Jerzy Świerkot

Rola subpopulacji limfocytów pomocniczych Th1, Th17 i Treg w patogenezie reumatoidalnego zapalenia stawów z uwzględnieniem przeciwzapalnego działania cytokin Th1

Dysregulation in the immune system plays an important role in the pathogenesis of rheumatoid arthritis (RA). The persistent nature of arthritis strengthens the suggestion of immune dysfunction, consisting in predominance of the pro-inflammatory response. It seems that both local and systemic immune abnormalities, including PBMC secreting abnormal levels of pro- and anti-inflammatory cytokines, may be involved in the evolution of the disease. Helper T cells (Th) differentiate towards Th1, Th2, Th17, and Treg cells according to the cytokine microenvironment. Active RA results from the imbalance in distribution of functional pro-inflammatory Th17 and anti-inflammatory Treg cells. Affected Th1 cytokine secretion observed in the course of RA contributes to the increase in IL-17 production and Th17 infiltration in the synovial tissue. Current studies have demonstrated that pro-inflammatory Th1 cytokines may also exert an anti-inflammatory action on the balance between Th17 and Treg cells by promotion of Treg differentiation. In the paper, we also show the influence of TNF-alpha and its inhibitors on the distribution of Th subpopulations in RA patients.

The Activity of JAK/STAT and NF-κB in Patients with Rheumatoid Arthritis.

BACKGROUND Research is still being conducted in order to determine the mechanisms responsible for the initiation of rheumatoid arthritis (RA) as well as for its persistence and progression. OBJECTIVES The aim of this work was to establish the expression of the signal transducer and activator of transcription (STAT) transcription factors and the nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB) transcription factor in peripheral blood leukocytes and synovial fluid cells. The correlations between the activation level of the transcription factors and the activity of the disease were also analyzed. MATERIAL AND METHODS In total, the study included 34 RA patients and 19 healthy individuals as controls. The expression of NFκB, STAT1, STAT3, STAT4, STAT5 and STAT6 in peripheral blood leukocytes and synovial fluid cells was established. The immunocytochemistry method was used to determine the degree of activation of STAT and NF-κB transcription factors. For the location of the factors, primary polyclonal anti-STATs and monoclonal anti-NF-κB antibodies were used. RESULTS The expression of STAT1, STAT3, STAT4, STAT5, STAT6 and NFκB was significantly higher in the group of RA patients than in the controls. No statistically significant differences were found between the expression of STATs in peripheral blood leukocytes and synovial fluid cells. CONCLUSIONS In comparison with the control group, the expression of the STAT and NFκB transcription factors in RA patients was higher, which may be helpful in better understanding the etiopathogenesis of the disease in the future, and may potentially have important therapeutic implications.

Polymorphisms within Genes Involved in Regulation of the NF-κB Pathway in Patients with Rheumatoid Arthritis

Genes involved in regulation of the nuclear factor-κB (NF-κB)—pathway are suggested to play a role in pathogenesis of rheumatoid arthritis (RA). In the present study, genetic polymorphisms of TLR2, TLR4, TLR9 and NF-κB1 genes were investigated to assess their associations with RA susceptibility, progression and response to anti-TNF-α therapy. A group of 110 RA patients and 126 healthy individuals were genotyped for TLR2 (rs111200466), TLR4 (rs4986790, rs4986791), TLR9 (rs5743836, rs187084) and NF-κB1 (rs28362491) alleles. The presence of the TLR9 −1486 T variant (p < 0.0001) and its homozygosity (p < 0.0001) were found to be associated with disease susceptibility. The TLR9 −1237 C allele was associated with predisposition to RA in females only (p = 0.005). Moreover, the TLR4 rs4986791 G (rs4986790 T) alleles were more frequently detected among patients with the stage IV disease (p = 0.045), and were associated with more effective response to anti-TNF-α therapy (p = 0.012). More efficient response to anti-TNF-α treatment was also observed in patients with del within the NF-κB1 gene (p = 0.047), while for the TLR9 −1486 T homozygotes, the treatment was ineffective (p = 0.018). TLR polymorphisms affect disease susceptibility and response to therapy with TNF-α inhibitors in RA patients of Caucasian origin.

Polymorphisms within the human leucocyte antigen‐E gene and their associations with susceptibility to rheumatoid arthritis as well as clinical outcome of anti‐tumour necrosis factor therapy

Involvement of the non‐classical human leucocyte antigen‐E (HLA‐E) in both innate and acquired immune response suggests its possible role in development of autoimmune pathologies. This study was undertaken to investigate relationships between the HLA‐E gene single nucleotide polymorphisms (SNPs) and a risk of rheumatoid arthritis (RA), as well as to evaluate a potential of these polymorphisms to modulate clinical outcome of anti‐tumour necrosis factor (TNF) treatment in female patients. A total of 223 female patients with RA receiving anti‐TNF biological therapy and 134 female healthy subjects were enrolled into the study. Genotypings for two SNPs within the HLA‐E gene (rs1264457 HLA‐E*01:01/01:03; rs1059510 HLA‐E*01:03:01/01:03:02) were performed using a polymerase chain reaction (PCR) amplification employing LightSNiP assays. Clinical response was evaluated according to the European League Against Rheumatism (EULAR) criteria at 12 and 24 weeks after initiation of the therapy. The frequency of the HLA‐E*01:01/01:01 genotype was decreased significantly in RA patients in comparison to controls (P = 0·031). The presence of the HLA‐E*01:01/01:01 genotype in patients correlated with better EULAR response after 12 weeks of anti‐TNF treatment, while 01:03 allele carriers were generally unresponsive to the treatment (P = 0·014). The HLA‐E*01:03/01:03 genotype was also over‐represented among non‐responding patients in comparison to HLA‐E*01:01/01:01 homozygotes (P = 0·021). With respect to the HLA‐E rs1059510 variation, a better response after 12 weeks was observed more frequently in patients carrying the HLA‐E*01:03:01/01:03:01 genotype than other genotypes (P = 0·009). The results derived from this study imply that HLA‐E polymorphisms may influence RA susceptibility and affect clinical outcome of anti‐TNF therapy in female RA patients.

Significance of association of HLA-C and HLA-E with psoriatic arthritis

Psoriatic arthritis (PsA) is a complex genetic disorder that results from an interplay between multiple genetic and environmental factors. The aim of the study was to assess the significance of the association between the HLA-C and HLA-E allelic groups and PsA. Our results confirm the association between HLA-C(∗)06 and PsA (OR=5.16, p<0.0001). Furthermore, HLA-C(∗)06-positive patients develop more severe disease (p<0.01) and more frequently present with polyarticular pattern of PsA (p=0.08). Additionally our study revealed that the HLA-C(∗)02 allele was more frequently observed in PsA patients (OR=5.40, p<0.0005) and also that the HLA-E(∗)01:01 allele was significantly over-represented among HLA-C(∗)02-negative patients in comparison to healthy individuals (OR=6.44, p=0.045). Therefore these results suggest that the HLA-E and HLA-C(∗)02 molecules may also play an important role in determination immune response contributing to the PsA development.

Influence of NKG2D Genetic Variants on Response to Anti-TNF Agents in Patients with Rheumatoid Arthritis

A natural killer group 2 member D (NKG2D) acts as a powerful activating and co-stimulatory receptor on immune effector cells including NK and T cells. Disruptions within the NKG2D signalling pathway may trigger an exacerbated immune response and promote autoimmune reactions. The objective of the study was to evaluate a plausible role of polymorphisms within the NKG2D gene as a predictor of how effective anti-tumor necrosis factor (TNF) therapy is in rheumatoid arthritis (RA) patients. A total of 280 RA patients receiving anti-TNF therapy were genotyped for NKG2D rs2255336 (A > G), rs1049174 (C > G), and rs1154831 (C > A). Clinical response was evaluated according to the European League against Rheumatism (EULAR) criteria at the 12th and 24th week. Both the NKG2D rs225336 and rs1049174 polymorphisms were significantly associated with efficacy of TNF inhibitors. Inefficient therapy was more frequently observed in patients with rs2255336 GG or rs1049174 CC genotype as compared to other genotypes (p-value = 0.003 and p-value = 0.004, respectively). The presence of the rs2255336 G or the rs1049174 C allele correlated with a worse EULAR response (p-value = 0.002, p-value = 0.031, respectively). Moreover, patients carrying the rs2255336 or rs1049174 heterozygous genotype achieved better EULAR responses than patients with homozygous genotypes (p-value = 0.010 and p-value = 0.002, respectively). Data from the present study provides evidence that NKG2D polymorphisms may affect response to anti-TNF inhibitors in RA patients.

Influence of CD94 and NKG2A variants on susceptibility to rheumatoid arthritis and efficacy of anti-TNF treatment

OBJECTIVES The present study aimed to investigate relationships between the CD94 and NKG2A gene polymorphisms and the risk of rheumatoid arthritis (RA) development as well as their association with the response to anti-TNF therapy. METHODS A total of 284 patients with RA receiving anti-TNF therapy and 124 healthy subjects were enrolled to the study. Genotypings for CD94 (rs2302489) and NKG2A (rs7301582, rs2734440, rs2734414) polymorphisms were performed using a polymerase chain reaction (PCR) amplification employing LightSNiP assays (TIB-MolBiol, Berlin, Germany). Clinical response was evaluated at 12th and 24th week after initiation of the therapy according to the EULAR response criteria. RESULTS The frequency of the CD94 AA genotype was significantly decreased in RA patients compared to controls (OR=0.44; P=0.016). The CD94 AA homozygotes were also more common among patients negative to anti-cyclic citrullinated peptide (anti-CCP) antibodies (OR=11.28; P=0.001) as compared to anti-CCP-positive patients and the presence of the CD94 allele A was associated with lack of anti-CCP antibodies (OR=5.00; P=0.0005). The CD94 rs2302489 TT genotype was over-represented in patients exhibiting worse EULAR response at 12th week (OR=3.33; P=0.017). Furthermore, the lack of response after 12 weeks was more frequent among patients carrying the NKG2A rs7301582C allele (OR=3.68; P=0.019) or the CC genotype (OR=3.58; P=0.035) in comparison to allele T or CT/TT genotypes, respectively. CONCLUSIONS These results indicate that CD94 and NKG2A polymorphisms may contribute to genetic susceptibility to RA or affect the response to anti-TNF therapy in patients of Caucasian origin.

Cytokine profiles in axial spondyloarthritis.

Objectives Current studies concentrate on the cytokine network and its role in the pathogenesis of spondyloarthritis (SpA). In this study, we analyzed whether the serum cytokine profile (interleukins: IL-10, IL-11, IL-12, IL-15, IL-17, IL-23 and IL-33) correlates with demographic data, clinical manifestations, disease activity and treatment outcome in a group of patients with axial spondyloarthritis. Material and methods Forty-nine patients with an established diagnosis of axial spondyloarthritis (aSpA) and 19 healthy volunteers as controls were enrolled in the study. Clinical evaluation included patients medical history, 44 joint count, back pain intensity and global disease activity in the preceding week (VAS), the duration of morning stiffness and blood tests. Disease activity was assessed using BASDAI and ASDAS-CRP. Serum concentration of IL-10, IL-11, IL-12, IL-15, IL-17, IL-23 and IL-33 was determined. Results In patients with aSpA, elevated serum concentration of IL-10, IL-15, IL-17 and IL-23 was detected. In the aSpA group we detected higher values of serum concentration of IL-23 and IL-33 in the subgroup with anterior uveitis (83.1 ±184.0 pg/ml vs. 14.0 ±17.1 pg/ml, p < 0.0001 and 45.5 ±71.9 pg/ml vs. 18.4 ±14.3 pg/ml, p < 0.0001, respectively). Additionally, in the subgroup with peripheral arthritis, elevation of serum concentration of IL-12 (249.3 ±246.9 pg/ml vs. 99.9 ±105.9 pg/ml, p = 0.0001) was detected. Patients with preradiological SpA had higher serum concentration of IL-17 than patients with established diagnosis of AS (6.37 ±8.50 pg/ml vs. 2.04 ±2.98 pg/ml, p = 0.0295). No differences in serum concentration of analyzed cytokines were found between the subgroup with low to moderate disease activity and the subgroup with high to very high disease activity. Conclusions We report that in aSpA patients, compared to controls, elevated serum concentrations of IL-10, IL-15, IL-17 and IL-23 were observed. Some cytokines may predispose to a more severe course of aSpA.

Polymorphisms within the HLA‐E gene and their associations with susceptibility to rheumatoid arthritis as well as clinical outcome of anti‐TNF therapy

Involvement of the non‐classical human leucocyte antigen‐E (HLA‐E) in both innate and acquired immune response suggests its possible role in development of autoimmune pathologies. This study was undertaken to investigate relationships between the HLA‐E gene single nucleotide polymorphisms (SNPs) and a risk of rheumatoid arthritis (RA), as well as to evaluate a potential of these polymorphisms to modulate clinical outcome of anti‐tumour necrosis factor (TNF) treatment in female patients. A total of 223 female patients with RA receiving anti‐TNF biological therapy and 134 female healthy subjects were enrolled into the study. Genotypings for two SNPs within the HLA‐E gene (rs1264457 HLA‐E*01:01/01:03; rs1059510 HLA‐E*01:03:01/01:03:02) were performed using a polymerase chain reaction (PCR) amplification employing LightSNiP assays. Clinical response was evaluated according to the European League Against Rheumatism (EULAR) criteria at 12 and 24 weeks after initiation of the therapy. The frequency of the HLA‐E*01:01/01:01 genotype was decreased significantly in RA patients in comparison to controls (P = 0·031). The presence of the HLA‐E*01:01/01:01 genotype in patients correlated with better EULAR response after 12 weeks of anti‐TNF treatment, while 01:03 allele carriers were generally unresponsive to the treatment (P = 0·014). The HLA‐E*01:03/01:03 genotype was also over‐represented among non‐responding patients in comparison to HLA‐E*01:01/01:01 homozygotes (P = 0·021). With respect to the HLA‐E rs1059510 variation, a better response after 12 weeks was observed more frequently in patients carrying the HLA‐E*01:03:01/01:03:01 genotype than other genotypes (P = 0·009). The results derived from this study imply that HLA‐E polymorphisms may influence RA susceptibility and affect clinical outcome of anti‐TNF therapy in female RA patients.

Replication study of polymorphisms associated with response to methotrexate in patients with rheumatoid arthritis

About 70 genetic studies have already addressed the need of biomarkers to predict the response of patients with rheumatoid arthritis (RA) to methotrexate (MTX) treatment. However, no genetic biomarker has yet been sufficiently validated. Here, we aimed to replicate a selection of 25 SNPs in the largest collection of patients up to date, which consisted of 915 patients treated with MTX. The change in disease activity (measured as ΔDAS28) from baseline was considered the primary outcome. In addition, response according to widely used criteria (EULAR) was taken as secondary outcome. We considered consistency between outcomes, P values accounting for the number of SNPs, and independence from potential confounders for interpretation of the results. Only the rs1801394 SNP in MTRR fulfilled the high association standards. Its minor allele was associated with less improvement than the major allele according to ΔDAS28 (p = 0.0016), and EULAR response (p = 0.004), with independence of sex, age, baseline DAS28, smoking, seropositivity, concomitant corticosteroid use or previous treatments. In addition, previous evidence suggests the association of this SNP with response to MTX in another autoimmune disease, juvenile idiopathic arthritis, and with high intracellular folate levels, which could contribute to poor response.