Monoiodoacetate-induced Arthritis Research Articles

TAP2, a peptide antagonist of Toll-like receptor 4, attenuates pain and cartilage degradation in a monoiodoacetate-induced arthritis rat model

Because inflammation in osteoarthritis (OA) is related to the Toll-like receptor 4 (TLR4) signaling cascades, TLR4 is a reasonable target for developing therapeutics for OA. Thus, we investigated whether TAP2, a peptide antagonist of TLR4, reduces the monoiodoacetate (MIA)-induced arthritic pain and cartilage degradation in rats. TLR4 expression of human OA chondrocytes and synoviocytes and the knee joint tissue of MIA-induced arthritis were evaluated. MIA-induced arthritic model using Sprague–Dawley rats (6 week-old-male) were treated with TAP2, a TLR4 antagonist, and evaluated with behavioral test, immunohistochemistry, and quantitative PCR. TLR4 was highly expressed in the knee joints of patients with OA and the MIA-induced rat model. Further, a single intraarticular injection of TAP2 (25 nmol/rat) molecules targeting TLR4 on day 7 after MIA injection dramatically attenuated pain behavior for about 3 weeks and reduced cartilage loss in the knee joints and microglial activation in the spinal dorsal horns. Likewise, the mRNA levels of TNFα and IL-1β, reactive oxygen species, and the expression of MMP13 in the knee joints of TAP2-treated rats was significantly decreased by TAP2 treatment compared with the control. Moreover, interestingly, the duration of OA pain relief by TAP2 was much longer than that of chemical TLR4 antagonists, such as C34 and M62812. In conclusion, TAP2 could effectively attenuate MIA-induced arthritis in rats by blocking TLR4 and its successive inflammatory cytokines and MMP13. Therefore, TAP2 could be a prospective therapeutic to treat patients with OA.

A peptide antagonist of toll-like receptor 4 attenuates pain and cartilage degradation in monoiodoacetate-induced arthritis rat model

Purpose: Because osteoarthritis (OA) is tightly related with Toll-like receptor (TLR) 4 signaling cascades, TLR4 is a reasonable target in developing therapeutics for OA. Here, we investigated TAP2, a peptide antagonist of TLR4, could reduce long-termly monoiodoacetate (MIA)-induced arthritic pain and cartilage damage in rats.

Chondroprotective effects of platelet lysate towards monoiodoacetate-induced arthritis by suppression of TNF-α-induced activation of NF-ĸB pathway in chondrocytes.

Platelet lysate (PL) contains a cocktail of growth factors that actively participates in cartilage repair. This study was designed to determine the effect and mechanism of PL on osteoarthritis (OA). An arthritis model was established to mimic human OA by intra-articular injection of monoiodoacetate (MIA) to Sprague Dawley (SD) rats. The model was weekly treated with PL by intra-articular injection. Thermal withdrawal latency, mechanical withdrawal threshold, and treadmill gait were tested for pain behavior observation. Histopathological and immunohistochemical analyses were conducted for evaluating cartilage degradation. Real time PCRs and Western blots were conducted to elucidate the mechanism of PL on primary chondrocytes. Results showed that, in vivo, PL significantly attenuated pain symptoms and exerted chondrocyte-protective and extracellular matrix (ECM)-modifying effect on the arthritic cartilage in a dose-dependent manner. The in situ expressions of type II Collagen (Col2) and matrix metalloproteinase 13 (Mmp13) in the arthritic cartilage was abnormal and was restored by PL. In vitro, PL significantly restored tumor necrosis factor α (TNF-α)-suppressed anabolic gene expression (Col2 and aggrecan) and TNF-α-increased catabolic gene expression (Col10, Mmp13, Adamts5, and Adamts9) in chondrocytes. The effects were mediated by TNF-α downstream signaling, including inhibition of NF-κB and c-Jun activities. This study provides certain knowledge of anti-OA effect and TNF signaling-related mechanism of PL, placing it as a promising and alternative option for OA therapy in the future.

Chondrocyte Cell-Sheet Transplantation for Treating Monoiodoacetate-Induced Arthritis in Rats.

Engineered cell sheets that are fabricated by using temperature-responsive culture dishes are now available in the clinical setting. Engineered chondrocyte cell sheets are used in clinical trials for treating focal articular cartilage defects. However, the efficacy of engineered chondrocyte cell sheets for treating diffusely degenerated articular cartilage remains unknown. In the present study, we assessed the utility of engineered chondrocyte cell sheets for treating diffusely degenerated articular cartilage in a monoiodoacetate (MIA)-induced arthritis animal model. At 1 and 3 weeks after intra-articular MIA injection into rat knees (1 w-MIA/3 w-MIA model), the degenerated cartilage at load-bearing area sized 1 × 3 mm was removed. Rats in each group were divided into three subgroups: Removal (R) group as control, Removal + Drilling (RD) group in which cartilage was removed followed by subchondral drilling, and Removal + Drilling + Sheet transplantation (RDS) group in which the removed cartilage area was covered with a 2 × 4 mm size of triple-layered chondrocyte cell sheet. In the histological assessment, the score of the RDS group in each model was significantly better than that of R and RD groups at 4 and 12 weeks, respectively, after the surgery. Our results demonstrated that the transplanted chondrocyte cell sheets improved cartilage repair even under such severe conditions as diffusely degenerated articular cartilage. Chondrocyte cell sheets appear to be useful as a treatment option for MIA-induced arthritis models.

Silencing of miR-101 Prevents Cartilage Degradation by Regulating Extracellular Matrix–related Genes in a Rat Model of Osteoarthritis

Osteoarthritis (OA) is a common, degenerative joint disease characterized by articular cartilage degradation. Currently, clinical trials based on microRNA therapy have been performed to treat various diseases. However, no treatment has been found for arthritis. This study investigated the functions of miR-101 in cartilage degradation in vivo and evaluated the feasibility of using miR-101 as a therapeutic agent for OA. Mono-iodoacetate-induced arthritis (MIA) rats were used as an animal model of OA. miR-101 mimic or miR-101 inhibitor was injected into the rats' knees to evaluate its effects on cartilage degradation. Cartilage degradation aggravated at 14 days after the injection of miR-101 mimic. By contrast, miR-101 silencing reduced cartilage degradation. Moreover, the administration of miR-101 mimic is sufficient to cause cartilage degradation in the normal cartilage of rats. By contrast, miR-101 inhibitor could prevent this change. Microarray and qPCR were used to investigate the different expressed genes after injecting miR-101 mimic and miR-101 inhibitor in the rats' articular cartilage. Several cartilage degradation-related genes were selected and validated to function in cartilage degradation with miR-101. Our results demonstrated the therapeutic effect of miR-101 inhibition on cartilage degradation in MIA rats by regulating several cartilage degradation–related genes.

AS1069562, the (+)-isomer of indeloxazine, exerts analgesic effects in rat models of nociceptive pain

Objectives:The (+)-isomer of indeloxazine AS1069562 has multiple pharmacological actions, such as serotonin (5-HIT) and norepinephrine (NE) reuptake inhibition and analgesic effects in animal models of neuropathic pain. Here, we investigated the analgesic effects of AS1069562 in rat models of inflammatory and noninflammatory nociceptive pain.Methods:Adjuvant-induced arthritis (AIA) and bradykinin-induced knee joint pain were used as rat models of inflammatory pain. The chronic phase of monoiodoacetate-induced arthritis (MIA) was used as a rat model of noninflammatory pain. Analgesic effects were evaluated by weight-bearing deficit in the AIA and MIA models and by pain response in the bradykinin-induced knee joint pain model.Results:In the AIA model and the bradykinin-induced knee joint pain model, AS1069562 significantly ameliorated the pain-related behavior of weight-bearing deficit and the pain response, respectively. AS1069562 also significantly improved the pain-related behavior of weight-bearing deficit in the chronic phase of the MIA model. Further, following monoiodoacetate injection, repeated administration of AS1069562 or duloxetine significantly improved weight-bearing deficit in the MIA model. Interestingly, the analgesic effect of AS1069562 was sustained for 24 hours after the last administration, although the plasma concentration of AS1069562 was reduced to undetectable levels. In contrast, the analgesic effect of duloxetine did not continue after treatment discontinuation.Discussion:AS1069562 exerts analgesic effects on inflammatory and noninflammatory nociceptive pain in rat models of arthritis pain, and repeated administration of AS1069562 exerts a more persistent analgesic effect on arthritis pain than duloxetine. These findings suggest that AS1069562 has an attractive analgesic profile for the treatment of nociceptive pain.

Inhibition of asporin signaling is critical in the prevention of cartilage damage by physiotherapies

Purpose. Physiotherapies are the most frequently recommended non-invasive treatment options recommended for arthritic diseases. Here, we examined the signaling pathways responsible for the beneficial effects of exercise on various stages of monoiodoacetate-induced arthritis (MIA). We demonstrate that both success and failure of the physiotherapy is mediated via discrete signaling pathways involving Asporin, a susceptibility gene in osteoarthritis (OA). Asporin-mediated regulation of matrix synthesis in turn may regulate Transforming Growth Factor-β (TGF-β) networks essential for cartilage matrix synthesis. Methods. Treadmill walking (TW), used as a means of exposing MIA afflicted knees to physiotherapy, was initiated in rats at different stages of MIA induced cartilage damage and compared to unexercised MIA afflicted knees. TW started at 1 day post-MIA induction, or after cartilage damage had progressed to Grade I or Grade II. The cartilage specimens were analyzed by macroscopic, microscopic, μCT imaging, transcriptome-wide gene expression analysis and activation/suppression of signaling networks. Data was further analyzed by Ingenuity Pathways Analysis to construct molecular functional networks and signaling pathways to dissect pathways regulated by exercise. One-way ANOVA and a post hoc Tukey test were used for statistical analysis. Results. Treadmill walking immediately following initiation of MIA (1 day post-MIA induction) demonstrated a significant prevention of MIA progression. However, the efficacy of this intervention was significantly reduced when implemented on knees showing close to Grade I or greater cartilage damage. On the contrary, TW accelerated damage in the knees with close to Grade II cartilage pathologies. Transcriptome-wide gene expression analysis revealed that exercise intervention started 1-day post-MIA inception significantly suppressed signaling networks that inhibit matrix synthesis. In parallel, TW upregulated gene networks associated with matrix synthesis. However, TW intervention following Grade I damage was less effective in preventing cartilage damage and regulating matrix synthesis. Interestingly, Asporin and networks associated with TGF-β expression and signaling were the major gene products that were regulated by TW to control matrix synthesis (Aggrecan, Collagen type II, Matrilin, FRZB, Col 9A2 and Col9A3) and prevent cartilage damage as evident by microscopic grading of cartilage. Conclusions. The findings demonstrate that Asporin might be one of the critical genes controlled by TW, that in turn controls expression of TGF-β family of molecules, essential for cartilage matrix synthesis. The findings underscore the importance of physiotherapies by showing suppression of Asporin, an OA susceptibility gene by TW. Nevertheless, the extent of cartilage damage at the initiation of physiotherapy is an important determinant for the effectiveness of TW.

The progression of monoiodoacetate-induced arthritis involves sequential expression/suppression of matrix associated genes

The progression of monoiodoacetate-induced arthritis involves sequential expression/suppression of matrix associated genes

Sequential alterations in catabolic and anabolic gene expression parallel pathological changes during progression of monoiodoacetate-induced arthritis.

Chronic inflammation is one of the major causes of cartilage destruction in osteoarthritis. Here, we systematically analyzed the changes in gene expression associated with the progression of cartilage destruction in monoiodoacetate-induced arthritis (MIA) of the rat knee. Sprague Dawley female rats were given intra-articular injection of monoiodoacetate in the knee. The progression of MIA was monitored macroscopically, microscopically and by micro-computed tomography. Grade 1 damage was observed by day 5 post-monoiodoacetate injection, progressively increasing to Grade 2 by day 9, and to Grade 3–3.5 by day 21. Affymetrix GeneChip was utilized to analyze the transcriptome-wide changes in gene expression, and the expression of salient genes was confirmed by real-time-PCR. Functional networks generated by Ingenuity Pathways Analysis (IPA) from the microarray data correlated the macroscopic/histologic findings with molecular interactions of genes/gene products. Temporal changes in gene expression during the progression of MIA were categorized into five major gene clusters. IPA revealed that Grade 1 damage was associated with upregulation of acute/innate inflammatory responsive genes (Cluster I) and suppression of genes associated with musculoskeletal development and function (Cluster IV). Grade 2 damage was associated with upregulation of chronic inflammatory and immune trafficking genes (Cluster II) and downregulation of genes associated with musculoskeletal disorders (Cluster IV). The Grade 3 to 3.5 cartilage damage was associated with chronic inflammatory and immune adaptation genes (Cluster III). These findings suggest that temporal regulation of discrete gene clusters involving inflammatory mediators, receptors, and proteases may control the progression of cartilage destruction. In this process, IL-1β, TNF-α, IL-15, IL-12, chemokines, and NF-κB act as central nodes of the inflammatory networks, regulating catabolic processes. Simultaneously, upregulation of asporin, and downregulation of TGF-β complex, SOX-9, IGF and CTGF may be central to suppress matrix synthesis and chondrocytic anabolic activities, collectively contributing to the progression of cartilage destruction in MIA.

Transcriptome-wide gene regulation by gentle treadmill walking during the progression of monoiodoacetate-induced arthritis

Physiotherapies are the most widely recommended conservative treatment for arthritic diseases. The present study was undertaken to examine the molecular mechanisms underlying the effects of gentle treadmill walking (GTW) on various stages of monoiodoacetate-induced arthritis (MIA) to elucidate the basis for the success or failure of such therapies in joint damage. Knees were obtained from untreated control rats, rats with MIA that did not undergo GTW, rats with MIA in which GTW regimens were started 1 day post-MIA induction, and rats with MIA in which GTW regimens were started after cartilage damage had progressed to grade 1 or grade 2. The cartilage was examined macroscopically, microscopically, and by microfocal computed tomography imaging. Transcriptome-wide gene expression analysis was performed, and microarray data were assessed by Ingenuity Pathways Analysis to identify molecular functional networks regulated by GTW. GTW intervention started on day 1 post-MIA induction significantly prevented the progression of MIA, but its efficacy was reduced when implemented on knees exhibiting close to grade 1 cartilage damage. GTW accelerated cartilage damage in knees with close to grade 2 damage. Transcriptome-wide gene expression analysis revealed that GTW intervention started 1 day post-MIA inception significantly suppressed inflammation-associated genes and up-regulated matrix-associated gene networks. However, delayed GTW intervention after grade 1 damage had occurred was less effective in suppressing proinflammatory genes or up-regulating matrix synthesis. The present findings suggest that GTW suppresses proinflammatory gene networks and up-regulates matrix synthesis to prevent progression of cartilage damage in MIA-affected knees. However, the extent of cartilage damage at the initiation of GTW may be an important determinant of the success or failure of such therapies.

Antinociceptive Effects of AS1892802, a Novel Rho Kinase Inhibitor, in Rat Models of Inflammatory and Noninflammatory Arthritis

Rho kinase (ROCK) is involved in various physiological functions, including cell motility, vasoconstriction, and neurite extension. Although a functional role of ROCK in nociception in the central nervous tissue has been reported in neuropathy, the peripheral function of this protein in hyperalgesia is not known. In this study, antinociceptive effects of AS1892802 [1-[(1S)-2-hydroxy-1-phenylethyl]-3-[4-(pyridin-4-yl)phenyl]urea], a novel and highly selective ROCK inhibitor, were investigated in two rat models of arthritis. Orally administered AS1892802 exhibited potent antinociceptive effect in both an adjuvant-induced arthritis (AIA) model (inflammatory arthritis model) and a monoiodoacetate-induced arthritis (MIA) model (noninflammatory arthritis model), with an ED(50) of 0.15 mg/kg (MIA model). Fasudil, a ROCK inhibitor, and tramadol were also effective in both models; however, diclofenac was effective only in the AIA model. The onset of antinociceptive effect of AS1892802 was as fast as those of tramadol and diclofenac. AS1892802 did not induce gastric irritation or abnormal behavior. Because AS1892802 rarely penetrates the central nervous tissue and is also effective by intra-articular administration, it seemed to function peripherally. These results suggest that AS1892802 has an attractive analgesic profile for the treatment of severe osteoarthritis pain.

Sustained Analgesic Effect of the Rho Kinase Inhibitor AS1892802 in Rat Models of Chronic Pain

To assess the pharmacological profile of AS1892802, a novel and selective Rho kinase (ROCK) inhibitor, we examined the effects of repeated dosing with AS1892802 on models of monoiodoacetate-induced arthritis and streptozotocin-induced neuropathy. Although single dosing of AS1892802 exerted a short-acting, moderate analgesic effect, repeated dosing exhibited a long-lasting and more potent analgesic effect in both models. Furthermore, the analgesic effect was sustained for seven days after the last administration. These results suggest that peripheral ROCK plays a crucial role in chronic pain maintenance and that AS1892802 may be useful in treating chronic pain.

Plasma 3-nitrotyrosine is a biomarker in animal models of arthritis: Pharmacological dissection of iNOS’ role in disease

The contribution of inducible nitric oxide synthase (iNOS) to oxidative/nitrative stress is well-documented in inflammation, but difficult to quantify. Using a novel, recently developed assay for 3-nitrotyrosine (3-NT), we characterized iNOS activity and its inhibition in preclinical models of inflammation. In particular, we utilized the 3-NT assay to assess the role of iNOS in the disease pathology as well as for proof of pharmacology of iNOS inhibitors in an acute endotoxin challenge model, in models of rheumatoid arthritis (RA) such as rat adjuvant- and collagen-induced arthritis (AIA and CIA) and a model of osteoarthritis (OA) such as rat sodium monoiodoacetate-induced arthritis (MIA). Quantification of nitrotyrosine was performed using immuno-affinity 2-D LC–MS/MS assay. This assay is a very specific and reproducible and is amenable to a number of biological fluids. Plasma levels of 3-NT were significantly elevated in an acute model of inflammation (rat LPS) and in models of rheumatoid arthritis (adjuvant- and collagen-induced arthritis), and osteoarthritis (monoiodoacetate-induced arthritis). Plasma 3-NT correlated with the severity of the inflammatory response; thus, a 20-fold increase was observed in the rat LPS model, a 10-fold increase in AIA, and only a 2.5-fold elevation in CIA. Pharmacological intervention with iNOS inhibitors decreased 3-NT levels and associated pathology. 3-NT determination allowed for better elucidation of the role of iNOS in RA and OA disease patholog y and provided proof of pharmacology for NOS inhibitors in animal models of RA and OA.

The utility of measuring C-terminal telopeptides of collagen type II (CTX-II) in serum and synovial fluid samples for estimation of articular cartilage status in experimental models of destructive joint diseases

To characterize and validate a novel, enzyme-linked immunoassay for measuring cross-linked dimer forms of C-terminal telopeptides of type II collagen (CTX-II) in serum and synovial fluid of rodents, and investigate whether CTX-II measurements can reflect joint status in two established animal models of destructive joint diseases. Firstly, the specificity, in vivo validity, antigen recovery, and reproducibility of the assay were investigated. Secondly, we induced arthritis in rats using either bovine collagen type II or mono-iodoacetate. CTX-II levels were measured in the serum and synovial fluid of the affected femoro-tibial joint and correlated with microscopic severity of joint lesions as determined by validated scoring systems. The F4601 monoclonal antibody (mAb) is highly specific for the EKGPDP sequence at the CTX-II. Strong CTX-II signals were detected during enzymatic degradation of articular cartilage explants by matrix metalloproteinase (MMP)-9 or MMP-13. The assay presented a good degree of precision and reproducibility (inter- and intra-assay CVs< 8.0%). In the collagen-induced arthritis (CIA) model, the assay indicated markedly increased levels of CTX-II in both the synovial fluid and the serum. Furthermore, CTX-II levels in both the synovial fluid (r = 0.76; P < 0.0001) and the serum (r = 0.85; P < 0.0001) showed strong correlations with the microscopic severity scores of joint lesions at Day 22. In the mono-iodoacetate-induced arthritis (MIA) model, CTX-II concentration in the synovial fluid (r = 0.53; P < 0.0001), but not in the serum, correlated with the microscopic severity score. The Preclinical CTX-II assay could provide a useful supplement to currently available methods for the non-invasive assessment of cartilage status. The utility of serum CTX-II to reflect joint status appeared to be limited to systemic forms of destructive joint diseases.