Experimental Inflammatory Arthritis Research Articles

IĸBζ as a Central Modulator of Inflammatory Arthritis Pathogenesis.

Current therapies targeting individual factors in inflammatory arthritis show variable efficacy, often requiring treatment with combinations of drugs, and are associated with undesirable side effects. NF-ĸB is critical for the production and function of most inflammatory cytokines. However, given its essential role in physiologic processes, targeting NF-ĸB is precarious. Hence, identifying pathways downstream of NF-ĸB that selectively govern the expression of inflammatory cytokines in inflammatory arthritis would be advantageous. We have previously identified IĸBζ as a unique inflammatory signature of NF-ĸB that controls the transcription of inflammatory cytokines only under pathologic conditions while sparing physiologic NF-ĸB signals. We generated mice harboring myeloid, lymphoid, and global deletion of Nfkbiz (the gene encoding IĸBζ). These models were subjected to serum transfer-induced arthritis. Additionally, pharmacologic inhibitors of IĸBζ were injected intraperitonially. Joint swelling, microcomputed tomography, immunohistochemistry, flow cytometry, and cytokine measurements were conducted using synovial tissue samples. Global deletion of Nfkbiz or depletion of neutrophils (vastly IĸBζ+ cells) reduced inflammatory synovial cells and increased anti-inflammatory and regenerative synovial cells, plummeted expression of inflammatory factors and ameliorated experimental mouse inflammatory arthritis. Further, expression of immune responsive gene-1, the enzyme responsible for itaconate production, was increased in synovial cells. Accordingly, the itaconate derivative dimethyl itaconate (DI) inhibited IĸBζ-mediated inflammatory factors. Further, in silico screen identified 8-hydroxyquinoline (HQ) as a putative inhibitor of IĸBζ not affecting physiologic NF-ĸB activity. Congruently, systemic administration of either DI or HQ inhibited joint swelling and damage. Our study positions IĸBζ as an inflammation-specific target for therapeutic consideration in rheumatoid arthritis because its inhibition spares the beneficial functions of NF-ĸB.

Sinapic acid-pullulan based inflammation responsive nanomicelles for the local treatment of experimental inflammatory arthritis

Rheumatoid arthritis (RA) is a chronic inflammatory disorder of joints. It is one of the major causes of disability and morbidity worldwide. Administration of conventional drugs through the systemic route restricts the bioavailability of drugs, systemic toxicity, and reduced efficacy. We have introduced Rebamipide (Reb)-loaded Sinapic acid (SA)-Pullulan (PL) nanomicelles (Reb@SA-PL NMs), a nanotechnology based drug delivery system for the treatment of inflammatory arthritis. PL is a polysaccharide obtained from the fungus Aureobasidium pullulans, and SA is a bioactive polyphenol found in various plants. Both are classified by US-FDA Generally Recognised as Safe (GRAS) materials. Reb@SA-PL NMs found to be cytocompatible. Subsequently, intra-articular administration of Reb@SA-PL NMs enhances the anti-arthritic potential compared to free Reb drug in collagen-induced experimental inflammatory arthritis rat model. Reb@SA-PL NMs reduced the expression of RANKL receptor and Nf-κB. Reb@SA-PL NMs reverses the breakdown of type II collagen, MMP-13, and inhibits the pro-inflammatory markers. Reb@SA-PL NMs prevented bone erosion, cartilage degradation, joint oedema, and synovial inflammation. The results of the study demonstrated that Reb@SA-PL NMs, an enzyme-responsive drug delivery system, has excellent potential for alleviating inflammatory arthritis by blocking MMP-13 and RANKL.

Molecular Determinants of Neutrophil Extracellular Vesicles That Drive Cartilage Regeneration in Inflammatory Arthritis.

This study was undertaken to establish the potential therapeutic profile of neutrophil-derived extracellular vesicles (EVs) in experimental inflammatory arthritis and associate pharmacological activity with specific EV components, focusing on microRNAs. Neutrophil EVs were administered intra-articularly through a prophylactic or therapeutic protocol to male C57BL/6 mice undergoing serum-transfer-induced inflammatory arthritis. Transcriptomic analysis of knees was performed on joints following EV administration, naive and arthritic mice (untreated; n = 4/group) and EV-treated diseased mice (intra-articular administration) with contralateral (vehicle-treated; n = 8/group). Comparison of healthy donor and patients with rheumatoid arthritis (RA) neutrophil EVs was performed. EVs afforded cartilage protection with an increase in collagen-II and reduced collagen-X expression within the joint. To gain mechanistic insights, RNA sequencing of the arthritic joints was conducted. A total of 5,231 genes were differentially expressed (P < 0.05), with 257 unique to EV treatment. EVs affected key regenerative pathways involved in joint development, including Wnt and Notch signaling. This wealth of genomic alteration prompted to identify microRNAs in EVs, 10 of which are associated with RA. As a proof of concept, we focused on miR-455-3p, which was detected in both healthy donor and RA EVs. EV addition to chondrocyte cultures elevated miR-455-3p and exerted anticatabolic effects upon interleukin-1β stimulation; these effects were blocked by actinomycin or miR-455-3p antagomir. Neutrophils from patients with RA yielded EVs with composition, efficacy, and miR-455-3p content similar to those of healthy volunteers, suggesting that neutrophil EVs could be developed as an autologous treatment to protect and repair joint tissue of patients affected by inflammatory arthritides.

Endothelial cell sphingosine 1-phosphate receptor 1 restrains VE-cadherin cleavage and attenuates experimental inflammatory arthritis.

In rheumatoid arthritis, inflammatory mediators extravasate from blood into joints via gaps between endothelial cells (ECs), but the contribution of ECs is not known. Sphingosine 1-phosphate receptor 1 (S1PR1), widely expressed on ECs, maintains the vascular barrier. Here, we assessed the contribution of vascular integrity and EC S1PR1 signaling to joint damage in mice exposed to serum-induced arthritis (SIA). EC-specific deletion of S1PR1 or pharmacological blockade of S1PR1 promoted vascular leak and amplified SIA, whereas overexpression of EC S1PR1 or treatment with an S1PR1 agonist delayed SIA. Blockade of EC S1PR1 induced membrane metalloproteinase-dependent cleavage of vascular endothelial cadherin (VE-cadherin), a principal adhesion molecule that maintains EC junctional integrity. We identified a disintegrin and a metalloproteinase domain 10 (ADAM10) as the principal VE-cadherin "sheddase." Mice expressing a stabilized VE-cadherin construct had decreased extravascular VE-cadherin and vascular leakage in response to S1PR1 blockade, and they were protected from SIA. Importantly, patients with active rheumatoid arthritis had decreased circulating S1P and microvascular expression of S1PR1, suggesting a dysregulated S1P/S1PR1 axis favoring vascular permeability and vulnerability. We present a model in which EC S1PR1 signaling maintains homeostatic vascular barrier function by limiting VE-cadherin shedding mediated by ADAM10 and suggest this signaling axis as a therapeutic target in inflammatory arthritis.

Macrophage-derived human resistin promotes perivascular adipose tissue dysfunction in experimental inflammatory arthritis

Cardiovascular disease (CVD) is the leading cause of death in rheumatoid arthritis (RA). Resistin is an adipokine that induces adipose tissue inflammation and activation of monocytes/macrophages via adenylate cyclase-associated protein-1 (CAP1). Resistin levels are increased in RA and might cause perivascular adipose tissue (PVAT) dysfunction, leading to vascular damage and CVD. This study aimed to investigate the role of resistin in promoting PVAT dysfunction by increasing local macrophage and inflammatory cytokines content in antigen-induced arthritis (AIA). Resistin pharmacological effects were assessed by using C57Bl/6J wild-type (WT) mice, humanized resistin mice expressing human resistin in monocytes-macrophages (hRTN+/-/-), and resistin knockout mice (RTN-/-) with AIA and respective controls. We investigated AIA disease activity and functional, cellular, and molecular parameters of the PVAT. Resistin did not contribute to AIA disease activity and its concentrations were augmented in the PVAT and plasma of WT AIA and hRTN+/-/- AIA animals. In vitro exposure of murine arteries to resistin impaired vascular function by decreasing the anti-contractile effect of PVAT. WT AIA mice and hRTN+/-/- AIA mice exhibited PVAT dysfunction and knockdown of resistin prevented it. Macrophage-derived cytokines, markers of types 1 and 2 macrophages, and CAP1 expression were increased in the PVAT of resistin humanized mice with AIA, but not in knockout mice for resistin. This study reveals that macrophage-derived resistin promotes PVAT inflammation and dysfunction regardless of AIA disease activity. Resistin might represent a translational target to reduce RA-driven vascular dysfunction and CVD.

A bioactive and biodegradable vitamin C stearate-based injectable hydrogel alleviates experimental inflammatory arthritis.

Rheumatoid arthritis (RA) is a chronic autoimmune inflammatory joint disorder affecting nearly 1% of the global population. In RA, synovial joints are infiltrated by inflammatory mediators and enzymes, leading to articular cartilage deterioration, joint damage, and bone erosion. Herein, the 9-aminoacridine-6-O-stearoyl-L-ascorbic acid hydrogel (9AA-SAA hydrogel) was formulated by the heat-cool method and further characterized for surface charge, surface morphology, rheology, and cytocompatibility. Furthermore, we evaluated the therapeutic efficacy of the 9AA-SAA hydrogel, an enzyme-responsive drug delivery system with on-and-off switching capabilities based on disease severity against collagen-induced experimental arthritis in Wistar rats. The anti-inflammatory action of the US FDA-approved drug 9-aminoacridine (9AA) was revealed which acted through nuclear receptor subfamily 4 group A member 1 (NR4A1), an anti-inflammatory orphan nuclear receptor that inhibits nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB). Furthermore, we have explored the role of ascorbic acid, an active moiety of 6-O-stearoyl-L-ascorbic acid (SAA), in promoting the production of collagen production through ten-eleven translocation-2 (TET2) upregulation. Targeting through NR4A1 and TET2 could be the probable mechanism for the treatment of experimental arthritis. The combination of 9AA and ascorbic acid demonstrated enhanced therapeutic efficacy in the 9AA-SAA hydrogel, significantly reducing the severity of experimental arthritis. This approach, in contrast to existing treatments with limited effectiveness, presents a promising and more effective strategy for RA treatment by mitigating inflammation in experimental arthritis.

Biomaterial-based combinatorial approach of aescin-comprised zein-coated gelatin nanoparticles alleviates synovial inflammation in experimental inflammatory arthritis.

Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease that mostly affects joints. Although RA therapy has made significant progress, difficulties including extensive medication metabolism and its quick clearance result in its inadequate bioavailability. The anti-inflammatory effect of zein was reported with other medications, but it has certain limitations. There are reports on the anti-oxidant and anti-inflammatory effect of aescin, which exhibits low bioavailability for the treatment of rheumatoid arthritis. Also, the combinatorial effect of zein with other effective drug delivery systems is still under investigation for the treatment of experimental collagen-induced rheumatoid arthritis. The focus of this study was to formulate and define the characteristics of zein-coated gelatin nanoparticles encapsulated with aescin (Ze@Aes-GNPs) and to assess and contrast the therapeutic effectiveness of Ze@Aes-GNPs towards collagen-induced RA in Wistar rats. Nanoprecipitation and the layer-by-layer coating process were used to fabricate Ze@Aes-GNPs and their hydrodynamic diameter was determined to be 182 nm. Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) were used to further validate the size, shape, and surface morphology of Ze@Aes-GNPs. When tested against foreskin fibroblasts (BJ), these nanoparticles demonstrated significantly high cytocompatibility. Both Aes and Ze@Aes-GNPs were effective in treating arthritis, as shown by the decreased edoema, erythema, and swelling of the joints, between which Ze@Aes-GNPs were more effective. Further, it was demonstrated that Aes and Ze@Aes-GNPs reduced the levels of oxidative stress (articular elastase, lipid peroxidation, catalase, superoxide dismutase and nitric oxide) and inflammatory indicators (TNF-α, IL-1β and myeloperoxidase). The histopathology findings further demonstrated that Ze@Aes-GNPs considerably reduced the infiltration of inflammatory cells at the ankle joint cartilage compared to Aes. Additionally, immunohistochemistry examination showed that treatment with Ze@Aes-GNPs suppressed the expression of pro-inflammatory markers (COX-2 and IL-6) while increasing the expression of SOD1. In summary, the experiments indicated that Aes and Ze@Aes-GNPs lowered the severity of arthritis, and critically, Ze@Aes-GNPs showed better effectiveness in comparison to Aes. This suppression of oxidative stress and inflammation was likely driven by Aes and Ze@Aes-GNPs.

Tracking Cardiovascular Comorbidity in Models of Chronic Inflammatory Disease.

Immune-mediated inflammatory diseases (IMIDs) are commonly associated with complex coexisting conditions, and cardiovascular comorbidities are a common cause of mortality in systemic inflammation. Experimental models of disease provide an opportunity to dissect inflammatory mechanisms that promote damage to vascular tissues affected by comorbidity. Here, we describe methods to recover the thoracic aorta from mice during experimental inflammatory arthritis and assess vascular constriction responses by isometric tension myography. To complement the assessment of functional changes in the vasculature during inflammatory arthritis, we also outline a method to characterize vascular inflammation by immunohistochemistry.

Consequences of collagen induced inflammatory arthritis on circadian regulation of the gut microbiome.

The gut microbiota is important for host health and immune system function. Moreover autoimmune diseases, such as rheumatoid arthritis, are associated with significant gut microbiota dysbiosis, although the causes and consequences of this are not fully understood. It has become clear that the composition and metabolic outputs of the microbiome exhibit robust 24 h oscillations, a result of daily variation in timing of food intake as well as rhythmic circadian clock function in the gut. Here, we report that experimental inflammatory arthritis leads to a re-organization of circadian rhythmicity in both the gut and associated microbiome. Mice with collagen induced arthritis exhibited extensive changes in rhythmic gene expression in the colon, and reduced barrier integrity. Re-modeling of the host gut circadian transcriptome was accompanied by significant alteration of the microbiota, including widespread loss of rhythmicity in symbiont species of Lactobacillus, and alteration in circulating microbial derived factors, such as tryptophan metabolites, which are associated with maintenance of barrier function and immune cell populations within the gut. These findings highlight that altered circadian rhythmicity during inflammatory disease contributes to dysregulation of gut integrity and microbiome function.

Bioflavonoid Robinin from Astragalus falcatus Lam. Mildly Improves the Effect of Metothrexate in Rats with Adjuvant Arthritis.

Anti-inflammatory potential of orally administrated bioflavonoid-robinin, active sub-stance of original drug Flaroninum™ (FL), was investigated in the combination with methotrexate (MTX) and in monotherapy in rats suffering from adjuvant-induced arthritis (AA). Robinin (kaempferol-3-O-robinoside-7-O-rhamnoside) was isolated from the aerial parts of Astragalus falcatus Lam. The monotherapy with robinin was not efficient in alleviating symptoms of AA. The combination of MTX with robinin was similarly active as MTX alone in reducing the hind paw volume and change of body weight during the whole experiment. The combination, however, reduced plasma levels of Interleukin-17Aand activity of gamma-glutamyl transferase in joint more efficiently then MTX alone. Our results demonstrate that the novel combination of robinin and MTX mildly improved the reduction of inflammation in experimental arthritis.

NIRF-Molecular Imaging with Synovial Macrophages-Targeting Vsig4 Nanobody for Disease Monitoring in a Mouse Model of Arthritis.

Nanobody against V-set and Ig domain-containing 4 (Vsig4) on tissue macrophages, such as synovial macrophages, could visualize joint inflammation in multiple experimental arthritis models via single-photon emission computed tomography imaging. Here, we further addressed the specificity and assessed the potential for arthritis monitoring using near-infrared fluorescence (NIRF) Cy7-labeled Vsig4 nanobody (Cy7-Nb119). In vivo NIRF-imaging of collagen-induced arthritis (CIA) was performed using Cy7-Nb119. Signals obtained with Cy7-Nb119 or isotope control Cy7-NbBCII10 were compared in joints of naive mice versus CIA mice. In addition, pathological microscopy and fluorescence microscopy were used to validate the arthritis development in CIA. Cy7-Nb119 accumulated in inflamed joints of CIA mice, but not the naive mice. Development of symptoms in CIA was reflected in increased joint accumulation of Cy7-Nb119, which correlated with the conventional measurements of disease. Vsig4 is co-expressed with F4/80, indicating targeting of the increasing number of synovial macrophages associated with the severity of inflammation by the Vsig4 nanobody. NIRF imaging with Cy7-Nb119 allows specific assessment of inflammation in experimental arthritis and provides complementary information to clinical scoring for quantitative, non-invasive and economical monitoring of the pathological process. Nanobody labelled with fluorescence can also be used for ex vivo validation experiments using flow cytometry and fluorescence microscopy.

Galectins-1 and-3 Increase in Equine Post-traumatic Osteoarthritis.

Galectins are potent regulators of cell adhesion, growth and apoptosis in diverse cell types, including chondrocytes and synovial fibroblasts. Elevations in synovial fluid galectin-3 have been observed in rheumatoid arthritis, juvenile idiopathic arthritis and experimental inflammatory arthritis in animal models, whereas galectin-1 is thought to be protective. Less is known about galectins-1 and-3 in osteoarthritis (OA). Therefore, the purpose of this study was: (1) to determine whether galectin-1 and-3 synovial fluid concentrations and synovial membrane and cartilage histochemical staining were altered following osteochondral injury in an experimental equine osteoarthritis (OA) model and (2) to measure galectin-1 and-3 mRNA expression and synovial fluid concentrations in naturally occurring equine carpal OA. Synovial fluid galectin-1 and-3 concentrations were quantified using custom ELISAs in two research horse cohorts undergoing experimental OA induction (n = 5 and 4) and in a cohort of horses with naturally occurring carpal OA (n = 57). Galectin mRNA expression in synovial membrane and cartilage tissue obtained from carpal joints of horses with naturally occurring OA was measured using RT-qPCR, and galectin immunostaining was assessed in synovial membrane and osteochondral tissues in the experimental model (n = 5). Synovial fluid galectin-1 and-3 concentrations increased following experimental carpal osteochondral fragmentation. Cartilage galectin-1 mRNA expression increased with OA severity in naturally occurring disease. The superficial zone of healthy articular cartilage stained intensely for galectin-3 in sham-operated joints, whereas galectin-1 staining was nearly absent. Chondrocyte galectin-1 and-3 immunoreactivity increased following cartilage injury, particularly in galectin-1 positive chondrones. Galectins-1 and-3 are present in healthy equine synovial fluid and increase following post-traumatic OA. Healthy superficial zone chondrocytes express galectin-3, whereas galectin-1 chondrocyte staining is limited predominantly to chondrones and injured cartilage. Further work is needed to clarify the functions of galectins-1 and-3 in healthy and OA joints.

Brain Stimulation Differentially Modulates Nociception and Inflammation in Aversive and Non-aversive Behavioral Conditions

Inflammation and pain are major clinical burdens contributing to multiple disorders and limiting the quality of life of patients. We previously reported that brain electrical stimulation can attenuate joint inflammation in experimental arthritis. Here, we report that non-aversive electrical stimulation of the locus coeruleus (LC), the paraventricular hypothalamic nucleus (PVN) or the ventrolateral column of the periaqueductal gray matter (vlPAG) decreases thermal pain sensitivity, knee inflammation and synovial neutrophilic infiltration in rats with intra-articular zymosan. We also analyzed the modulation of pain and inflammation during aversive neuronal stimulation, which produces defensive behavioral responses such as freezing immobility to avoid predator detection. Electrical stimulation with higher intensity to induce freezing immobility in rats further reduces pain but not inflammation. However, tonic immobility further reduces pain, knee inflammation and synovial neutrophilic infiltration in guinea pigs. The duration of the tonic immobility increases the control of pain and inflammation. These results reveal survival behavioral and neuromodulatory mechanisms conserved in different species to control pain and inflammation in aversive life-threatening conditions. Our results also suggest that activation of the LC, PVN, or vlPAG by non-invasive methods, such as physical exercise, meditation, psychological interventions or placebo treatments may reduce pain and joint inflammation in arthritis without inducing motor or behavioral alterations.

Selective inhibition of Tfh cells by a small molecule inhibitor abrogates progression of experimental inflammatory arthritis

Abstract Background Rheumatoidarthritis (RA) is an inflammatory autoimmune disease characterized by T cell infiltration in the joints and autoantibody production. T follicular helper (Tfh) cells are a unique subset of CD4+ T cells regulating antibody production in B cell follicle. Our previous studies have showed that increased circulating Tfh cells were correlated with anti-CCP antibody titer and disease activity in active RA patients, indicating that Tfh cells may play an important role in RA pathogenesis. Here we investigate the therapeutic potential of a small molecule inhibitor targeting Tfh cells (SMI-Tfh) in mice with collagen-induced arthritis (CIA). Methods CIA was induced in DBA/1 mice by immunization with chicken type II collagen. Following the onset of clinical arthritis, mice were treated with SMI-Tfh (50mg/kg/day) for 10 days. Arthritis progression was monitored daily and recorded by the paw swollenness. Blood, spleen, and affected paws were collected at the end of the study. Tfh cells were examined and defined by CD4+CXCR5+ICOS+via flow cytometry and further confirmed by immunohistochemistry staining in spleen. Results Mice developed arthritis four weeks after immunization with type II collagen. Treatment with SMI-Tfh significantly reduced the disease progression/activity in mice with CIA. SMI-Tfh significantly inhibited the frequency of Tfh cells in spleen (P&amp;lt;0.01), but not the frequency of circulating Tfh cells in CIA mice (P&amp;gt;0.05). Conclusion The small molecule inhibitor SMI-Tfh selectively inhibits Tfh cells and abrogates progression/activity of inflammatory arthritis in CIA mouse model. Treatment with SMI-Tfh in CIA mice provides a potential strategy for joint protection and may be beneficial in RA patients.

Baroreflex stimulation attenuates central but not peripheral inflammation in conscious endotoxemic rats

We previously reported that activation of the baroreflex, a critical physiological mechanism controlling cardiovascular homeostasis, through electrical stimulation of the aortic depressor nerve attenuates joint inflammation in experimental arthritis. However, it is unknown whether baroreflex activation can control systemic inflammation. Here, we investigate whether baroreflex activation controls systemic inflammation in conscious endotoxemic rats. Animals underwent sham or electrical aortic depressor nerve stimulation initiated 10 min prior to a lipopolysaccharide (LPS) challenge, while inflammatory cytokine levels were measured in the blood, spleen, heart and hypothalamus 90 min after LPS treatment. Baroreflex activation did not affect LPS-induced levels of pro-inflammatory (tumor necrosis factor, interleukin 1β and interleukin 6) or anti-inflammatory (interleukin 10) cytokines in the periphery (heart, spleen and blood). However, baroreflex stimulation attenuated LPS-induced levels of all these cytokines in the hypothalamus. Notably, these results indicate that the central anti-inflammatory mechanism induced by baroreflex stimulation is independent of cardiovascular alterations, since aortic depressor nerve stimulation that failed to induce hemodynamic changes was also efficient at inhibiting inflammatory cytokines in the hypothalamus. Thus, aortic depressor nerve stimulation might represent a novel therapeutic strategy for neuroprotection, modulating inflammation in the central nervous system.

Inhibition of spinal p38 MAPK prevents articular neutrophil infiltration in experimental arthritis via sympathetic activation.

The central nervous system controls the innate immunity by modulating efferent neuronal networks. Recently, we have reported that central brain stimulation inhibits inflammatory responses. In the present study, we investigate whether spinal p38 mitogen-activated protein kinase (MAPK) affects joint inflammation in experimental arthritis. Firstly, we observed that intra-articular administration of zymosan in mice induces the phosphorylation of the spinal cord p38 MAPK. In addition, we demonstrated that spinal p38 MAPK inhibition with intrathecal injection of SB203580, a conventional and well-characterized inhibitor, prevents knee joint neutrophil recruitment, edema formation, experimental score and cytokine production. This local anti-inflammatory effect was completely abolished with chemical sympathectomy (guanethidine) and beta-adrenergic receptors blockade (nadolol). In conclusion, our results suggest that pharmacological strategies involving the modulation of spinal p38 MAPK circuit can prevent joint inflammation via sympathetic networks and beta-adrenoceptors activation.

PEGylated TRAIL ameliorates experimental inflammatory arthritis by regulation of Th17 cells and regulatory T cells

TNF-related apoptosis-inducing ligand (TRAIL) is a death ligand that can induce apoptosis in cells expressing its cognate death receptors (DRs). Previously, we demonstrated the therapeutic potential of recombinant human TRAIL in experimental rheumatoid arthritis (RA) models. However, the mechanisms of how DR-mediated apoptosis elicits these actions is not known. Here, we show that systemically administering a potent, long-acting PEGylated TRAIL (TRAILPEG) is profoundly anti-rheumatic against two complementary experimental RA mouse models, collagen-induced arthritis (CIA) and collagen antibody-induced arthritis (CAIA), via targeting IL-17 secreting Th17 cells and regulatory T cells (Treg). Systemic administration of TRAILPEG after disease onset ameliorated the severity of inflammatory arthritis including arthritis indices, paw thickness, cartilage damage and neutrophil infiltration in both CIA and CAIA models. Additionally, the levels of inflammatory molecules (p-p65, ICAM-1, Cox-2, MMP3, and iNOS), pro-inflammatory cytokines (TNF-α, IL-1β, IFN-γ, IL-6, IL-17) and accumulation of activated macrophages were significantly reduced after the TRAILPEG treatment. Importantly, TRAILPEG decreased the number of pro-inflammatory Th17 cells in inflamed arthritic joints through TRAIL-induced apoptosis while increasing anti-inflammatory Treg population in vivo. These results suggest that TRAILPEG ameliorates autoimmunity by targeting the Th 17-Tregs axis, making it a promising candidate drug for the treatment of RA.

Deconstructing the Lectin Pathway in the Pathogenesis of Experimental Inflammatory Arthritis: Essential Role of the Lectin Ficolin B and Mannose-Binding Protein-Associated Serine Protease 2.

Complement plays an important role in the pathogenesis of rheumatoid arthritis. Although the alternative pathway (AP) is known to play a key pathogenic role in models of rheumatoid arthritis, the importance of the lectin pathway (LP) pattern recognition molecules such as ficolin (FCN) A, FCN B, and collectin (CL)-11, as well as the activating enzyme mannose-binding lectin-associated serine protease-2 (MASP-2), are less well understood. We show in this article that FCN A-/- and CL-11-/- mice are fully susceptible to collagen Ab-induced arthritis (CAIA). In contrast, FCN B-/- and MASP-2-/-/sMAp-/- mice are substantially protected, with clinical disease activity decreased significantly (p < 0.05) by 47 and 70%, respectively. Histopathology scores, C3, factor D, FCN B deposition, and infiltration of synovial macrophages and neutrophils were similarly decreased in FCN B-/- and MASP-2-/-/sMAp-/- mice. Our data support that FCN B plays an important role in the development of CAIA, likely through ligand recognition in the joint and MASP activation, and that MASP-2 also contributes to the development of CAIA, likely in a C4-independent manner. Decreased AP activity in the sera from FCN B-/- and MASP-2-/-/sMAp-/- mice with arthritis on adherent anti-collagen Abs also support the hypothesis that pathogenic Abs, as well as additional inflammation-related ligands, are recognized by the LP and operate in vivo to activate complement. Finally, we also speculate that the residual disease seen in our studies is driven by the AP and/or the C2/C4 bypass pathway via the direct cleavage of C3 through an LP-dependent mechanism.

Specificity Evaluation and Disease Monitoring in Arthritis Imaging with Complement Receptor of the Ig superfamily targeting Nanobodies.

Single-photon emission computed tomography combined with micro-CT (SPECT/μCT) imaging using Nanobodies against complement receptor of the Ig superfamily (CRIg), found on tissue macrophages such as synovial macrophages, has promising potential to visualize joint inflammation in experimental arthritis. Here, we further addressed the specificity and assessed the potential for arthritis monitoring. Signals obtained with 99mTc-labelled NbV4m119 Nanobody were compared in joints of wild type (WT) versus CRIg−/− mice with collagen-induced arthritis (CIA) or K/BxN serum transfer-induced arthritis (STIA). In addition, SPECT/μCT imaging was used to investigate arthritis development in STIA and in CIA under dexamethasone treatment. 99mTc-NbV4m119 accumulated in inflamed joints of WT, but not CRIg−/− mice with CIA and STIA. Development and spontaneous recovery of symptoms in STIA was reflected in initially increased and subsequently reduced joint accumulation of 99mTc-NbV4m119. Dexamethasone treatment of CIA mice reduced 99mTc-NbV4m119 accumulation as compared to saline control in most joints except knees. SPECT/μCT imaging with 99mTc-NbV4m119 allows specific assessment of inflammation in different arthritis models and provides complementary information to clinical scoring for quantitatively and non-invasively monitoring the pathological process and the efficacy of arthritis treatment.

The effect of intra-articular vanilloid receptor agonists on pain behavior measures in a murine model of acute monoarthritis.

Arthritis is the most common cause of disability in the US, and the primary manifestation of arthritis is joint pain that leads to progressive physical limitation, disability, morbidity, and increased health care utilization. Capsaicin (CAP) is a vanilloid agonist that causes substance P depletion by interacting with vanilloid receptor transient receptor potential V1 on small unmyelinated C fibers. It has been used topically for analgesia in osteoarthritis with variable success. Resiniferatoxin (RTX) is an ultra potent CAP analog. The aim of this study was to measure the analgesic effects of intra-articular (IA) administration of CAP and RTX in experimental acute inflammatory arthritis in mice. Evoked pain score (EPS) and a dynamic weight bearing (DWB) device were used to measure nociceptive behaviors in a murine model of acute inflammatory monoarthritis. A total of 56 C57B16 male mice underwent EPS and DWB testing – 24 nonarthritic controls and 32 mice with carrageenan-induced arthritis. The effects of pretreatment with 0.1% CAP, 0.0003% RTX, or 0.001% RTX were measured. Nociception was reproducibly demonstrated by increased EPS and reduced DWB measures in the affected limb of arthritic mice. Pretreatment with 0.001% RTX resulted in statistically significant improvement in EPS and DWB measures when compared with those observed in carrageenan-induced arthritis animals. Pretreatment with IA 0.0003% RTX and IA 0.01% CAP resulted in improvement in some but not all of these measures. The remaining 24 mice underwent evaluation following treatment with 0.1% CAP, 0.0003% RTX, or 0.001% RTX, and the results obtained were similar to that of naïve, nonarthritic mice.