Adjuvant-induced Arthritis In Rats Research Articles

Effect of ethanol and aqueous extracts of seed pod of Copaifera salikounda (Heckel) on complete Freund's adjuvant-induced rheumatoid arthritis in rats.

The antirheumatoid arthritis potential of ethanol and aqueous extracts of seed pod of Copaifera salikounda (SPCS) was evaluated using the chicken collagen/complete Freund's adjuvant-induced arthritic rats model. Adjuvat-induced rats were treated with varied doses of the extracts (400, 600, and 800mg/kg body weight) and with reference drug, indomethacin for 21days. Antiarthritic evaluation was done through measurement of body weight, paw size, inflammatory makers, hematological parameters, cytokines, antioxidant enzymes, reduced glutathione, lipid peroxidation as well as histopathological examinations. Treatment with the ethanol and aqueous extracts of SPCS markedly inhibited the paw size and caused weight gain. The extracts considerably modulated the hematological as well as the antioxidant parameters. Likewise, the extract restored the altered lipid peroxidation, pro-inflammatory mediators, and inflammatory factors which further accentuate the implication in adjuvant-induced arthritis. Thus, the ethanol and aqueous extracts of SPCS showed a significant antiarthritic activity that was statistically analogous to that of indomethacin. Practical applications Copaifera salikounda (Heckel) has been used in treatment of different ailments including rheumatoid arthritis in folklore medicine. This is the first reported proof of the antiarthritic potential of the seed pod. Oxidative stress has been implicated in rheumatoid arthritis. Ethanol extract of SPCS has been shown to be predominantly rich in phenols, terpenoids, alkaloids, and flavonoids which are natural antioxidant. The present study has demonstrated that ethanol and aqueous extracts of SPCS can exert antioxidative and antiinflammatory effects, thus strengthening its antiarthritic potentials.

Penta-acetyl geniposide induces apoptosis of fibroblast-like synoviocytes from adjuvant-induced arthritis rats in vitro, associated with inhibition of NF-κB activation.

Approaches promoting fibroblast-like synoviocytes (FLS) apoptosis are considered as a meaningful strategy for rheumatoid arthritis (RA) treatment. We have previously reported the anti-arthritic effect of penta-acetyl geniposide ((Ac)5GP, an active derivative of geniposide) on adjuvant-induced arthritis (AIA) rats in vivo. The present study aimed to investigate the pro-apoptotic effect of (Ac)5GP on AIA FLS in vitro and the underlying molecular mechanisms. Rat AIA was induced by complete Freund's adjuvant, and FLS were primary-cultured from synovial tissues. AIA FLS were treated with (Ac)5GP (50, 100 and 200 μM) for 48 h and cell proliferation and apoptosis were respectively examined. The involvement of apoptosis-related proteins (Bax, Bcl-2 and caspase 3) and nuclear factor kappa B (NF-κB) signaling pathway was checked. (Ac)5GP inhibited the viability of AIA FLS and reduced the percentage of Ki67-positive cells in AIA FLS. Particularly, (Ac)5GP promoted AIA FLS apoptosis in vitro by inducing apoptotic nuclear morphology, facilitating DNA ladder formation and increasing percentages of both early and late apoptotic cells. (Ac)5GP treatment on AIA FLS decreased Bcl-2 protein level whereas increased the levels of Bax and caspase 3 proteins. Moreover, (Ac)5GP reduced the degradation and phosphorylation of IκBα, down-regulated NF-κB p65 protein level in nucleus and inhibited NF-κB p65 nuclear translocation. (Ac)5GP had a potent pro-apoptotic effect on AIA FLS in vitro, which is associated with regulating apoptosis-related proteins and inhibiting NF-κB activation.

Delivery of siRNA using folate receptor-targeted pH-sensitive polymeric nanoparticles for rheumatoid arthritis therapy

Systemic delivery of siRNA to target tissues is difficult to achieve owing to its limited cellular uptake and poor serum stability. Herein, polymeric nanoparticles were developed for systemic administration of siRNA to inflamed tissues. The polymeric nanoparticles were composed of PK3 as a pH-sensitive polymer, folate-polyethyleneglycol-poly(lactide-co-glycolide) as a targeting ligand, and a DOTAP/siRNA core. The polymeric nanoparticles had a mean particle size of 142.6 ± 0.61 nm and a zeta potential of 3.6 ± 0.43 mV. In vitro studies indicated pH-dependent siRNA release from polymeric nanoparticles, with accelerated release at pH 5.0. Cellular uptake was efficient and gene silencing was confirmed by Western blot. In vivo, polymeric nanoparticles were shown to have inflammation-targeting activity and potent therapeutic effects in an adjuvant-induced arthritis rat model. These results suggest that pH-sensitive and folate receptor-targeted nanoparticles are a promising drug carrier for siRNA delivery for rheumatoid arthritis.

Eccentric training enhances the αB-crystallin binding to the myofibrils and prevents skeletal muscle weakness in adjuvant-induced arthritis rat.

Patients with rheumatoid arthritis (RA) frequently suffer from muscle weakness. We examined whether eccentric training prevents skeletal muscle weakness in adjuvant-induced arthritis (AIA) rat, a widely used animal model for RA. AIA was induced in the knees of Wistar rats by injection of complete Freund's adjuvant. To induce eccentric contractions (ECCs), neuromuscular electrical stimulation (45 V) was applied to the plantar flexor muscles simultaneously with forced dorsiflexion of the ankle joint (0-40°) and was given every 6 s. ECC exercise was applied every other day for a total of 11 sessions and consisted of 4 sets of 5 contractions. There was a significant reduction in in vitro maximum Ca2+-activated force in skinned fibers in gastrocnemius muscle from AIA rats. These changes were associated with reduced expression levels of contractile proteins (i.e., myosin and actin), increased levels of inflammation redox stress-related biomarkers (i.e., TNF-α, malondialdehyde-protein adducts, NADPH oxidase 2, and neuronal nitric oxide synthase), and autolyzed active calpain-1 in AIA muscles. ECC training markedly enhanced the steady-state levels of αB-crystallin, a small heat shock protein, and its binding to the myofibrils and prevented the AIA-induced myofibrillar dysfunction, reduction in contractile proteins, and inflammation-oxidative stress insults. Our findings demonstrate that ECC training preserves myofibrillar function without muscle damage in AIA rats, which is at least partially attributable to the protective effect of αB-crystallin on the myofibrils against oxidative stress-mediated protein degeneration. Thus ECC training can be a safe and effective intervention, counteracting the loss of muscle strength in RA patients. NEW & NOTEWORTHY Eccentric contractions (ECCs) are regarded as an effective way to increase muscle strength. No studies, however, assess safety and effectiveness of ECC training on muscle weakness associated with rheumatoid arthritis. Here, we used adjuvant-induced arthritis (AIA) rats to demonstrate that ECC training prevents intrinsic contractile dysfunction without muscle damage in AIA rats, which may be attributed to the protective effect of αB-crystallin on the myofibrils against inflammation-oxidative stress insults.

Over-Expression of PTEN Suppresses the Proliferation and Migration of Fibroblast-Like Synoviocytes in Adjuvant-Induced Arthritis.

Rheumatoid arthritis (RA) is characterized by a tumor-like expansion of the synovium and the subsequent destruction of adjacent articular cartilage and bone. Recent studies have shown that phosphatase and tension homolog deleted on chromosome 10 (PTEN) might contribute to the surviva of fibroblast-like synoviocytes (FLSs) and the production of pro-inflammatory cytokines in RA. The purpose of this study was to explore the functions and underlying mechanisms of PTEN in the proliferation and migration of FLSs. FLSs were obtained from adjuvant-induced arthritis (AIA) and normal rats. The expression levels of PTEN, c-Myc, cyclin D1, PCNA, and MMP-9 were detected by quantitative-real-time-PCR and western blot assay. A BrdU proliferation assay, cell cycle analysis, and a wound-healing assay were used to study the role of PTEN in FLSs treated with PTEN inhibitor bpv, specific small interfering RNA targeting PTEN (PTEN-RNAi) or a PTEN over-expression vector (PTEN-GV141). Chromatin immunoprecipitation and methylation-special PCR assays were used to study the expression of PTEN mRNA in the presence of DNA methylation. PTEN expression was downregulated in AIA FLSs in comparison to normal rats. Moreover, inhibition of PTEN expression by bpv or PTEN-RNAi could promote the proliferation and migration of FLSs, and increase the expression of c-Myc, cyclin D1, PCNA, and MMP-9 in AIA FLSs, but had no effect on TIMP-1 expression.In addition, transfection of AIA FLSs with PTEN-GV141 reduced their proliferation and migration. Further study indicated that DNA methylation could regulate PTEN expression in AIA. Our findings suggest that PTEN might play a pivotal role in the proliferation and migration of FLSs through the activation of the AKT signaling pathway. Additionally, PTEN expression may be regulated by DNA methylation in the pathogenesis of AIA.

Deciphering the pharmacological mechanism of Guan-Jie-Kang in treating rat adjuvant-induced arthritis using omics analysis.

Traditional Chinese medicine (TCM) formulas have attracted increasing attention worldwide in the past few years for treating complex disease including rheumatoid arthritis. However, their mechanisms are complex and remain unclear. Guan-Jie-Kang (GJK), a prescription modified from "Wu Tou Decoction," was found to significantly relieve arthritis symptoms in rats with adjuvant-induced arthritis after 30-day treatment, especially in the 24 g/kg/day group. By analyzing 1749 targets related to 358 compounds in the five herbs of GJK, we identified the possible anti-arthritis pathways of GJK, including the calcium signaling and metabolic pathways. Bone damage levels were assessed by micro-computed tomography, and greater bone protective effect was observed with GJK treatment than with methotrexate. Receptor activator of nuclear factor κB ligand (RANKL)-RANK signaling, which is related to calcium signaling, was significantly regulated by GJK. Moreover, a target metabolomics assay of serum was conducted; 17 metabolic biomarkers showed significant correlations with treatment. An integrated pathway analysis revealed that pyruvate metabolism, purine metabolism, and glycolysis metabolism were significantly associated with the effects of GJK in arthritis treatment. Thus, this study establishes a new omics analytical method integrated with bioinformatics analysis for elucidating the multi-pathway mechanisms of TCM.

Cannabinoid Receptor 2 Agonist JWH-015 Inhibits Interleukin-1β-Induced Inflammation in Rheumatoid Arthritis Synovial Fibroblasts and in Adjuvant Induced Arthritis Rat via Glucocorticoid Receptor.

Management of pain in the treatment of rheumatoid arthritis (RA) is a priority that is not fully addressed by the conventional therapies. In the present study, we evaluated the efficacy of cannabinoid receptor 2 (CB2) agonist JWH-015 using RA synovial fibroblasts (RASFs) obtained from patients diagnosed with RA and in a rat adjuvant-induced arthritis (AIA) model of RA. Pretreatment of human RASFs with JWH-015 (10–20 μM) markedly inhibited the ability of pro-inflammatory cytokine interleukin-1β (IL-1β) to induce production of IL-6 and IL-8 and cellular expression of inflammatory cyclooxygenase-2 (COX-2). JWH-015 was effective in reducing IL-1β-induced phosphorylation of TAK1 (Thr184/187) and JNK/SAPK in human RASFs. While the knockdown of CB2 in RASFs using siRNA method reduced IL-1β-induced inflammation, JWH-015 was still effective in eliciting its anti-inflammatory effects despite the absence of CB2, suggesting the role of non-canonical or an off-target receptor. Computational studies using molecular docking and molecular dynamics simulations showed that JWH-105 favorably binds to glucocorticoid receptor (GR) with the binding pose and interactions similar to its well-known ligand dexamethasone. Furthermore, knockdown of GR using siRNA abrogated JWH-015's ability to reduce IL-1β-induced IL-6 and IL-8 production. In vivo, administration of JWH-015 (5 mg/kg, daily i.p. for 7 days at the onset of arthritis) significantly ameliorated AIA in rats. Pain assessment studies using von Frey method showed a marked antinociception in AIA rats treated with JWH-015. In addition, JWH-015 treatment inhibited bone destruction as evident from micro-CT scanning and bone analysis on the harvested joints and modulated serum RANKL and OPG levels. Overall, our findings suggest that CB2 agonist JWH-015 elicits anti-inflammatory effects partly through GR. This compound could further be tested as an adjunct therapy for the management of pain and tissue destruction as a non-opioid for RA.

Alpha7 nAChR Expression Is Correlated with Arthritis Development and Inhibited by Sinomenine in Adjuvant-Induced Arthritic Rats.

Sinomenine (SIN) is the active ingredient of the Chinese herb Sinomenium acutum that has been used to treat rheumatoid arthritis (RA) for about 30 years in China. Marked expression of the alpha7 nicotinic acetylcholine receptor (α7nAChR) in the joint synovium of RA patients suggested a relationship between α7nAChR and RA. This study investigated the relationship between α7nAChR and RA development and the effects of SIN on α7nAChR expression in vivo and in vitro. Sprague-Dawley rats were injected with complete Freund's adjuvant to induce arthritis and then treated with SIN or methotrexate (MTX) from day 0 to day 30. Four clinical parameters—paw volume, arthritic index (AI), serum TNF-α concentration, and erythrocyte sedimentation rate (ESR)—were measured. Splenic lymphocytes were isolated for Bacille Calmette Guerin (BCG) stimulation. α7nAChR expression in tissues and cells was examined by RT-PCR, western blot, immunofluorescence, flow cytometry, and immunohistochemistry. Cell proliferation was evaluated by the CCK-8 assay. The relationship between α7nAChR expression and the four clinical parameters was analyzed by single-factor correlation analysis. Our results showed that the paw volume, AI, TNF-α concentration, and ESR in adjuvant-induced arthritic (AIA) rats were reduced by SIN or MTX treatment. SIN decreased α7nAChR expression in tissues and cells compared to the model group, while MTX had no significant effect on α7nAChR expression. Moreover, there was a positive relationship between α7nAChR expression and paw swelling, AI, and TNF-α concentration. Splenic lymphocyte activation was accompanied by increased α7nAChR expression, while SIN treatment inhibited cell activation and downregulated α7nAChR expression. α7nAChR expression showed a positive correlation with the progression of RA in AIA rats that may involve lymphocyte activation. Different from MTX, the inhibition of SIN on α7nAChR expression might contribute to its antiarthritic effect, suggesting that SIN could be an important supplement to the treatment strategy for RA.

Cervus and cucumis peptides ameliorates bone erosion in experimental arthritis by inhibiting osteoclastogenesis

ObjectiveRheumatoid arthritis is an autoimmune disease characterised by inflammation and bone loss, leading to joint destruction and deformity. The cervus and cucumis polypeptide (CCP) injection, one of the traditional Chinese...

Increased lymphatic circulation decreases CD4 counts in the lymph nodes of rats in adjuvant-induced arthritis

Abstract Rheumatoid arthritis (RA) is a disorder characterized by chronic inflammation. Rat adjuvant-induced arthritis (AIA) is a RA model with dysfunctional lymphatic circulation. We hypothesized that increasing lymphatic circulation would decrease arthritic inflammation. In order to determine if increasing lymphatic flow would decrease arthritic inflammation, we utilized osteopathic lymphatic manipulation (Lymphatic pump treatment (LPT)) during either the onset or during arthritis. LPT was performed by gradually increasing compression on the rat chest wall over 5 seconds followed by rapid release for 5 seconds. This technique was repeated for 1 minute 3X/day for 7 consecutive days either before the onset of inflammation (preventative treatment) or after measurable inflammation (therapeutic treatment). Articular index scoring, ankle caliper measurements, and peripheral blood, spleens, and popliteal lymph nodes were collected. In both the preventative and therapeutic studies, the LPT-treated rats had reduced inflammation. Therapeutically treated animals also had significantly decreased CD4+ lymphocytes in the draining popliteal lymph nodes, with increases in CD4+ cells in the spleen. These results suggest that LPT can reduce arthritis and alter lymphocyte trafficking in secondary lymphoid organs during inflammation. These data suggest that forced lymphatic circulation through osteopathic manipulation may serve to enhance current treatments to decrease inflammatory diseases.

AK106-001616, a Potent and Selective Inhibitor of Cytosolic Phospholipase A2: In Vivo Efficacy for Inflammation, Neuropathic Pain, and Pulmonary Fibrosis.

3-[3-Amino-4-(indan-2-yloxy)-5-(1-methyl-1H-indazol-5-yl)-phenyl]-propionic acid (AK106-001616) is a novel, potent, and selective inhibitor of the cytosolic phospholipase A2 (cPLA2) enzyme. Unlike traditional nonsteroidal anti-inflammatory drugs and selective cyclooxygenase-2 inhibitors, AK106-001616 reduced prostaglandin E2 (PGE2) and leukotriene B4 (LTB4) production by stimulated cells. The suppression of PGE2 and LTB4 production was also confirmed using an air pouch model in rats administered a single oral dose of AK106-001616. AK106-001616 alleviated paw swelling in a rat adjuvant-induced arthritis (AIA) model. The maximum effect of the inhibitory effect of AK106-001616 was comparable with that of naproxen on paw swelling in a rat AIA model. Meanwhile, the inhibitory effect of AK106-001616 was more effective than that of naproxen in the mouse collagen antibody-induced arthritis model with leukotrienes contributing to the pathogenesis. AK106-001616 dose dependently reversed the decrease in paw withdrawal threshold not only in rat carrageenan-induced hyperalgesia, but also in a rat neuropathic pain model induced by sciatic nerve chronic constriction injury (CCI). However, naproxen and celecoxib did not reverse the decrease in the paw withdrawal threshold in the CCI model. Furthermore, AK106-001616 reduced the disease score of bleomycin-induced lung fibrosis in rats. In addition, AK106-001616 did not enhance aspirin-induced gastric damage in fasted rats, increase blood pressure, or increase the thromboxane A2/ prostaglandin I2 ratio that is thought to be an underlying mechanism of thrombotic cardiovascular events increased by selective cyclooxygenase-2 inhibitors. Taken together, these data demonstrate that oral AK106-001616 may provide valuable effects for wide indications without attendant gastrointestinal and cardiovascular risks.

Guggulipid ameliorates adjuvant-induced arthritis and liver oxidative damage by suppressing inflammatory and oxidative stress mediators

BackgroundArthritis is a common degenerative joint disease characterized by deterioration of articular cartilage, subchondral bone, and associated with immobility, pain and inflammation. The incessant action of reactive oxygen species (ROS) during progressive arthritis causes severe oxidative damage to vital organs and circulatory system. PurposeIn this study we investigated the ability of guggulipid (GL), a lipid rich extract from the gum resin of the plant Commiphora whighitii to suppress the progressive arthritis and associated liver oxidative stress both in vivo and in vitro. Study design/MethodsThe anti-arthritic ability of GL was demonstrated in vitro using IL-1β stimulated bovine nasal cartilage model and in vivo Freund's complete adjuvant-induced arthritic rat model. Collagen/proteoglycan degradation and pro-inflammatory mediators were monitored in the harvested culture medium of nasal cartilage by estimating the levels of matrix metalloproteinases (MMPs), hydroxy proline, glycosaminoglycans and inflammatory mediators. Further, anti-arthritic ability of GL was evaluated in vivo by measuring enzymatic and non-enzymatic mediators of cartilage degradation, inflammation and oxidative stress markers. ResultsGL significantly inhibited the IL-1β stimulated cartilage degradation in vitro by mitigating the MMPs activity, collagen degradation and secretion of pro-inflammatory mediators. Further, GL significantly reduced the adjuvant-induced paw swelling and body weight loss in vivo. GL remarkably reduced the MMPs and hyaluronidases activities in serum and bone homogenate along with altered hematological parameters. GL also mitigated the elevated bone resorbing enzymes cathepsins, exoglycosidases and phosphatases. Additionally, GL effectively mitigated ROS and oxidative stress-mediators recuperating the altered serum/liver oxidative stress and liver damage incurred during arthritic progression. ConclusionIn summary, the study clearly demonstrates the protective efficacy of GL against arthritis and its associated oxidative stress, particularly, liver oxidative damage. Hence, GL could be a potential alternative and complementary medicine to treat inflammatory joint diseases.

Effect of umbelliferone on adjuvant-induced arthritis in rats by MAPK/NF-κB pathway.

PurposeUmbelliferone (Umb), a member of coumarin family, is found in many plants and is a promising molecule with potential anti-inflammatory, anti-oxidative, and anti-tumor activities. However, the effect of Umb on arthritis remains unclear.MethodsA rat model with Freund’s complete adjuvant (FCA)-induced arthritis was developed and used to test the efficacy of Umb on arthritis rats. Rats were given an intragastric injection of Umb (20 and 40 mg/kg) once daily from days 21 to 28 after the administration of FCA. Hind paw volume was assessed using a volume meter. The pro-inflammatory cytokine levels and prostaglandin E2 (PEG2) level in serum and synovial fluid were detected by ELISA. HE staining was used to determine representative histological changes in joint tissues, and Western blot analysis was employed to study the effects of Umb on MAPK/NF-κB signaling pathway.ResultsOur results showed that Umb suppressed the release of IL-6, IL-1β, tumor necrosis factor-alpha, and PEG2. In addition, Umb could also dramatically ameliorate the pathological changes observed in rat joints. Based on the results of Western blot, we also observed that Umb could strikingly suppress the expression of MAPK/NF-κB pathway molecules.ConclusionThese results proved that treatment with Umb is very effective for arthritis and inhibiting the MAPK/NF-κB signaling pathway may be a potential therapeutic target for treatment of arthritis.

Venlafaxine alleviates complete Freund's adjuvant-induced arthritis in rats: Modulation of STAT-3/IL-17/RANKL axis

AimsRheumatoid arthritis is usually accompanied by various comorbidities especially on the psychological side such as depression. This study aimed at revealing the potential curative effects of venlafaxine (VFX), a serotonin/norepinephrine reuptake inhibitor (SNRI), on experimentally-induced arthritis in rats. MethodsArthritis was induced by injecting complete Freund's adjuvant (CFA, 0.1 ml, s.c.). One day thereafter, VFX (50 mg/kg, p.o.) was given for 21 days. Methotrexate was used as a standard disease modifying anti-rheumatic drug. Key findingsCFA injection caused prominent arthritis evident by the increase in the hind paw and ankle diameter accompanied by elevating tumor necrosis factor-alpha, interleukin-6, interleukin-17 and matrix metalloproteinase-3 levels, effects that were diminished by VFX. Moreover, VFX down regulated gene expressions of receptor activator of nuclear factor kappa-B (NF-кB) ligand and signal transducer and activator of transcription-3 beside hampering immunohistochemical expression of vascular endothelial growth factor and NF-кB. This SNRI also improved the oxidant status of the hind limb as compared to the arthritic group. Nonetheless, MTX was better in amendment of arthritis authenticated by its effect on some inflammatory and oxidative stress biomarkers. SignificanceThis study provides a novel therapeutic use of VFX as a considerable anti-arthritic drug and offers an incentive to expand its use in RA.

The effects of laser acupuncture on the modulation of cartilage extracellular matrix macromolecules in rats with adjuvant-induced arthritis.

ObjectivesArticular cartilage damage related to irreversible physical disability affects most patients with chronic rheumatoid arthritis (RA). Strategies targeting the preservation of cartilage function are needed. Laser acupuncture (LA) can be an emerging alternative therapy for RA; however, its molecular mechanism underlying the beneficial effect on cartilage has not been elucidated. This study aimed to examine the potential chondroprotective effects of LA on extracellular matrix (ECM) macromolecules and proinflammatory cytokines in the articular cartilage of adjuvant-induced arthritis (AIA) rats and explore its related mechanisms.DesignMonoarthritis was induced in adult male Sprague–Dawley rats (250–300 g) via intraarticular injection of complete Freund’s adjuvant (CFA) into the tibiotarsal joint. Animals were treated with LA at BL60 and KI3 acupoints three days after CFA administration with a 780 nm GaAlAs laser at 15 J/cm2 daily for ten days. The main outcome measures including paw circumference, paw withdrawal threshold, histopathology and immunoassays of tumor necrosis factor-α (TNF-α), collagen type II (CoII), cartilage oligomeric matrix protein (COMP) were analyzed.ResultsLA significantly reduced ankle edema and inflammation-induced hyperalgesia in AIA rats (P < 0.05). Moreover, the TNF-α levels were significantly decreased while CoII, COMP and proteoglycans proteins were significantly enhanced following LA stimulation of the AIA cartilage compared to those treated with sham-LA (P < 0.05).ConclusionsLA attenuates cartilage degradation in AIA rat by suppressing TNF-α activation and up-regulating ECM macromolecules, suggesting LA might be of potential clinical interest in RA treatment.

Investigation of Efficacy Enhancing and Toxicity Reducing Mechanism of Combination of Aconiti Lateralis Radix Praeparata and Paeoniae Radix Alba in Adjuvant-Induced Arthritis Rats by Metabolomics.

Combination of Aconiti Lateralis Radix Praeparata (FZ) and Paeoniae Radix Alba (BS) shows a significant effect in rheumatoid arthritis (RA). This study aimed to investigate the efficacy enhancing and toxicity reducing mechanism of combination of them in adjuvant-induced arthritis (AIA) rats by metabolomics. Rats were randomly divided into seven groups, including A (healthy control), B (model control), C1 (therapy group), C2 (efficacy enhancing group), D1 (toxicity group), and D2 (toxicity reducing group), and dexamethasone group was used as positive control. The plasma biochemical indexes showed that therapeutic dose of lipid-soluble alkaloids of FZ could significantly inhibit the concentrations of IL-1β, TNF-α, and IFN-γ in AIA rats, and combination with total glucosides of peony could further reduce the concentration of IL-1β. Then, UPLC-LTQ/Orbitrap MS with untargeted metabolomics was performed to identify the possible metabolites and pathways. Through multivariate data analysis of therapeutic dose groups (A vs. B vs. C1 vs. C2) and multivariate data analysis of toxic dose groups (A vs. B vs. D1 vs. D2), 10 and 7 biomarkers were identified based on biomarker analysis, respectively. After inducing AIA model, the plasma contents of spermidine, vanillylmandelic acid, catechol, and linoleate were increased significantly, and the contents of citric acid, L-tyrosine, L-phenylalanine, leucine, L-tryptophan, and uridine 5'-monophosphate (UMP) were decreased significantly. High dose of lipid-soluble alkaloids of FZ could increase the plasma contents of L-lysine, L-arginine, and deoxycholic acid, while the plasma contents of UMP, carnitine, N-formylanthranilic acid, and adenosine were decreased significantly. The pathway analysis indicated that therapeutic dose of lipid-soluble alkaloids of FZ could regulate energy and amino acid metabolic disorders in AIA rats. However, toxic dose could cause bile acid, fat, amino acid, and energy metabolic disorders. And combination with total glucosides of peony could enhance the therapeutic effects and attenuate the toxicity induced by lipid-soluble alkaloids of FZ.

Vascular Arginase Is a Relevant Target to Improve Cerebrovascular Endothelial Dysfunction in Rheumatoid Arthritis: Evidence from the Model of Adjuvant-Induced Arthritis.

Emerging data revealed that rheumatoid arthritis (RA) is associated with higher risk of cerebrovascular diseases. Whereas cerebral endothelial dysfunction is acknowledged as a critical aspect of cerebrovascular diseases, its presence in RA and the mechanisms involved are currently unknown. By using the model of rat adjuvant-induced arthritis (AIA), the present study investigated cerebrovascular reactivity in pressurized middle cerebral arteries (MCA) on day 33 post-immunization. The results revealed that arthritis induced a dramatic decrease in the vasodilatory response to acetylcholine (ACh), ADP, and bradykinin (n = 7-9 arteries, p < 0.0001). By using nor-NOHA, L-NAME, BH4, and Tempol, the results showed that the reduced response to ACh relied on arginase overactivation (n = 8), low NOS activity (n = 8), BH4 deficiency (n = 9), and excessive superoxide production (n = 9). Immunohistological analysis revealed an endothelial upregulation of arginase 2 (p < 0.05, n = 5-6) and NADPH oxidase (p < 0.05, n = 5-7) while eNOS expression was unchanged in AIA (n = 6). To assess whether arginase inhibition may be a relevant therapeutic, AIA rats were treated with an arginase inhibitor (nor-NOHA, 40mg/kg/day, i.p., n = 20 rats) daily from day 10 to day 33 post-immunization. The treatment alleviated the impaired response of MCA to endothelium-dependent agonists, through an increase in NOS signaling and a suppression of BH4 deficiency and superoxide overproduction. By contrast, it did not change the course of arthritis. In conclusion, arthritis induced a cerebrovascular endothelial dysfunction involving an imbalance in the arginase/NOS pathway. Arginase inhibition appears as a promising therapy beyond anti-rheumatic drugs for reducing the risk of cerebrovascular diseases in RA.

Therapeutic effect of Parmotrema tinctorum against complete Freund’s adjuvant-induced arthritis in rats and identification of novel Isophthalic ester derivative

The present study reveals the anti-arthritic potential of traditionally used Parmotrema tinctorum (Pt) on experimental rats and purification of novel Isophthalic ester derivative. Arthritis was induced in rats using Freund’s complete adjuvant (CFA) and subsequently treated with Pt extract (100 & 200 mg/kg.b.w). Assessment of antiarthritic activity was carried out using paw volume, arthritic score, haematological, biochemical, tissue antioxidant, histopathology and radiological analyses of ankle joints. The results revealed that continuous administration of Pt reduces the complication associated with arthritis by inhibiting the edema formation and arthritic score significantly (P < 0.05). The altered changes in biochemical parameters were brought back with an improvement in free radical scavenging ability after treatment with Pt significantly. Further purification of Pt using conventional column chromatography led to the isolation of four compounds and the structure of these isolated compounds were elucidated on the basis of spectral data’s FT-IR, 1H NMR, 13C NMR, DEPT-NMR spectroscopy, COSY & HSQC-NMR spectroscopy and LC–MS. The spectral data revealed that the three compounds were found to be Methyl-γ-Orsellinate, Atranorin, and usnic acid (CI-III) along with a novel secondary metabolite, (C-IV)4-Hydroxy-5-methyl-isophthalicacid3-(3,4-dihydroxy-5-methyl-phenyl)ester (C16H14O7,318.1amu). The present study reveals that Parmotrema tinctorum reduces complications associated with arthritis and the compounds were isolated for the first time.

Anti-inflammatory and antioxidative properties of honey bee venom on Freund's Complete Adjuvant-induced arthritis model in rats

The aim of this study was to assess anti-inflammatory, anti-oxidant, anti-genotoxic and immunomodulatory effects of honey bee venom (HBV) on adjuvant-induced arthritis in rats. Thirty-five rats were equally divided into a negative control (NC), a positive control (PC) and low, moderate and high doses (2, 4 and 20 mg/kg, respectively) of HBV treatment groups. Freund's Complete Adjuvant (FCA) was given to the rats to form arthritis. The treatment groups were treated with HBV for 3 consecutive weeks. After the treatment, plasma IL-1β, IL-6, TNF-α, IFN-γ and TGF-β1, total oxidant status (TOS), total antioxidant status (TAS) and myeloperoxidase (MPO) activities and mononuclear leukocyte (MNL) DNA damage levels were measured. Oxidative stress index (OSI) was calculated. IL-1β, IL-6, TNF-α, TGF-β1, IFN-γ, TOS, OSI, DNA damage levels and MPO activities were significantly higher and TAS levels were lower in the PC group than the NC. After low-doses of HBV treatment IL-1β, IL-6, TNF-α, TGF-β1, TOS, OSI, MPO and MNL-DNA damage levels significantly decreased according to the PC, while IFN-γ and TAS levels increased. The differences in moderate and high-dose HBV treatment groups were not as significant as low HBV doses. Low-doses of HBV has been shown to treat RA with anti-inflammatory and antioxidant effects, by preventing DNA damage. However, these effects have not been observed as strong at higher doses of HBV. In summary, HBV may be an effective option to ameliorate RA, but the optimization of the therapeutic dose has a crucial role.

Methotrexate improves the anti-arthritic effects of Paeoniflorin-6'-O-benzene sulfonate by enhancing its pharmacokinetic properties in adjuvant-induced arthritis rats

Paeoniflorin-6'-O-benzene sulfonate (CP-25), is a novel derivative of paeoniflorin (Pae) with improved anti-arthritic effects. The aim of this study was to evaluate the therapeutic effects and pharmacokinetics of CP-25 when co-administered with MTX in adjuvant-induced arthritis (AA) rats. AA rats were randomly divided into 6 groups and treated as follows: TGP (50 mg/kg/day, ig), CP-25 (50 mg/kg/day, ig), MTX (0.5 mg/kg, 3 times/week, ig), TGP (50 mg/kg/day, ig) + MTX (0.5 mg/kg, 3 times/week, ig) and CP-25 (50 mg/kg/day, ig) + MTX (0.5 mg/kg, 3 times/week, ig) from days 14 - 35 after induction. Clinical, biochemical, and histological scores were used to assess the severity of arthritis while novel ultra-performance liquid chromatography mass spectrometry (UPLC-MS/MS) was used to detect plasma concentration of CP-25. Co-administration of CP-25 and MTX significantly reduced clinical signs of inflammation, suppressed the serum production of IL-17 compared to TGP (50 mg/kg/day, ig) and MTX (0.5 mg/kg, 3 times/week, ig). Pharmacokinetics studies showed increased AUC0-t, Cmax, and t1/2 of CP-25 while V/F and CL/F decreased when co-administered with MTX. We observed an increase concentration and longer exposure with decreased clearance of CP-25 when combined with MTX, that should be further explored for clinical RA therapy.