This study aims to investigate the effect and mechanism of total triterpenes of Euphorbium in treating rheumatoid arthritis(RA). The rat model of RA was established with Freund's complete adjuvant(FCA). Male rats were randomly assigned into control, model, Tripterygium glycosides(7.5 mg·kg~(-1)), and low-, medium-, and high-dose total triterpenes of Euphorbium(32, 64, and 128 mg·kg~(-1), respectively) groups, with 10 rats in each group. In other groups except the control group, 0.2 mL FCA was injected into the right hind toe. Rats in the intervention groups were administrated with corresponding drugs by gavage, and the control group and the model group with the same volume of 0.5% CMC-Na solution once a day. During the treatment period, the swelling degree of the hind paw was measured and the arthritis was scored until day 30. At the end of drug administration, the pathological changes of the joint tissue were observed by hematoxylin-eosin staining. The content of malondialdehyde(MDA), glutathione(GSH), and Fe~(2+) and the activity of superoxide dismutase(SOD) in the joint tissue were measured by biochemical colorimetry. RT-PCR was performed to determine the mRNA levels of nuclear factor erythroid 2-related factor 2(Nrf2), glutathione peroxidase 4(GPX4), and acyl-CoA synthetase long chain family member 4(ACSL4) in the joint tissue. Western blot was employed to determine the protein levels of Nrf2, Kelch-like ECH-associated protein 1(Keap1), heme oxygenase-1(HO-1), NAD(P)H quinone oxidoreductase 1(NQO1), SOD2, GPX4, and ACSL4 in the joint tissue. The results showed that the treatment with Tripterygium glycosides(7.5 mg·kg~(-1)) and total triterpenes of Euphorbium(32, 64, and 128 mg·kg~(-1)) alleviated the swelling degree of bilateral hind limbs, decreased the arthritis score, reduced joint tissue lesions and the content of MDA and Fe~(2+) in the joint tissue, and increased GSH content and SOD activity. Furthermore, the interventions up-regulated the protein and mRNA levels of Nrf2 and GPX4, down-regulated the protein and mRNA levels of ACSL4, and up-regulated the protein levels of Keap1, NQO1, HO-1, and SOD2 in the Nrf2/HO-1/GPX4. In summary, the total triterpenes of Euphorbium can treat RA by inhibiting lipid peroxidation and abnormal ferroptosis, which may involve the Nrf2/HO-1/GPX4 signaling pathway.
Rats ; Male ; Animals ; Rats, Sprague-Dawley ; NF-E2-Related Factor 2/metabolism* ; Kelch-Like ECH-Associated Protein 1/metabolism* ; Triterpenes/pharmacology* ; Oxidative Stress ; Arthritis, Rheumatoid/genetics* ; Glutathione ; Superoxide Dismutase/metabolism* ; Glycosides/pharmacology* ; RNA, Messenger/metabolism*

This study aimed to explore the molecular mechanism of Juanbi Qianggu Formula(JBQGF), an empirical formula formulated by the prestigious doctor in traditional Chinese medicine, in the treatment of rheumatoid arthritis based on network pharmacology and cell function experiments. The main active components and targets of JBQGF were obtained through Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP) and Encyclopedia of Traditional Chinese Medicine(ETCM), and the core targets underwent functional enrichment analysis and signaling pathway analysis. Cytoscape 3.6.0 was used to construct a visualized "active component-target-signaling pathway" network of JBQGF. After screening, nine potential pathways of JBQGF were obtained, mainly including G protein-coupled receptor signaling pathway and tyrosine kinase receptor signaling pathway. As previously indicated, the fibroblast growth factor receptor 1(FGFR1) signaling pathway was highly activated in active fibroblast-like synoviocytes(FLS) in rheumatoid arthritis, and cell and animal experiments demonstrated that inhibition of the FGFR1 signaling pathway could significantly reduce joint inflammation and joint destruction in collagen-induced arthritis(CIA) rats. In terms of the tyrosine kinase receptor signal transduction pathway, the analysis of its target genes revealed that FGFR1 might be a potential target of JBQGF for rheumatoid arthritis treatment. The biological effect of JBQGF by inhibiting FGFR1 phosphorylation was preliminarily verified by Western blot, Transwell invasion assay, and pannus erosion assay, thereby inhibiting matrix metalloproteinase 2(MMP2) and receptor activator of nuclear factor-κB ligand(RANKL) and suppressing the invasion of fibroblasts in rheumatoid arthritis and erosive effect of pannus bone. This study provides ideas for searching potential targets of rheumatoid arthritis treatment and TCM drugs through network pharmacology.
Rats ; Animals ; Synoviocytes ; Matrix Metalloproteinase 2/metabolism* ; Network Pharmacology ; Receptor, Fibroblast Growth Factor, Type 1/therapeutic use* ; Arthritis, Rheumatoid/genetics* ; Signal Transduction ; Fibroblasts ; Drugs, Chinese Herbal/therapeutic use*

OBJECTIVE: To investigate the therapeutic effect of gentisic acid (GA) on rheumatoid arthritis (RA) based on the miR-19b-3p/RAF1 axis. METHODS: The cell counting kit-8 method was used to detect the growth inhibitory effect of different concentrations of GA on MH7A cells, and the drug concentration of GA was determined in the experiment. The quantificational real-time polymerase chain reaction (qRT-PCR) was used to detect the expression of miR-19b-3p and RAF1. RAF1, extracellular regulated protein kinases1/2 (ERK1/2) and phospho-ERK1/2 (p-ERK1/2) were examined by Western blotting. Three methods (dual-luciferase assay, qRT-PCR and Western blot analysis) were used to verify miR-19b-3p targeting RAF1. Flow cytometry was performed to detect MH7A cell apoptosis. Transwell and wound healing assays were used to determine the invasion and migration capacities of MH7A cells. RESULTS: The growth of MH7A cells was gradually inhibited with increasing GA concentration. When the GA concentration exceeded 80 mmol/L, GA was significantly cytotoxic to MH7A cells, so the half maximal inhibitory concentration of GA for MH7A cells was calculated as 67.019 mmol/L. GA upregulated miR-19b-3p expression, downregulated RAF1 expression, inhibited ERK1/2 phosphorylation, induced MH7A cell apoptosis and suppressed MH7A cell invasion and migration (P<0.05 or P<0.01). RAF1 was identified as the target of miR-19b-3p and reversed inhibitory effects on miR-19b-3p expression (P<0.05 or P<0.01). The miR-19b-3p inhibitor upregulated RAF1 expression and ERK1/2 phosphorylation, suppressed MH7A cell apoptosis and induced MH7A cell invasion and migration (P<0.01). CONCLUSION GA regulated miR-19b-3p/RAF1 axis to mediate ERK pathway and inhibit the development of RA.
Humans ; Cell Proliferation ; MicroRNAs/metabolism* ; Arthritis, Rheumatoid/genetics* ; Gentisates/pharmacology* ; Cell Movement/genetics*

Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease caused by inflammatory cells. Various inflammatory cells involved in RA include fibroblast-like synoviocytes, macrophages, CD4⁺T-lymphocytes, B lymphocytes, osteoclasts and chondrocytes. The close interaction between various inflammatory cells leads to imbalance of immune response and disorder of the expression of mRNA in inflammatory cells. It helps to drive production of pro-inflammatory cytokines and stimulate specific antigen-specific T- and B-lymphocytes to produce autoantibodies which is an important pathogenic factor for RA. Competing endogenous RNA (ceRNA) can regulate the expression of mRNA by competitively binding to miRNA. The related ceRNA network is a new regulatory mechanism for RNA interaction. It has been found to be involved in the regulation of abnormal biological processes such as proliferation, apoptosis, invasion and release of inflammatory factors of RA inflammatory cells. Understanding the ceRNA network in 6 kinds of RA common inflammatory cells provides a new idea for further elucidating the pathogenesis of RA, and provides a theoretical basis for the discovery of new biomarkers and effective therapeutic targets.
Humans ; Arthritis, Rheumatoid/genetics* ; MicroRNAs/metabolism* ; Synoviocytes/pathology* ; Cytokines/metabolism* ; RNA, Messenger/metabolism* ; Fibroblasts/pathology* ; Cell Proliferation

OBJECTIVES: Rheumatoid arthritis (RA) is a chronic autoimmune disease. MicroRNA has been shown to play an important role in RA. MicroRNA-124a (miR-124a) has anti-proliferative and anti-inflammatory effects in RA fibroblast synovial cells. This study aims to explore the effects of miR-124a overexpression on arthritis in collagen-induced arthritis (CIA) mice and the underlying mechanisms. METHODS: Bovine type II collagen and complete Ferris adjuvant were used to induce CIA model from DBA/1 mice. Twenty-eight days after initial immunization (D28), CIA mice were randomly divided into a model group, a miR-124a treatment group, and a negative control (NC) group. Physiological saline, miR-124a agomir, and miR-124a agomir NC were injected into the skin at the tail root of mice every 3 days for 4 times, respectively. The degree of joint swelling and arthritis index of mice were recorded accordingly. Sixty-three days after initial immunization (D63), the mice were sacrificed to obtain the synovial tissue of ankle joint. HE staining was used to observe the proliferation of synovial cell, infiltration of inflammatory cell, pannus, and bone erosion of synovial tissues; TUNEL staining was used to detect cell apoptosis; qRT-PCR was used to detect the mRNA expression of miR-124a, phosphatidylinositol-3-kinase catalytic subunit alpha (PIK3CA) and its downstream genes Bcl-2 and Bax. Immunohistochemistry was used to detect the protein expression of PIK3CA, Bcl-2, and Bax protein in synovial tissues of each group. RESULTS: Different degrees of swelling presented in the paws of DBA/1 mice at D28, which indicated the CIA model was constructed successfully. Forty-eight days after initial immunization (D48), the paws of mice in the miR-124a treatment group were only slightly red and swollen, while the paws of mice in the model group and the NC group were obviously red and swollen. The arthritis index of mice in the miR-124a treatment group were decreased significantly compared to the NC group at D51, D53, D59, and D62 (51, 53, 59, 62 days after initial immunization) (all P<0.05). Sixty-three days after initial immunization (D63), HE staining indicated that the scores of synovial cell proliferation, inflammatory cell infiltration, synovial pannus, and bone erosion were significantly reduced in the miR-124a treatment group (P<0.05 or P<0.01), while cell apoptosis was increased in the miR-124a treatment group compared with the model group and NC group (P<0.01 or P<0.001). Besides, the expression of miR-124a and Bax in the synovial tissue in miR-124a treatment group was significantly higher than those in the model group and NC group (P<0.01 or P<0.001), while the expressions of PIK3CA and Bcl-2 were decreased (P<0.05 or P<0.01 or P<0.001), and the ratio of Bcl-2 to Bax was significantly decreased (P<0.01 or P<0.001). CONCLUSIONS Overexpression of miR-124a can reduce arthritis in CIA mice bacause it could promote synovial cell apoptosis and inhibit synovial cell proliferation via targeting PIK3CA and regulating its downstream pathways.
Animals ; Arthritis, Experimental/metabolism* ; Arthritis, Rheumatoid/genetics* ; Cattle ; Cell Proliferation ; Class I Phosphatidylinositol 3-Kinases/metabolism* ; Mice ; Mice, Inbred DBA ; MicroRNAs/metabolism* ; Proto-Oncogene Proteins c-bcl-2/metabolism* ; Synovial Membrane ; bcl-2-Associated X Protein/metabolism*

OBJECTIVES: Rheumatoid arthritis (RA) is an autoimmune disease characterized by chronic inflammation and joint destruction. Both inflammatory response and oxidative stress contribute to the pathogenesis of RA. Oxidative damage can induce and aggravate the imbalance of immune inflammation and promote cell and tissue damage. In this study, the expression of long non-coding RNA (lncRNA) LINC00638 in peripheral blood of patients with RA damp-heat arthralgia syndrome was observed, and the correlation between LINC00638 and disease activity, immune inflammation and oxidative stress indicator was investigated. Subsequently, the mechanisms for LINC00638 in regulating the inflammatory response and oxidative stress in RA fibroblast-like synoviocyte (FLS) under the condition of overexpression and interference were further explored. METHODS: In this study, 48 RA patients with damp-heat arthralgia syndrome and 27 normal healthy subjects, who came from Department of Rheumatology, First Affiliated Hospital of Anhui University of Chinese Medicine, were included; and they were divided into a RA group and a control group. The expression of LINC00638 in peripheral blood mononuclear cells (PBMC) from the subjects was detected by real-time PCR. Enzyme linked immunosorbent assay (ELISA) was used to detect serum interleukin (IL)-10, IL-17, tumor necrosis factor-α (TNF-α), malondialdehyde (MDA), heme oxygenase 1 (HO-1) and superoxide dismutase 2 (SOD2) expression. Spearman method was used to study the relationship between LINC00638 and erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), rheumatoid factor (RF), anti-cyclic citrullinated peptide antibody (anti-CCP), and to observe the relation between LINC00638 and the Disease Activity Score of 28 Joint (DAS28), Quantitative Score of Damp Heat Syndrome, Visual Analogue Scale (VAS), Self-rating Anxiety Scale (SAS) and Self-rating Depression Scale (SDS). RA-FLS was induced by RA-PBMC, and the RA in vitro cell experimental model was established. LINC00638 overexpression plasmid and small interfering RNA (siRNA) were constructed and transfected into RA-FLS. The cell experiments were divided into 4 groups: a pcDNA3. 1- control group, a pcDNA3.1-LINC00638 group, a siRNA-control group, and a siRNA-LINC00638 group. The transfection efficiency of overexpression plasmid and siRNA was detected by real-time PCR, the expression of TNF-α and IL-10 was detected by ELISA, and the expression of antioxidant proteins HO-1 and SOD2 was detected by immunofluorescence. RESULTS: Compared with the control group, the expression of LINC00638 in the RA group was lower (P<0.01). The area under the curve (AUC) of the receiver operating characteristic (ROC) curve of LINC00638 was 0.9271. The DAS28 in RA group was 5.70 (5.40-6.50), the Quantitative Score of Damp-heat Syndrome was 20.0 (17.0-23.0), and the VAS score was 7.0 (6.3-8.0). Compared with the control group, the ESR, CRP, RF, anti-CCP, SAS and SDS scores in the RA group were significantly increased (all P<0.01). Spearman correlation analysis showed that: LINC00638 was negatively correlated with ESR (r=-0.532, P<0.01), CRP (r=-0.367, P<0.05), TNF-α (r=-0.375, P<0.01), MDA (r= -0.295, P<0.05), DAS28 (r=-0.450, P<0.01), and which was positively correlated with SOD2 (r=0.370, P<0.05). After the induction of RA-FLS, the expression level of LINC00638 was significantly decreased (P<0.01), indicating that the stimulation of PBMC could effectively reduce the expression of LINC00638 in RA-FLS, so the experimental model of RA-FLS-induced by PBMC was utilized. Compared with the pcDNA3.1-control group, the expressions of LINC00638, IL-10, SOD2, and HO-1 in the pcDNA3.1-LINC00638 group were significantly increased (all P<0.01), and the expression of TNF-α was decreased (P<0.01). Compared with siRNA-control group, LINC00638, IL-10, SOD2 and HO-1 in the siRNA-LINC00638 group were significantly decreased (all P<0.01), and TNF-α was significantly increased (P<0.01). CONCLUSIONS LINC00638 is down-regulated in the peripheral blood of RA patients with damp-heat arthralgia syndrome, which is correlated with disease activity, immune inflammation and oxidative stress. Overexpression of LINC00638 can down-regulate pro-inflammatory factors, up-regulate anti-inflammatory factors, and increase antioxidant enzyme activity, thereby improving inflammation and oxidative stress in RA. LINC00638 is the differential lncRNA obtained by the research group's previous high-throughput sequencing of the whole transcriptome of peripheral blood PBMCs in RA patients and validation of clinical samples. In order to deepen the molecular biology research of this gene, the microRNA and mRNA targeted by LINC00638 can be further studied from the perspective of competing endogenous RNAs.
Anti-Citrullinated Protein Antibodies/metabolism* ; Antioxidants ; Arthralgia/metabolism* ; Arthritis, Rheumatoid ; C-Reactive Protein ; Hot Temperature ; Humans ; Inflammation/genetics* ; Interleukin-10/metabolism* ; Leukocytes, Mononuclear ; Oxidative Stress ; RNA, Long Noncoding/metabolism* ; RNA, Small Interfering ; Tumor Necrosis Factor-alpha/metabolism*

The present study investigated the pharmacodynamic material basis of Laportea bulbifera in the treatment of rheumatoid arthritis. Firstly, human rheumatoid arthritis fibroblast-like synoviocyte line MH7A was cultured in vitro and treated with tumor necrosis factor alpha(TNF-α, 50 ng·mL~(-1)). The proliferation and the levels of inflammatory cytokines such as prostaglandin E2(PGE2), interleukin-1β(IL-1β), and interleukin-6(IL-6) of the MH7A cells exposed to the serum containing L. bulbifera were determined to evaluate the anti-rheumatoid arthritis effects of the serum. Furthermore, the ultra-performance liquid chromatography tandem mass spectrometry fingerprints of the L. bulbifera crude extract, the drug-containing serum, and the drug-free serum were compared to identify the compounds newly generated in the serum after oral administration of the extract. According to the peak areas of common peaks and the results of anti-rheumatoid arthritis effect test, the active components were identified. The serum containing L. bulbifera significantly inhibited the proliferation of the MH7A cells activated by TNF-α and the expression of PGE2, IL-6, and IL-1β. Thirty newly generated compounds were detected in the drug-containing serum. Among them, neochlorogenic acid, cryptochlorogenic acid, chlorogenic acid, rutin, isoquercitrin, luteoloside, kaempferol-3-O-rutinoside, and quercitrin were also present in the crude extract. Twelve characteristic peaks(3, 7, 8, 14, 18, 19, 21, 23, 24, m6, m7, and m15) were significantly correlated with the pharmaceutical effect. According to the correlations, neochlorogenic acid, cryptochlorogenic acid, and chlorogenic acid had great contributions to the anti-rheumatoid arthritis activity. This study preliminarily clarified the potential pharmacodynamic substances of L. bulbifera in the treatment of rheumatoid arthritis, which laid a theoretical and experimental foundation for further development and application of the medicinal plant.
Animals ; Arthritis, Experimental/drug therapy* ; Arthritis, Rheumatoid/drug therapy* ; Chlorogenic Acid/analogs & derivatives* ; Cytokines/metabolism* ; Dinoprostone ; Humans ; Interleukin-1beta/genetics* ; Interleukin-6 ; Plant Extracts/therapeutic use* ; Quinic Acid/analogs & derivatives* ; Rutin ; Tumor Necrosis Factor-alpha/metabolism* ; Urticaceae/chemistry*

This paper aims to explore the activity of Codonopsis canescens extract against rheumatoid arthritis(RA) based on the Toll-like receptors(TLRs)/mitogen-activated protein kinases(MAPKs)/nuclear factor kappa B(NF-κB) signaling pathways and its mechanism. The ultra-performance liquid chromatography-quadrupole time-of-flight/mass spectrometry(UPLC-Q-TOF-MS) was used to identify the components of C. canescens extract. Forty-eight male SD rats were randomly divided into six groups, namely the normal group, the model group, the methotrexate(MTX) tablet group, and the low, medium, and high-dose C. canescens extract(ZDS-L, ZDS-M, and ZDS-H) groups, with 8 rats in each group. The model of collagen-induced arthritis in rats was induced by injection of bovine type Ⅱ collagen emulsion. MTX(2.5 mg·kg~(-1)), ZDS-L, ZDS-M, and ZDS-H(0.3 g·kg~(-1), 0.6 g·kg~(-1), and 1.2 g·kg~(-1)) were administrated by gavage. Rats in the normal group and the model group received distilled water. MTX was given once every three days for 28 days, and the rest medicines were given once daily for 28 days. Body weight, degree of foot swelling, arthritis index, immune organ index, synovial histopathological changes, and serum levels of tumor necrosis factor-α(TNF-α), interleukin-1β(IL-1β), and interleukin-6(IL-6) were observed. Protein expressions of TLR2, TLR4, NF-κB p65, p38 MAPK, and p-p38 MAPK in rats were determined by Western blot. Thirty-four main components were identified by UPLC-Q-TOF-MS, including 15 flavonoids, 7 phenylpropanoids, 4 terpenoids, 4 organic acids, 2 esters, and 2 polyalkynes. As compared with the normal group, the body weight of the model group was significantly decreased(P<0.01), and foot swelling(P<0.05, P<0.01), arthritis index(P<0.01), and the immune organ index(P<0.01) were significantly increased. The synovial histopathological injury was obviously observed in the model group. The serum levels of inflammatory factors TNF-α, IL-1β, and IL-6 were significantly increased(P<0.01), and the protein expression levels of TLR2, TLR4, NF-κB p65, p-p38 MAPK/p38 MAPK in the synovial tissue were significantly increased(P<0.01) in the model group. As compared with the model group, the body weights of the ZDS dose groups were increased(P<0.01), and the degree of foot swelling(P<0.01) and the arthritis index were decreased(P<0.05, P<0.01). The immune organ index was decreased(P<0.01) in the ZDS dose groups, and the synovial tissue hyperplasia and inflammatory cell infiltration were alleviated. The serum levels of TNF-α, IL-1β, and IL-6 were significantly decreased(P<0.05, P<0.01), and the protein expression levels of TLR2, TLR4, NF-κB p65, p-p38 MAPK/p38 MAPK were decreased(P<0.05, P<0.01) in the ZDS dose groups. C. canescens extract containing apigenin, tricin, chlorogenic acid, aesculin, ferulic acid, caffeic acid, and oleanolic acid has a good anti-RA effect, and the mechanism may be related to the inhibition of TLRs/MAPKs/NF-κB signaling pathways.
Animals ; Cattle ; Male ; Rats ; Arthritis, Experimental/drug therapy* ; Arthritis, Rheumatoid/drug therapy* ; Body Weight ; Codonopsis/chemistry* ; Interleukin-6/blood* ; NF-kappa B/genetics* ; p38 Mitogen-Activated Protein Kinases/metabolism* ; Plant Extracts/therapeutic use* ; Rats, Sprague-Dawley ; Signal Transduction ; Toll-Like Receptor 2/metabolism* ; Toll-Like Receptor 4/metabolism* ; Tumor Necrosis Factor-alpha/pharmacology*

OBJECTIVE: To investigate the expression of leukocyte immunoglobulin-like receptor A3 (LILRA3) in CD14 monocytes of patients with rheumatoid arthritis (RA). METHODS: Fifty three RA patients admitted in the Second Affiliated Hospital of Zhejiang University School of Medicine from February 2017 to August 2017, and 21 healthy subjects were enrolled in the study. The expression of LILRA3 in CD14 monocyte subset was determined by flow cytometry, and its correlations with clinical features, laboratory examination results, antibodies and disease activity were analyzed. RESULTS: LILRA3 percentage in the CD14 monocyte subset of RA patients was higher than that in the healthy controls (<0.01). The percentage of LILRA3 was positively correlated with number of tenderness joints, number of swollen joints and erythrocyte sedimentation rate (=0.280, 0.371, 0.341, <0.05 or <0.01), but was not correlated with the age, course of disease, Sharp score, C reactive protein, blood routine index and immunoglobulin (all >0.05). In addition, the percentages of LILRA3 in the monocytes of rheumatoid factor (RF)-positive or anti-cyclic citrullinated peptide (CCP) antibody-positive patients were significantly higher than those of the RF-or anti-CCP antibody-negative patients (all < 0.05); and the percentage of LILRA3 in patients with DAS28>5.1 was higher than that in patients with DAS28 ≤ 5.1 (<0.05). CONCLUSIONS The expression of LILRA3 is up-regulated in CD14 monocyte subset isolated from RA patients, and it is correlated with disease activity.
Arthritis, Rheumatoid ; blood ; physiopathology ; Autoantibodies ; blood ; Biomarkers ; blood ; Flow Cytometry ; Humans ; Monocytes ; metabolism ; Receptors, Immunologic ; genetics ; Up-Regulation

OBJECTIVE: To investigate the expression of apoptosis related protein cellular Fas associated death domain like interleukin 1 converting enzyme inhibitory protein (c-FLIP) in peripheral blood mononuclear cells (PBMCs) of patients with rheumatoid arthritis and its relation with extrinsic apoptotic pathway. METHODS: Sixty patients with rheumatoid arthritis were collected from Zhangzhou Affiliated Hospital of Fujian Medical University during January 2014 and June 2015, including 22 patients with low activities (DAS28<3.2), 20 patients with middle activities (3.2 ≤ DAS28 ≤ 5.1), and 18 patients with high activities (DAS28>5.1). And 25 healthy controls were also collected. The mRNA and protein expression levels of c-FLIP and the extrinsic apoptotic pathway related proteins Fas-associated protein with death domain (FADD), caspase-8 in PBMCs were detected by real-time RT-PCR and Western blot, respectively. Correlations between c-FLIP and FADD, caspase-8 in PBMCs were analyzed by pearson test. RESULTS: mRNA expression levels of c-FLIP, FADD and caspase-8 in PBMCs of patients with rheumatoid arthritis were all higher than those of healthy controls (all <0.05). mRNA expression levels of FADD and caspase-8 in patients with middle activities were significantly higher than those in patients with low activities (all <0.05), but the mRNA expression level of c-FLIP was not significantly higher than that in patients with low activities. mRNA expression level of c-FLIP in patients with high activities was higher than those in patients with middle or low activities (all <0.05), while the mRNA expression level of caspase-8 was lower than those in patients with middle or low activities (all <0.05). mRNA expression level of FADD in patients with high activities was higher than those in patients with low activities (<0.05). Pearson analysis showed that there was a positive correlation between c-FLIP and FADD mRNA expression (=0.323, <0.05), and negative correlation between c-FLIP and caspase-8 mRNA expression (-1.104, <0.05). The protein expression levels of c-FLIP and FADD in patients with middle activities were significantly higher than those in control group and patients with low or high activities (<0.05 or 0.01). The protein expression levels of caspase-8 in patients with middle and high activities were significantly higher than those in control group and patients with low activities (<0.05 or <0.01), and the protein expression level of caspase-8 in patients with high activities was higher than that in patients with middle activities (<0.05). CONCLUSIONS c-FLIP may be involved in the extrinsic apoptotic pathway in rheumatoid arthritis, and can provide reference for the evaluation of disease activities.
Apoptosis ; genetics ; Arthritis, Rheumatoid ; blood ; CASP8 and FADD-Like Apoptosis Regulating Protein ; genetics ; Gene Expression Profiling ; Gene Expression Regulation ; Humans ; Leukocytes, Mononuclear ; metabolism