Chronic Inflammatory Joint Research Articles

High percentages and activity of synovial fluid NK cells present in patients with advanced stage active Rheumatoid Arthritis

Rheumatoid Arthritis (RA) causes chronic inflammation of joints. The cytokines TNFα and IFNγ are central players in RA, however their source has not been fully elucidated. Natural Killer (NK) cells are best known for their role in elimination of viral-infected and transformed cells, and they secrete pro-inflammatory cytokines. NK cells are present in the synovial fluids (SFs) of RA patients and are considered to be important in bone destruction. However, the phenotype and function of NK cells in the SFs of patients with erosive deformative RA (DRA) versus non-deformative RA (NDRA) is poorly characterized. Here we characterize the NK cell populations present in the blood and SFs of DRA and NDRA patients. We demonstrate that a distinct population of activated synovial fluid NK (sfNK) cells constitutes a large proportion of immune cells found in the SFs of DRA patients. We discovered that although sfNK cells in both DRA and NDRA patients have similar phenotypes, they function differently. The DRA sfNK secrete more TNFα and IFNγ upon exposure to IL-2 and IL-15. Consequently, we suggest that sfNK cells may be a marker for more severely destructive RA disease.

Association of interleukin-4 gene polymorphism and rheumatoid arthritis in Egyptian patients

Introduction Rheumatoid arthritis (RA) is a progressive disease characterized by chronic joint inflammation and subsequent structural damage. Interleukin (IL)-4-590 promoter polymorphism (rs2243250), a C-to-T base substitution, has been suggested to be associated with RA and has become of great interest to be investigated. Aim The aim of this study was to find the relationship between IL-4-590 promoter polymorphism and RA in Egyptians, and also to study the relationship of this gene with clinical and laboratory features of the disease. Patients and methods This study was carried on 180 subjects divided into two groups. The first group included 120 patients with RA and the second group were 60 apparently healthy individuals as controls. Genomic DNA was extracted from blood leukocytes of both groups and genotyped by PCR for amplification of IL-4 gene followed by restriction fragment length polymorphism. Results IL-4-590 (TT) genotype was significantly more frequent in patients with RA than controls (10 vs. 1.70%, P=0.027, odd ratio (OR)=7.543 and Confidence interval (CI)=0.947–60.049). IL-4-590 (CT) genotype showed no significant difference between patients with RA and controls (31.70 vs. 25%, P=0.195 OR=1.592 and CI=0.786–3.228), whereas IL-4-590 (CC) genotype was significantly less frequent in patients with RA than controls (58.30 vs. 73.30%, P=0.048). Regarding the distribution of different alleles, the frequency of T allele was significantly more in patients with RA than controls (P<0.01). In patients with RA, there were significant differences in some clinical and laboratory parameters of RA disease between different IL-4-590 genotypes (e.g. number of tender and swollen joints, duration of morning stiffness, disease activity score 28, serum rheumatoid factor, serum C-reactive protein, and serum anticyclic citrullinated peptide levels), all were higher in TT genotype, which means patients with RA with TT genotype may have more aggressive course of the disease. Conclusion The T allele and the TT genotype at position −590 of IL-4 gene may be related to development of RA in Egyptians and may be associated with the disease activity.

Antibody modified gold nanoparticles for fast colorimetric screening of rheumatoid arthritis.

Rheumatoid arthritis (RA) is a systemic autoimmune disease characterized by chronic joint inflammation and one of the main causes of chronic disability worldwide with high prevalence in the ageing population. RA is characterized by autoantibody production, synovial inflammation and bone destruction, and the most accepted biomarker is rheumatoid factor (RF) autoantibodies. In this work, we developed a low-cost approach for the detection and quantification of the RF marker. This colorimetric immunosensor is based on gold nanoprobe crosslinking that results in extensive aggregation in the presence of the pentameric IgM RF. Aggregation of the nanoconjugates yields a color change from red to purple that can be easily observed by the naked eye. The interaction between nanoconjugates and the specific target was confirmed via dynamic light scattering (DLS), Raman spectroscopy and atomic force microscopy (AFM) imaging. This conceptual system shows a LOD of 4.15 UA mL-1 IgM RF (clinical threshold is set for 20 IU mL-1). The one-step biosensor strategy herein proposed is much faster than conventional detection techniques, without the need for secondary antibodies, additional complex washing or signal amplification protocols. To the best of our knowledge this is the first report on target induced aggregation of gold nanoprobes for quantitative colorimetric autoantibody detection.

Temporomandibular Disorders and Oral Features in Early Rheumatoid Arthritis Patients: An Observational Study.

Aims: Temporomandibular disorders (TMD) represent a heterogeneous group of inflammatory or degenerative diseases of the stomatognatic system, with algic and/or dysfunctional clinical features involving temporomandibular joint (TMJ) and related masticatory muscles. Rheumatoid Arthritis (RA) is an autoimmune polyarthritis characterized by the chronic inflammation of synovial joints and oral implications such as hyposalivation, difficulty in swallowing and phoning, feeling of burning mouth, increased thirst, loss of taste or unpleasant taste and smell, dental sensitivity.The aim of this observational study was to investigate the prevalence of TMD symptoms and signs as well as oral implications in patients with Early Rheumatoid Arthritis (ERA), that is a RA diagnosed within 12 months, compared with a control group.Methods: The study group included 52 ERA patients (11 men, 41 women) diagnosed according to the 2010 ACR/EULAR Classification Criteria for Rheumatoid Arthritis. A randomly selected group of 52 patients not affected by this disease, matched by sex and age, served as the control group. The examination for TMD signs and symptoms was based on the standardized Research Diagnostic Criteria for Temporomandibular Disorders (RDC/TMD) by means of a questionnaire and through clinical examination.Results: Regarding the oral kinematics, the left lateral excursion of the mandible was restricted in statistically significant way in ERA patients (p=0.017). The endfeel values were significantly increased in ERA group (p=0.0017), thus showing the presence of a higher muscle contracture. On the other side, the study group complained less frequently (67.3%) of TDM symptoms (muscle pain on chewing, pain in the neck and shoulders muscles, difficulty in mouth opening, arthralgia of TMJ, tinnitus) than controls (90.4%) (χ2= 8.301 p=0.0039). The presence of TMJ noises was significantly lower in the study group (χ2= 3.869 p=0.0049), as well as presence of opening derangement (χ2= 14.014 p=0.0002).The salivary flow was significantly decreased in the study group respect to the control one (p<0.0001).Conclusions: The data collected show a weak TMJ kinematic impairment, a paucisymptomatic muscle contracture (positive endfeel) and a remarkable reduction of salivary flow in ERA patients. Myofacial pain (MP) evoked by palpation was more frequent and severe in the control group than in the study one, this result being highly significant.

Baicalin Ameliorates Collagen-Induced Arthritis Through the Suppression of Janus Kinase 1 (JAK1)/Signal Transducer and Activator of Transcription 3 (STAT3) Signaling in Mice.

BackgroundRheumatoid arthritis is an autoimmune disease that causes chronic joint inflammation and there is no cure. Baicalin, as an ingredient in the roots of Scutellaria baicalensis, is supposed to possess an anti-inflammatory effect. However, the protective effect of baicalin on collagen-induced arthritis requires further investigation.Material/MethodsA model of rheumatoid arthritis was established in 20 mice (8- to 10-weeks old). The mice were randomly divided into 2 groups after modeling and then injected with saline or baicalin, respectively. The synovial fluids and tissues were collected, and the pressure pain threshold and clinical arthritis score were measured. The levels of tumor necrosis factor (TNF)-α, interlukin-1β (IL-1β), IL-6, matrix metalloproteinases (MMP)-2, MMP-9, nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2) and their downstream inflammatory mediators Janus kinase 1 (JAK1)/signal transducer and activator of transcription 3 (STAT3), extracellular signal-regulated kinases 1/2 (ERK1/2), p38, Jun N-terminal kinases (JNK) activation were detected using enzyme-linked immunosorbent assay (ELISA), and western blotting analyses. The mononuclear cells apoptosis ratio was calculated by flowcytometry analyses.ResultsBaicalin significantly reduced disease activities in a rheumatoid arthritis mouse model, which were reflected by pressure pain thresholds and clinical arthritis scores. Relevant proinflammatory cytokines such as TNF-α, IL-1β, IL-6, gelatinases (MMP-2, MMP-9) and inducible enzymes (iNOS, COX-2) were generally suppressed. Moreover, baicalin treatment induced cells apoptosis in synovial fluid monocytes and markedly down regulated JAK1/STAT3 but not mitogen-activated protein kinases (MAPKs) expressions in synovium of arthritis.ConclusionsThese observations confirm the relief of rheumatoid arthritis by baicalin. Our results indicate the effect is related with the modulation of decreased proinflammatory cytokines and inflammatory markers. And the apoptosis promotion of monocytes in synovial fluid were also inhibited. Moreover, the molecular mechanism implies suppressed JAK1/STAT3 signaling with baicalin treatment.

Studies on the design of a composition of gel for the treatment of inflammatory diseases of the joints

The aim: the aim of our work was to experimentally substantiate the effectiveness of antimicrobial preservatives in the design of the composition of the combined gel for the treatment of acute and chronic inflammation of the joints. Methods: biological pharmacopoeial method of research was used (research of the effectiveness of antimicrobial preservatives). Results. All drugs must meet the requirements of regulatory documents on indicators Microbiological purity. To ensure the microbiological stability of the preparations, it is necessary to eliminate the factors associated with microbial contamination, therefore we carried out experimental studies on the choice of preservative and its concentration for the developed gel. According to the results of experimental studies on the effectiveness of antimicrobial preservatives in gel samples, the following data were obtained. Samples of the studied gel with sodium preservatives benzoate 0.5 % and nipagin 0.2 % meet the criterion A according to the requirements of SPHU for drugs for skin application. But, according to research results, the antimicrobial efficacy of the gel with the preservative nipagin 0.2 % was slightly higher (lg reduction in the number of viable Staphylococcus aureus ATCC 6538 cells was 3.32 and 4.81; Pseudomonas aeruginosa ATCC 9027 - 3.28 and 4.66 ; Candida albicans ATCC 885-653 - 3.50 and 4.09; Aspergillus brasiliensis ATCC 16404 - 3.10 and 4.00 (2 and 7 days, respectively) and the spectrum of antimicrobial action is wider, which will contribute to the quality of the rotated gel also during storage. Therefore, for further research, we will use nipagin 0.2 % as a preservative. Conclusions: The expediency of using nipagin 0.2 % as a part of a combined gel for the treatment of acute and chronic joint inflammation has been theoretically proved and experimentally proved

The role of Cutibacterium acnes in auto-inflammatory bone disorders.

Chronic nonbacterial osteomyelitis (CNO) and SAPHO (synovitis, acne, pustulosis, hyperostosis and osteitis) syndrome are auto-inflammatory disorders manifesting as chronic inflammation of bones and joints, which in SAPHO is often accompanying by skin changes. The aetiology of these diseases is unknown, but includes genetic, infectious and immunological components. It has been proposed that Cutibacterium (formerly Propionibacterium) acnes plays a role in the pathogenesis. In this review, we summarise reported cases of CNO or SAPHO syndrome in which C. acnes has been isolated from bones. To identify cases, a search was done in May 2018 using the MEDLINE Ovid interface (1946 to present). We found 14 publications reporting 98 patients with auto-inflammatory bone disorders, of whom 48 (49%) had positive bone biopsies for C. acnes. This bacteriumwas more frequently isolated from open biopsies than percutaneous ones (43/69 (62%) vs 1/7 (14%); p = 0.04) and biopsies were more frequently positive in patients who presented with simultaneous skin manifestations (19/36 (53%) vs 4/12 (33%); p = 0.03).Conclusion: In patients with CNO or SAPHO, C. acnes can be isolated from open biopsies suggesting that in these patients, C. acnes might be a pathogen rather than a contaminant. The fact that biopsies are more frequently positive in patients who present with simultaneous skin manifestations suggests that these individuals might have a genetic predisposition for impaired clearance of C. acnes. What is known • Chronic nonbacterial osteomyelitis (CNO) and SAPHO (synovitis, acne, pustulosis, hyperostosis and osteitis) syndrome are auto-inflammatory disorders manifesting as inflammation of bones. Both diseases are an important differential diagnosis in children who present with symptoms of (multifocal) osteomyelitis. • The pathogenesis of CNO and SAPHO is multifactorial emcompassinggenetic, infectious and immunological components, including interleukin (IL)-1 dysregulation. There is a controversy as to whether Cutibacterium (formerly Propionibacterium) acnes plays a role in the aetiology of CNO and SAPHO. It has been postulated that the presence of C. acnes might trigger auto-inflammatory chronic inflammation in genetically predisposed individuals. What is new • In patients with CNO or SAPHO, C. acnes can be isolated more frequently from open biopsies, than from percutaneous ones, suggesting that C. acnes might be a pathogen rather than a contaminant. • Biopsies are more frequently positive in patients who present with simultaneous skin manifestations suggesting that these individuals might have a genetic predisposition for impaired clearance of C. acnes. Impaired C. acnes clearance likely leads to increased IL-1 beta (β) production by skin cells, bone cells and phagocytes, which is one of the main cytokines underlying chronic inflammatory bone disorders.

Polymorphism of Foxp3 gene affects the frequency of regulatory T cells and disease activity in patients with rheumatoid arthritis in Iranian population

Rheumatoid arthritis (RA) is a chronic autoimmune disease that mainly affects joints and characterized by chronic joint inflammation and infiltration of various immune cells in the synovium. Forkhead box P3 (Foxp3)-expressing regulatory T cells (Tregs) play a crucial role in preventing autoimmunity and undesirable T cell responses. However, there are controversial reports regarding the defective function or frequency of these cells in various studies, which may be in part related to different polymorphisms of FoxP3 and influence of ethnicity on these differences. Therefore, the main subject of this study was to evaluate the association of Foxp3 gene polymorphism and Treg frequency in Iranian patients with RA. Accordingly, 240 RA patients diagnosed according to American college of rheumatology 2010 criteria and 240 normal subjects were recruited for this study. Genomic DNA was genotyped for –3279 C/A Foxp3 gene SNP using the PCR-RFLP. The frequency of Tregs and serum levels of interleukin (IL)-10, transforming growth factor (TGF)-β, anti-cyclic citrullinated peptide (CCP) and rheumatoid factor (RF) were determined by flow cytometry and ELISA methods, respectively. The results showed a significant association of Foxp3 −3279 A allele with augmented risk of RA in Iranian patients compared to wild-type allele. While the frequencies of CA and AA genotypes were significantly higher in patients, RA patients with AA genotype had a significant lower frequency of Tregs compared to patients with CC and CA genotypes. Consistently, TGF-β and IL-10 significantly diminished in patients with AA genotype compared to patients with CA and CC genotypes. Our findings indicated that the AA genotype of Foxp3 in RA patients is associated with downregulation of Tregs and susceptibility to RA in the Iranian population.

Genome-wide association study of response to tumour necrosis factor inhibitor therapy in rheumatoid arthritis

Rheumatoid arthritis (RA) is characterised by chronic synovial joint inflammation. Treatment has been revolutionised by tumour necrosis factor alpha inhibitors (TNFi) but each available drug shows a significant non-response rate. We conducted a genome-wide association study of 1752 UK RA TNFi-treated patients to identify predictors of change in the Disease Activity Score 28 (DAS28) and subcomponents over 3–6 months. The rs7195994 variant at the FTO gene locus was associated with infliximab response when looking at a change in the swollen joint count (SJC28) subcomponent (p = 9.74 × 10−9). Capture Hi-C data show chromatin interactions in GM12878 cells between rs2540767, in high linkage disequilibrium with rs7195994 (R2 = 0.9) and IRX3, a neighbouring gene of FTO. IRX3 encodes a transcription factor involved in adipocyte remodelling and is regarded as the obesity gene at the FTO locus. Importantly, the rs7195994 association remained significantly associated following adjustment for BMI. In addition, using capture Hi-C data we showed interactions between TNFi-response associated variants and 16 RA susceptibility variants.

Divergent effects on macrophage biomarkers soluble CD163 and CD206 in axial spondyloarthritis

The chronic joint inflammation in axial spondyloarthritis (axSpA) is characterized by infiltration of activated macrophages. The haptoglobin–hemoglobin receptor CD163 and the mannose receptor CD206 are strongly expressed on M2c and M2a macrophages, respectively. We measured the soluble forms of the receptors (sCD163 and sCD206) in plasma (PL) in two axSpA cohorts. All patients fulfil the 2009 Assessment of SpondyloArthritis International Society (ASAS) classification criteria for axSpA and/or the 1984 modified New York criteria for ankylosing spondylitis. The first cohort included anti-TNF-α treated patients with active axSpA (n = 30); the second cohort included patients in early disease stages (n = 38). Plasma sCD163 and sCD206 were both within the reference interval of healthy controls (HC), but sCD163 decreased slightly during anti-TNF-α treatment [baseline: 1.49 mg/L (IQR: 1.22–1.77 mg/L, 12 weeks: 1.29 (IQR: 1.09–1.57) mg/L, 20 weeks: 1.25 (IQR: 0.99–1.75) mg/L, 52 weeks: 1.39 (IQR: 1.15–1.78) mg/L], while sCD206 increased [baseline: 0.17 (IQR: 0.13–0.21) mg/L, 12 weeks: 0.19 (0.16–0.23) mg/L, 20 weeks: 0.20 (0.14–0.24) mg/L, 52: 0.19 (IQR: 0.14–0.23) mg/L], pointing toward a shift in polarization of involved macrophages. Plasma levels of sCD206 proved significantly higher in patients with early disease stages and definite radiological sacroiliitis (n = 10). This was not the case for sCD163. A significant increase in response to anti-TNF-α treatment, could suggest sCD206 as a marker of response to anti-TNF-α treatment, however, the potential for the two macrophage markers as diagnostic and prognostic indicators of disease in axSpA is weak.

The Anti-Inflammatory Mediator, Vasoactive Intestinal Peptide, Modulates the Differentiation and Function of Th Subsets in Rheumatoid Arthritis.

Genetic background, epigenetic modifications, and environmental factors trigger autoimmune response in rheumatoid arthritis (RA). Several pathogenic infections have been related to the onset of RA and may cause an inadequate immunological tolerance towards critical self-antigens leading to chronic joint inflammation and an imbalance between different T helper (Th) subsets. Vasoactive intestinal peptide (VIP) is a mediator that modulates all the stages comprised between the arrival of pathogens and Th cell differentiation in RA through its known anti-inflammatory and immunomodulatory actions. This “neuroimmunopeptide” modulates the pathogenic activity of diverse cell subpopulations involved in RA as lymphocytes, fibroblast-like synoviocytes (FLS), or macrophages. In addition, VIP decreases the expression of pattern recognition receptor (PRR) such as toll-like receptors (TLRs) in FLS from RA patients. These receptors act as sensors of pathogen-associated molecular pattern (PAMP) and damage-associated molecular pattern (DAMP) connecting the innate and adaptive immune system. Moreover, VIP modulates the imbalance between Th subsets in RA, decreasing pathogenic Th1 and Th17 subsets and favoring Th2 or Treg profile during the differentiation/polarization of naïve or memory Th cells. Finally, VIP regulates the plasticity between theses subsets. In this review, we provide an overview of VIP effects on the aforementioned features of RA pathology.

Association between microalbuminuria and metabolic syndrome in patients with rheumatoid arthritis

ContextRheumatoid arthritis (RA) is an autoimmune, symmetrical polyarticular disease characterized by chronic inflammation of the synovial joints. Microalbuminuria (MA) occurs as leakage of small amounts of albumin into the urine. Metabolic syndrome (MetS) describes the risk factors for cardiovascular diseases such as dyslipidemia, obesity, hypertension, and diabetes.AimThe aim of this study was to detect the prevalence of MA in patients with RA and study its correlation with disease activity and severity. Our aim extends to identify the association of MA with MetS in RA.Patients and methodsThis study was carried out on 30 adult RA patients, 30% of them were men and 70% were women (mean±SD: 42.27±10.99 years). Their mean disease duration was 12.8±7.06 years. A total of 20 apparently healthy adults, age-matched and sex-matched served as a control group. All the patients were subjected to full history taking, full clinical examination, laboratory investigations, and assessment of disease activity using the disease activity score for 28joints score. Urinary microalbumin level was measured in all participants in early morning samples by the immunoturbidometry method. MetS was assessed in all participants according to Grundy’s criteria.ResultsThe frequency of MetS was highly statistically significant in patients with RA compared with the control group. The RA patients’ group had highly significantly elevated mean values of urinary microalbumin and urinary albumin to creatinine ratio compared with the control group.ConclusionMA and MetS are frequent in RA, particularly in those with long-standing disease. Early detection of albuminuria allows early intervention with the goal of reducing inflammation development in RA, cardiovascular risk. MetS is frequent in RA patients with MA.

The Effect of Bone Marrow Mesenchymal Stem Cells Versus Methotrexate on the Knee Joint in A Rat Model of Rheumatoid Arthritis. A Histological Study

Background and objectives: Rheumatoid arthritis (RA) is an autoimmune disease characterized by chronic progressive joint inflammation with subsequent cartilage destruction. This study was conducted to evaluate and compare the effect of bone marrow mesenchymal stem cells (BM-MSCs) versus Methotrexate (MTX) on knee joint in a rat model of RA.Materials and Methods: Thirty-five adult male albino rats were divided into two groups. Group I: control group (15 rats). Group II (20 rats) in which RA was induced, and then the rats were subdivided into four subgroups. Subgroups IIa and IIb were sacrificed two and four weeks after induction of RA respectively. Subgroups IIc and IId were treated by MTX and BM-MSCs respectively after two weeks of induction of RA and were sacrificed after further two weeks. The knee joints were collected, decalcified and processed for histological, histochemical, immunehistochemical and morphometric studies.Results: Histological examination of the knee joints revealed that RA resulted in thickening of the intimal lining of the synovial membrane, infiltration, congestion and increased collagen content of the subintima. The articular cartilage showed erosions, thinning, cell and ground substance loss and increased expression of inducible nitric oxide synthase (iNOS). Injection of BM-MSCs resulted in improvement of the structure of the synovial membrane and the articular cartilage of the knee joint whereas, injection of MTX was relatively less effective.Conclusion: Intra- articular injection of BM-MSCs resulted in a significant improvement in the histological structure of the knee joint in comparison to MTX in a rat model of RA.

Autoimmune Th17 Cells Induced Synovial Stromal and Innate Lymphoid Cell Secretion of the Cytokine GM-CSF to Initiate and Augment Autoimmune Arthritis

SummaryDespite the importance of Th17 cells in autoimmune diseases, it remains unclear how they control other inflammatory cells in autoimmune tissue damage. Using a model of spontaneous autoimmune arthritis, we showed that arthritogenic Th17 cells stimulated fibroblast-like synoviocytes via interleukin-17 (IL-17) to secrete the cytokine GM-CSF and also expanded synovial-resident innate lymphoid cells (ILCs) in inflamed joints. Activated synovial ILCs, which expressed CD25, IL-33Ra, and TLR9, produced abundant GM-CSF upon stimulation by IL-2, IL-33, or CpG DNA. Loss of GM-CSF production by either ILCs or radio-resistant stromal cells prevented Th17 cell-mediated arthritis. GM-CSF production by Th17 cells augmented chronic inflammation but was dispensable for the initiation of arthritis. We showed that GM-CSF-producing ILCs were present in inflamed joints of rheumatoid arthritis patients. Thus, a cellular cascade of autoimmune Th17 cells, ILCs, and stromal cells, via IL-17 and GM-CSF, mediates chronic joint inflammation and can be a target for therapeutic intervention.

Gambaran Aktivitas Fungsional Penderita Osteoarthritis Lutut Yang Menggunakan Knee Support

Abstract : Osteoarthritis, Functional Activity, Knee Support. Knee osteoarthritis is a chronic inflammation of the knee joint characterized by a decrease in muscle value and an impact on decreased knee joint stability resulting in decreased ability of a person to perform functional activities. The ability of functional activity is directly related to quality of life and its assessment is very important in clinical practice. The decrease in functional activity of knee osteoarthritis patient can be minimized by the use of knee support. The purpose of this study was to determine the effect of knee support on functional activity in patient with knee osteoarthritis. This study used qualitative research method with one subject of knee osteoarthritis. The result of this research there are some progress of the subject when performing functional activity that she could not do. The conclusion obtained is the use of knee support within six weeks can increase the functional activity of patients with knee osteoarthritis degree three. As for suggestions that can be given for researchers who want to continue this research is to add research subjects and also pay attention to uncontrolled factors cause knee osteoarthritis.

Pathophysiology of rheumatoid arthritis is associated with polymorphisms in Protein Tyrosine Phosphatase Non-receptor type 2 and 22 (PTPN2/22) and susceptibility to Mycobacteria

Abstract Single nucleotide polymorphisms (SNPs) in T-cell regulator genes such as Protein Tyrosine Phosphatase Non-receptor type 2 and 22 (PTPN2/22) have been associated with many inflammatory autoimmune disorders including rheumatoid arthritis (RA). Along with these SNPs and an environmental trigger of a mycobacterial infection from Mycobacterium avium subspecies paratuberculosis (MAP), we hypothesis that this will lead to chronic inflammation of the synovial joints of RA. We gathered 132 consented subjects blood samples, where genotyping was done for nine SNPs found in PTPN2/22 using TaqMan™ genotyping. Quantification of the effect of the SNPs on gene expression of PTPN2/22 and IFN-γ expression was determined through real time PCR. T-cell proliferation assays were done to examine the response of phytohematoagglutonin (PHA) and mycobacterial antigens in subject samples by the use of BrdU proliferation assays. Detection of MAP bacterial IS900 DNA was done by nested PCR. Out of the nine SNPs examined, PTPN2:rs478582/PTPN22:rs2476601 were found significant in RA compared to the healthy controls (p-values &amp;lt; 0.05). Gene expression of PTPN2/22 was downregulated in RA overall compared to healthy controls. IFN-γ expression was upregulated in samples with significant SNPs. T-cell proliferation increased when T-cells tested had significant SNPs (p-values &amp;lt; 0.05). IS900 MAP DNA was detected more significantly in RA than in healthy controls, where samples with either significant SNPs had higher presence of MAP (p-values &amp;lt; 0.05). This study suggests that SNPs in PTPN2/22 dysregulates T-cell regulation and increases susceptibility to Mycobacteria, leading to increased proliferation of T-cells and higher amounts of IFN-γ expression in RA.

Manganese dioxide nanoparticles mitigate oxidative stress in osteoarthritic cartilage

Purpose: Osteoarthritis (OA) is associated with chronic joint inflammation, whereby pro-inflammatory cytokines such as IL-1β upregulate reactive oxygen species (ROS) production while downregulating anti-oxidants in cells. The resulting oxidative stress leads to extracellular matrix degradation, joint inflammation, and chondrocyte death and senescence. The role of oxidative stress in OA etiology makes it a therapeutic target to ameliorate some of the devastating symptoms of the disease. The overall goal of this research is to develop a bioactive nanoparticle (NP) system that scavenges ROS in cartilage to modulate the impact of joint inflammation. Manganese dioxide (MnO2) catalyzes the breakdown of hydrogen peroxide (H2O2) and is currently being evaluated for scavenging ROS in inflammatory diseases such as atherosclerosis. We hypothesize that MnO2 NPs will protect cartilage from inflammation-induced oxidative stress. To do this, we have engineered chondroprotective MnO2 NPs with properties that enable them to penetrate into the cartilage. Methods: MnO2 NPs were synthesized by the oxidation of potassium permanganate with polyallyamine hydrochloride and further stabilized with polyethylene glycol (PEG). The NPs were characterized for size and zeta potential in phosphate buffer saline (PBS) by Dynamic Light Scattering (DLS) and polydispersity by transmission electron microscopy (TEM). Scavenging capacity of MnO2 NPs was evaluated by a H2O2 colorimetric detection kit. Uptake of Alexa Fluor 488 conjugated NPs by bovine chondrocytes in 2-D culture and cartilage explants were evaluated by fluorescent imaging after 24 hours of incubation. The cytotoxicity of NPs to chondrocytes was tested using the MTS colorimetric assay. Fresh bovine cartilage biopsies were challenged with human recombinant IL-1β, to mimic OA-like cartilage, in the presence of MnO2 NPs and evaluated for loss of glycosaminoglycans (GAG), a key component of cartilage extracellular matrix, and production of nitric oxide (NO), a key reactive nitrogen species (RNS) related to oxidative stress in OA, over a 9-day period. The loss of GAG was quantified using spectrophotometric Dimethylmethylene Blue Assay while NO concentrations were evaluated by the Griess reaction. Articular joint retention of Alexa Fluor 750 conjugated NPs in rats was monitored using an in vivo IVIS Imaging System. Results: The PEG stabilized MnO2 NPs had a hydrodynamic size of 15 nm (number weighted) and TEM size of 8 nm. The NPs had neutral charge. The NPs effectively scavenged H2O2, with 5ug/mL MnO2 NPs neutralizing 55% of 100uM H2O2. Chondrocytes in monolayer showed uptake of MnO2 NPs, which escaped the endosomes and localized perinuclearly without cytotoxicity (Fig. 1). The MnO2 NPs penetrated through the depth of the cartilage biopsies and remained in the tissue matrix. Interestingly, the NPs were also endocytosed by resident chondrocytes (Fig. 2). In cytokine challenged cartilage explants, supplementation with MnO2 NPs decreased NO production by 80% and reduced GAG loss by 55% compared to controls (Fig. 3A and B). The MnO2 NPs after in vivo injection in articular joints displayed a linear decline (R2 = 0.933) in retention with time. Quantification of fluorescent signals showed that 78% of the initial amount of NPs remained in joints after 5 days (Fig. 4A and B). Conclusions: MnO2 NPs have the potential to reduce oxidative stress in osteoarthritic cartilage. Given their joint retention time and ROS scavenging capacity, these NPs could target oxidative stress to treat or prevent OA. As the NPs show intracellular localization in chondrocytes, they could also deliver other chondroprotective agents including nucleic acids to target multiple pathways in the OA pathology. Further studies will focus on the therapeutic impact of MnO2 NPs to joint tissues in vivo.View Large Image Figure ViewerDownload Hi-res image Download (PPT)View Large Image Figure ViewerDownload Hi-res image Download (PPT)View Large Image Figure ViewerDownload Hi-res image Download (PPT)

Mesenchymal stem cells in chronic joint inflammation

Mesenchymal stem cells in chronic joint inflammation

Laboratory diagnostics and pathogenesis of rheumatoid arthritis — the past and the present

Rheumatoid arthritis (RA) is one of the most common systemic connective tissue diseases of autoimmune origin and is characterised by chronic inflammation of joints. The aetiology of this disease has not been fully understood yet. A crucial role in the development of RA is played by multiple genetic (shared epi-tope, HLA-DR genes, family predisposition) and environmental factors (smoking, female-specific factors, bacteria, viruses and mucosal inflammation, periodontal and lung diseases). The purpose of this article is to review the latest data on the pathogenesis of this disease and biomarkers used in the diagnostics of RA. RA is associated with the production of autoantibodies, among which rheumatoid factor (RF) and anti-citrullinated protein (anti-CCP) antibodies are included in the new classification criteria of RA. So far, anti-CCP is the best marker of RA; its high sensitivity and specificity have been helpful in diagnostics and monitoring disease activity and the development of more aggressive disease, as well as the pharmacological treatment used. Recently, anti-carbamylated (anti-carP) antibodies and calprotectin have been described in RA, which also appears to be promising in the diagnostics of this disease.

Expression of IGF-1, IL-27 and IL-35 Receptors in Adjuvant Induced Rheumatoid Arthritis Model.

IGF-1 and certain other cytokines have been shown to exert inflammatory/anti-inflammatory roles in chronic joint diseases. To assess the effect of IGF-1, IL-27 and IL-35, their interaction and their receptor expression in a rheumatoid arthritis model. Freund's adjuvant-induced chronic joint inflammation was operated on 160 male rats. Animals were divided into histopathology and receptor expression groups, each composed of 10 subgroups including; control, vehicle, IGF-1, IL-27, IL-35, their antagonists, IGF-1+IL-27 antagonist and IGF-1+IL-35 antagonist. After two weeks, vehicle or agonist/antagonists were injected into the joint space every other day until day 28 where joint histopathology was performed. The expression of IGF-1, IL-27 and IL-35 receptors were assessed by western blot analysis. IGF-1 did not show pro- or anti- inflammatory functions; endogenous IL-27 and IL-35, on the other hand, exerted inflammatory effects. IL-27 and IL-35 antagonists exerted the highest anti-inflammatory effects. The total inflammation scores were 0.55 ± 0.06, 4.63 ± 0.40, 3.63 ± 0.60, 2.50 ± 0.38 and 1.63 ± 0.40 regarding control, vehicle, IGF-1 Ant., IL-27 Ant. and IL-35 Ant., respectively. IGF-1 receptor expression was reduced in chronic joint inflammation and all three antagonists augmented the IGF-1 receptor expression. IL-27 and IL-35 receptors were up-regulated by chronic joint inflammation. Overall, the results demonstrated the pro-inflammatory role of endogenous IL-27 and IL-35 along with the over expression of their receptors in chronic joint inflammation. IL-27 and IL-35 antagonists exerted the most anti-inflammatory effects and increased IGF-1 receptor expression. These two antagonists may be potential agents for new treatment strategies in chronic joint inflammatory diseases.