The development of anticyclic citrullinated peptide (anti-CCP) antibody following severe COVID-19.

OBJECTIVES: The dysregulated immune response is one of the cardinal features of severe coronavirus disease 2019 (COVID-19). This study was conducted to clarify the occurrence of autoantibodies (AABs) associated with systemic autoimmune rheumatic diseases (SARDs) in hospitalized patients with a moderate, severe, and critical form of COVID-19. METHODS: The serum samples obtained from 176 hospitalized COVID-19 patients were investigated in this study, including patients with moderate (N = 90), severe (N = 50), and critical (N = 36) forms of COVID-19. Also, the serum samples collected from healthy subjects before the COVID-19 pandemic were used as controls (N = 176). The antinuclear antibodies (ANAs), antidouble-stranded DNA (anti-dsDNA), cytoplasmic-anti neutrophil cytoplasmic antibody (c-ANCA), perinuclear ANCA (p-ANCA), antiphospholipid antibodies (aPLs), and anticyclic citrullinated peptide (anti-CCP) occurrence was evaluated using a solid-phase enzyme-linked immunosorbent assay (ELISA). RESULTS: The results showed that the occurrence of ANAs, anti-dsDNA, anti-CCP, c-ANCA, and p-ANCA was significantly higher in the COVID-19 patients compared to serum obtained from healthy subjects (p < .0001, p < .0001, p < .0001, p < .05, and p < .001, respectively). The positive number of anti-CCP tests increased significantly in severe COVID-19 compared to the moderate group (p < .01). CONCLUSION: Our study further supports the development of autoantibodies related to systemic autoimmune rheumatologic diseases. To the best of our knowledge, this is the first study with a large sample size that reported the occurrence of anti-CCP in a severe form of COVID-19.

Overexpression of Synoviolin and miR-125a-5p, miR-19b-3p in peripheral blood of rheumatoid arthritis patients after treatment with conventional DMARDs and methylprednisolone.

PURPOSE: SYVN1 is an endoplasmic reticulum (ER)-resident E3 ubiquitin ligase that has an essential function along with SEL1L in rheumatoid arthritis (RA) pathogenesis. This study aimed to investigate the changes in the expression of peripheral blood ncRNAs and SYVN1-SEL1L affected by DMARDs treatment. METHODS: Twenty-five newly diagnosed RA patients were randomly assigned to receive conventional DMARDs (csDMARDs) and methylprednisolone for six months. The peripheral blood gene expression of SYVN1 and SEL1L and possible regulatory axes, NEAT1, miR-125a-5p, and miR-19b-3p, were evaluated before and after qRT-PCR. We also compared differences between the patients and healthy controls (HCs), and statistical analyses were performed to determine the correlation between ncRNAs with SYVN1-SEL1L and the clinical parameters of RA. RESULTS: Expression of NEAT1 (P = 0.0001), miR-19b-3p (P = 0.007), miR-125a-5p (P = 0.005), and SYVN1 (P = 0.036) was significantly increased in newly diagnosed patients compared to HCs; also, miR-125a-5p, miR-19b-3p, and SYVN1 were significantly overexpressed after treatment (P = 0.001, P = 0.001, and P = 0.005, respectively). NEAT1 was positively correlated with SYVN1, and miR-125a-5p had a negative correlation with anti-cyclic citrullinated peptides. The ROC curve analysis showed the potential role of selected ncRNAs in RA pathogenesis. CONCLUSION: The results indicate the ineffectiveness of the csDMARDs in reducing SYVN1 expression. The difference in expression of ncRNAs might be useful markers for monitoring disease activity and determining therapeutic responses in RA patients. Key Points • The expression of NEAT1 is significantly upregulated in RA patients compared to HC subjects. • miR-19b-3p, miR-125a-5p, and SYVN1 are significantly upregulated in RA patients compared to HC subjects. • The expression of miR-19b-3p and miR-125a-5p is significantly increased in RA patients after treatment with DMARDs and methylprednisolone. • NEAT1 is positively correlated with SYVN1.

CXCL9 and NT-proBNP, a notable link between inflammatory mediator and cardiovascular disease biomarker in rheumatoid arthritis.

INTRODUCTION: Cardiovascular disease (CVD) is the most critical extra-articular manifestation of rheumatoid arthritis, and inflammatory molecules contribute to its pathogenesis. Recently, CXCL9 has been considered an inflammatory chemokine associated with the pathogenesis of CVD. Here, we evaluated the association of plasma CXCL9 with well-established cardiac biomarkers, including HS-CRP (High sensitivity C-reactive protein) and NT-ProBNP (N-terminal pro-B-type natriuretic peptide), in newly diagnosed and under-treatment RA patients. METHODS: Thirty newly diagnosed patients, 30 under-treatment RA patients, and 30 healthy subjects were recruited. The plasma concentration of CXCL9 and NT-ProBNP was measured using the ELISA method. The HS-CRP levels was measured in plasma samples using latex-enhanced immunoturbidimetric test. RESULTS: We found increased plasma levels of CXCL9, HS-CRP, and NT-proBNP in RA patients compared to healthy subjects, besides that the concentration of CXCL9, HS-CRP, and NT-ProBNP showed elevated levels in newly diagnosed RA patients compared to under-treatment group. The mean plasma concentration of CXCL9, NT-proBNP, and HS-CRP were statistically different among healthy subjects, newly diagnosed, and under-treatment RA patients (p < 0.001, p = 0.016, and p < 0.001, respectively). We also found a significant positive correlation between CXCL9 and DAS-28 (p = 0.0005, r = 0.436) in the patients' group (new-case + under-treatment). There was a significantly positive correlation between CXCL9 and NT-proBNP in newly diagnosed and under-treatment patients (p = 0.020, r = 0.424; p < 0.0001, r = 0.853, respectively). In the patient's group (new-case + under-treatment), there was a significantly positive correlation between CXCL9 with NT-proBNP (p < 0.001, r = 0.703) and CXCL9 with HS-CRP (p = 0.015, r = 0.313). CONCLUSION: CXCL9 correlates significantly with well-established cardiovascular biomarkers, including HS-CRP and NT-ProBNP in RA patients. Key Points • CXCL9 is an inflammatory marker in RA. • CXCL9 has correlated with DAS-28. • There is a strong correlation between CXCL9 with NT-proBNP and HS-CRP.

Increased gene expression of CCR6 and RORγt in peripheral blood cells of rheumatoid arthritis patients and their correlation with anti-cyclic citrullinated peptide and disease activity.

OBJECTIVES: The significance of T helper 17 (Th17) cells in the pathogenesis of rheumatoid arthritis (RA) has recently been demonstrated in many studies. Retinoic acid receptor-related orphan receptor γt (RORγt) is a transcription factor that is specifically involved in the generation of Th17 cells. Besides, the chemokine receptor CCR6, the receptor for CCL20, is characteristically expressed by these cells. Considering the pivotal roles of Th17 cells in RA pathogenesis, in this study, we assessed the gene expression of CCR6 and RORγt in the peripheral blood leukocytes of new case RA patients. Also, we evaluated their association with anticyclic citrullinated peptide (anti-CCP) antibodies and disease activity. METHODS: Forty-five new case RA patients and 45 healthy persons have been recruited in this investigation. The gene expression of CCR6 and RORγt was evaluated by quantitative real-time PCR (qRT-PCR), and anti-CCP antibodies plasma levels were measured using the enzyme-linked immunosorbent assay (ELISA) technique. Disease activity was measured according to the disease activity score-28 (DAS-28) formula. RESULTS: The gene expression of CCR6 and RORγt increased remarkably in new case RA patients compared to healthy controls (p < .05 and p < .01, respectively). Moreover, there was a positive correlation between RORγt gene expression and parameters, including gene expression of CCR6 (p = .001, r = .461), plasma levels of CCL20 (p = .0009, r = .477), ESR (p = .004, r = .419), DAS-28 (p = .006, r = .402), anti-CCP (p = .019, r = .346), and RF (p = .001, r = .451). Also, CCR6 gene expression was positively associated with the DAS-28 (p = .037, r = .310), plasma levels of anti-CCP (p = .037, r = .312), and ESR (p = .029, r = .327). CONCLUSION: Increased gene expression of CCR6 and RORγt in peripheral blood leukocytes of new case RA patients may contribute to the exacerbation and pathogenesis of RA.

Assessing the gene expression of the adenosine 5'-monophosphate-activated protein kinase (AMPK) and its relation with the IL-6 and IL-10 plasma levels in COVID-19 patients.

BACKGROUND: Metabolic dysregulation and excessive inflammation are implicated in the pathogenesis of the highly infectious disease of coronavirus disease 2019 (COVID-19), which is caused by a newly emerging coronavirus (i.e., severe acute respiratory syndrome-coronavirus 2; SARS-CoV-2). The adenosine 5'-monophosphate-activated protein kinase (AMPK), an energy sensor regulating the metabolic pathways in diverse cells, exerts a regulatory role in the immune system. This study aims to examine the mRNA expression level of AMPK and the plasma levels of interleukin-6 (IL-6) and IL-10 cytokines in patients with different grades of COVID-19. METHODS: Peripheral blood was collected from 60 patients with COVID-19 (Moderate, severe, and critical). The plasma levels of IL-6 and IL-10 were quantified by enzyme-linked immunosorbent assay (ELISA), and the mRNA expression level of AMPK was determined using real-time PCR. RESULTS: The results showed that the plasma levels of IL-6 increased significantly in critical and severe patients compared to moderate cases of COVID-19 (P < 0.001). Moreover, IL-10 plasma concentrations were significantly higher in critical and severe cases than in moderate cases of COVID-19 (P < 0.01 and P < 0.05, respectively). Also, the gene expression of AMPK was meaningfully enhanced in critical patients relative to moderate and severe cases of COVID-19, in order (P < 0.001 and P < 0.01, respectively). There was a positive association between AMPK gene expression and plasma levels of IL-6 and IL-10 (P = 0.006, r = 0.348, P = 0.028, r = 0.283, respectively). CONCLUSION: Increasing AMPK gene expression is likely a necessary effort of the immune system to inhibit inflammation in critical COVID-19. However, this effort seems to be inadequate, probably due to factors that induce inflammation, like erythrocyte sedimentation rate (ESR) and IL-6.

CXCL9 and its Receptor CXCR3, an Important Link Between Inflammation and Cardiovascular Risks in RA Patients.

Cardiovascular disease (CVD) is the most common cause of mortality in rheumatoid arthritis (RA), and Inflammation has a decisive role in its pathogenesis. CXCL9 contributes to multi aspects of inflammatory reactions associated with the pathogenesis of CVD. In the current study, we evaluated the association of plasma CXCL9 and CXCR3 gene expression with Cardiovascular risk factors in RA patients for the first time. Thirty newly diagnosed, 30 on-treatment RA patients, and 30 healthy subjects were recruited in this study. The plasma concentration of CXCL9 and CXCR3 gene expression were measured using ELISA and Real-Time PCR, respectively. The CVD risk was evaluated using Framingham Risk Score (FRS) and Systematic Coronary Risk Evaluation (SCORE). The plasma levels of CXCL9 were significantly higher in the newly diagnosed and on-treatment RA patients compared to the control group (P < 0.0001 and P < 0.001, respectively). Also, The CXCR3 gene expression was strongly elevated in newly diagnosed and on-treatment patients (P < 0.001 and P < 0.01, respectively). The CXCL9 and CXCR3 were significantly associated with RA disease activity (P = 0.0005, r = 0.436; P = 0.0002, r = 0.463, respectively). The FRS was remarkably higher in newly diagnosed and on-treatment patients (P = 0.014 and P = 0.035, respectively). The CXCR3 gene expression significantly correlated with age, systolic blood pressure, FRS, and SCORE (P = 0.020, r = 0.298; P = 0.006, r = 0.346; P = 0.006, r = 0.349; P = 0.007, r = 0.341, respectively). The CXCL9 plasma concentration had a significant negative correlation with plasma HDL and LDL levels (P = 0.033, r = -0.275; P = 0.021, r = -0.296, respectively). CXCL9 and CXCR3 correlates with different variables of CVD in RA.

Investigating the correlation of the NF-κB and FoxP3 gene expression with the plasma levels of pro- and anti-inflammatory cytokines in rheumatoid arthritis patients.

INTRODUCTION: Rheumatoid arthritis (RA) is a chronic inflammatory systemic autoimmune disease. Cytokines regulate a wide range of inflammatory processes involved in RA pathogenesis. Anti-inflammatory cytokines (i.e., TGF-ß and lL-10) and pro-inflammatory cytokines, like IL-6, were found to be potentially implicated in RA pathogenesis. Besides, NF-κB and FoxP3 are critical transcription factors regulating the inflammatory events occurring in RA patients. This study intends to assess the plasma levels of IL-6, IL-10, and TGF-ß1 cytokines, as well as the expression of NF-κB and FoxP3 genes in RA patients, compared to the healthy controls. METHODS: Peripheral blood was collected from 50 RA patients (25 new case and 25 under-treatment) and 25 age- and gender-matched healthy subjects. The disease activity was determined using the DAS-28 and ESR criteria. Also, plasma levels of TGF-ß1, lL-10, and IL-6 were measured by enzyme-linked immunosorbent assay (ELISA) technique, and the gene expression of NF-κB and FoxP3 was evaluated using the real-time PCR method. RESULTS: Our results showed a significant up-regulation of Rel-A and NF-κB1, and also a down-regulation of FoxP3 gene expression in under-treatment RA patients compared to the controls (P=0.031, P=0.014, and P=0.011, respectively). Moreover, there was a significant reduction of Rel-A and FoxP3 in the under-treatment RA patients compared to new case RA patients (P=0.005 and P=0.015, respectively). Also, plasma levels of TGF-ß1 were significantly increased in both the new case and under-treatment RA patients relative to controls (P<0.001). CONCLUSION: In conclusion, classical NF-κB (P65/P50) and FoxP3 may have significant pro- and anti-inflammatory roles in RA pathogenesis, respectively. Key Point • NF-κB (P65/P50) has a contribution to the early phase of RA.

Partners in crime: Autoantibodies complicit in COVID-19 pathogenesis.

Autoantibodies (AABs) play a critical role in the pathogenesis of autoimmune diseases (AIDs) and serve as a diagnostic and prognostic tool in assessing these complex disorders. Viral infections have long been recognized as a principal environmental factor affecting the production of AABs and the development of autoimmunity. COVID-19 has primarily been considered a hyperinflammatory syndrome triggered by a cytokine storm. In the following, the role of maladaptive B cell response and AABs became more apparent in COVID-19 pathogenesis. The current review will primarily focus on the role of extrafollicular B cell response, Toll-like receptor-7 (TLR-7) activation, and neutrophil extracellular traps (NETs) formation in the development of AABs following SARS-CoV-2 infection. In the following, this review will clarify how these AABs dysregulate immune response to SARS-CoV-2 by disrupting cytokine function and triggering neutrophil hyper-reactivity. Finally, the pathologic effects of these AABs will be further described in COVID-19 associate clinical manifestations, including venous and arterial thrombosis, a multisystem inflammatory syndrome in children (MIS-C), acute respiratory distress syndrome (ARDS), and recently described post-acute sequelae of COVID-19 (PASC) or long-COVID.

Efficacy of DMARDs and methylprednisolone treatment on the gene expression levels of HSPA5, MMD, and non-coding RNAs MALAT1, H19, miR-199a-5p, and miR-1-3p, in patients with rheumatoid arthritis.

BACKGROUND: Rheumatoid arthritis (RA) is a systemic autoimmune disease with chronic inflammation characterized by joint damage and even extra-articular involvement. In this study, the gene expression levels of MALAT1, H19 and their possible downstream microRNAs, miR-199a-5p, miR-1-3p, and the predicted targets of these miRNAs, HSPA5 and MMD, were examined. METHODS: Twenty-five newly diagnosed RA patients and 25 healthy individuals were included. For six months, patients were treated with conventional disease-modifying antirheumatic drugs (DMARDs) and Methylprednisolone (mPRED). Blood samples were obtained from healthy controls and patients (before and after treatment). After RNA extraction, the RT-qPCR technique was used to evaluate the expression level of the studied genes. RESULTS: Data showed that the expression level of MALAT1, H19, miR-199a-5p, and miR-1-3p was significantly higher in the newly diagnosed patients with RA than the healthy subjects, but the increase in the expression level of HSPA5 and MMD genes in the new cases was not significant compared to healthy controls. After treatment, except for the expression level of lncRNAs, the expression level of miRNAs, HSPA5, and MMD significantly increased. Based on ROC curve analysis of MALAT1, H19, miR-199a-5p and miR-1-3p have a high ability to identify patients from healthy individuals (AUC = 0.986, AUC = 0.995, AUC = 0.855, AUC = 0.675, respectively). CONCLUSION: MALAT1 and H19 may be candidates as potential biomarkers for the discrimination between RA patients and controls. DMARDs plus mPRED therapy do not have a desirable effect on reducing inflammatory responses and ER stress.

Evaluation serum levels of Insulin Growth Factor-1 (IGF-1) and its association with clinical parameters in severe COVID-19.

BACKGROUND: Severe coronavirus disease-2019 (COVID-19) is associated with dysregulated immune response and extreme inflammatory injury. Considering the role of insulin growth factor-1 (IGF-1) in immune-mediated and inflammatory reactions, this study was conducted to investigate the IGF-1 contribution to the pathogenesis of severe form of COVID-19. MATERIAL AND METHODS: Sixty-two patients with severe COVID-19 and 52 healthy subjects were enrolled in this study. The serum levels of IGF-1 were measured using a solid-phase enzyme-linked chemiluminescent immunoassay on an Immulite 2000 system (Siemens Healthcare Diagnostics. RESULT: The serum levels of IGF-1 had no significant difference in COVID-19 patients compared to the healthy subjects (p = 0.359). There was a positive correlation between IGF-1 and age in the severe COVID-19 patients, while a negative correlation was observed for the serum levels of IGF-1 and age in the control group (r = 0.364, p = 0.036, r = - 0.536, p = 0.001, respectively). Moreover, IGF-1 was remarkably associated with hypertension, neurogenic disease, shock, and nausea in patients with the severe form of COVID-19 (p = 0.031, p = 0.044, p = 0.01, p = 0.03, respectively). CONCLUSION: Our results pointed to the complex role of IGF-1 in the severe form of COVID-19, and its association with clinical parameters, and some risk factors in the severe form of COVID-19.

Increased plasma levels of CCL20 in peripheral blood of rheumatoid arthritis patients and its association with clinical and laboratory parameters.

INTRODUCTION: Rheumatoid arthritis (RA) is a systemic autoimmune disease that primarily affects small joints. The impaired chemokine and cytokine responses are essential pathological mechanisms for the RA clinical presentation. Given the role of chemokines and inflammatory reactions in RA pathogenesis, we evaluate the association between the plasma concentration of CCL20 with the clinical and laboratory parameters in newly diagnosed RA patients. MATERIAL AND METHODS: Forty-five newly diagnosed RA patients and forty-five healthy subjects were enrolled in this study. The plasma levels of CCL20, rheumatoid factor, and anti-citrullinated peptide antibodies were measured using the enzyme-linked immunosorbent assay (ELISA) technique. RESULT: The plasma levels of CCL20 were increased significantly in RA patients compared to the healthy controls (p < 0.0001). There was a positive correlation between CCL20 and RF, anti-CCP, ESR, and DAS-28 (p < 0.0001, r = 0.669; p < 0.015, r = 0.358; p < 0.0001, r = 0.586; p < 0.0001, r = 0.769). CONCLUSION: The increased plasma levels of CCL20 in newly diagnosed RA patients may contribute to RA pathogenesis, and it is in association with clinical and laboratory parameters. Key Points • CCL20 has a contribution to the early phase of RA.

A review on the immune responses against novel emerging coronavirus (SARS-CoV-2).

Coronavirus disease 2019 (COVID-19) is a highly contagious disease caused by a newly identified coronavirus called the severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) which was initially emerged in Wuhan, China in late December 2019 and then rapidly extended to other countries worldwide. COVID-19 is now known as a pandemic threat to global public health. It possesses a wide spectrum of clinical manifestations, ranging from asymptomatic infection to mild, moderate, and ultimately severe pneumonia accompanied by multi-organ system dysfunction that can cause the death of the afflicted patients. The host immune system plays a critical role in defending against potentially pathogenic microorganisms such as coronaviruses, and it eliminates and eradicates these invading agents by triggering effective immune responses. However, there exists evidence indicating that in critically ill cases of the COVID-19, dysregulated immune responses and hyper-inflammation lead to acute respiratory distress syndrome (ARDS) and multi-organ failure. Achieving a profound understanding of the pathological immune responses involved in the pathogenesis of COVID-19 will boost our comprehending of disease pathogenesis and its progression toward severe form, contributing to the identification and rational design of effective therapies. In this review, we have tried to summarize the current knowledge regarding the role of immune responses against SARS-CoV-2 and also give a glimpse of the immune evasion strategies of this virus.

The Association between the Plasma Sugar and Lipid Profile with the Gene Expression of the Regulatory Protein of mTOR (Raptor) in Patients with Rheumatoid Arthritis.

BACKGROUND: Rheumatoid arthritis (RA) is an autoinflammatory and self-perpetuating disease with both articular and extra-articular manifestations, such as cardiovascular complications, which are the leading cause of mortality and morbidity in RA patients. Impaired sugar and lipid metabolism are considered as the critical risk factors for cardiovascular disease (CVD). Regarding the regulatory function of Raptor in the immunometabolism, in this study, we evaluated the association between plasma sugar and lipid profiles with the gene expression of Raptor and the cytokine tumor necrosis factor-α (TNF-α), as an inflammatory mediator, in peripheral blood leukocyte of RA patients. MATERIAL AND METHODS: Thirty-five RA patients who received combinational disease modified anti-rheumatoid drugs (DMARD) regimen and thirty healthy subjects enrolled in this study. The gene expression of Raptor was assessed by the real-time PCR method, and the Plasma levels of glucose and lipids, as well as TNF-α, were obtained using Hitachi device and enzyme-linked immunosorbent assay (ELISA) technique, respectively. RESULTS: The gene expression of Raptor was reduced significantly in RA patients compared to the healthy subjects (p = .001). The plasma level of HDL was significantly higher in RA patients than the control group (p = .001), while the plasma level of LDL was reduced significantly in these patients (p = .001). CONCLUSION: In our study, the reduced gene expression of Raptor may contribute to the impaired immunometabolism in RA patients, which is independent of plasma sugar and lipid profile.

The impaired gene expression of adenosine monophosphate-activated kinase (AMPK), a key metabolic enzyme in leukocytes of newly diagnosed rheumatoid arthritis patients.

The disturbed immune homeostasis is involved in the pathogenesis of an array of autoimmune diseases like rheumatoid arthritis (RA). The adenosine monophosphate-activated protein kinase (AMPK) with a pivotal role in immunometabolism process, also plays a regulatory function in the immune system. This study aims to evaluate the alteration of AMPK gene expression in peripheral blood leukocytes of RA patients and its effects on disease severity as well as plasma levels of anti-inflammatory cytokines. 60 RA patients, including 30 newly diagnosed and 30 patients whose disease were under controlled with the combinational disease-modifying anti-rheumatic drug (DMARD), as well as 30 healthy subjects, were enrolled in our study. The gene expression of AMPK was evaluated using real-time PCR method. The plasma concentrations of IL-10 and TGF-ß1 were measured using sandwich ELISA. The gene expression of AMPK was significantly lower in the newly diagnosed RA patients in comparison with the control group (P = 0.049). Inversely, in RA patients who received DMARD therapy, the gene expression of AMPK was significantly higher than the control group (P = 0.003). There was no significant correlation between AMPK gene expression and plasma levels of IL-10 and TGF-ß1. The plasma levels of TGF-ß1 was significantly higher in both newly diagnosed and under-treatment patients compared with healthy subjects (P < 0.001). The impaired gene expression of AMPK in peripheral blood leukocytes and elevated levels of plasma TGF-ß1 can be contributed in RA pathogenesis.