Experimental Arthritis Model Research Articles

Role of Preclinical Arthritis Models for the Clinical Translation of Anti-Arthritic Therapeutics

The overcomplicated and elusive pathophysiology with unclear etiology, have been the main driving force of the medical scientists all over the world to develop a predictive, reliable, robust and reproducible simulation model of arthritis. This review highlights osteoarthritis and rheumatoid arthritis with distinct conditions pertaining to each type of disease. The advances in various in vitro and in vivo experimental models of osteoarthritis and rheumatoid arthritis have been presented along with their pros and cons for antiarthritic drug discovery and formulation development. Additionally, the ethical issues to be considered while selecting animal models and handling them have been covered briefly. The current status quo on clinical trials of antiarthritic therapeutic interventions has also been covered.

Reset of Inflammatory Priming of Joint Tissue and Reduction of the Severity of Arthritis Flares by Bromodomain Inhibition.

We have recently shown that priming of synovial fibroblasts (SFs) drives arthritis flares. Pathogenic priming of SFs is essentially mediated by epigenetic reprogramming. Bromodomain and extraterminal motif (BET) proteins translate epigenetic changes into transcription. Here, we used a BET inhibitor (I-BET151) to target inflammatory tissue priming and to reduce flare severity in a murine experimental arthritis model. BALB/c mice were treated by intraperitoneal injection or by local injection in the paw with I-BET151, which blocks the interaction of BET proteins with acetylated histones. We assessed the effects of I-BET151 on acute arthritis and/or inflammatory tissue priming in a model of repeated injections of monosodium urate crystals or zymosan into the mouse paw. I-BET151 was given before arthritis induction, at peak inflammation, or after healing of the first arthritis bout. We performed transcriptomic (RNA-Seq), epigenomic (ATAC-Seq), and functional (invasion, cytokine production, migration, senescence, metabolic flux) analyses of murine and human SFs treated with I-BET151 in vitro or in vivo. Systemic I-BET151 administration did not affect acute inflammation but abolished inflammatory tissue priming and diminished flare severity in both preventive and therapeutic treatment settings. I-BET151 was also effective when applied locally in the joint. BET inhibition also inhibited osteoclast differentiation, while macrophage activation in the joint was not affected. Flare reduction after BET inhibition was mediated, at least in part, by rolling back the primed transcriptional, metabolic, and pathogenic phenotype of SFs. Inflammatory tissue priming is dependent on transcriptional regulation by BET proteins, making them promising therapeutic targets for prevention of arthritis flares in previously affected joints.

Acute effects of photobiomodulation applied on the dorsal root ganglion in gout model-induced rats.

Gouty arthritis is an inflammatory disease that triggers symptoms such as pain, swelling, and joint stiffness. Since its main therapy is medication, research on other forms of treatment that do not generate side effects is necessary. Given this, the objective of this research was to evaluate the effects of combined photobiomodulation (LASER and LED) applied on the dorsal root ganglion (DRG) in an experimental model of gouty arthritis. For this, 40 Wistar rats were randomized into 4 groups: simulation of the model with saline injection, without treatment (CTL; n=10); gout simulation with photobiomodulation treatment (CTL-PBM; n=10); gout model with the injection of monosodium urate crystals (1.25mg) in the femorotibial joint, without treatment (GOT; n=10); or gout model with photobiomodulation treatment (GOT-PBM; n=10). After 7h of gout induction, photobiomodulation was performed with a cluster of 4 diodes applied to the GRD region in animals from the CTL-PBM and GOT-PBM groups. After analysing the results, it was concluded that the therapy favored the reduction of edema and joint incapacity, as well as the increase in the nociceptive threshold and plantar grip strength. Furthermore, PBM stimulated an increase in the inflammatory response (with increased levels of IL-1β and greater recruitment of leukocytes) and greater activation of the antioxidant system. Therefore, PBM can be considered an effective therapeutic alternative to improve the functional status in this model of joint disease.

PPARβ/δ-Interfering Peptide Enhanced Mesenchymal Stromal Cell Immunoregulatory Properties.

Mesenchymal stem/stromal cells (MSCs) have been widely used for their therapeutic properties in many clinical applications including osteoarthritis. Despite promising preclinical results showing the ability of MSC to reduce the clinical severity of osteoarthritis (OA) in experimental animal models, the benefits of intra-articular injection of MSC in OA patients are limited to the short term. In this regard, it is anticipated that improving the properties of MSC may collectively enhance their long-term beneficial effects on OA. Recently, we have shown that PPARβ/δ inhibition using a commercially available antagonist in murine MSC increases their immunoregulatory potential in vitro as well as their therapeutic potential in an experimental murine arthritis model. Here, we relied on an innovative strategy to inhibit PPARβ/δ:NF-κB TF65 subunit interaction in human MSC by designing and synthesizing an interfering peptide, referred to PP11. Through RT-qPCR experiments, we evidenced that the newly synthesized PP11 peptide reduced the expression level of PDK4, a PPARβ/δ target gene, but did not modify the expression levels of ACOX1 and CPT1A, PPARα target genes, and FABP4, a PPARγ target gene compared with untreated human MSC. Moreover, we showed that human MSCs pretreated with PP11 exhibit a significantly higher capacity to inhibit the proliferation of activated PBMC and to decrease the frequency of M1-like macrophages. We designed and synthesized an interfering peptide that potently and specifically blocks PPARβ/δ activity with concomitant enhancement of MSC immunoregulatory properties.

Advances in experimental models of rheumatoid arthritis.

Rheumatoid arthritis (RA) is a chronic systemic autoimmune disease characterized by persistent articular inflammation and joint damage. RA was first described over 200 years ago; however, its etiology and pathophysiology remain insufficiently understood. The current treatment of RA is mainly empirical or based on the current understanding of etiology with limited efficacy and/or substantial side effects. Thus, the development of safer and more potent therapeutics, validated and optimized in experimental models, is urgently required. To improve the transition from bench to bedside, researchers must carefully select the appropriate experimental models as well as draw the right conclusions. Here, we summarize the establishment, pathological features, potential mechanisms, advantages, and limitations of the currently available RA models. The aim of the review is to help researchers better understand available RA models; discuss future trends in RA model development, which can help highlight new translational and human-based avenues in RA research.

Traditional Chinese Medicine Rhodiola Sachalinensis Borissova from Baekdu Mountain (RsBBM) for Rheumatoid Arthritis: Therapeutic Effect and Underlying Molecular Mechanisms.

The lack of effective rheumatoid arthritis (RA) therapies is a persistent challenge worldwide, prompting researchers to urgently evaluate traditional Chinese medicines (TCMs) as potential clinical RA treatments. The present investigation was conducted to evaluate the therapeutic effects and potential molecular mechanisms of the active components isolated from TCM Rhodiola sachalinensis Borissova from Baekdu Mountain (RsBBM) using an experimental adjuvant arthritis model induced by injection of rats with Freund’s complete adjuvant. After induction of the adjuvant arthritis rat model, the extract-treated and untreated groups of arthritic rats were evaluated for RsBBM therapeutic effects based on comparisons of ankle circumferences and ELISA-determined blood serum inflammatory factor levels (TNF-α, IL-1β, and PGE2). In addition, the joint health of rats was evaluated via microscopic examination of hematoxylin-eosin-stained synovial tissues. Furthermore, to explore whether NF-κB and RANK/RANKL/OPG signaling pathways participated in observed therapeutic effects from a molecular mechanistic viewpoint, mRNA and protein levels related to the expression of nuclear factor kappa-B (NF-κB), osteoprotegerin (OPG), and receptor activator of nuclear factor kappa-Β ligand (RANKL) were analyzed via quantitative RT-PCR and Western blot analysis, respectively. Treatment of arthritic rats with the extract of RsBBM was shown to reduce ankle swelling, reduce blood serum levels of inflammatory factors, and alleviate arthritis-associated synovial inflammation and joint damage. Moreover, an RsBBM 50% ethanol extract treatment inhibited bone destruction by up-regulating OPG-related mRNA and protein expression and down-regulating RANKL-related mRNA and protein expression, while also reducing inflammation by the down-regulating of the NF-κB pathway activity. The results clearly demonstrated that the extract of RsBBM alleviated adjuvant arthritis-associated joint damage by altering activities of inflammation-associated NF-κB and the RANK/RANKL/OPG signaling pathways. Due to its beneficial effects for alleviating adjuvant arthritis, this RsBBM 50% ethanol extract should be further evaluated as a promising new therapeutic TCM treatment for RA.

Trans-anethole, the major constituent of the essential oil of anise, improves the antiarthritic effects of methotrexate in rats

The aim of this study was to evaluate the effect of trans-anethole in combination with methotrexate, compared to monotherapy with trans-anethole or methotrexate, on inflammatory parameters and oxidative stress of arthritic rats. The experimental model of arthritis induced by Freund's complete adjuvant (AIA) in rats was used. Treatment with trans-anethole and methotrexate, in combination or monotherapy, was started on the day of AIA induction and continued for 21 days. The association of anethole (62.5 mg/Kg) to methotrexate (6 mg/Kg) therapy was able to further reduce both the oxidative stress induced by arthritis and the inflammatory events that characterize the disease, as demonstrated by the following indicators: 1) decrease in the edema of the hind legs, in the score of secondary lesions, in the recruitment of leukocytes to the articular cavity and in the plasma myeloperoxidase activity; 2) increase in plasma content of reduced thiols and total antioxidant capacity; 3) diminution of protein carbonylation in plasma, liver and kidneys; 4) decrease in lipid peroxidation in the liver; 5) reduction in the hepatic ROS content; 6) enhancement of the activity of the antioxidant enzymes and GSH/GSSG ratio. In general, the combination anethole + methotrexate presented a substantially higher effect than the monotherapy with anethole or methotrexate (at the same doses). The data obtained in this work allow us to conclude that the use of trans-anethole in combination with methotrexate suppressed arthritic progression in rats, with the main advantage of reduction of methotrexate dose, attenuating adverse effects caused by high dose therapy.

Crocin-loaded nanoliposomes: Preparation, characterization, and evaluation of anti-inflammatory effects in an experimental model of adjuvant-induced arthritis

This study aimed to develop nanoliposomal crocin formulations for the treatment of rheumatoid arthritis. Three crocin-loaded formulations including hydrogenated soy phosphatidylcholine (HSPC), 1,2- distearoyl-sn-glycero-3-phospho-(1′-rac-glycerol) (DSPG) and cholesterol (Chol) were made at different molar ratios using a solvent evaporation method plus extrusion. Two formulations contained methoxy-polyethylene glycol (MW 2000)-distearoylphosphatidylethanolamine. Based on the characterization tests, the crocin encapsulation efficiency of the formulations was between 85.3% and 95.6%, and the crocin release of the formulations was less than 40%, with the stability of about 12 weeks at 4 °C storage. To evaluate the anti-arthritic activity of nanoliposomal formulations, an adjuvant-induced arthritis (AIA) model was used. Rats were randomly divided into six groups. Changes in paw edema, biochemical parameters, and histopathology were then evaluated. The results showed that F1 nanoliposomes reduced serum levels of TNFα, IL1β, and IL17 cytokines, biochemical parameters (ALP, ALT, and AST), and paw edema in comparison to the AIA rats in the control groups. In conclusion, crocin-loaded liposomes can serve as a favorable agent in enhancing the anti-arthritic efficacy of crocin by decreasing the symptoms and controlling proinflammatory molecules.

RvD1 disrupts nociceptor neuron and macrophage activation and neuroimmune communication, reducing pain and inflammation in gouty arthritis in mice.

Gouty arthritis is characterized by an intense inflammatory response to monosodium urate crystals (MSU), which induces severe pain. Current therapies are often ineffective in reducing gout-related pain. Resolvin D1 (RvD1) is a specialized pro-resolving lipid mediator with anti-inflammatory and analgesic proprieties. In this study, we evaluated the effects and mechanisms of action of RvD1 in an experimental mouse model of gouty arthritis, an aim that was not pursued previously in the literature. Male mice were treated with RvD1 (intrathecally or intraperitoneally) before or after intraarticular stimulation with MSU. Mechanical hyperalgesia was assessed using an electronic von Frey aesthesiometer. Leukocyte recruitment was determined by knee joint wash cell counting and immunofluorescence. IL-1β production was measured by ELISA. Phosphorylated NF-kB and apoptosis-associated speck-like protein containing CARD (ASC) were detected by immunofluorescence, and mRNA expression was determined by RT-qPCR. CGRP release was determined by EIA and immunofluorescence. MSU crystal phagocytosis was evaluated by confocal microscopy. RvD1 inhibited MSU-induced mechanical hyperalgesia in a dose- and time-dependent manner by reducing leukocyte recruitment and IL-1β production in the knee joint. Intrathecal RvD1 reduced the activation of peptidergic neurons and macrophages as well as silenced nociceptor to macrophage communication and macrophage function. CGRP stimulated MSU phagocytosis and IL-1β production by macrophages. RvD1 downmodulated this phenomenon directly by acting on macrophages, and indirectly by inhibiting CGRP release and CGRP-dependent activation of macrophages. This study reveals a hitherto unknown neuro-immune axis in gouty arthritis that is targeted by RvD1.

Experimental model of periodontitis and rheumatoid arthritis potentiates the deleterious effects on functional capacity, leukocyte migration, synovial and periodontal tissues in Wistar rats

Objective This study aimed to evaluate whether ligature-induced periodontitis and rheumatoid arthritis (RA) potentiate the deleterious effects on functional capacity, periodontal and synovial tissues, leukocyte migration, and interleukin 17 (IL-17) levels, and to investigate the repercussions of single Freund’s Complete Adjuvant (FCA) injection associated with periodontitis. Materials and methods Fifty-one male Wistar rats were randomised into six groups: control (CG, n = 8), RA (RAG, n = 9), periodontitis (PG, n = 9), periodontitis and RA (PRAG, n = 9), periodontitis and intradermal injection (PIDG, n = 9), and periodontitis and intra-articular injection (PIAG, n = 7). The animals underwent ligature placement and one or two injections with FCA to induce RA. Motor disability, nociceptive threshold, joint edema, and muscle strength were assessed, and the animals were euthanized on day 30. Synovial fluid, hemimandibles, and knee joints were collected. Results PRAG showed no reduction of edema or improvement of muscle strength, whereas it showed most significant changes in leukocyte migration, morphological analyses of the synovial membrane (SM), and radiographic and histometric analyses of the jaw. The PIAG showed some alterations, though not permanent. Conclusion Ligature-induced periodontitis and RA induced by two FCA injections accentuated the deleterious effects on functional capacity, leukocyte migration, synovial and periodontal tissues.

AB0064 PHARMACOLOGICAL INHIBITION OF IL-6/JAK/STAT AXIS INCREASES MUSCLE MASS IN AN EXPERIMENTAL MODEL OF SARCOPENIA ASSOCIATED TO RHEUMATOID ARTHRITIS

BackgroundSarcopenia is a frequent comorbidity of rheumatoid arthritis (RA) due to an imbalance in muscle remodelling with dominance of catabolism over anabolism. IL-6/JAK/STAT axis is of particular importance in this regulation because of its dual activity. According to local and acute factors, it promotes muscle healing and hypertrophy, whereas in chronic inflammation, this signalling substantially increases muscle catabolism activating pro-atrophic pathways involving atrogene expression and ubiquitin proteasome system (UPS). JAK/STAT blockade suppresses muscle wasting induced by IL-6, through STAT3 activation. Interestingly, different clinical trials have shown that JAK inhibitors produce an asymptomatic increase in serum creatine kinase (CK) and creatinine levels in RA patients, suggesting an impact on muscle.ObjectivesTo evaluate the effect of JAK inhibition and its effect in muscle remodelling in an experimental model that accurately mimics human rheumatoid sarcopenia.MethodsAn experimental model of antigen-induced arthritis (AIA) was carried out in 14 rabbits by immunization against ovalbumin followed by 4 intra-articular (i.a) injections of this protein. One week after the first i.a injection, 7 of these rabbits received tofacitinib (TOFA, orally 10mg/kg/day) for 2 wk. Animals were euthanized one day after the last i.a injection, when tibialis anterior (TA), extensor digitorum longus (EDL) and gastrocnemius (GN) were isolated from both hind limbs. Histological changes and ATPase staining were analysed in TA, while the number of myonuclei was evaluated in EDL fibres by immunofluorescence studies. C-reactive protein (CRP) and myostatin (MSTN) serum concentration were determined by ELISA. The gene expression of proinflammatory cytokines (IL-1β, IL-6, TNF-α, MCP-1), atrogenes (Atrogin-1, MuRF-1) and MSTN was measured by quantitative PCR, while proliferative muscle marker (PAX-7), differentiation muscle markers (MyoD, Myogenin), pSTAT3, pSTAT1, MSTN and CK protein expression was analysed by western blot in GN. Creatine presence in GN was analysed by a colorimetric assay.ResultsA significant increase in body weight was observed in the AIA+TOFA group vs. AIA at the end of the study. A significant increase in TA cross-sectional area and diameter in the AIA+TOFA, in comparison to AIA, was found, while the decrease in the area of type II fibres observed in the AIA animals was not noticed in AIA+TOFA. Moreover, the number of myonuclei in the EDL that was increased in the AIA group when compared to healthy animals, was significantly decreased in AIA+TOFA. However, systemic inflammation measured by CRP was not modified by TOFA treatment at the end of the study, in comparison to AIA, and so did serum MSTN concentration. TOFA evoked a significant reduction in the gene expression of IL-6, MCP-1, atrogin-1 and MuRF-1 in GN in comparison to untreated AIA rabbits. Notably, AIA+TOFA showed higher protein levels of CK and lower creatine compared to AIA in muscle. Simultaneously, no differences in the proliferative marker PAX-7 were found between groups, while AIA rabbits showed an increase in the differentiation markers MyoD and Myogenin in the muscle that was prevented in AIA+TOFA.ConclusionThese data provide novel insights into the effects of JAK inhibitors in muscle remodelling during rheumatoid cachexia. Despite an elevated systemic inflammatory status, JAK inhibition was able to rapidly increase muscle mass through attenuating IL-6/JAK/STAT activation, decreasing atrogene expression, and restoring to baseline the muscle cell differentiation markers in the tissue. The increase in muscle mass was accompanied by an increase in CK presence, supporting the role of CK as a valuable marker of muscle gain following treatment with JAK inhibitors. However, the low levels of MCP-1 and the disorganization of muscle fibre myonuclei raise the question of whether these treatments could trigger a complete regeneration of muscle fibre cell or just hypertrophy of cells already present.Disclosure of InterestsIsmael Bermejo: None declared, Sandra Pérez-Baos: None declared, Juan Pablo Medina: None declared, Paula Gratal: None declared, Ivan Prieto-Potín: None declared, Raquel Largo: None declared, Gabriel Herrero-Beaumont Grant/research support from: The Bone and Joint Research Unit has received funding from several pharmaceutical laboratories (Pfizer, LILLY...).

Physical Exercise and Low-Level Laser Therapy Systemic Effects on the Ankle Joint in an Experimental Rheumatoid Arthritis Model

ObjectiveThe purpose of this study was to analyze the systemic effects of stair climbing exercise, low-level laser therapy (LLLT), and the association of both treatments on pelvic member functionality and ankle joint histomorphometric aspects of Wistar rats submitted to experimental rheumatoid arthritis (RA) protocol. MethodsMale Wistar rats were randomly distributed into the following 8 groups: control; control LLLT; control exercise; control LLLT and exercise; arthritis group; arthritis LLLT; arthritis exercise; and arthritis LLLT and exercise, with n = 8 for functional and n = 5 for histomorphometrical tests. The experimental RA was induced by complete Freund adjuvant injection in the knee joint cavity. Functionality was evaluated by proprioception and motor function using Sciatic Functional Index and maximum angle reached at an inclined plane. Histomorphometrical aspects were evaluated in the ankle joint after histological routine. ResultsThe arthritis LLLT and exercise group had positive effects in Sciatic Functional Index (F [3.96] = 11.3, P < .001) and in inclined plane (F [3.4] = 36.1, P < .001). The arthritis exercise group presented a greater number of chondrocytes in the tibia (Wald [1; 6605.6] = 25.2, P < .001) and talus (Wald [1; 15958.6] = 19.8, P = .006) in relation to the other groups. The arthritis group morphology showed significant degenerative lesions as subintima with angiogenesis, inflammatory cells, flocculated articular cartilage, chondrocytes disorganization and pannus. Even with the higher chondrocytes number, the arthritis exercise group had morphological characteristics more similar to the control group. ConclusionLow-level laser therapy and exercise restored functionality, and exercise restored morphological aspects of tissues in experimentally induced RA in rats.

Antibodies to Cartilage Oligomeric Matrix Protein Are Pathogenic in Mice and May Be Clinically Relevant in Rheumatoid Arthritis

ObjectiveCartilage oligomeric matrix protein (COMP) is an autoantigen in rheumatoid arthritis (RA) and experimental models of arthritis. This study was undertaken to investigate the structure, function, and relevance of anti‐COMP antibodies.MethodsWe investigated the pathogenicity of monoclonal anti‐COMP antibodies in mice using passive transfer experiments, and we explored the interaction of anti‐COMP antibodies with cartilage using immunohistochemical staining. The interaction of the monoclonal antibody 15A11 in complex with its specific COMP epitope P6 was determined by x‐ray crystallography. An enzyme‐linked immunosorbent assay and a surface plasma resonance technique were used to study the modulation of calcium ion binding to 15A11. The clinical relevance and value of serum IgG specific to the COMP P6 epitope and its citrullinated variants were evaluated in a large Swedish cohort of RA patients.ResultsThe murine monoclonal anti‐COMP antibody 15A11 induced arthritis in naive mice. The crystal structure of the 15A11–P6 complex explained how the antibody could bind to COMP, which can be modulated by calcium ions. Moreover, serum IgG specific to the COMP P6 peptide and its citrullinated variants was detectable at significantly higher levels in RA patients compared to healthy controls and correlated with a higher disease activity score.ConclusionOur findings provide the structural basis for binding a pathogenic anti‐COMP antibody to cartilage. The recognized epitope can be citrullinated, and levels of antibodies to this epitope are elevated in RA patients and correlate with higher disease activity, implicating a pathogenic role of anti‐COMP antibodies in a subset of RA patients.

Role of Lysine-Specific Demethylase 1 in Metabolically Integrating Osteoclast Differentiation and Inflammatory Bone Resorption Through Hypoxia-Inducible Factor 1α and E2F1.

Hypoxia occurs in tumors, infections, and sites of inflammation, such as in the affected joints of patients with rheumatoid arthritis (RA). It alleviates inflammatory responses and increases bone resorption in inflammatory arthritis by enhancing osteoclastogenesis. The mechanism by which the hypoxia response is linked to osteoclastogenesis and inflammatory bone resorption is unclear. This study was undertaken to evaluate whether the protein lysine-specific demethylase 1 (LSD1) metabolically integrates inflammatory osteoclastogenesis and bone resorption in a state of inflammatory arthritis. LSD1-specific inhibitors and gene silencing with small interfering RNAs were used to inhibit the expression of LSD1 in human osteoclast precursor cells derived from CD14-positive monocytes, with subsequent assessment by RNA-sequencing analysis. In experimental mouse models of arthritis, inflammatory osteolysis, or osteoporosis, features of accelerated bone loss and inflammatory osteolysis were analyzed. Furthermore, in blood samples from patients with RA, cis-acting expression quantitative trait loci (cis-eQTL) were analyzed for association with the expression of hypoxia-inducible factor 1α (HIF-1α), and associations between HIF-1α allelic variants and extent of bone erosion were evaluated. In human osteoclast precursor cells, RANKL induced the expression of LSD1 in a mechanistic target of rapamycin-dependent manner. Expression of LSD1 was higher in synovium from RA patients than in synovium from osteoarthritis patients. Inhibition of LSD1 in human osteoclast precursors suppressed osteoclast differentiation. Results of transcriptome analysis identified several LSD1-mediated hypoxia and cell-cycle pathways as key genetic pathways involved in human osteoclastogenesis. Furthermore, HIF-1α protein, which is rapidly degraded by the proteasome in a normoxic environment, was found to be expressed in RANKL-stimulated osteoclast precursor cells. Induction of LSD1 by RANKL stabilized the expression of HIF-1α protein, thereby promoting glycolysis, in conjunction with up-regulation of the transcription factor E2F1. Analyses of cis-eQTL revealed that higher HIF-1α expression was associated with increased bone erosion in patients with RA. Inhibition of LSD1 decreased pathologic bone resorption in mice, both in models of accelerated osteoporosis and models of arthritis and inflammatory osteolysis. LSD1 metabolically regulates osteoclastogenesis in an energy-demanding inflammatory environment. These findings provide potential new therapeutic strategies targeting osteoclasts in the management of inflammatory arthritis, including in patients with RA.

Evaluation of Antiarthritic and Antinociceptive Effects of Cedrol in a Rat Model of Arthritis.

Pharmacological studies revealed that cedrol, a natural sesquiterpene, has antioxidant, anti-inflammatory, and analgesic properties. This study is aimed at evaluating the potential antiarthritic activity of cedrol in a rat experimental model of arthritis induced by using complete Freund's adjuvant (CFA). Arthritis was induced in Wistar rats by CFA (0.1 ml) injection. Cedrol (10 and 20 mg/kg) and indomethacin (5 mg/kg) were orally administered from day one and continued for 21 days. The antiarthritic activity was assessed through mechanical allodynia and thermal hyperalgesia responses, paw edema assessment, and arthritis scores. Serum TNF-α and IL-1β levels were measured for the evaluation of inflammation. Furthermore, serum oxidative stress markers, including malondialdehyde (MDA) and thiol levels, as well as superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities, were also assessed. Oral administration of cedrol and indomethacin significantly decreased paw edema and arthritis score. Besides, cedrol and indomethacin significantly decreased pain responses. In the serum of the CFA group, TNF-α, IL-1β, and MDA were higher, while thiol and SOD and GPx were lower than the control group. Treatment by cedrol and indomethacin corrected the biochemical parameters in the serum. In this study, cedrol offers potential antiarthritic properties through its anti-inflammatory and antioxidant effects.

Toxic Effects of Endocrine Disruptor Exposure on Collagen-Induced Arthritis.

Endocrine disruptors (EDs) are chemical substances capable of affecting endocrine system functioning and interfering with organ morphogenesis and physiological functions. The development and regeneration of bone tissues have a complex hormonal regulation, and therefore, bone tissue cells can be considered potential targets for endocrine disruptors. In that regard, the aim of this research was to investigate the impact of ED exposure on the inflammatory response and oxidative stress in an experimental model of collagen-induced arthritis (CIA). Arthritis was induced by an emulsion of type II collagen (CII) and complete Freund’s adjuvant, which was administered intradermally on days 0 and 21. Mice from day 21 to day 35 received the following EDs by oral gavage: cypermethrin (CP), diethyl phthalate (DEP), vinclozolin (VCZ), 17α-ethinylestradiol (EE), perfluorooctanesulfonic acid (PFOS) and atrazine (ATR). ED exposure caused worsening of clinical signs (erythema and edema in the hind paws), histological and radiographic changes, as well as behavioral deficits, induced by CII injections. Furthermore, ED exposure significantly increased the degree of inflammation and oxidative damage induced by arthritis; this upregulation was more evident after exposure to ATR than to other EDs. The results from our study suggest that exposure to EDs may play a deleterious role in the progression of RA; therefore, exposure to EDs should be limited.

Synergistic Effect of L-Carnosine and Hyaluronic Acid in Their Covalent Conjugates on the Antioxidant Abilities and the Mutual Defense against Enzymatic Degradation.

Hyaluronic acid (Hy) is a natural linear polymer that is widely distributed in different organisms, especially in the articular cartilage and the synovial fluid. During tissue injury due to oxidative stress, Hy plays an important protective role. All the beneficial properties of Hy make the polymer attractive for many biomedical uses; however, the low stability and short biological half-life limit Hy application. To overcome these problems, the addition of small antioxidant molecules to Hy solution has been employed to protect the molecular integrity of Hy or delay its degradation. Carnosine (β-alanyl-L-histidine, Car) protects cells from the damage due to the reactive species derived from oxygen (ROS), nitrogen (RNS) or carbonyl groups (RCS). Car inhibits the degradation of hyaluronan induced by free radical processes in vitro but, like Hy, the potential protective action of Car is drastically hampered by the enzymatic hydrolysis in vivo. Recently, we conjugated Hy to Car and the derivatives (HyCar) showed protective effects in experimental models of osteoarthritis and rheumatoid arthritis in vivo. Here we report the antioxidant activity exerted by HyCar against ROS, RNS and RCS. Moreover, we tested if the covalent conjugation between Hy and Car inhibits the enzymatic hydrolysis of the polymer and the dipeptide backbone. We found that the antioxidant properties and the resistance to the enzymatic hydrolysis of Hy and Car are greatly improved by the conjugation.

Discovery proteomics for the detection of putative markers for eradication of infection in an experimental model of equine septic arthritis using LC-MS/MS

Septic arthritis (SA) is a life-threatening condition in horses, and identifying eradication of infection in equine SA is challenging. This study explored the discovery of putative biomarkers for the eradication of joint infection in horses. We performed proteomics analysis of synovial fluid (SF) and plasma from horses with experimental SA, non-septic lipopolysaccharide-induced arthritis, and controls. The point of eradication of infection in horses with SA was determined previously. We compared spectral intensities between groups as well as before and after the eradication of infection. Twenty-six differentially abundant proteins were identified, which were upregulated in SF of horses with SA compared to the other groups, as well as compared to the same horses post-eradication of infection. In plasma, we did not identify differentially abundant proteins. Differentially abundant proteins in SF were of cellular origin and their biological functions included ubiquitination, signal transduction, apoptosis etc. The difference in their relative abundance between experimental groups was ≥10-fold compared to the abundance expected based on the difference in cell count alone (2-fold). Since most of cells in joints with bacterial infection are neutrophils, we suggest that the variable abundance of neutrophil- and cell-associated proteins represent potential biomarkers of eradication of infection in equine SA. SignificanceSeptic arthritis is an important condition in horses, which can be life-threatening. At present, identifying eradication of infection in cases of equine septic arthritis is challenging. In this study, we performed a global proteomics analysis of synovial fluid and plasma in horses with experimental septic arthritis and identified 26 differentially abundant proteins compared to non-septic arthritis and post eradication of infection. The results of this study provide the basis for further characterization of the differentially abundant proteins and identification of clinically relevant biomarkers of septic arthritis in horses.

CdSe magic-sized quantum dots attenuate reactive oxygen species generated by neutrophils and macrophages with implications in experimental arthritis

The biological applicability of nanomaterials has been limited due to cytotoxicity. Studies have described the effects of nanomaterials on different tissues and cell types, but their actions on immune cells are less elucidated. This study describes unprecedented in vitro and in vivo antioxidant activities of cadmium selenide magic-sized quantum dots (CdSe MSQDs) with implications on rheumatoid arthritis. While the generation of ROS induced by nanomaterials is linked to cytotoxicity, we found that CdSe MSQDs reduced ROS production by neutrophils and macrophages following opsonized-zymosan stimuli, and we did not find cytotoxic effects. Interestingly, inherent antioxidant properties of CdSe MSQDs were confirmed through DPPH, FRAP, and ORAC assays. Furthermore, CdSe MSQDs reduced ROS levels generated by infiltrating leukocytes into joints in experimental model of rheumatoid arthritis. Briefly, we describe a novel application of CdSe MSQDs in modulating the inflammatory response in experimental rheumatoid arthritis through an unexpected antioxidant activity.

Methotrexate-loaded nanoparticles ameliorate experimental model of autoimmune arthritis by regulating the balance of interleukin-17-producing T cells and regulatory T cells

BackgroundRheumatoid arthritis (RA) is a progressive systemic autoimmune disease that is characterized by infiltration of inflammatory cells into the hyperplastic synovial tissue, resulting in subsequent destruction of adjacent articular cartilage and bone. Methotrexate (MTX), the first conventional disease-modifying antirheumatic drug (DMARD), could alleviate articular damage in RA and is implicated in humoral and cellular immune responses. However, MTX has several side effects, so efficient delivery of low-dose MTX is important.MethodsTo investigate the efficacy of MTX-loaded nanoparticles (MTX-NPs) against experimental model of RA, free MTX or MTX-NPs were administered as subcutaneous route to mice with collagen-induced arthritis (CIA) at 3 weeks after CII immunization. The levels of inflammatory factors in tissues were determined by immunohistochemistry, confocal microscopy, real-time PCR, and flow cytometry.ResultsMTX-NPs ameliorated arthritic severity and joint destruction in collagen-induced arthritis (CIA) mice compared to free MTX-treated CIA mice. The levels of inflammatory cytokines, including interleukin (IL)-1β, tumor necrosis factor-α, and vascular endothelial growth factor, were reduced in MTX-NPs-treated mice. Number of CD4 + IL-17 + cells decreased whereas the number of CD4 + CD25 + Foxp3 + cells increased in spleens from MTX- NPs-treated CIA mice compared to MTX-treated CIA mice. The frequency of CD19 + CD25 + Foxp3 + regulatory B cells increased in ex vivo splenocytes from MTX-loaded NPs-treated CIA mice compared to MTX-treated CIA mice.ConclusionThe results suggest that MTX-loaded NPs have therapeutic potential for RA.