Xingfu Li

Glucocorticoid downregulates expression of IL-12 family cytokines in systemic lupus erythematosus patients

Objective This study aims to investigate expression of interleukin 12 (IL-12) family cytokines IL-12, IL-23, IL-27 and IL-35 in systemic lupus erythematosus (SLE) patients and the effect of glucocorticoid (GC) treatment on their expression. Methods Plasma concentration of IL-12, IL-23, IL-27, IL-35, IL-6 and anti-double-stranded DNA (dsDNA) antibodies in 30 newly diagnosed severe SLE patients and 30 matched healthy subjects was measured by enzyme-linked immunosorbent assay. The correlation between the levels of IL-12 family cytokines and the levels of IL-6 or anti-dsDNA antibodies was analyzed by Spearman rank correlation. Results Significantly higher levels of plasma IL-12, IL-23, IL-27, IL-35, IL-6 and anti-dsDNA antibodies were observed in SLE patients compared with healthy controls (p<0.05), and after prednisone treatment, the serum levels of IL-12 family cytokines decreased significantly. Moreover, serum levels of IL-12, IL-23, IL-27 and IL-35 were correlated with serum levels of IL-6 and anti-dsDNA antibodies in pre-treatment as well as post-treatment SLE patients. Conclusions SLE patients have increased plasma levels of IL-12 family cytokines and GCs can downregulate the expression of them in SLE patients. Therefore, members of the IL-12 family may be involved in the pathophysiological process of SLE.

Expression level of the growth factor progranulin is related with development of systemic lupus erythematosus

BackgroundThis study is to investigate the expression of progranulin (PGRN) in systemic lupus erythematosus (SLE) patients and the effect of glucocorticoid (GC) treatment on its expression.MethodsThirty newly diagnosed severe SLE patients and 30 healthy subjects were enrolled in this study. The serum levels of PGRN and the inflammatory factors of SLE were detected by ELISA and the mRNA expression of these proteins were detected by real-time PCR.ResultsThe serum levels of PGRN, IL-6, PR3, TNFR, TNF-α and anti-dsDNA antibody in SLE patients were increased significantly compared with healthy controls (P < 0.05). The relative expression of PGRN mRNA was increased by 4.88-fold in pre-treatment SLE patients compared with controls (P < 0.05). After prednisone treatment, the serum levels of PGRN decreased significantly, and the relative expression of PGRN mRNA was decreased by 1.34-fold compared with the untreated controls (P < 0.01). Moreover, Serum concentration of PGRN was correlated with serum levels of IL-6, TNF-α, TNFR and anti-dsDNA antibody in both pre-treatment and post-treatment SLE patients.ConclusionsPGRN is up-regulated in the SLE patients and is correlated with pro-inflammatory cytokines and anti-dsDNA antibody. Glucocorticoids can down-regulate the expression of PGRN in SLE patients.Virtual slideshttp://www.diagnosticpathology.diagnomx.eu/vs/1562484036905973

Using SELDI‐TOF MS to identify serum biomarkers of rheumatoid arthritis

Objectives: No satisfactory biomarkers are currently available to screen for rheumatoid arthritis (RA). We have developed and evaluated surface‐enhanced laser desorption/ionization time‐of‐flight mass spectrometry (SELDI‐TOF MS) for detection and analysis of multiple proteins for distinguishing individuals with RA from control individuals. Methods: A total of 156 serum samples from 90 RA patients, 30 patients with ankylosing spondylitis (AS), and 36 healthy individuals were examined by SELDI technology. Spectral data were analysed by the support vector machine (SVM) approach and potential biomarkers were chosen for system training and were used to construct a diagnostic model. Results: Pattern 1, consisting of four protein peaks with m/z values of 3899, 4594, 7566, and 13 842, distinguished RA from the healthy samples with sensitivity of 90.0% and a specificity of 91.7%. Pattern 2, consisting of m/z peaks 4287 and 6471, distinguished RA from AS with a sensitivity of 86.7% and a specificity of 85.0%. Conclusion: The combination of SELDI‐TOF MS and SVM could facilitate the discovery of better biomarkers for RA and also provide a useful tool for molecular diagnosis in the future.

Combined measurement of multiple acute phase reactants to predict relapse of rheumatoid arthritis.

Acute phase reactants (APRs), such as serum C‐reactive protein (CRP), ferritin, plasma fibrinogen and platelet count, are common biomarkers used to monitor the status of inflammatory diseases. The aim of this study was to determine whether APRs are predictive markers of relapse in rheumatoid arthritis (RA).

Increased Tim-3 expression on peripheral T lymphocyte subsets and association with higher disease activity in systemic lupus erythematosus

BackgroundBoth the T cell immunoglobulin domain- and mucin domain-containing molecule-3 (Tim-3) and the death receptor Fas contribute to the pathogenesis of various autoimmune diseases, including systemic lupus erythematosus (SLE). The aim of the present study was to determine whether Tim-3 and Fas are co-expressed on certain peripheral T lymphocyte subsets, and whether this expression is associated with greater disease activity in SLE.MethodsPeripheral blood mononuclear cells were isolated from 46 patients newly diagnosed with SLE and 28 age- and sex-matched healthy controls (HCs). Expression of Tim-3 and Fas on T subsets was analyzed by flow cytometry, while mRNA levels of the Tim-3 ligand galectin-9 and Fas ligand FasL were assayed using real-time RT-PCR.ResultsThe proportions of CD3+CD4+ and CD3+CD4- T cells expressing Tim-3+ and Tim+Fas+ were significantly higher in patients than in HCs (p < 0.05), while the proportions of these subtypes expressing Fas were similar for the two groups. Patients with active SLE, as defined by their score on the SLE Disease Activity Index, had lower proportions of CD3+CD4+ T cells and higher proportions of CD3+CD4+Tim-3+ and CD3+CD4+Tim-3+Fas+ T cells than did patients with stable SLE. Serum levels of complement C3 and C4 proteins, considered as a marker of SLE activity, correlated negatively with proportions of CD3+CD4+ and CD3+CD4- T cells expressing Tim-3.ConclusionsExpression of Tim-3 and co-expression of Tim-3 and Fas on certain peripheral T subsets are associated with disease activity in SLE patients. Future research should examine whether the same is true of other T subsets implicated in SLE, and should explore the potential role(s) of Tim-3 in the disease pathway.Virtual slideshttp://www.diagnosticpathology.diagnomx.eu/vs/1855527845145188

The role of Act1, a NF-κB-activating protein, in IL-6 and IL-8 levels induced by IL-17 stimulation in SW982 cells

Abstract Context: Rheumatoid arthritis (RA) is a systemic autoimmune disease characterized by chronic inflammation in the synovial membrane of affected joints. It has been shown that several kinds of cytokine were increased in synovial fluid, while the underlying mechanism remains poorly understood. Objectives: NF-κB activator 1 (Act1) is a recently identified protein binding to the IκB kinase complex. Our study aimed to investigate the expression of Act1 induced by cytokine IL-17 stimulation in SW982 cells. Materials and methods: The human synovial sarcoma cell line SW982 and primary cultured RA fibroblast-like synovial cells were used. RT-PCR and Western blot assays were selected to investigate the genetic and protein expression of Act1. Additionally, four independent Act1 small interfering RNA (siRNA) oligonucleotides were designed and obtained according to the GenBank cDNA, the sequence of Act1 (Traf3ip2). Finally, enzyme-linked immunosorbent assay (ELISA) double antibody sandwich was used to assay supernatant IL-6 and IL-8 concentrations. Results: The Act1 mRNA expression level increased significantly after stimulation with IL-17 (5–100 ng/ml) in SW982 cells. Additionally, the level of Act1 mRNA expression correlated positively with the concentration of IL-17 (p < 0.01). IL-17 induced IL-6 and IL-8 in SW982 cells was in a concentration- and time-dependent way. Furthermore, ELISA assay revealed that IL-17 (20 ng/ml) significantly increased IL-6 (1927.4 ± 288.77 versus 786.5 ± 172.42 ng/ml, p < 0.01) and IL-8 levels (984.8 ± 95.09 ng/ml versus 307.1 ± 90.83 ng/ml, p < 0.01) compared with control group after stimulation for 24 h. However, transfection of Traf3ip2 siRNA markedly decreased IL-6 (995.9 ± 115.30 ng/ml versus 1816.1 ± 273.27 ng/ml, p < 0.01) and IL-8 levels (575.6 ± 65.96 ng/ml versus 929.4 ± 124.39 ng/ml, p < 0.01) compared to transfection negative control. These findings suggested that IL-6 and IL-8 level induced by IL-17 in SW982 cells could be reversed by down-regulation of Act1 expression level with Traf3ip2 siRNA. Conclusion: Our results suggested that Act1 might play a key role in the pathophysiology and the treatment of RA.

Upregulated expression level of the growth factor, progranulin, is associated with the development of primary Sjögren's syndrome

The aim of the present study was to investigate the expression and effect of progranulin (PGRN) in patients with primary Sjögren’s syndrome (pSS). In total, 26 newly diagnosed pSS patients and 26 healthy subjects were enrolled in this study. The serum levels of PGRN and the inflammatory factor, interleukin-6 (IL-6), were detected using ELISA. In addition, the mRNA expression levels of these molecules were detected by quantitative polymerase chain reaction. The serum levels of PGRN and IL-6 in the pSS patients increased significantly compared with the healthy controls (P<0.05). During the remission stages, the levels of PGRN and IL-6 were comparable to those of the healthy controls. The serum level of PGRN in the pSS patients was shown to correlate with that of IL-6 in the pre-treatment and post-treatment stages. PGRN was upregulated in the pSS patients, indicating a possible role of PGRN in the pathogenesis and development of pSS.

Noradrenaline regulates substance P release from rat dorsal root ganglion neurons in vitro

To determine whether activation and/or inhibition of α-adrenoreceptors influences substance P (SP) release from dorsal root ganglion (DRG) primary sensory neurons in vitro. DRGs were dissected from 15-day embryonic Wistar rats. DRG neurons were dissociated and cultured for 2 d and then exposed to noradrenaline (NA) alone (1×10−4 mol/L), or along with the α1-adrenoreceptor antagonist prazosin (1×10−6 mol/L) or the α2-adrenoreceptor antagonist yohimbine (1×10−5 mol/L) for 4 d. Then, RT-PCR was used to determine the levels of preprotachykinin (PPT) mRNA encoding for SP and Western blot to assess the protein levels of SP. Basal and capsaicin (CAP)-evoked SP release were measured by enzyme-linked immunosorbent assay (ELISA). CAP-evoked SP release was sensitized by NA and this effect was inhibited by pre-incubation with prazosin but not with yohimbine. The levels of PPT mRNA, SP peptide, and basal SP release did not change significantly in any of the experimental conditions. NA may significantly increase CAP-evoked SP release through activation of α-adrenoreceptors, which may contribute to noradrenergic pain modulation. 观察激活或抑制α-肾上腺素受体是否影响体外培养的背根神经节(dorsal root ganglion, DRG)神经元 P物质(substance P, SP)的释放。 胎龄15天的Wistar大鼠DRG神经元培养2天后, 分别用去甲肾上腺素(noradrenaline, NA)(1×10−4 mol/L)、 α1-受体拮抗剂哌唑嗪(1×10−6 mol/L)+NA(1×10−4 mol/L)、 α2-受体拮抗剂育亨宾(1×10−5 mol/ L)+NA(1×10−4 mol/L)孵育4天。 用RT-PCR法检测DRG神经元编码SP蛋白的PPT mRNA表达水平, 用Western blot 法检测DRG神经元SP蛋白的表达水平, 用酶联免疫吸附测定法检测SP的基础释放量和辣椒素刺激后的释放量。 NA单独孵育显著增加了DRG神经元辣椒素刺激后的SP释放量, α1-受体拮抗剂哌唑嗪预处理可阻断NA的效应, 而α2-受体拮抗剂育亨宾不产生此作用。 在各种实验条件下, PPT mRNA水平、 SP蛋白表达水平和SP的基础释放量没有显著性差异。 NA通过激活α1-受体增加了DRG神经元辣椒素刺激后的SP释放量, 这一作用可能与去甲肾上腺素能的疼痛调节有关。ObjectiveTo determine whether activation and/or inhibition of α-adrenoreceptors influences substance P (SP) release from dorsal root ganglion (DRG) primary sensory neurons in vitro.MethodsDRGs were dissected from 15-day embryonic Wistar rats. DRG neurons were dissociated and cultured for 2 d and then exposed to noradrenaline (NA) alone (1×10−4 mol/L), or along with the α1-adrenoreceptor antagonist prazosin (1×10−6 mol/L) or the α2-adrenoreceptor antagonist yohimbine (1×10−5 mol/L) for 4 d. Then, RT-PCR was used to determine the levels of preprotachykinin (PPT) mRNA encoding for SP and Western blot to assess the protein levels of SP. Basal and capsaicin (CAP)-evoked SP release were measured by enzyme-linked immunosorbent assay (ELISA).ResultsCAP-evoked SP release was sensitized by NA and this effect was inhibited by pre-incubation with prazosin but not with yohimbine. The levels of PPT mRNA, SP peptide, and basal SP release did not change significantly in any of the experimental conditions.ConclusionNA may significantly increase CAP-evoked SP release through activation of α-adrenoreceptors, which may contribute to noradrenergic pain modulation.摘要目的观察激活或抑制α-肾上腺素受体是否影响体外培养的背根神经节(dorsal root ganglion, DRG)神经元 P物质(substance P, SP)的释放。方法胎龄15天的Wistar大鼠DRG神经元培养2天后, 分别用去甲肾上腺素(noradrenaline, NA)(1×10−4 mol/L)、 α1-受体拮抗剂哌唑嗪(1×10−6 mol/L)+NA(1×10−4 mol/L)、 α2-受体拮抗剂育亨宾(1×10−5 mol/ L)+NA(1×10−4 mol/L)孵育4天。 用RT-PCR法检测DRG神经元编码SP蛋白的PPT mRNA表达水平, 用Western blot 法检测DRG神经元SP蛋白的表达水平, 用酶联免疫吸附测定法检测SP的基础释放量和辣椒素刺激后的释放量。结果NA单独孵育显著增加了DRG神经元辣椒素刺激后的SP释放量, α1-受体拮抗剂哌唑嗪预处理可阻断NA的效应, 而α2-受体拮抗剂育亨宾不产生此作用。 在各种实验条件下, PPT mRNA水平、 SP蛋白表达水平和SP的基础释放量没有显著性差异。结论NA通过激活α1-受体增加了DRG神经元辣椒素刺激后的SP释放量, 这一作用可能与去甲肾上腺素能的疼痛调节有关。

Analysis of 15 patients with systemic lupus erythematosus manifesting with negative immunofluorescence anti-nuclear antibodies after treatment

The purpose of this study was to investigate the clinical and laboratorial characteristics of patients with systemic lupus erythematosus (SLE) manifesting with negative immunofluorescence anti-nuclear antibodies (IFANA) after treatment for the better understanding of negative conversion of IFANA. Demographic characteristics, clinical and laboratory data of hospitalized SLE patients between March 2006 and May 2011 were retrospectively reviewed. Fifteen cases with negative IFANA were identified in 960 patients. All of the 15 patients were severe, 11 patients manifested with nephritic range proteinuria and hypoalbuminemia, 8 patients were complicated with severe infection and all of the patients had been treated with glucocorticoid and immunosuppressant. Anti-ENA antibodies were positive in 4 of 15 patients. Eight patients died after average 1-year follow-up. Collectively, negative IFANA is mainly attributed to nephritic-range proteinuria; and large-dose glucocorticoid, immunosuppressant and severe infection are also important factors for negative IFANA. Antinuclear antibody can be detected in some SLE patients with negative IFANA by changing the detection method and titer. Negative conversion of IFANA often indicates unfavorable prognosis for severe patients.