Objective
To investigate the role of scavenger receptor class B type 1(SR-B1) signaling pathway in serum amyloid A(SAA)-induced angiogenesis in rheumatoid arthritis(RA).
Methods
The expression and location of SR-B1 in RA and osteoarthritis(OA) synovial tissues were observed by immun-ohistochemistry. And SR-B1 expression in the resting human umbilical vein endothelial cells(HUVECs) was detected by immunoflourescence. Wound repair assessement and tube formation assessement were employed to evaluate the effect on cell migration and tube formation stimulated by SAA and/or anti-SR-B1 antibody. The t-test and one-way analysis of variance(ANOVA) were used for statistical analysis.
Results
① SR-B1 was significantly highly expressed in RA tissue samples(A=6 788±819) when compared to the minimal expression in OA(A=31 849±6 977,t=3.567,P<0.01). Positive staining of SR-B1 was observed in RA synovial vascular endothelial cells and perivascular areas. ② Strong staining for SR-B1 was observed in all HUVECs tested. ③ Significant wound healing induced by SAA(MI=2.50±0.17) was found compared with the untreated controls(MI=1.00±0.09, q=14.38,P<0.01), and the effects were inhibited in the presence of anti-SR-B1 antibody(MI=1.16±0.14, q=13.02,P<0.01). ④ Compared to the untreated group(branch point number: 6.6±0.8), there was an enhanced formation of branched and capillary-like tube structure followed by SAA stimulation(branch point number: 19.0±1.1,q=25.04,P<0.01) after culturing for 72 h, whereas, tube formation decreased markedly upon pre-treated with anti-SR-B1 antibody(branch point number: 7.6±1.3, vs SAA,q=23.32,P<0.01).
Conclusion
Our present study suggests that serum amyloid A may induce angiogenesis via SR-B1 signaling pathway in RA.
Key words:
Serum amyloid A protein; Arthritis, rheumatoid; Scavenger receptor class B type 1; Angiogenesis
Objective
Fibroblast-like synoviocytes (FLS) play important roles in the pathogenesis of rheumatoid arthritis (RA). The present study was undertaken to investigate the mechanism of calreticulin (CRT) to promote FLS survival in RA.
Methods
FLS were isolated by enzymatic digestion of synovial tissue specimens obtained from RA and osteoarthritis (OA) patients and cultured in vitro. The expression of Bcl-XL and Mcl-1 in FLS at mRNA and protein level was detected by quantitative-polymerase chain reaction (q-PCR), Western blotting and immunofluorescence respectively. RA and OA FLS were cultured with different concentrations of recombinant human CRT for 48-72 h, the expression of Bcl-XL and Mcl-1 was detected by q-PCR and Western blotting. The proliferation of RA FLS following CRT stimulation was determined by MTT assay.
Results
① Compared with FLS from OA patients (1.00 ± 0.39; 1.00 ± 0.46), the anti-apoptotic Bcl-XL and Mcl-1 mRNA expression (14.51 ± 2.20; 12.82 ± 1.80) was significantly higher in the FLS from RA patients (t=10.47, 11.02; P 0.05); Western blotting results showed elevated protein levels of both Bcl-XL and Mcl-1 in RA FLS after CRT treatment at a concentration dependent manner. However, neither Bcl-XL nor Mcl-1 expression was significantly changed in OA FLS. ③ MTT assay showed that CRT had no significant effect on the proliferation of RA FLS (F=2.88, P> 0.05).
Conclusion
Our results indicate that CRT-mediated up-regulation of anti-apoptotic Bcl-XL and Mcl-1 may inhibit apoptosis and promote the survival of FLS from RA patients.
Key words:
Calreticulin; Arthritis, rheumatoid; Synovial membrane
Objective
To investigate the anti-apotoptic effects of calreticulin (CRT) on tumour necrosis factor-related apoptosis-inducing ligand (TRAIL)-mediated T cells apoptosis in rheumatoid arthritis (RA).
Methods
Levels of CRT in the synovial fluid from patients with RA (23 cases) and osteoarthritis (OA) (36 cases) were detected with enzyme-linked immunosorbent assay (ELISA); the expression and localization of CRT in RA and OA synovial tissue were detected by immunohistochemistry. The capacity of CRT binding to TRAIL was measured by ELISA; the effect of TRAIL on Jurkat T cell proliferation was determined by the methylthiazolyl tetrazolium (MTT) assay, CRT on TRAIL-mediated apoptosis by flow cytometry analysis. The t-test and SNK-q test were used for statistical analysis.
Results
① CRT were significantly up-regulated both in synovial fluid and tissue from RA than those from OA [ (5.6±2.5) ng/ml and (3.2±1.3) ng/ml, t=6.233, P<0.05; 32 787±18 294 and 7 945±2 765, t=15.662, P<0.01]. ② Binding rate of CRT-TRAIL increased to (117±9)% and(155±14)% with TRAIL stimulation (1 μg/ml and 2 μg/ml, respectively) compared to the controls (q=3.858, 8.647; P<0.01). ③ The proliferation of Jurkat T cell was inhibited after co-cultured with TRAIL, at 12 hours, 100ng/ml TRAIL treated cell showed a significant decreased survival rate and at 24 hours(q=7.532, P<0.05), and the similar results were shown both with 50 ng/ml and 100 ng/ml TRAIL (q=9.211, 12.879; P<0.01). ④ T cell early apoptosis was suppressed by the combination of CRT and TRAIL: with CRT (4, 8 and 16 ng/ml), the corresponding apoptotic rates were (30.7±1.1)%, (24.5±1.1)% and (22.3±1.5)% respectively, which showed significant differences compared with the control group (32.5±1.4)% (q=3.82, 12.17, 15.39; P<0.05).
Conclusion
Increased levels of CRT are detected in the synovial fluid and tissue in RA; and CRT is expressed in inflammatory cells in synovial tissue from RA patients. In addition, CRT can partly inhibit TRAIL-mediated apoptosis of Jurkat T cells via binding to TRAIL, which may suggest its potential role in the pathogenesis especially in inflammation of RA.
Key words:
Arthritis, rheumatoid; Calreticulin; Tumor necrosis factor-related apoptosis-inducing ligand
Calprotectin S100A8/A9, a heterodimer composed of S100A8 and S100A9, is the main component of cytoplasmic proteins in neutrophils. It plays multiple roles in the immuno-inflammatory reactions intracellularly and extracellularly. Recent studies find that S100A8/A9 is closely related with the initiation and progression of periodontal inflammatory diseases. S100A8/A9 is expected to be a new biomarker for diagnosing periodontal inflammatory diseases, monitoring inflammatory activities in patients with periodontitis, evaluating the outcome of periodontal treatments and predicting the susceptibility of individual patient to periodontitis. In this literature review, we summarize the clinical research progress on the relation between S100A8/A9 and periodontal inflammatory diseases.S100A8/A9蛋白是由S100A8与S100A9蛋白组成的异二聚体，属于钙结合蛋白S100家族，是中性粒细胞的主要胞质蛋白，在免疫炎症反应中发挥多种胞内、胞外的生物学功能。近年研究发现，S100A8/A9与牙周炎症性疾病的发生发展关系密切。S100A8/A9在诊断牙周炎症性疾病、监测牙周炎症活动性、评价牙周治疗效果以及预测牙周炎易感性等方面有望成为新的生物标志物。本文就S100A8/A9蛋白与牙周炎症性疾病关系的临床研究进展作一文献综述。.
Proliferative gingivitis was studied by electron microscopy. The crevicular epithelium adjacent to inflamed gingival connective tissue was primarily characterized by its relatively larger intercellular spaces which contained a variety of emigrating cells and granular precipitates resembling plasmic substances. Neutrophils and lymphocytes migrated between epithelial cells so that the intercellular spaces enlarged. The interruption of basement membrane was associated with inflammatory cells. Among the inflammatory cells plasma cells were predominant and displayed a variety of morphological characteristics. They consisted primarily of tightly packed, flattened cisternae of rough endoplasmic reticulum (rER) and a prominent paranuclear golgi zone. Fragments of cytoplasm apparently originating from the plasma cells were lying free in the surrounding intercellular space, in which collagen fibrils were impaired, disrupted and dissolved. All of these changes in plasma cells may relate to the synthesis and release of antibody. It is suggested that the inflammatory destructive features in connective tissue were mainly contributed to plasma cells.
Objective
To investigate the neutrophil extracellular traps (NETs) formation and their molecular mechanisms induced by serum amyloid A (SAA) in rheumatoid arthritis (RA).
Methods
Neutrophils were isolated from peripheral blood of RA and healthy volunteers. ① Neutrophils were cultured in vitro, the formation of NETs was observed and their percentage was calculated. ② Neutrophils were cultured in vitro, divided into six groups: control, SAA, [SAA+anti-Toll like receptro4 (TLR4)-Ab], LPS, (LPS+anti-TLR4-Ab) and anti-TLR4-Ab. Appropriate stimulation was conducted for each group. NETs formation and their percentages were investigated. The concentration of DNA in supernatant was detected by fluorescent staining. F test and t test were used for statistical analysis.
Results
① The purification of isolated neutrophils was higher than 95%. The network which was collocated with the spreading neutrophils nucleus and neutrophil elastase under the microscope, was NETs. In the RA group, the formation of NETs induced by SAA was significantly more than control [(19.1±0.8) vs (7.4±0.5), t=12.30, P<0.05]. ② However, after pretreated with anti-TLR4 antibody, NETs formation was significantly less than the SAA group [(5.7±0.4) vs (14.7±1.1), t=7.825, P<0.05]. Moreover, the fluorescence intensity of DNA in supernatant was significantly higher in SAA group than that of anti-TLR4-Ab pretreatment group[(18.7±0.7) vs (12.9±0.8), t=5.552, P<0.05]. The concentration of DNA in supernatant of SAA group was higher than that of anti-TLR4-Ab pretreatment group[(36.9±1.3) μg/ml and(16.3±0.6) μg/ml, t=14.41, P<0.05].
Conclusion
SAA can induce the formation of NETs from neutrophils by binding to TLR4 in RA.
Key words:
Arthritis, rheumatoid; Serum amyloid A protein; Toll-like receptor 4; Neutrophil extra-cellular traps
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