Inactive Rheumatoid Arthritis Group Research Articles

TH SUBSETS AND SERUM CYTOKINES IN THE PATHOGENESIS OF RHEUMATOID ARTHRITIS

Rheumatoid arthritis (RA) is a systemic inflammatory disease. It is characterized by damage of cartilages and bones which results in destruction of joint function. Multi-organic disorders of lung, heart and kidney can be developed in time. Complex network of immune and stromal cells, cytokines and chemokines produced by these cells participate in onset of the disease, in the systemic active phase and during the transition to more localized inactive disease after the treatment. To evaluate the Th1, Th17, Tregs and CD4+CD39+ cells pattern and pro- and anti-inflammatory cytokines in peripheral blood of patients with RA, 47 RA patients and 20 healthy individuals were included in the study. RA patients were divided into active and inactive RA groups. Frequencies of circulating Th1, Th17, Tregs and CD4+CD39+ cells were analyzed by flow cytometry. Serum levels of cytokines: IL-6, IL-10, IL-4, IL-17, TNF-a and TGF-b1 were detected by ELISA. The results demonstrated an increase of Th1, Th17 cell frequencies in active RA patients, whereas Tregs remain unchanged in RA groups. CD39 marker expression shows significant decline in active RA patients in comparison with inactive RA group and controls. Concentrations of innate cytokines IL-6, TNF-a in the peripheral blood was significantly increased in active RA while according to IL-17/IL-21 serum concentration this group was divided into two subgroups. Anti-inflammatory cytokines TGF-b, IL-4 shows decrease in active RA group compared to healthy controls. Study demonstrated that development of RA is associated with changes of serum pro- and anti-inflammatory cytokines which influence balance of T helpers subsets and could provide therapeutic opportunities.

OSTEOPROTEGERIN (OPG) AND SOLUBLE RECEPTOR ACTIVATOR OF NUCLEAR FACTOR KAPPA B LIGAND (S-RANKL) IN PATIENTS WITH RHEUMATOID ARTHRITIS

Background: Rheumatoid arthritis (RA) is a chronic inflammatory disease that may result in debilitating joint deformities with destruction of bone and cartilage. Inflammation is still considered the pivotal inducer of both components of joint damage. A mechanism of bone destruction that could be dissociated from inflammation was proposed in which osteoclasts activation play a prominent role in bone resorption. RANKL and its natural decoy receptor, osteoprotegerin (OPG), play key roles in osteoclast activation. Objective: To evaluate osteoprotegrin (OPG) and soluble RANKL (s-RANKL) level in serum of rheumatoid arthritis patients, and to correlate their levels in serum to disease activity and radiological findings (bone loss). Patients and methods: Fifty five rheumatoid arthritis patients were included in the study. They were classified according to disease activity into 2 groups: The first group was active RA group, and the second was inactive RA group. In addition, RA group was classified according to bone erosions into 2 groups: The first group was RA group with bone erosions and the second group was RA group without bone erosions. The study included 25 healthy individuals of matched age and sex as a control group. All participants were exposed to complete clinical examination, radiological examination, and full laboratory evaluation including osteoclast activity markers (OPG/ RANKL ratio), inflammation markers (ESR, CRP and DAS-28 score), immunological markers (RF and anti-CCP), biochemical markers (calcium, phosphorus and alkaline phosphatase) and complete blood picture (CBC). Results: OPG/ RANKL ratio was significantly lower in RA group compared to control group. OPG/RANKL ratio was significantly lower in active RA group compared to inactive RA group. OPG/ RANKL ratio was significantly lower in RA group with bone erosion compared to RA group without bone erosions. OPG/ RANKL ratio showed significant negative correlation with both disease duration (in all RA groups) and inflammatory markers (in some not all RA groups). OPG/ RANKL ratio did not correlate with biochemical markers in all RA groups. Conclusion: OPG/ RANKL ratio could be used to monitor joint damage (bone erosions) progression in patients with RA. Progression of joint damage (bone erosions) due to osteoclast over activity could, at least in part, be dissociated from inflammation. Therefore, targeting OPG/ RANKL ratio may be effective in preventing bone damage in RA patients.

The importance of red cell distribution width and neutrophil-lymphocyte ratio as a new biomarker in rheumatoid arthritis

Objectives: Rheumatoid arthritis (RA) is a long-lasting autoimmune disorder that primarily affects the joints. Various biomarkers have been used for the prognosis and clinical follow-up. There are few studies that have investigated whether or not neutrophil-lymphocyte ratio (NLR) and red cell distribution width (RDW) are good indicators of systemic inflammation. The present study aims to explore the prognostic value of RDW and NLR in rheumatoid arthritis (RA) as a new inflammatory marker. Methods: RA patients (n = 124) who presented to the Rheumatology outpatient clinic in our hospital between March 2015 and May 2015 were included in this study retrospectively. As a first group, 47 clinically active RA patients who had high acute phase proteins were included. In the second group, 73 clinically in-remission RA patients who had normal acute phase proteins were included. Fifty-five healthy volunteers constituted the control group. Results: The mean RDW was found to be 15.2 ± 2.9 in the active group; 14.6 ± 2 in the inactive group and 13.4 ± 1.4 in the control group (p < 0.01). The mean NLR was found to be 3.7 ± 2.2 in the active group; 3.7 ± 1.6 in the inactive group and 3.2 ± 0.9 in the control group (p = 0.190). There were statistically significant differences between the RDW values of the active-period RA patients with the control group (p < 0.01). There was statistically significant difference between RDW values of active RA and inactive RA patients (p < 0.01). The NLR results between the RA group and the control group (p = 0.700); the active RA group, and the inactive RA group (p = 0.169) were similar. There was not statistically difference between the NLR values of active RA patients with the control group (p = 0.360). There was statistically difference between the NLR values of inactive RA patients with the control group (p = 0.047). Conclusion: RDW was found higher in all RA group than control, additionally was also higher in active RA group than remission group. NLR values of remission group was higher than control.

Vitamin D level in Rheumatoid arthritis and its correlation with the disease activity

Objective Vitamin D has an immunomodulatory and anti-inflammatory action, and its deficiency may cause several autoimmune disorders, including rheumatoid arthritis (RA). The relationship between vitamin D level and the severity of RA is of mere interest to several researchers. Patients and methods This prospective study included 40 cases of RA and 20 healthy controls, of age group between 40–70 years. Serum Vitamin D levels were measured and compared in RA patients and controls. Vitamin D levels in RA patients were measured in three different groups; active RA group, inactive RA group and control. Results Sixty-five percent patients of active RA were Vitamin D deficient versus only 40% of inactive RA patients. The serum Vitamin D levels were also significantly lower in the RA patients (mean value of 18.09 ± 8.99 ng/ml), as compared to the controls (mean value of 29.67 ± 11.34 ng/ml). There was an inverse significant correlation between serum Vitamin D levels and RA disease activity as measured by DAS28. The mean serum Vitamin D levels were 18.81 ± 6.679 ng/ml, 14.42 ± 6.856 ng/ml, 9.83 ± 6.791 ng/ml, in low disease activity, moderate disease activity, and high disease activity groups, respectively. Conclusion Vitamin D deficiency is more common in autoimmune diseases as RA and may be one of the leading cause of increased disease activity.

Association of MiRNA-146a, MiRNA-499, IRAK1 and PADI4 Polymorphisms with Rheumatoid Arthritis in Egyptian Population

Background/Aims: Rheumatoid arthritis (RA) is a systemic autoimmune disease affecting up to 1% of the population worldwide. The aim of the present study was to investigate whether miRNA-146a rs2910164, miRNA-499 rs3746444, IRAK1 rs3027898 and PADI4 rs1748033 polymorphisms are associated with susceptibility to RA in Egyptians and whether they influence disease severity and activity. Methods: The study was performed on 104 unrelated RA patients and 112 healthy subjects. RA patients were further subdivided into active and inactive RA groups. Polymorphisms were genotyped by using real-time polymerase chain reaction with TaqMan allelic discrimination assay. Results: Significant differences in the frequency of miRNA-146a rs2910164, miRNA-499 rs3746444, IRAK1 rs3027898 and PADI4 rs1748033 alleles and genotypes were observed between RA patients and controls. Only CA and AA genotypes of IRAK1 rs3027898 shows a significant difference between active and inactive subgroups. MiRNA-146a rs2910164 and IRAK1 rs3027898 polymorphisms were a risk factor for predisposition to RA in codominant and dominant tested inheritance models, while, the miRNA-499 rs3746444 and PADI4 rs1748033 polymorphisms were a risk factor in codominant and recessive one. CG and GG genotypes of miRNA-146a rs2910164 were associated with positive erosions. CA genotype of IRAK1 rs3027898 was associated with low disease activity and negative erosions, while, the AA genotype was associated with high disease activity. CC genotype of PADI4 rs1748033 was associated with negative rheumatoid factor. Conclusion: The 4 studied SNPs were likely to play an important role in the susceptibility to RA and can influence disease severity and activity in Egyptian population.

Upregulated Expression of microRNA-16 Correlates with Th17/Treg Cell Imbalance in Patients with Rheumatoid Arthritis

To explore the correlation between miR-16 expression in T cells of peripheral blood mononuclear cells (PBMCs) and Th17/Treg imbalance in rheumatoid arthritis (RA) patients. Forty RA patients were recruited as the case group and further grouped as active RA and inactive RA groups; 21 healthy individuals were selected as the control group. Th17 and Treg were measured by flow cytometry, and their related cytokines were measured by FlowCytomix. RORγt, FoxP3 mRNA, and miR-16 expression in T cells was determined by real-time quantitative polymerase chain reaction. Western blotting was performed to measure RORγt and FoxP3 protein expression. RA patients showed upregulated Th17 and RORγt mRNA and protein expression compared with the controls (all p < 0.05); active RA patients showed lower Treg and FoxP3 mRNA and protein expression compared with inactive RA patients and controls (all p < 0.05). Secretion levels of Th17-related cytokines were higher in active RA patients than in inactive RA patients and controls (all p < 0.05); whereas those of Treg-related cytokines were lower in active RA patients than in controls (all p < 0.05). Active RA patients showed increased miR-16 expression in Th17 cells and decreased miR-16 expression in Treg cells of PBMCs (both p < 0.05). Pearson's test showed that in the PBMCs of the RA patients, miR-16 expression in the Th17 cells was positively related with RORγt mRNA expression, and miR-16 expression in the Treg cells was positively related with FoxP3 mRNA expression (both p < 0.05). The expression of miR-16 in Th17 and Treg cells of PBMCs in RA patients was closely associated with the expression of RORγt and FoxP3. MiR-16 may be involved in Th17/Treg imbalance of RA patients by affecting the expression of RORγt and FoxP3.

Clinical Significance of Red Blood Cell Distribution Width and Inflammatory Factors for the Disease Activity in Rheumatoid Arthritis.

To evaluate the significance of red blood cell distribution width (RDW) and other factors for the disease activity in rheumatoid arthritis (RA). Each patient underwent clinical examination and blood sampling for assessment of serum high-sensitivity C-reactive protein (hs-CRP) levels, erythrocyte sedimentation rate (ESR), hemoglobin concentration (Hb), hematocrit (HCT), RDW, and other erythrocyte parameters (mean corpuscular volume [MCV], mean cell Hb [MCH], mean corpuscular Hb concentration [MCHC]). Rheumatoid factors (RF-IgA, -IgG, -IgM) and anti-cyclic citrullinated protein antibodies (anti-CCP) were purified from the plasma and detected by ELISA. RA patients were divided into two groups based on DAS28 scores: active RA group (DAS28 > 5.1) and inactive RA group (DAS28 2.6 - 3.2). Patient samples were within normal ranges for Hb (15.2 ± 1.3 g/L), HCT (29.9 ± 2.2%), and other red blood cell (RBC) parameters (MCV 80.3 ± 12.1 fL, MCH 26.6 ± 3.5 pg, MCHC 323 ± 25 g/L). The RDW was higher in the active RA group than in the inactive group (16.5 ± 3.2 vs. 13.9 ± 1.5%, p < 0.01). Similarly, the proportion of patients positive for RF and anti-CCP was higher in the active group than in the inactive group (RF 62 vs. 53%, CCP 83 vs. 61%). RF and anti-CCP were present at higher titers in patients with active RA. The bone erosion rate in the 110 RA patients was 67%. Patients with erosion had higher RDW than those without erosion (17.1 ± 2.2 vs. 13.9 ± 3.5%). High titers of anti-CCP and RF-IgM, -IgG, and -IgA were also associated with high bone erosion rates (67%, 77%, 67%, and 73%, respectively). Our results suggest an association between RDW and levels of inflammatory factors and autoantibodies in RA. This association may be linked to the underlying proinflammatory state and increased oxidative stress, both of which correlate with impaired erythrocyte maturation. RDW, RF, and anti-CCP are key players in the proinflammatory and proatherogenic status of RA, and they may represent useful markers to improve characterization of disease activity in RA patients, thereby helping the clinician to define more appropriate therapeutic strategies.

Chemokine receptors expression on peripheral CD4-lymphocytes in rheumatoid arthritis: Coexpression of CCR7 and CD95 is associated with disease activity

Rheumatoid arthritis (RA) is a chronic autoimmune disease characterized by synovial inflammation triggered by infiltrating CD4 lymphocytes. The positioning and activation of lymphocyte in inflamed synovial tissues are dependent on a number of factors including their chemokine receptor expression profile. We aimed to investigate which chemokine receptors pattern correlate with serum cytokine levels and with disease activity. Forty patients with RA (34 female and 6 male) with age range from 21 to 68years were included. Twenty healthy volunteers (16 female and 4 male) with matched age (range 21–48years) were served as healthy controls (HCs). Expression of chemokine receptors (CCR5, CX3CR1 and CCR7) together with the apoptosis-related marker (CD95) was analyzed using three-color flow cytometry analysis after gating on CD4+ peripheral blood lymphocytes. Plasma levels of IL-6, IL-10, IL-12 and TNF-α cytokines were measured in all participants using ELISA. Disease activity score (DAS28-CRP) system was assessed and active disease was defined as DAS28 ⩾3.2. Twenty-five (62.4%) patients were classified as active RA (ARA) and 15 (37.5%) patients with inactive RA (IRA). Percentages of CD4+ lymphocytes expressing CD95 with either of CCR7 or CCR5 were significantly higher in ARA compared to IRA and HCs groups, while the expression of CX3CR1 on T-cells was found significantly lower in both CD95− and CD95+ T-cells in RA groups than HC. Percentages of CD4+CD95+CCR7+ cells correlated positively with IL-6 (r=0.390). Whereas CD4+CD95+CX3CR1+ were negatively correlated with TNF-α (r=−0.261). Correlation of CD4+CD95+CCR7+ T cell subset with disease activity and inflammatory cytokines suggests a role for this cell subset in the pathogenesis of RA. Further investigation will be required to fully characterize this cell subset and its role in disease progression.

Elevated Levels of T Helper 17 Cells Are Associated with Disease Activity in Patients with Rheumatoid Arthritis

BackgroundInterleukin-17 (IL-17)-producing T helper (Th) 17 cells are considered as a new subset of cells critical to the development of rheumatoid arthritis (RA). We aimed to investigate the distribution of Th1 and Th17 cells and their association with disease activity, and determine the Th17-related cytokine levels in the peripheral blood of RA patients.MethodsPeripheral blood mononuclear cells from 55 RA and 20 osteoarthritis (OA) patients were stimulated with mitogen, and the distributions of CD4+Interferon (INF)+IL-17- (Th1 cells) and CD4+INF-IL-17+ (Th17 cells) were examined by flow cytometry. Serum levels of IL-6, IL-17, IL-21, IL-23, and tumor necrosis factor (TNF)-α were measured by ELISA. Erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) were recorded. The 28-joint disease activity score (DAS28) was also assessed.ResultsThe median percentage of Th17 cells was higher in RA patients than in OA patients (P=0.04), and in active than in inactive RA (P=0.03), whereas that of Th1 cells was similar in both groups. Similarly, the levels of IL-17, IL-21, and IL-23 were detected in a significantly higher proportion of RA patients than OA patients and the frequencies of detectable IL-6, IL-17, and IL-21 were higher in active RA than in inactive RA group. The percentage of Th17 cells positively correlated with the DAS28, ESR, and CRP levels.ConclusionsThese observations suggest that Th17 cells and Th17-related cytokines play an important role in RA pathogenesis and that the level of Th17 cells in peripheral blood is associated with disease activity in RA.

Circulating Hepcidin in the Crossroads of Anemia and Inflammation Associated with Rheumatoid Arthritis

To investigate the role of circulating hepcidin, which is a homeostatic regulator of iron metabolism and a mediator of inflammation, in anemia associated with rheumatoid arthritis. Forty patients with rheumatoid arthritis (19 with anemia and 21 without anemia), 12 patients with iron deficiency anemia and 14 healthy adults were studied. Serum hepcidin levels were analyzed with hepcidin prohormone solid phase enzyme-linked immunosorbent assay. The mean serum hepcidin levels were significantly higher in patients with rheumatoid arthritis with anemia compared to healthy adults and those with iron deficiency anemia. The active rheumatoid arthritis group had significantly higher mean serum hepcidin levels than the inactive rheumatoid arthritis group. In the rheumatoid arthritis group, serum hepcidin levels were positively correlated with disease activity, but inversely correlated with hemoglobin levels. The serum hepcidin level was found to be a significant predictor for hemoglobin level. Serum hepcidin levels are closely associated with disease activity in rheumatoid arthritis patients and might play a role in the pathobiology of chronic disease anemia associated with rheumatoid arthritis.

Increased platelet activation markers in rheumatoid arthritis: Are they related with subclinical atherosclerosis?

Atherosclerotic cardiovascular mortality is increased in rheumatoid arthritis (RA) patients. We evaluated the association of inflammatory response with platelet, endothelial, coagulation activation parameters; and subclinical atherosclerosis in RA patients. We included 27 RA patients (21 female; six male) and 19 healthy subjects (14 female; five male). Disease activity score (DAS28) in RA patients was calculated; and patients were divided into two groups as active and inactive. Flow cytometry was used to determine platelet CD62P expression, platelet microparticles (PMP), platelet-monocyte (PMC) and platelet-neutrophil complexes (PNC). Plasma E-selectin, thrombin-antithrombin (TAT) complex, and serum sCD40L levels were determined by ELISA. The intima-media thickness (IMT) of carotid arteries was determined by B-mode ultrasonography. In RA patients, platelet CD62P expression (p < 0.001), PMC (p = 0.037) and sCD40L (p < 0.001) levels were increased when compared to the control group. PNC (p = 0.07) and TAT levels (p = 0.1) were non-significantly higher, and PMP level (p = 0.075) was nonsignificantly lower in RA patients. Soluble E-selectin level was significantly higher in the active RA group than in the inactive RA group (p = 0.009). There was no correlation between carotid IMT and activity markers, the evaluated parameters (p > 0.05).The increase in markers of active platelets, CD62P and sCD40L, and PMC levels might be associated with the increased cardiovascular mortality in RA. Nevertheless, none of these parameters were associated with carotid IMT: this suggests that one cross-sectional value might not be a good marker for atherosclerosis.

Investigation of protein oxidation and lipid peroxidation in patients with rheumatoid arthritis

We aimed to determine the importance of neutrophil activation and the source of oxidative stress in the pathogenesis of rheumatoid arthritis (RA) by quantification of advanced oxidation protein products (AOPP) and total thiol levels as markers of oxidative protein damage, malondialdehyde (MDA) levels as a marker of lipid peroxidation and myeloperoxidase (MPO) activity as a marker of neutrophil activation in patients with RA. Fifty-seven rheumatoid arthritis patients were included in the study and sub-grouped according to disease activity (active, n = 31; inactive, n = 26) and compared with healthy controls (n = 25). Serum MPO activity, AOPP, MDA, and thiol levels were measured by an enzymic spectrophotometric method. Serum MPO activity (p < 0.001), AOPP (p < 0.001), MDA (p < 0.001) and levels of thiol (p < 0.002), were higher in the patient group than the controls. Active and inactive RA groups were compared with the control group and there were significant differences between each parameter. MPO activity, AOPP, MDA and thiol levels were significantly higher in both active and inactive RA patients than the controls. On the other hand, when a comparison was made between active and the inactive stage, a statistically significant difference was present only in MDA (p < 0.05) and AOPP levels (p < 0.05). There was also a significant positive correlation between all parameters. These data strongly suggest that neutrophils, which constitute the most important source of chlorinated oxidants due to their high MPO content, may be involved in serum AOPP formation and therefore the production of a novel class of pro-inflammatory mediators of oxidative stress in RA patients and that protein oxidation could play an important role in the pathogenesis of RA as does lipid peroxidation.

Assessment of paraoxonase 1 activity and malondialdehyde levels in patients with rheumatoid arthritis

Objectives: We aimed to evaluate antioxidant paraoxonase 1 activity together with malondialdehyde (MDA) (an oxidative stress parameter) levels in patients with rheumatoid arthritis. Design and methods: Fifty-seven rheumatoid arthritis patients were included in the study and subgrouped according to disease activity (active, n = 31; inactive, n = 26) and compared with healthy controls ( n = 25). Serum paraoxonase 1 activity and MDA levels were measured according to an enzymatic spectrophotometric method. Results: Serum MDA level was higher ( P = 0.001) whereas paraoxonase 1 activity was lower ( P = 0.001) in the patient group than the controls. When active and inactive subgroups were compared with the control group, there was a statistically significant difference between each parameter. Serum MDA levels were significantly higher, while paraoxonase 1 activity was lower in the active and inactive rheumatoid arthritis groups than the control group. But there was not any difference between active and inactive patients with RA. There was a negative correlation between MDA levels and paraoxonase 1 activity. Conclusions: Increased reactive oxygen species levels in rheumatoid arthritis may result in a pro-oxidation environment, which in turn could result in decreased antioxidant paraoxonase 1 activity and increased MDA levels.