Intra-articular Injection Of Monoiodoacetate Research Articles

Green Light Exposure Reduces Primary Hyperalgesia and Proinflammatory Cytokines in a Rodent Model of Knee Osteoarthritis: Shedding Light on Sex Differences

Knee osteoarthritis (OA) often causes chronic pain that disproportionately affects females. Proinflammatory cytokines TNF-α, IL-1β, and IL-6 are key effectors of OA pathological changes. Green light shows potential as an alternative intervention for various pain conditions. However, no studies have investigated green light′s analgesic effects in both sexes in chronic knee OA. We induced unilateral knee OA with intra-articular injection of monoiodoacetate (MIA) in male and female Sprague-Dawley rats. Two days post-injection, the rats were exposed to green-light-emitting diodes (GLED) or ambient room light eight hours daily for 24 days. Knee mechanical sensitivity was assessed using a small animal algometer. Blood serum concentrations of TNF-α, IL-1β, IL-6, and IL-10 were quantified at baseline and 23 days post-injection. MIA injection decreased the knee mechanical thresholds of the male and female rats. GLED exposure attenuated mechanical hypersensitivity in both sexes compared to the controls; however, GLED-induced analgesia occurred sooner and with greater magnitude in males than in females. In both sexes, the analgesic effects of green light lasted 5 days after the final GLED session. Finally, GLED exposure reversed the elevation of serum proinflammatory cytokines. These findings suggest that GLED exposure reduces primary hyperalgesia in OA, potentially by lowering proinflammatory cytokines, and indicate sex differences in GLED-induced analgesia.

S-Equol Ameliorates Menopausal Osteoarthritis in Rats through Reducing Oxidative Stress and Cartilage Degradation.

Osteoarthritis (OA) is a chronic degenerative disease leading to articular cartilage destruction. Menopausal and postmenopausal women are susceptible to both OA and osteoporosis. S-equol, a soy isoflavone-derived molecule, is known to reduce osteoporosis in estrogen-deficient mice, but its role in OA remains unknown. This study aimed to explore the effect of S-equol on different degrees of menopausal OA in female Sprague-Dawley (SD) rats induced by estrogen deficiency caused by bilateral ovariectomy (OVX) combined with intra-articular injection of mono-iodoacetate (MIA). Knee joint histopathological change; serum biomarkers of bone turnover, including N-terminal propeptide of type I procollagen (PINP), C-terminal telopeptide of type I collagen (CTX-I) and N-terminal telopeptide of type I collagen (NTX-I); the cartilage degradation biomarkers hyaluronic acid (HA) and N-terminal propeptide of type II procollagen (PIINP); and the matrix-degrading enzymes matrix metalloproteinases (MMP)-1, MMP-3 and MMP-13, as well as the oxidative stress-inducing molecules nitric oxide (NO) and hydrogen peroxide (H2O2), were assessed for evaluation of OA progression after S-equol supplementation for 8 weeks. The results showed that OVX without or with MIA injection induced various severity levels of menopausal OA by increasing pathological damage, oxidative stress, and cartilage matrix degradation to various degrees. Moreover, S-equol supplementation could significantly reduce these increased biomarkers in different severity levels of OA. This indicates that S-equol can lessen menopausal OA progression by reducing oxidative stress and the matrix-degrading enzymes involved in cartilage degradation.

Changes in the Serum Metabolome in an Inflammatory Model of Osteoarthritis in Rats.

Osteoarthritis (OA) is a pathology of great impact worldwide. Its physiopathology is not completely known, and it is usually diagnosed by imaging techniques performed at advanced stages of the disease. The aim of this study was to evaluate early serum metabolome changes and identify the main metabolites involved in an inflammatory OA animal model. This study was performed on thirty rats. OA was induced in all animals by intra-articular injection of monoiodoacetate into the knee joint. Blood samples were taken from all animals and analyzed by mass spectrometry before OA induction and 28, 56, and 84 days following induction. Histological evaluation confirmed OA in all samples. The results of this study allow the identification of several changes in 18 metabolites over time, including organic acids, benzenoids, heterocyclic compounds, and lipids after 28 days, organic acids after 56 days, and lipid classes after 84 days. We conclude that OA induces serological changes in the serum metabolome, which could serve as potential biomarkers. However, it was not possible to establish a relationship between the identified metabolites and the time at which the samples were taken. Therefore, these findings should be confirmed in future OA studies.

The Effect of Activated Growth Factor (AGF) from Platelets on Alpha-SMA Levels in Osteoarthritis: Invivo Study

Background: The most prevalent joint condition and leading contributor to disability is osteoarthritis (OA), resulting in a significant socio-economic impact. It occurs due to an imbalance of pro-inflammatory and anti-inflammatory mediators, influencing anabolic and catabolic activities. This study aims to determine the effect of activated growth factor (AGF) on alpha-SMA levels.
 Methods: This research is an in vivo post-test-only control group experimental study conducted at the Eureka Research Laboratory. Thirty male Wistar rats were divided into five treatment groups: normal control group, negative control group, AGF I group (TGF-β 100 pg/mL), AGF II group (TGF-β 1000 pg/mL), and AGF III group (TGF-β 10,000 pg/mL). AGF was obtained from rat blood intravenously and centrifuged at a predetermined speed. Growth factor activation was performed by adding 10% CaCl2, and then TGF-β levels were measured. All groups of rats were acclimatized for 7 days. After that, osteoarthritis was induced with intra-articular injection of monoiodoacetate (MIA) 4.8 mg/60μL in all groups of rats except the normal control group. Next, rats were given treatment according to the group for 21 days. On the 21st day, rats were euthanized, and alpha-SMA levels were measured using the sandwich method ELISA kit.
 Results: Sequentially, the mean Alpha-SMA levels for each group in pg/mL were: 91.495 ± 2.36; 10.682 ± 1.09; 30.502 ± 2.00; 52.892 ± 1.29; and 76.180 ± 2.65. In the AGF group, there was an increase in alpha-SMA levels directly proportional to the dose of TGF-β injected.
 Conclusion: AGF has an effect on increasing alpha-SMA levels in the joints of rats with a model of osteoarthritis.

Description of Novel Molecular Factors in Lumbar DRGs and Spinal Cord Factors Underlying Development of Neuropathic Pain Component in the Animal Model of Osteoarthritis.

Osteoarthritis (OA) is one of the most common joint disorder, with pain accompanied by functional impairment, as the most pronounced clinical symptom. Currently used pharmacotherapy involves symptomatic treatment that do not always provide adequate pain relief. This may be due to concomitance of central sensitization and development of neuropathic features in OA patients. Here we performed studies in the animal model of OA to investigate of the neuropathic component. Intraarticular injection of monoiodoacetate (MIA, 1mg) was used to induce OA in Wistar male rats. Development of pain phenotype was assessed by behavioral testing (PAM test and von Frey's test), while corresponding changes in dorsal root ganglia (DRGs L3-L5) and spinal cord (SC) gene expression were assessed by means of qRT-PCR technique. We also performed microtomography of OA-affected knee joints to correlate the level of bone degradation with observed behavioral and molecular changes. We observed gradually developing remote allodynia after MIA treatment, indicating the presence of neuropathic component. Our results showed that, among DRGs innervating knee joint, development of central sensitization is most likely due to peripheral input of stimuli through DRG L5. In SC, development of secondary hypersensitivity correlated with increased expression of TAC1 and NPY. Our studies provided molecular records on abnormal activation of pain transmission markers in DRG and SC during development of OA that are responsible for the manifestation of neuropathic features. The obtained results increase insight into molecular changes occurring in the neuronal tissue during OA development and may contribute to readdressing treatment paradigms.

Comparing Effectiveness of Hyaluronic Acid-Chitosan Nanoparticles Encapsulation Versus Hyaluronic Acid Monotherapy in Osteoarthritis Rat Model: Microarray Screening for miR-140.

Osteoarthritis is a debilitating, progressive joint disease linked to lower quality of life and higher health care costs. This study compared hyaluronic acid-chitosan nanoparticle encapsulation to hyaluronic-acid monotherapy in a rat model of knee osteoarthritis. Four groups of 40 adult male albino rats were designed. Group (Gp) I: control; Gp II (osteoarthritis model): intra-articular injection of monoiodoacetate; Gp III (hyaluronic acid-treated): intra-articular injections of hyaluronic-acid on days 14 and 21 after monoiodoacetate injection; and Gp IV (hyaluronic acid-chitosan nanoparticle-treated): intra-articular injections of hyaluronic acid-chitosan nanoparticle on days 14 and 21 after monoiodoacetate injection. After 28 days, knee joints were examined using H&E, Safranin O, and immunohistochemistry for nuclear factor kappa beta (NF-κB), inducible nitric oxide synthase (iNOS), and matrix metalloproteinase (MMP)-13. Quantification for gene expression of collagen-II, aggrecan, and micro-RNA-140; ELISA for interleukin (IL)-1β and IL-8; and western blotting for IKBα and NF-κB was estimated. Osteoarthritis-knee joints showed a severe cartilage damage and synovial inflammation with increased NF-κB, iNOS, and MMP-13 immunostaining, decreased miR-140, collagen II, and aggrecan levels, and increased inflammatory markers' gene expressions. The hyaluronic acid-chitosan nanoparticle significantly improved knee joint structure and reduced inflammatory cytokines compared to hyaluronic acid monotherapy. Intra-articular injection of hyaluronic acid-chitosan nanoparticle encapsulation revealed a significant improvement in the knee joint structure compared to hyaluronic-acid in a rat model of osteoarthritis.

Pain Intensity and Trajectory Following Intra-Articular Injection of Mono-Iodoacetate in Experimental Osteoarthritis: A Meta-Analysis of In Vivo Studies.

Although most frequently used in experimental osteoarthritis (OA) pain induction, intra-articular mono-iodoacetate (MIA) injection lacks concluded references for dose selection and timing of intervention. Herein, we aimed to compare the pain intensity of rats induced by different doses of MIA and explored the trajectory of pain. PubMed, Embase, and Web of Science were searched up to June 2021 for literatures involving MIA experiments investigating OA pain. Pain intensity was measured based on weightbearing distribution (WBD) and paw withdrawal thresholds (PWT), and the pain trajectory was constructed by evaluating pain intensity at a series of time points after MIA injection. A conventional meta-analysis was conducted. A total of 140 studies were included. Compared with saline, MIA injections caused significantly higher pain intensity for WBD and PWT. Dose-response relationships between different doses of MIA and pain intensity were observed (P-for-trend<0.05). A pronounced increase in pain occurred from day 0 to day 7, but the uptrend ceased between day 7 and day 14, after which the pain intensity continued to rise and reached the maximum by day 28. Pain intensity after intra-articular MIA injection increased in a dose-dependent manner and the pain trajectory manifested a specific pattern consistent with the pathological mechanisms of MIA-induced pain, providing possible clues for proper dose selection and timing of specific OA pain interventions.

Pilot comparison of outcome measures across chemical and surgical experimental models of chronic osteoarthritis in the rat (Rattus norvegicus).

Relatively little work has evaluated both the disease of osteoarthritis (OA) and clinically-relevant pain outcome measures across different OA models in rats. The objective of this study was to compare sensitivity, pain, and histological disease severity across chemical and surgical models of OA in the rat. Stifle OA was induced in Sprague-Dawley rats via intraarticular injection of monoiodoacetate (MIA) or surgical transection of anterior cruciate ligament and/or destabilization of medial meniscus (ACL+DMM or DMM alone). Reflexive (e.g., mechanical and thermal stimuli) measures of sensitivity and non-reflexive assays (e.g., lameness, static hindlimb weight-bearing asymmetry, dynamic gait analysis) of pain were measured over time. Joint degeneration was assessed histologically. Six-weeks post OA-induction, the ACL+DMM animals had significantly greater visually observed lameness than MIA animals; however, both ACL+DMM and MIA animals showed equal pain as measured by limb use during ambulation and standing. The MIA animals showed increased thermal, but not mechanical, sensitivity compared to ACL+DMM animals. Joint degeneration was significantly more severe in the MIA model at 6 weeks. Our pilot data suggest both the ACL+DMM and MIA models are equal in terms of clinically relevant pain behaviors, but the MIA model is associated with more severe histological changes over time potentially making it more suitable for screening disease modifying agents. Future work should further characterize each model in terms of complex pain behaviors and biochemical, molecular, and imaging analysis of the sensory system and joint tissues, which will allow for more informed decisions associated with model selection and investigative outcomes.

Effects of Combined Allogenic Adipose Stem Cells and Hyperbaric Oxygenation Treatment on Pathogenesis of Osteoarthritis in Knee Joint Induced by Monoiodoacetate.

The beneficial effects of HBO in inflammatory processes make it an attractive type of treatment for chronic arthritis. In addition, the effects of combination therapy based on adipose stem cells and HBO on OA progression have not been fully investigated. The current study explored the efficacy of intra-articular injection of allogeneic adipose-derived mesenchymal stem cells (ADMSCs) combined with hyperbaric oxygenation treatment (HBO) in a rat osteoarthritis (OA) model. The rat OA model was induced by intra-articular injection of monoiodoacetate (MIA) and 7 days after application of MIA rats were divided into five groups: healthy control (CTRL), osteoarthritis (OA), ADMSCs (ADS), the HBO+ADS21day and HBO+ADS28day groups. A single dose of 1 × 106 allogeneic ADMSCs suspended in sterile saline was injected into the knee joint alone or in combination with HBO treatment. Rats were sacrificed at 3 or 4 weeks after MIA injection. Treatment outcomes were evaluated by radiographic, morphological and histological analysis and by specific staining of articular cartilage. We also measured the level of inflammatory and pro/antioxidative markers. We confirmed that combined treatment of ADMSCs and HBO significantly improved the regeneration of cartilage in the knee joint. Rtg score of knee joint damage was significantly decreased in the HBO+ADS21day and HBO+ADS28day groups compared to the OA. However, the positive effect in the HBO+ADS28day group was greater than the HBO+ADS21day group. The articular cartilage was relatively normal in the HBO+ADS28day group, but moderate degeneration was observed in the HBO+ADS21day compared to the OA group. These findings are in line with the histopathological results. A significantly lower level of O2−. was observed in the HBO+ADS28day group but a higher NO level compared to the HBO+ADS21day group. Moreover, in the HBO+ADS28day group significantly higher concentrations of IL-10 were observed but there was no significant difference in proinflammatory cytokine in serum samples. These results indicate that a single intra-articular injection of allogeneic ADMSCs combined with HBO efficiently attenuated OA progression after 28 days with greater therapeutic effect compared to alone ADMSCs or after 3 weeks of combined treatment. Combined treatment might be an effective treatment for OA in humans.

Intra-articular injection of monoiodoacetate induces diverse hip osteoarthritis in rats, depending on its dose

BackgroundMonoiodoacetate (MIA)-induced arthritis models are used widely in osteoarthritis (OA) research to develop effective conservative treatments for hip OA, as an alternative to joint replacement surgery. In joint OA models, such as the MIA-induced knee OA model, various doses of MIA are utilized, depending on the purpose of the research. So far, only 2 mg of MIA has been used for MIA-induced hip OA research. We hypothesized that the amount of MIA should be adjusted according to the osteoarthritis model under investigation. We performed radiographic and histological evaluations in rats for hip OA models induced by different doses of MIA.MethodsOne hundred and eighty right hips of six-week-old, male Sprague–Dawley rats (n = 30 rats per group) were treated with either a single intra-articular injection of various doses of MIA (0.25, 0.5, 1.0, 2.0, and 4.0 mg) dissolved in 25 μl of sterile saline (MIA group), or with 25 μl of sterile saline alone (Sham group). Radiographic and histological evaluations of the hip joint were performed at one, two, four, eight, and 12 weeks after administration (n = 6 rats per group per time point).ResultsOA changes progressed from 1 week after administration in the 1.0-mg, 2.0-mg, and 4.0-mg MIA groups. The degree of OA changes increased as the dose of MIA increased. The 0.25-mg and 0.5-mg MIA groups presented fewer OA changes than the 2.0-mg and 4.0-mg MIA groups during the entire study period (up to 12 weeks). The administration of 0.25 mg and 0.5 mg of MIA-induced both radiographic and histological OA changes in a time-dependent manner, whereas more than 2 mg of MIA provoked end-stage OA at 8 weeks after injection. Absolute, dose-dependent histopathological OA changes were observed 4 weeks after MIA administration.ConclusionsIntra-articular MIA injection to the hip joints of rats induced diverse OA changes dose-dependently. Research for developing novel conservative treatments for hip OA and intractable pain should consider the pathological condition when determining the dose of MIA to be employed.

Changes of Grip Strength, Articular Cartilage and Subchondral Bone in Monoiodoacetate-Induced Osteoarthritis in Rats

Osteoarthritis is a degenerative disease affecting articular cartilage among the elderly. The intra-articular monoiodoacetate injection is one of the most widely used methods to induce osteoarthritis in animals. While the effects of monoiodoacetate on cartilage are well-characterized, its effects on subchondral bone remodeling are less studied. The purpose of this study was to determine the changes of the grip strength, articular cartilage structure and subchondral bone remodeling in monoiodoacetate-induced osteoarthritis in rats. Three-month-old male Wistar rats were assigned to normal control (n=6) and osteoarthritis group (n=6), which received intra-articular injection of 4 mg/50 µL monoiodoacetate solution once at the left knee of hindlimb. The rats were monitored for four weeks. The grip strength test was performed before injection and every week after injection. After four weeks, the femurs with intact cartilage were harvested for histomorphological analysis. Grip strength was reduced significantly in the osteoarthritic rats compared to the normal rats (p&lt;0.05). Food intake was reduced significantly one week following monoiodoacetate-induction (p&lt;0.05), but it stabilized afterwards. Monoiodoacetate injection increased cartilage erosion and osteoclast number in the subchondral bone of the osteoarthritic rats compared to the normal rats (p&lt;0.05). However, it did not affect body weight, subchondral bone osteoblast activity, mineralization and microstructure of osteoarthritic rats (p&gt;0.05). As a conclusion, monoiodoacetate-induced osteoarthritis affects the cartilage and increases osteoclast formation in the subchondral bone of rats.

Antinociceptive and chondroprotective effects of prolonged β-caryophyllene treatment in the animal model of osteoarthritis: Focus on tolerance development

Osteoarthritis (OA) is a chronic joint disease in which cartilage degeneration leads to chronic pain. The endocannabinoid system has attracted attention as an emerging drug target for OA. However, the therapeutic potential of cannabinoids is limited by psychoactive side-effects related to CB1 activation and tolerance development for analgesic effects. β-Caryophyllene (BCP) is a low-efficacy natural agonist of CB2 and a common constituent of human diet with well-established anti-inflammatory properties. The results presented herein show the anti-nociceptive and chondroprotective potential of BCP in an animal model of OA induced by intra-articular injection of monoiodoacetate (MIA). Behavioural assessment included pressure application measurement and kinetic weight bearing tests. Histological assessment of cartilage degeneration was quantified using OARSI scoring. Experiments established the dose-response effects of BCP and pharmacological mechanisms of the antinociceptive action dependent on CB2 and opioid receptors. Chronic BCP treatment was able to hamper cartilage degeneration without producing tolerance for the analgesic effects. The data presented herein show that BCP is able to produce both acute and prolonged antinociceptive and chondroprotective effects. Together with the safety profile and legal status of BCP, these results indicate a novel and promising disease-modifying strategy for treating OA.This article is part of the Special Issue on ‘Advances in mechanisms and therapeutic targets relevant to pain’.

Pharmacological characterization of the chronic phase of the monoiodoacetate-induced rat model of osteoarthritis pain in the knee joint.

For patients with osteoarthritis (OA) of the knee, pain is the most debilitating symptom. Although it has been proposed that the chronic phase of the monoiodoacetate (MIA)-induced rodent model of knee joint pain may be superior to other chronic or acute OA models for assessing the analgesic efficacy of novel molecules, relatively few pharmacological studies have been conducted in the chronic phase of this model. Hence, this study was designed to use pharmacological methods to characterize the chronic phase of the MIA-induced rat model of knee joint OA pain. Rats received a single intraarticular injection of MIA at 2.5mg or vehicle (saline) into the left (ipsilateral) knee joint. Pain behaviour was assessed by measuring paw withdrawal thresholds (PWTs) in the hindpaws pre-MIA injection and twice-weekly until study completion on day 42. Mechanical allodynia was fully developed in the ipsilateral hindpaws (PWTs ≤6g) from day 7 and it persisted until day 42. MIA-injected rats with PWTs ≤6g in the ipsilateral hindpaws received single doses of one of four clinically available drugs that represent four distinct pharmacological classes, viz gabapentin, amitriptyline, meloxicam and morphine, according to a 'washout' protocol with at least 48hours between successive doses. Gabapentin evoked dose-dependent anti-allodynia as did morphine whereas amitriptyline and meloxicam were inactive. Our findings are aligned with clinical data showing that gabapentin and morphine alleviated OA pain in the knee. The lack of efficacy of amitriptyline is consistent with the loss of descending diffuse noxious inhibitory controls reported by others in this model.

Proline-Serine-Threonine Phosphatase-Interacting Protein 2 Alleviates Diabetes Mellitus-Osteoarthritis in Rats through Attenuating Synovial Inflammation and Cartilage Injury.

ObjectiveTo explore the possible way of proline‐serine–threonine phosphatase‐interacting protein 2 (PSTPIP2) influencing diabetes mellitus‐osteoarthritis (DM‐OA) progression.Methods In vivo, eight‐week‐old male Sprague Dawley rats were induced with DM‐OA by intraperitoneal injection of streptozotocin with high‐fat diet feeding and intra‐articular injection of monoiodoacetate. PSTPIP2 overexpression was achieved by intra‐articular injection of lentivirus vectors. PSTPIP2 expression was verified by real‐time polymerase chain reaction and Western blotting. Histological changes were examined by hematoxylin/eosin and safranin‐O/fast‐green staining. In vitro, rat synovial fibroblasts were induced DM‐OA by stimulation of high glucose (HG) and interleukin (IL)‐1β. PSTPIP2 overexpression was achieved by lentivirus infection. U0126 was added as an ERK inhibitor. Levels of tumor necrosis factor (TNF)‐α, IL‐6, and IL‐1β were detected using enzyme‐linked immunosorbent assay. Expression of matrix metalloproteinase (MMP)‐3, MMP‐13, aggrecanase‐2 (ADAMTS‐5), intercellular cell adhesion molecule (ICAM)‐1, extracellular regulated protein kinase (ERK) and phospho‐ERK (p‐ERK) was detected by Western blotting.ResultsIn DM‐OA rats, PSTPIP2 relative messenger RNA (mRNA) level was significantly decreased compared to control rats. The protein expression was also decreased obviously. Inflammation score in synovium was dramatically increased, accompanying with increased TNF‐α, IL‐6, and IL‐1β levels. Osteoarthritis research society international (OARSI) score in cartilage was markedly increased, along with increased MMP‐3, MMP‐13, ADAMTS‐5, ICAM‐1, ERK and p‐ERK expression. In PSTPIP2‐overexpressed DM‐OA rats, PSTPIP2 mRNA level and protein expression was increased compared to DM‐OA rats received negative‐control lentivirus vectors. The inflammation score, as well as TNF‐α, IL‐6, and IL‐1β levels were dramatically decreased. Also, the OARSI score and protein expression of MMP‐3, MMP‐13, ADAMTS‐5, ICAM‐1, ERK and p‐ERK were decreased. In HG+IL‐1β‐treated rat synovial fibroblasts, PSTPIP2 protein expression was decreased compared to normal glucose (NG)‐treated cells. Levels of TNF‐α, IL‐6, and IL‐1β, as well as expression of MMP‐3, MMP‐13, ADAMTS‐5, ICAM‐1, ERK and p‐ERK were increased. After cells were infected with PSTPIP2‐overexpressed lentivirus, levels of TNF‐α, IL‐6, and IL‐1β, and expression of MMP‐3, MMP‐13, ADAMTS‐5, ICAM‐1, ERK and p‐ERK were obviously decreased compared to cells infected with NC lentivirus. In addition, ERK inhibitor U0126 treatment also decreased the TNF‐α, IL‐6, and IL‐1βlevels and MMP‐3, MMP‐13, ADAMTS‐5, ICAM‐1, ERK and p‐ERK expression in HG + IL‐1β treated rat synovial fibroblasts.ConclusionOverexpression of PSTPIP2 alleviates synovial inflammation and cartilage injury during DM‐OA progression via inhibiting ERK phosphorylation.

Artemin and its cognate receptor, GFRΑ3, Play a function role in osteoarthritis pain

Purpose: Osteoarthritis associated pain (OA-pain) is a significant global problem. OA-pain limits limb use and mobility, and is associated with widespread sensitivity. Therapeutic options are limited, and the ones that are available are often associated with side or adverse effects. The lack of therapeutic options is partly due to a lack of understanding of clinically relevant underlying neural mechanisms of OA-pain. In previous work in naturally occurring OA-pain in dogs, we identified potential signaling molecules (artemin/GFRα3) that were upregulated. Artemin/GFRα3 signaling is known to play a role in various pain states (mainly models of pain), including a recently reported inflammatory bone pain model. Artemin/GFRα3 is thought to potentiate pain via the upregulation and/or sensitization of various transient receptor potential (TRP) receptors. To date, no work has been performed to evaluate the role of artemin/GFRα3 in OA-pain. The purpose of this study was to explore the functional role of artemin/GFRα3 signaling in OA-pain. Methods: To examine the functional role of artemin/GFRα3 signaling in OA-pain, we used a mouse model of stifle (knee) OA based on the intra-articular injection of mono-iodoacetate (MIA). Following induction of OA using MIA, we assessed sensitivity to heat, mechanical and cold pain, and evaluated limb use, weekly to 28 days or longer. At the same timepoints, heat, mechanical, and cold sensitivity was assessed in TRPV1 KO mice with MIA arthritis to define the role of TRPV1 in MIA-induced sensitivity. Using artemin injections into the footpad, we evaluated the acute changes in heat, mechanical, and cold sensitivity. In male and female mice with MIA-induced OA, we assessed the anti-hypersensitivity effects of an anti-artemin antibody at day 28, and assessed the effects of an anti-artemin antibody on limb use measured while walking and standing (Catwalk assay, and pronograde incapacitance meter) at day 40. Results: We found the mono-iodoacetate (MIA)-induced OA model in mice is associated with significantly decreased limb use (both kinetic and static measures) and hypersensitivity to heat, mechanical, and cold. Mechanical and cold sensitivity in TRPV1 KO mice with MIA-induced OA were not significantly different from wild type mice with MIA-induced OA; there was some mitigation of heat hypersensitivity in the TRPV1 KO animals. Using immunohistochemistry, we found that GFRα3 expression in sensory neurons of the dorsal root ganglions serving the stifle joints was increased from ∼20% of neurons to ∼40% in mice with MIA OA. In naïve mice, exogenous artemin induced heat, cold and mechanical hypersensitivity for between 2 and 6 hours following injection. An anti-artemin monoclonal antibody administered intraperitoneal on day 28 following MIA-induction of OA reversed mechanical and cold hypersensitivity for between 2 and 4 hours following injection, and between 2 and 6 hours for heat hypersensitivity. Approximately 40 days following MIA-induction of OA, an anti-artemin antibody restored limb use to normal (both kinetic and static) in mice with MIA-induced OA pain for ∼48 hours. Conclusions: Overall, we present the first evidence of a potential functional role of artemin/GFRα3 in chronic OA-pain. However, there is much to understand about the potential role of artemin, including the mechanisms leading to artemin release and which cells types in the joints are responsible for artemin release; whether artemin is involved in the induction and/or maintenance of OA-pain; and whether artemin acts through only GFRα3, or other receptors (such as NCAM) are involved. Also, the degree to which artemin may drive the ultimate experience of OA pain needs to be fully elucidated. Although we have found that artemin’s cognate receptor, GFRα3, was upregulated in MIA induced OA-pain, paralleling what we found in the naturally occurring OA model in the dog, work needs to be done to determine the contribution of GFRα3 upregulation and/or activation in OA pain, and further, the downstream targets and signaling mechanisms need to be unraveled.

Undenatured Type II Collagen Ameliorates Inflammatory Responses and Articular Cartilage Damage in the Rat Model of Osteoarthritis.

Osteoarthritis (OA) is an age-related joint disease that includes gradual disruption of the articular cartilage and the resulting pain. The present study was designed to test the effects of undenatured type II collagen (UC-II®) on joint inflammation in the monoiodoacetate (MIA) OA model. We also investigated possible mechanisms underlying these effects. Female Wistar rats were divided into three groups: (i) Control; (ii) MIA-induced rats treated with vehicle; (iii) MIA-induced rats treated with UC-II (4 mg/kg BW). OA was induced in rats by intra-articular injection of MIA (1 mg) after seven days of UC-II treatment. UC-II reduced MIA-induced Kellgren-Lawrence scoring (53.3%, P < 0.05). The serum levels of inflammatory cytokines [IL-1β (7.8%), IL-6 (18.0%), TNF-α (25.9%), COMP (16.4%), CRP (32.4%)] were reduced in UC-II supplemented group (P < 0.0001). In the articular cartilage, UC-II inhibited the production of PGE2 (19.6%) and the expression of IL-1β, IL-6, TNF-a, COX-2, MCP-1, NF-κB, MMP-3, RANKL (P < 0.001). The COL-1 and OPG levels were increased, and MDA decreased in UC-II supplemented rats (P < 0.001). UC-II could be useful to alleviate joint inflammation and pain in OA joints by reducing the expression of inflammatory mediators.

Osteoarthritis Pain Model Induced by Intra-Articular Injection of Mono-Iodoacetate in Rats

Osteoarthritis Pain Model Induced by Intra-Articular Injection of Mono-Iodoacetate in Rats

Osteoarthritis Pain Model Induced by Intra-Articular Injection of Mono-Iodoacetate in Rats.

The current animal models of osteoarthritis (OA) can be divided into spontaneous models and induced models, both of which aim to simulate the pathophysiological changes of human OA. However, as the main symptom in the late stage of OA, pain affects the patients' daily life, and there are not many available models. The mono-iodoacetate (MIA)-induced model is the most widely used OA pain model, mainly used in rodents. MIA is an inhibitor of glyceraldehyde-3-phosphate dehydrogenase, which causes chondrocyte death, cartilage degeneration, osteophyte, and measurable changes in animal behavior. Besides, expression changes of matrix metalloproteinase (MMP) and pro-inflammatory cytokines (IL1 β and TNF α) can be detected in the MIA-induced model. Those changes are consistent with OA pathophysiological conditions in humans, indicating that MIA can induce a measurable and successful OA pain model. This study aims to describe the methodology of intra-articular injection of MIA in rats and discuss the resulting pain-related behaviors and histopathological changes.

Use of magnetic capture to identify elevated levels of CCL2 following intra-articular injection of monoiodoacetate in rats

ABSTRACT Purpose Synovial fluid biomarkers help evaluate osteoarthritis (OA) development. Magnetic capture, our new magnetic nanoparticle-based technology, has proven to be effective for determining extracellular matrix fragment levels in two rat OA models. Here, the feasibility of magnetic capture for detecting monocyte chemoattractant protein-1 (MCP-1 or CCL2) is demonstrated after intra-articular injection of monoiodoacetate (MIA) in the rat knee. Methods Forty-eight male Lewis rats received a right hind limb, intra-articular injection of MIA (1 mg in 25 µl of saline) or 25 µl of saline. Magnetic capture and lavage were performed at 7 days after injection (n = 6 per treatment per procedure), with magnetic capture additionally performed at 14 and 28 days post-injection (n = 6 per treatment per time point). CCL2 was also assessed in serum. Results Serum CCL2 levels revealed no difference between MIA and saline animals (p = 0.0851). In contrast, magnetic capture and lavage detected a significant increase of CCL2 in the MIA-injected knee, with the MIA-injected knee having elevated CCL2 compared to contralateral and saline-injected knees (p = 0.00016 (contralateral) and p = 0.00016 (saline) for magnetic capture; p = 0.00023 (contralateral) and p = 0.00049 (saline) for lavage). Conclusions Magnetic capture of CCL2 was successfully developed and applied to determine levels of CCL2 in a rat knee. Magnetic capture detected a statistically significant increase of CCL2 in MIA-injected knees compared to controls, and CCL2 levels stayed relatively stable from week 1 through week 4 post-MIA injection.

The GLP-1 agonist, liraglutide, ameliorates inflammation through the activation of the PKA/CREB pathway in a rat model of knee osteoarthritis

BackgroundInflammation is a common pathological phenomenon of osteoarthritis (OA). Accumulated evidence indicates that ameliorating or suppressing inflammation might be a promising and effective therapeutic strategy for the treatment of OA. Notably, glucagon-like peptide-1 (GLP-1)-based drugs are being successfully used to control glucose levels in patients with diabetes mellitus. In addition, recent findings have indicated that GLP-1 agonists, such as liraglutide have therapeutic potential in preventing inflammation-related disorders through the regulation of protein kinase A (PKA)/ cyclic adenosine monophosphate (cAMP) response element-binding protein (CREB) signals. Intra-articular injection of monoiodoacetate (MIA) has been widely used to induce OA. Thus, the present study aimed to investigate whether liraglutide has anti-inflammatory effects on MIA-induced OA rats and uncover its underlying molecular mechanisms.MethodsIntra-articular injection of MIA was used to induce knee OA in a rat model. Subcutaneous injection of liraglutide was used to upregulate the expression of GLP-1 receptor (GLP-1R). Western blot analysis was utilized to measure the expression of GLP-1R, PKA/CREB pathway components and inflammation-related proteins, such as tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β), and IL-6. Immunoprecipitation techniques were used to detect the interactions between GLP-1R and the PKA/CREB pathway.ResultsThe levels of GLP-1R decreased significantly in the knees of OA rats, accompanied by the downregulation of PKA /CREB signals and upregulation of inflammation-related proteins. We also found that GLP-1R interacted with the PKA/CREB pathway and that liraglutide could activate PKA/CREB signals, thereby inhibiting the expression of inflammation-related proteins.ConclusionsTogether our results suggest that liraglutide exhibits anti-inflammatory activity through the activation of the PKA/CREB pathway in an OA rat model.