Adjuvant-induced Arthritis In Rats Research Articles

Traditional Chinese Medicine Aconiti Radix Cocta Improves Rheumatoid Arthritis via Suppressing COX-1 and COX-2.

According to Traditional Chinese Medicine (TCM), Aconiti Radix Cocta (AC) is clinically employed to expel wind, remove dampness, and relieve pain. We evaluated the antirheumatoid arthritis (RA) activities and underlying mechanisms of AC. The chemical constituents of AC were analyzed by high-performance liquid chromatography (HPLC) using three reference compounds (benzoylaconitine, benzoylmesaconine, and benzoylhypacoitine). The anti-RA effects of AC were evaluated in adjuvant-induced arthritis (AIA) rats by hind paw volume and histopathological analysis. The effects of AC on inflammatory cytokines (IL-1β and IL-17A) were determined by enzyme-linked immunosorbent assay. The regulation of cyclooxygenases (COX-1 and/or COX-2) was determined by Western blot and real-time quantitative reverse transcription polymerase chain reaction analyses. AC significantly reduced paw swelling, attenuated the inflammation and bone destruction in joint tissues, and reduced IL-1β and IL-17A in the serum. Moreover, AC downregulated the expression of COX-1 and COX-2 in the synovial tissues. We also identified that AC possesses significant anti-RA activities on AIA, which may be ascribed to the regulation of inflammatory cytokines IL-1β and IL-17, as well as to the inhibition of arachidonic acid signaling pathways. Our findings provide theoretical support for AC as an effective nature-derived therapeutic agent for RA treatment.

Styphnolobium japonicum (L.) Schott Flower Extract Alleviates Oxidative Stress and Inflammatory Factors in the Adjuvant-Induced Arthritis Rat Model.

IntroductionThis research was to evaluate the beneficial effects of Styphnolobium japonicum (L.) Schott flower extract (SJF) on the adjuvant-induced arthritis rat model.MethodsArthritis was evoked by injection of complete Freund’s adjuvant (CFA) in the hind paw. SJF (150 or 300 mg/kg/day) or Celecoxib (5 mg/kg/day) were administered intragastrically from the 0th day to the 28th day. The arthritis symptoms (paw edema, arthritic scores, mechanical hyperalgesia, and thermal hyperalgesia), inflammation biomarkers (RT and CRP), related enzymes (MMP1 and MMP13), oxidative stress markers (CAT, SOD, GPx, and MDA), and inflammatory cytokines (IL-6, IFN-γ, TNF-α, and IL-1β) of SJF-treated CFA rats were evaluated.ResultsCFA rats exhibited severe arthritis symptoms, increased oxidative stress, and inflammatory cytokines. Interestingly, SJF treatment relieving arthritis symptoms and restored body weight gain compared with those in the CFA group. SJF treatment decreased the levels of CRP, RF, MMP1, and MMP13 in the CFA group. Besides, SJF treatment increased the activities of antioxidant enzymes and decreased the MDA content and inflammatory cytokines compared with those in the CFA group. Moreover, SJF could increase the mRNA expression of GPx-1 and CAT and inhibit the mRNA expression of IL-6 and TNF-α in the ankle tissue of CFA rats.

MiR-143-3p Inhibits the Differentiation of Osteoclast Induced by Synovial Fibroblast and Monocyte Coculture in Adjuvant-Induced Arthritic Rats.

Osteoclast, which mediates overactive bone resorption, is one of the key factors for bone destruction in rheumatoid arthritis (RA). Existing studies have shown that abnormal miR-143-3p expression was observed in both RA patients and arthritis animals, which can participate in osteoclast differentiation, and mitogen-activated protein kinase (MAPK) signaling pathway was closely related to osteoclast differentiation. The primary objective of the current study was to determine the role of miR-143-3p in the progression of osteoclast differentiation and its relationship with MAPK signaling pathways. The results showed that miR-143-3p inhibited osteoclast differentiation and decreased the levels of M-CSF and RANKL during osteoclast differentiation. miR-143-3p inhibited the expression of MAPK signaling proteins, which is ERK1/2 in the early stage and JNK in the later stage of osteoclast differentiation. It was also observed that MAPK inhibitors upregulated miR-143-3p expression in osteoclast differentiation. Taken together, our results suggested that miR-143-3p could inhibit the differentiation of osteoclast, which was related to inhibiting MAPK signaling pathways. This may provide a novel strategy for curing RA.

Hesperidin mitigates inflammation and modulates ectoenzymes activity and some cellular processes in complete Freund's adjuvant-induced arthritic rats.

This study was aimed at assessing the anti-arthritic effects of hesperidin on the inflammatory markers in serum/plasma, ectoenzymes activity in platelet, reactive oxygen species (ROS), apoptosis and cell cycle in bone marrow cells of a rat model of arthritis. Fifty-six adult female Wistar rats (245-274 g) were grouped into eight of seven rats each: control rats given normal saline or 40 mg/kg of hesperidin or 80 mg/kg of hesperidin, 0.2 mg/kg of dexamethasone, arthritic rats given normal saline, or 40 mg/kg of hesperidin or 80 mg/kg of hesperidin, and 0.2 mg/kg of dexamethasone. Myeloperoxidase and nitrate plus nitrite levels were evaluated in the plasma and serum, respectively. The ecto-nucleoside triphosphate diphosphohydrolases, ecto-5'-nucleotidase and ecto-adenosine deaminase activities were assessed in platelets. Subsequently, the cells of the bone marrow were obtained, and the assays for ROS, apoptosis and cell cycle were evaluated using flow cytometry. The results showed that hesperidin mitigated inflammation, modulated adenosine nucleotides and nucleoside hydrolysing enzymes and levels, minimized ROS intracellularly, attenuated apoptotic process and activated cell cycle arrest in arthritic rat. This study suggests that hesperidin could be a natural and promising anti-inflammatory compound for the management of arthritis.

Sinomenine increases adenosine A2A receptor and inhibits NF-κB to inhibit arthritis in adjuvant-induced-arthritis rats and fibroblast-like synoviocytes through α7nAChR.

Sinomenine (SIN) is a clinical drug for treating rheumatoid arthritis (RA) in China. Our previous study found SIN inhibited inflammation via alpha7 nicotinic acetylcholine receptor (α7nAChR) in macrophages in vitro. Adenosine receptor A2A has anti-inflammatory and immunosuppressive function. However, the mechanisms of SIN acting on α7nAChR and the effect on adenosine A2A receptor (A2A R) in RA are not clear. In the present study, the effects of SIN on adjuvant-induced-arthritis (AIA) rats in vivo and on fibroblast-like synoviocytes (FLSs) in vitro were investigated. Indomethacin (Indo) and methotrexate (MTX), the clinical anti-arthritis drugs, were used as controls. Nicotine (Nic), a specific agonist of α7nAChR, was used as a control for targeting α7nAChR. Alpha-bungarotoxin (α-BTX), the antagonist of α7nAChR or small interference RNA (siRNA) was used to block or knock down α7nAChR. Results showed that SIN decreased arthritis index, hind paw volume, erythrocyte sedimentation (ESR) and serum TNF-α in AIA rats, and α-BTX attenuated the earlier-mentioned effects of SIN and Nic, but not Indo and MTX. The expressions of A2A R in synovium declined in AIA rats, but remarkably increased after the intervention of SIN. The expression of A2A R decreased by LPS or TNF-α, but increased by SIN; cAMP also increased by SIN in FLSs in vitro. SIN inhibited the expression of MCP-1, IL-6, and vascular endothelial growth factor in LPS-induced FLSs. SIN inhibited the activation of NF-κB. Meanwhile, α-BTX or α7nAChR siRNA blocked the earlier-mentioned effects of SIN in FLSs. Results suggested the expressions of A2A R in synovium and FLSs are negatively correlated with the arthritis progression of AIA rats and the activation of FLSs. SIN increases A2A R and inhibits the activation of NF-κB pathway via α7nAChR in AIA rats and FLSs.

S-Propargyl-Cysteine Remodels the Gut Microbiota to Alleviate Rheumatoid Arthritis by Regulating Bile Acid Metabolism.

BackgroundRheumatoid arthritis (RA) is a long-term autoimmune disorder characterized by chronic inflammation that results in swollen and painful joints and even cartilage and bone damage. The gut microbiota, a novel anti-inflammatory target, is considered an important environmental factor in the development of RA. S-propargyl-cysteine (SPRC), an amino acid analogue, exerts anti-inflammatory, cardioprotective effects, and neuroprotective effects on various diseases. In recent studies, an SPRC treatment exerted anti-inflammatory effects on RA. Meanwhile, gut microbiome dysbiosis in individuals with RA has also been reported by many researchers. However, the relationship between SPRC and gut microbiota in individuals with RA remains unclear.MethodsThirty male Sprague-Dawley (SD) rats were randomly divided into three groups of 10 each, including the Control, Model, and SPRC groups. Adjuvant-induced arthritis (AIA) rats in SPRC group were treated with SPRC. Measurement of paw volume and serum tumor necrosis factor-α (TNF-α) and interleukin 6 (IL-6) levels were applied to evaluate the inflammatory status. Fecal samples were collected on the 14th day and 28th day. Gut microbiota were analyzed using 16S ribosomal RNA (rRNA) gene amplicon sequencing. Untargeted metabolomics on plasma samples was applied to investigate the metabolic changes induced by the altered gut microbiota by using derivatization-UHPLC-Q-TOF/MS.FindingsUsing 16S rRNA amplicon sequencing, we found that SPRC significantly altered the gut microbiota structure in AIA rats. In particular, Bifidobacterium, a genus of BSH (Bile Salt Hydrolase)-producing microbes, was overrepresented in SPRC-treated AIA rats. Additionally, a subsequent metabolomics analysis indicated that bile acid metabolism was also altered by SPRC treatment. Interestingly, glycochenodeoxycholic acid (GCDCA) and glycocholic acid (GCA), which are formed with the participation of BSH-producing microbes in the intestine, were identified as crucial biomarkers responding to SPRC treatment with significantly lowered levels.InterpretationA mechanistic link between the gut microbiota and plasma metabolites was revealed in this study, which provides insights into the mechanism of SPRC treatment for RA from the perspective of the gut microbiota.

Clinical Evidence-Guided Anti-rheumatoid Arthritis Study of Shuji Tablet in Adjuvant-Induced Arthritis Rats and Mechanism Exploration via Network Pharmacological Approach.

Background: Rheumatoid arthritis (RA) is a kind of chronic autoimmune disease with several tissues damaged. Shuji tablet (SJT) is a prescription approved for treating lumbago and leg pain in the clinic. However, the efficacy of SJT against RA is still unknown. This study aims to evaluate the therapeutic effect of SJT on adjuvant-induced arthritis (AIA) rats and explore the mechanism via a network pharmacological approach.Methods: AIA rats were treated with SJT for 30 days at the dosages of 3.6, 1.8, and 0.9 g/kg, respectively, and the anti-RA effect was determined by measuring paw swelling, systemic symptoms score, arthritis index, and histopathological change. ELISA assay was used to evaluate the level of inflammatory cytokines in serum. The mechanism exploration and target prediction of SJT against RA were performed via a network pharmacological approach.Results: SJT showed excellent alleviation on AIA rats, with evidence of reducing paws swelling, decreasing systemic symptoms score, and arthritis index. Furthermore, SJT significantly reduced the serum cytokines of IL-6, IL-1β, TNF-α in AIA rats. Histopathological examination showed SJT remarkably reduced synovial hyperplasia, cartilage damage, and inflammatory infiltration in the secondary-side paws. According to network pharmacological analysis, 208 candidate compounds and 445 potential targets of SJT were identified, and 4465 RA therapy-related targets were searched out. Subsequently, 292 target genes of SJT were speculated to be associated with RA treatment, among which the top 5 “response values” targets were STAT3, AKT1, JUN, HSP90AA1, TNF. GO and KEGG enrichment analysis suggested that 45 signaling pathways were associating with SJT treating RA. The top 10 signaling pathways were PI3K-Akt, MAPK, AGE-RAGE pathway in diabetic complications, Ras, HIF-1, TNF, Chemokine, IL-17, FoxO, and Rap1.Conclusion: Our experimental study showed that SJT significantly alleviated rheumatoid arthritis of AIA rats. Network pharmacology showed that the key targets of SJT against RA probably were STAT3, AKT1, JUN, HSP90AA1, TNF, and the potential mechanism was associated with modulation on the signaling pathways of PI3K-Akt, MAPK, Ras, AGE-RAGE, HIF-1, TNF, chemokine, IL-17, FoxO, Rap 1. Our study strongly provides evidence for Shuji tablet in RA therapy and would enlarge its application in the clinic.

Increased brain penetration of diphenhydramine and memantine in rats with adjuvant-induced arthritis

Brain penetration of cationic drugs is an important determinant of their efficacy and side effects. However, the effects of alterations in the activity of uptake transporters in the brain under inflammatory conditions on the brain penetration of cationic drugs are not fully understood. The aim of this study was to examine changes in brain penetration of cationic drugs, including diphenhydramine (DPHM), memantine (MMT), and cimetidine (CMD), and changes in the expression of uptake transporters such as organic cation transporter (Oct) in brain microvascular endothelial cells (BMECs) under inflammatory conditions. To clarify the effects of inflammation on the brain penetration of DPHM, MMT, and CMD, we performed brain microdialysis studies in a rat model of adjuvant-induced arthritis (AA). Further, differences in transporter mRNA expression levels between BMECs from control and AA rats were evaluated. Brain microdialysis showed that the unbound brain-to-plasma partition coefficient (Kp,uu,brain) for DPHM and MMT was significantly lower in AA rats compared with control rats. OCT mRNA levels were increased and proton-coupled organic cation (H+/OC) antiporter mRNA levels were decreased in AA rats compared with control rats. Taken together, our findings suggest that inflammation decreases the brain penetration of H+/OC antiporter substrates such as DPHM and MMT.

PTEN Methylation Promotes Inflammation and Activation of Fibroblast-Like Synoviocytes in Rheumatoid Arthritis

Rheumatoid arthritis (RA) is characterized by a tumor-like expansion of the synovium and subsequent destruction of adjacent articular cartilage and bone. In our previous work we showed that phosphatase and tension homolog deleted on chromosome 10 (PTEN) contributes to the activation of fibroblast-like synoviocytes (FLS) in adjuvant-induced arthritis (AIA), but the underlying mechanism is not unknown. In this study, we show that PTEN is downregulated while DNA methyltransferase (DNMT)1 is upregulated in FLS from RA patients and a rat model of AIA. DNA methylation of PTEN was increased by administration of tumor necrosis factor (TNF)-α in FLS of RA patients, as determined by chromatin immunoprecipitation and methylation-specific PCR. Treatment with the methylation inhibitor 5-azacytidine suppressed cytokine and chemokine release and FLS activation in vitro and alleviated paw swelling in vivo. PTEN overexpression reduced inflammation and activation of FLS via protein kinase B (AKT) signaling in RA, and intra-articular injection of PTEN-expressing adenovirus into the knee of AIA rats markedly reduced inflammation and paw swelling. Thus, PTEN methylation promotes the inflammation and activation of FLS in the pathogenesis of RA. These findings provide insight into the molecular basis of articular cartilage destruction in RA, and indicate that therapeutic strategies that prevent PTEN methylation may an effective treatment.

α-Mangostin Alleviated Inflammation in Rats With Adjuvant-Induced Arthritis by Disrupting Adipocytes-Mediated Metabolism-Immune Feedback.

A previously identified anti-rheumatic compound α-mangostin (MAN) possesses notable metabolism regulatory properties. In this study, we investigated the immune implication of MAN-altered fat metabolism on adjuvant-induced arthritis (AIA) in rats. Seven days after AIA induction, the rats received oral treatment of MAN at 50 mg/kg/day for 30 days. Metabolic indicators and basic clinical parameters were evaluated using samples collected on day 20 and 38 since immunization. Expression of nicotinamide phosphoribosyltransferase (NAMPT), sirtuin 1 (SIRT1), peroxisome proliferator activated receptor gamma (PPAR-γ), stearoyl-coa desaturase 1 (SCD-1), toll like receptor 4 (TLR4), prostaglandin-endoperoxide synthase 2 (COX-2), (p)-JNK, (p)-p65 and IL-1β were investigated by either RT-qPCR or immunobloting methods. In in vitro experiments, we treated (pre)-adipocytes with monocytes/macrophages and MAN, and investigated the changes of macrophages brought by pre-adipocytes co-culture. Generally, MAN restored the impaired fat anabolism in AIA rats, indicated by increased fat reservoir, leptin and adiponectin secretion, and PPAR-γ and SCD-1 expression. Meanwhile, it decreased circulating IL-1β and IL-6 levels, restored serological lipid profile changes, and relieved oxidative stresses, demonstrating potent therapeutic effects on AIA. AIA rats-derived monocytes inhibited mRNA PPAR-γ and SCD-1 expression in pre-adipocytes. Contrarily, MAN facilitated adipocyte differentiation in vitro, and increased free fatty acids production. It also significantly increased PPAR-γ and SCD-1 expression, which can be abrogated by PPAR-γ inhibitor T0070907. Similarly, lipopolysaccharide-primed macrophages inhibited PPAR-γ expression in the co-cultured pre-adipocytes, which was reversed by MAN. In the same co-culture system, lipopolysaccharide-induced inflammation was amplified by the co-existence of pre-adipocytes. More secretion of IL-1β and IL-6 and higher levels expression of COX-2, p-JNK, p-p65 and TLR4 were observed in lipopolysaccharide-treated macrophages when co-cultured by pre-adipocytes. The intensified inflammatory situation was eased by MAN. The treatment with pre-adipocytes culture medium achieved similar effects. Medium from lipopolysaccharide-treated adipocytes promoted IL-1β, IL-6 and MCP-1 production in separately cultured macrophages, and COX-2, p-JNK, p-p65 and TLR4 expression were increased at the meantime. MAN treatment on pre-adipocytes impaired these changes. It suggests that fat anabolism in AIA rats was deficient due to increased energy expenditure caused by inflammatory conditions. MAN restored fat metabolism homeostasis by up-regulating PPAR-γ, and reshaped secretion profile of adipocytes.

METTL3 Promotes Activation and Inflammation of FLSs Through the NF-κB Signaling Pathway in Rheumatoid Arthritis.

Rheumatoid arthritis (RA), a common autoimmune disease, is extremely damaging to human health. Fibroblast-like synoviocytes (FLSs) have a vital role in the occurrence and development of RA. Methyltransferase-like 3 (METTL3), which is a crucial component of the N6-methyladenosine (m6A) methyltransferase complex, is involved in the progression of many diseases. In this study, we explored the role of METTL3 in the inflammatory response and proliferation, invasion, and migration of FLSs. We used human RA synovial tissues and the adjuvant-induced arthritis (AIA) animal model of RA. Experimental results revealed that METTL3 expression was significantly upregulated in human RA synovial tissues and in the rat AIA model. METTL3 knockdown suppressed interleukin (IL)-6, matrix metalloproteinase (MMP)-3, and MMP-9 levels in human RA-FLSs and rat AIA-FLSs. In contrast, they were increased by METTL3 overexpression. Additionally, we found that, in FLSs, METTL3 may activate the nuclear factor (NF)-κB signaling pathway. The experimental results showed that METTL3 may promote FLS activation and inflammatory response via the NF-κB signaling pathway.

Qing-Luo-Yin Alleviated Monocytes/Macrophages-Mediated Inflammation in Rats with Adjuvant-Induced Arthritis by Disrupting Their Interaction with (Pre)-Adipocytes Through PPAR-γ Signaling

BackgroundThe Chinese herbal formula Qing-Luo-Yin (QLY) has been successfully used in rheumatoid arthritis treatment for decades. It exhibits notable immune and metabolism regulatory properties. Thereby, we investigated its effects on the interplay between (pre)-adipocytes and monocytes/macrophages under adjuvant-induced arthritis (AIA) circumstances.MethodsFat reservoir and histological characteristics of white fat tissues (WAT) in AIA rats receiving QLY treatment were examined upon sacrifice. Metabolic parameters, clinical indicators, and oxidative stress levels were determined using corresponding kits, while mRNA/protein expression was investigated by PCR and immunoblotting methods. M1 macrophage distribution in WAT was assessed by flow cytometry. The effects of QLY on (pre)-adipocytes were further validated by experiments in vitro.ResultsCompared with normal healthy controls, body weight and circulating triglyceride were declined in AIA rats, but serological levels of free fatty acids and low-density lipoprotein cholesterol were increased. mRNA IL-1β and iNOS expression in white blood cells and rheumatoid factor, C-reactive protein, anti-cyclic citrullinated peptide antibody, MCP-1 and IL-1β production in serum/WAT were up-regulated. Obvious CD86+CD11b+ macrophages were enriched in WAT. Meanwhile, expression of PPAR-γ and SIRT1 and secretion of adiponectin and leptin in these AIA rats were impaired. QLY restored all these pathological changes. Of note, it significantly stimulated PPAR-γ expression in the treated AIA rats. Accordingly, QLY-containing serum promoted SCD-1, PPAR-γ, and SIRT1 expression in pre-adipocytes cultured in vitro. AIA rats-derived peripheral blood mononuclear cells suppressed PPAR-γ and SCD-1 expression in co-cultured pre-adipocytes, but serum from AIA rats receiving QLY treatment did not exhibit this potential. The changes on PPAR-γ expression eventually resulted in varied adipocyte differentiation statuses. PPAR-γ selective inhibitor T0070907 abrogated QLY-induced MCP-1 production decline in LPS-primed pre-adipocytes and reduced adiponectin secretion.ConclusionQLY was potent in promoting PPAR-γ expression and consequently disrupted inflammatory feedback in WAT by altering monocytes/macrophages polarization and adipocytes differentiation.

Anti-arthritic activity of Ricinus communis L. and Withania somnifera L. extracts in adjuvant-induced arthritic rats via modulating inflammatory mediators and subsiding oxidative stress.

Objective(s):This study aimed to evaluate the anti-arthritic activity of Ricinus communis leaves’ and Withania somnifera roots’ hydroalcoholic extracts in Complete Freund’s adjuvant-induced arthritis in Wistar rats. Materials and Methods:HPLC and FT-IR analysis detected pharmacologically important phytocompounds in both plant extracts. Oral treatments including methotrexate (MTX; 3 mg/kg twice a week) and extracts at 250 and 500 mg/kg/day were initiated after arthritis induction. Changes in paw swelling, arthritic score, body weight, organ indices (thymus and spleen), hematological and biochemical parameters, and pro-/anti-inflammatory cytokine expression using qRT-PCR were assessed. Oxidative stress markers in hepatic tissue were determined. Histopathological and radiological examinations were also performed. Results:RCE (R. communis extract) and WSE (W. somnifera extract) demonstrated a reduction in paw swelling, arthritic score, and restoration of body weight and organ indices. Hematological parameters, serum inflammatory markers such as CRP and RF, and liver function markers of arthritic rats were significantly (P<0.01) ameliorated with RCE and WSE treatment. Both plants persuasively down-regulated IL-1β, IL-6, IL-17a, TNF-α, and RANKL and up-regulated IL-4, INF-γ, and OPG relative expression as well as alleviating hepatic oxidative stress parameters. Histopathological and radiological findings revealed a marked reduction in tissue inflammation and bone erosion in extracts treated groups. Conclusion:The study findings suggest that R. communis leaves and W. somnifera roots have markedly subsided inflammation and improved health through modulating pro-/anti-inflammatory cytokine expression and reducing oxidative stress.

A Novel Natural Polymers Based Nanoparticles Gel Formulation for the Treatment of Rheumatoid Arthritis: Optimization and In-vivo Evaluation

Background:In 1988, the US Food and Drug Administration permitted low dose methotrexate for the treatment of rheumatoid arthritis that would change the progression of the disease. Methotrexate is a folic acid antagonist and its systemic use causes numerous side effects; including hepatic toxicity. It would be preferable to deliver methotrexate by the topical route to reduce side-effects along with ease of administration and reduced dosing frequency. So, nanoparticle gel is a hopeful approach to treat rheumatoid arthritis.Objective:The study aims to develop a nanoparticles gel containing novel natural polymer-based methotrexate nanoparticles and evaluate its therapeutic potential on Complete Freund’s Adjuvant– Induced Arthritis rat model and compare it to methotrexate and dexamethasone gel.Materials and Methods:The five batches of methotrexate nanoparticles gel were prepared viz. F1W2, F2W2, F3W2, F4W2 and methotrexate gel for the topical application by using different concentrations of Carbopol 934 base and characterized for their evaluation parameters: homogeneity, grittiness, pH, spread-ability, viscosity determination, and drug content studies. The arthritic potential of methotrexate-nanoparticles gel was evaluated by Complete Freund’s Adjuvant–Induced Arthritis rats model based on percent inhibition oedema and arthritic score.Result and Discussion:Methotrexate nanoparticles gel significantly reduced the percentage inhibition of oedema compared to methotrexate and dexamethasone gel. The therapeutic activity of nanoparticles gel was found to be F3W2 ≥ F2W2 ≥ F1W2 ≥ F4W2 ≥ MTX gel. So, the optimized nanoparticle gel formulation F3W2 can be effective in the treatment of rheumatoid arthritis.Conclusion:The developed novel nanoparticles gel formulation can be a promising alternative to existing methotrexate and dexamethasone gel.

Phytochemical and In-Vivo Anti-Arthritic Significance of Aloe thraskii Baker in Combined Therapy with Methotrexate in Adjuvant-Induced Arthritis in Rats.

Unlike other widely known Aloe species used for treatment of rheumatoid arthritis, this species suffers from a lack of sufficient studies on its biological and chemical characters. This is what drove us to perform this work to evaluate the in vivo anti-arthritic potential of its leaf ethanolic extract. The in vivo anti-arthritic activity of the leaf ethanolic extract at 100 and 200 mg/kg/day b.wt. was evaluated alone and in combination with methotrexate (MTX) using complete Freund’s adjuvant. Serum levels of rheumatoid factor, anti-cyclic citrullinated peptide (anti-CCP), cytokines pro-inflammatory marker, inflammatory mediator serum levels, and oxidative stress mediators were analyzed, in addition to liver function. Orientin, isoorientin, β-sitosterol, its palmitate and its glucoside were isolated. The combined therapy of MTX and the leaf ethanolic extract (especially at 200 mg/kg b.wt.) group showed better activity compared to MTX alone. Moreover, the combined therapy provided additional benefits in lowering the liver toxicity by comparison to MTX alone. We concluded that a synergetic combination of the leaf ethanolic extract and MTX is beneficial in the management of rheumatoid arthritis with fewer side effects on liver function, as well as the possibility of the leaf extract to stand alone as an effective natural anti-arthritic agent.

Nicotinamide mononucleotide-elicited NAMPT signaling activation aggravated adjuvant-induced arthritis in rats by affecting peripheral immune cells differentiation

Supplement of nicotinamide mononucleotide (NMN), the direct precursor of nicotinamide adenine dinucleotide (NAD+) has gained prominence due to the significant anti-aging potentials of nicotinamide phosphoribosyltransferas (NAMPT)/NAD+ signaling. Because over-expression of NAMPT is deeply implicated in inflammatory arthritis, we investigated the effects of NMN supplement on rats with adjuvant-induced arthritis (AIA). Tested rats were given oral treatment of NMN at 200 mg/kg/day for 25 days. Arthritis score and body weight were periodically recorded. Clinical outcomes were evaluated based on arthritic manifestations, ELISA analysis and histological examination. T cells subsets were analyzed by flow cytometry. Expressions of protein and mRNA were assessed by immunoblotting and PCR methods, respectively. Levels of CD172a, CD43, and NAMPT in peripheral blood mononuclear cells (PBMCs) were investigated by immunofluorescence approach. Obtained results were further validated by experiments in vitro. Generally, NMN exacerbated AIA severity in rats. It deteriorated MMP3-controlled tissues damages, and altered immune profile by increasing Th17/Treg cells ratio. The up-regulation of NAMPT in PBMCs from NMN-treated rats was confirmed by both immunofluorescence and PCR experiments, which was synchronized with significant increase in iNOS, MCP-1, IL-1β expression. NMN-primed AIA PBMCs were potent in up-regulating MCP-1, IL-1β, MMP3 and p-JNK expression in synovioblast. NMN stimulus barely affected Th17 cells count in in vitro cultured splenocytes, but it greatly potentiated the capability of AIA monocytes in inducing IL-17α secretion and Th17 cells differentiation in the co-cultured splenocytes. It suggested that long-term NMN supplement could exacerbate inflammatory arthritis by reshaping the immune milieu through the up-regulation of NAMPT.

Anti-inflammatory activity of different isolated sites of Chloranthus serratus in complete Freund's adjuvant-induced arthritic rats.

Chloranthus serratus is a traditional Chinese medicine for treating arthritis and bruises. The aim of the present study was to investigate the anti-arthritic activities and possible associated mechanisms of different isolated sites of Chloranthus serratus (DISC) in adjuvant-induced arthritis (AA) rats. The therapeutic effects of the extracts were assessed through changes in body weights, swelling rates, arthritis indexes (AI) and organ indexes. The levels of nitric oxide (NO), malondialdehyde and superoxide dismutase were determined using one-step method, TBA method and hydroxylamine method, respectively; the levels of TNF-α, IL-1β, IL-6, prostaglandin E2, macrophage inhibitor factor-1, VEGF, immunoglobulin (Ig) G, IgM and IFN-γ in serum were determined using ELISA. Pathological changes and positive expression of VEGF in the ankle joints were investigated using hematoxylin-eosin staining and immunohistochemical staining, respectively. DISC treatment increased the weight gains and thymus indexes, and decreased the swelling rates, spleen indexes and AI in AA rats. The water isolated site (WA) and ethyl acetate isolated site (EA) significantly reversed complete Freund's adjuvant (CFA)-induced changes in the levels of NO, IL-6, TNF-α, IgG and IFN-γ, while the n-butanol isolated site (NB) only reversed the changes in IL-6 and IgG contents. Some changes in the chloroform isolated site group showed the same trend as those in the model group. The extracts relieved synovial hyperplasia, inflammatory cell infiltration and articular surface defects, and reduced the positive expression rate of VEGF in the synovial tissues of the AA rats to varying degrees. The WA exhibited the most marked effects, followed by the EA and NB, indicating that WA had optimal therapeutic effects on CFA-induced arthritic rats, which may be mediated by the oxidative stress and inhibition of inflammatory factors. C. serratus may serve as a potential candidate for the treatment of rheumatoid arthritis.

Penta-acetyl Geniposide Suppresses Migration, Invasion, and Inflammation of TNF-α-Stimulated Rheumatoid Arthritis Fibroblast-Like Synoviocytes Involving Wnt/β-Catenin Signaling Pathway.

We previously reported that penta-acetyl geniposide ((Ac)5GP, an active derivative of geniposide) showed anti-arthritic effect on adjuvant-induced arthritis (AIA) rats by promoting the apoptosis of AIA fibroblast-like synoviocyte (FLS). This study aimed to demonstrate the effects of (Ac)5GP on migration, invasion, and inflammation of TNF-α-stimulated rheumatoid arthritis (RA) FLS (MH7A cell) and to explore the involved mechanisms. MTT assay was used to determine the applied non-cytotoxic doses of (Ac)5GP (12.5, 25, 50 μM) in vitro. Results of wound-healing, transwell, and phalloidin staining assays indicated that (Ac)5GP reduced the migration, invasion, and F-actin cytoskeletal reorganization of TNF-α-stimulated MH7A. Results of ELISA and western blot assays confirmed that (Ac)5GP reduced TNF-α-induced production of pro-inflammatory cytokines (like IL-1β, IL-6, IL-8) and matrix metalloproteinases (MMPs, such as MMP-2 and MMP-9). Moreover, (Ac)5GP inhibited TNF-α-induced activation of Wnt/β-catenin pathway, evidenced by reducing the protein levels of Wnt1, p-GSK-3β (Ser9), and β-catenin and preventing β-catenin nuclear translocation. Importantly, the combination of XAV939 (an inhibitor of Wnt/β-catenin) promoted the actions of (Ac)5GP on TNF-α-induced migration, invasion, and inflammation, further revealing the involvement of Wnt/β-catenin pathway underlying the therapeutic effects of (Ac)5GP on TNF-α-stimulated MH7A. In vivo, (Ac)5GP relieved the progression and severity of rat collagen-induced arthritis, related to reducing the levels of IL-1β, IL-6, IL-8, MMP-2, and MMP-9 as well as inhibiting Wnt/β-catenin pathway in synovial tissues. Collectively, (Ac)5GP could suppress TNF-α-induced migration, invasion, and inflammation in RA FLS involving Wnt/β-catenin pathway and (Ac)5GP might be as a candidate agent for RA treatment.

Angiotensin involvement in kidney injury induced by rheumatoid arthritis in rat.

Renal injury induced by rheumatoid arthritis is not clear and may be related to the angiotensin II. We aim to investigate the adjuvant-induced arthritis (AIA) injury in rat kidney, focusing the angiotensin II/AT1 pathway. Male Wistar rats were allocated in to three groups: Control, AIA and AIA plus losartan. The AIA was induced by injection of 100µL of an emulsion of dissected Mycobacterium tuberculosis (50mg/mL) on the paw. Treatment with losartan was initiated on the first day of immunization (daily subcutaneous injection, 1mg/kg). After 60days post immunization, we evaluated kidney function by plasma creatinine, urea and uric acid levels and creatinine depuration; kidney injury by apoptosis analysis and inflammation markers such as macrophages, transforming growth factor beta (TGF-β) and inducible nitric oxide synthase (iNOS) expression; oxidative stress by plasma thiobarbituric acid reactive substances (TBARS); renal expression of angiotensin receptors subtype 1 (AT1 ) and 2 (AT2 ) and plasma concentration of angiotensin II. AIA rats showed elevated plasma levels of creatinine, urea, uric acid, TBARS and Ang II and reduced creatinine depuration, and enhanced kidney macrophage number, TGF-β, caspase-3, iNOS and AT1 /AT2 receptors expression. The losartan reduced plasma creatinine and its clearance, reduced macrophages and the expression of TGF-β and iNOS in renal tissues, and reduced plasma TBARS. We conclude that AIA causes kidney injury by a physiopathological mechanism that involves AT1 stimulation in renal tissue, elevating the presence of macrophages, the expression of TGF-β and iNOS, as well the local oxidative stress, which contribute to renal function deterioration.

Antioxidant, anti-inflammatory and anti-arthritic activities of methanol extract of Tamus communis L. roots

Purpose: To evaluate the antioxidant, anti-inflammatory and anti‐arthritic effects of the methanol extract of Tamus communis roots.&#x0D; Methods: The total phenolic and flavonoid contents were determined using Folin–Ciocalteu’s reagent and aluminium chloride assays, respectively while the antioxidant capacity was determined using DPPH and β-carotene/linoleate tests. For anti-inflammatory activity, carrageenan-induced paw edema in rat was used. Formaldehyde-induced paw edema and adjuvant-induced arthritis in rats were used to evaluate the anti-arthritic effect of the extract.&#x0D; Results: The total phenolic and flavonoid contents of T. communis showed 55.2 ± 0.036 mg GAE/g dry weight and 1.191 ± 0.002 mg QEq/g dry weight, respectively while the antioxidant activity (DPPH) showed a half-maximal inhibitory concentration (IC50 of 0.128 ± 0.011 mg/mL. Using βcarotene/linoleate assay, the extract showed 88.13 ± 4.03 % inhibition. In carrageenan-induced paw edema in rat, all doses of T. communis did not show inhibition of edema. In contrast, formaldehyde induced paw edema decreased at the dose of 150 mg/kg days 2 and 4. However, doses of 300 and 600 mg/kg did not show any activity. In adjuvant-induced arthritis, all used doses caused no reduction in arthritic scores and in paw thickness except a significant decrease with the dose 150 mg/kg at the last day.&#x0D; Conclusion: T. communis extract exhibits high antioxidant activity related to the phenolic compound contents. However, the results of the anti-inflammatory and anti‐arthritic studies did not support its use in folk medicine in the treatment of rheumatic ailment except on low doses