one therapeutic course and one course for all. Another 20 healthy subjects were recruited as a normal control group. Coagulation parameters (APTT, PT, FIB,TT, D-D) were detected using automatic coagulation an- alyzer in the two groups before and after treatment as well as in the healthy control group. The expres- sions of peripheral blood cytokines (IL-1ß, TNF-α, IL-10) and NF-κB signaling pathway proteins (P65, P50, IκBα) were detected before and after treatment using ELISA. Erythrocyte sedimentation rate (ESR) was determined using Westergren method before and after treatment. High sensitivity C-reactive protein (hs-CRP) level was determined using automatic biochemical analyzer before and after treatment. Results Totally 44 SS patients had abnormal coagulation parameters, accounting for 75. 9% of the total. Com- pared with the healthy control group, the levels of D-D, FIB, IL-1 , TNF-α,P50, P65, IκBα, hs-CRP, and ESR increased (P <0.05, P <0. 01), and the IL-10 level decreased in SS patient groups (P <0.01). Spearman correlation analysis showed that coagulation parameters were significantly correlated with cy- tokines, NF-κB signal transduction pathway, and inflammation indices (P <0. 01 , P <0. 05). After drug in- tervention blood coagulation parameters and laboratory indices were partially improved in the two groups. The effective rate and the total effective rate were 59% and 86% in the treatment group, obviously higher than those of the control group with statistical difference (38% and 72%; P <0. 05). Besides, the treat- ment group was superior to the control group in reducing the levels of FIB, D-D, P50, P65, ESR, and hs- CRP, down-regulating levels of TNF-α and IL-1 ß, as well as up-regulating the expression of IL-10 (P < 0. 05, P <0. 01). Conclusions There is a hypercoagulable state in SS patients, which is related to abnor- mal activation of cytokines/NF-kappa B signaling pathway. XFC could effectively improve the hypercoagulative state of SS patients. Its mechanism might be related to inhibiting cytokines/NF-κB signaling pathway.

[Effect of Xinfeng Capsule in Improving Blood Stasis State of OA Patients].

Objective To observe the relationship between blood stasis state and activation of nuclear factor-κ-gene binding (NF-κB) signaling pathway/miRNA-146 in osteoarthritis (OA) patients and the effect of Xinfeng Capsule (XFC) on them. Methods Totally 70 OA patients were assigned to the treatment group and the control group according to random number table, 35 in each group. Patients in the treatment group took XFC, 3 pills each time, 3 times per day, while those in the control group took Glucosamine, 1 pill each time, 3 times per day. The therapeutic course for all was 3 months. Serum con- tents of p50, p65, inhibitor of nuclear factor κB O (IκBα) , nuclear factor kappa B activator 1 (Act1) , TGF-ß-activated kinase 1 (TAK1) , IL-1, IL-17, IL-10, and thromboxane A2(TXA2) , prostacycline (PGI2) were detected by ELISA. mRNA levels of Act1 , p65, p50, and TAK1 were detected using fluorescent quantitative PCR. The protein levels of p50 and p65 were detected using Western blot. The level of miRNA- 146 was detected using in one-step PCR. Results (1) Compared with pre-treatment in the same group, the levels of blood stasis score, platelets (PLT) , D-dimer (D-D) , TXA2, IL-1, IL-17, high-sensitivity C- reactive protein (hs-CRP), and erythrocyte sedimentation rate (ESR) all decreased; mRNA levels of p50, p65, Act1, and TAK1 were lowered; protein expressions of p50 and p65 decreased; serum levels of miRNA-146 decreased; activated partial thromboplastin time (APTT) , prothrombin time ( PT) , prosta- glandin 2 (PGI2), IL-10 increased in the two groups after treatment with statistical difference (P <0. 05, P <0. 01). Compared with the control group, the levels of blood stasis score, PLT, FIB,TXA2, IL-17, hs- CRP, and ESR were lowered; mRNA expressions of p65 and TAK1 were lowered; protein expressions of p50 decreased; levels of PT and PGI2 increased in the treatment group after treatment (P <0. 05, P < 0.01). Conclusion XFC could regulate the immunity and restore the equilibrium of cytokine network, and protect vascular endothelial cells possibly by up-regulating miRNA-146 expression and inhibiting acti- vation of NF-κB signaling pathway, thus improving blood stasis state of OA.

Efficacy and safety of Xinfeng capsule in patients with rheumatoid arthritis: a multi-center parallel-group double-blind randomized controlled trial.

OBJECTIVE: To evaluate the efficacy. and safety of Xinfeng capsule in patients suffering rheumatoid arthritis (RA). METHODS: A multi-center parallel-group designed, double-blind, randomized, controlled trial was conducted. Totally 304 RA patients were assigned to two groups: one group was administered Xinfeng capsule (XFC) plus the placebo of leflunomide and the other given leflunomide (LEF) plus the placebo of XFC for twelve weeks. The clinical and laboratory parameters were compared at baseline and fourth, eighth, and twelfth weeks. RESULTS: After twelve-week treatment, patients in two groups all showed some trend of effectiveness when compared in terms of American Rheumatism Association (ACR) recommended 20%, 50%, 70% improvement criteria, but it was insignificant. The validity in ameliorate modified disease activity score (DAS28) and laboratory indexes as erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), rheumatoid factor (RF) were also found no difference. The score of health assessment questionnaire (HAQ), self-rating anxiety scale (SAS), self-rating depression scale (SDS) and quality of life questionnaire with rheumatoid arthritis (RAQOL) both lower than the first week and the changes showed no difference. However, the score of SDS dropped more in XFC group than in the other. A total of 147 adverse reaction cases were reported, which shows no difference between the two groups. The most common adverse reactions were hepatic impairment, anemia, leukocytopenia, epigastric discomfort and phalacrosis. CONCLUSION: XFC demonstrated better improvement in the scores of SDS and compared with those of LEF group.