Adjuvant-induced Arthritis Mice Research Articles

Protective effect of marine animals hydrolysis-peptides on adjuvant-induced arthritis in mice by gut microbiota and SCFAs

Protective effect of marine animals hydrolysis-peptides on adjuvant-induced arthritis in mice by gut microbiota and SCFAs

Acupuncture alleviates inflammatory pain by activating CXCL1/CXCR2 signaling in the primary somatosensory cortex in adjuvant induced arthritis rats.

To observe whether acupuncture up-regulates chemokine CXC ligand 1 (CXCL1) in the brain to play an analgesic role through CXCL1/chemokine CXC receptor 2 (CXCR2) signaling in adjuvant induced arthritis (AIA) rats, so as to reveal its neuro-immunological mechanism underlying improvement of AIA. BALB/c mice with relatively stable thermal pain reaction were subjected to planta injection of complete Freund adjuvant (CFA) for establishing AIA model, followed by dividing the AIA mice into simple AF750 (fluorochrome) and AF750+CXCL1 groups (n=2 in each group). AF750 labeled CXCL1 recombinant protein was then injected into the mouse's tail vein to induce elevation of CXCL1 level in blood for simulating the effect of acupuncture stimulation which has been demonstrated by our past study. In vivo small animal imaging technology was used to observe the AF750 and AF750+CXCL1-labelled target regions. After thermal pain screening, the Wistar rats with stable pain reaction were subjected to AIA modeling by injecting CFA into the rat's right planta, then were randomized into model and manual acupuncture groups (n=12 in each group). Other 12 rats that received planta injection of saline were used as the control group. Manual acupuncture (uniform reinforcing and reducing manipulations) was applied to bilateral "Zusanli" (ST36) for 4×2 min, with an interval of 5 min between every 2 min, once daily for 7 days. The thermal pain threshold was assessed by detecting the paw withdrawal latency (PWL) using a thermal pain detector. The contents of CXCL1 in the primary somatosensory cortex (S1), medial prefrontal cortex, nucleus accumbens, amygdala, periaqueductal gray and rostroventromedial medulla regions were assayed by using ELISA, and the expression levels of CXCL1, CXCR2 and mu-opioid receptor (MOR) mRNA in the S1 region were detected using real time-quantitative polymerase chain reaction. The immune-fluorescence positive cellular rate of CXCL1 and CXCR2 in S1 region was observed after immunofluorescence stain. The immunofluorescence double-stain of CXCR2 and astrocyte marker glial fibrillary acidic protein (GFAP) or neuron marker NeuN or MOR was used to determine whether there is a co-expression between them. In AIA mice, results of in vivo experiments showed no obvious enrichment signal of AF750 or AF750+CXCL1 in any organ of the body, while in vitro experiments showed that there was a stronger fluorescence signal of CXCL1 recombinant protein in the brain. In rats, compared with the control group, the PWL from day 0 to day 7 was significantly decreased (P<0.01) and the expression of CXCR2 mRNA in the S1 region significantly increased in the model group (P<0.05), while in comparison with the model group, the PWL from day 2 to day 7, CXCL1 content, CXCR2 mRNA expression and CXCR2 content, and MOR mRNA expression in the S1 region were significantly increased in the manual acupuncture group (P<0.05, P<0.01). Immunofluorescence stain showed that CXCR2 co-stained with NeuN and MOR in the S1 region, indicating that CXCR2 exists in neurons and MOR-positive neurons but not in GFAP positive astrocytes. Acupuncture can increase the content of CXCL1 in S1 region, up-regulate CXCR2 on neurons in the S1 region and improve MOR expression in S1 region of AIA rats, which may contribute to its effect in alleviating inflammatory pain.

Protective effect of ethanol extract of Coriolopsis aspera fruiting bodies against adjuvant-induced arthritis mice

Coriolopsis aspera is a medicinal mushroom that exerts several pharmacological and biological activities. However, the protective effect of the ethanol extract of C. aspera against arthritis has not been studied yet. In this study, we estimated some bioactive compounds in the ethanol extract of C. aspera fruiting bodies and used the extract with a dose of 400 mg.kg-1 body weight or the reference drug Mobic (0.2 mg.kg-1 body weight) to treat an adjuvant-induced arthritis model. Arthritis severity was determined via the changes in the diameters and volumes of hind paws, arthritic scores, total leukocyte count, ESR, biochemical parameters such as CRP and RF, as well as histological features of ankle joints. We found the presence of polyphenols and flavonoids with a high content (37.79 ± 0.88 mg GAE/g DW and 8.56 ± 0.37 mg QE/g DW, respectively) in the extract. Furthermore, the results showed that Freund’s complete adjuvant-treated mice exhibited a severe arthritic score and histological features, an increase in the diameters and volumes of hind paws, along with an elevation of leukocyte count, erythrocyte sediment rate, and biochemical parameters such as rheumatoid factor and C-reactive protein compared to normal mice. On the contrary, treatment with ethanol extract at a dose of 400 mg.kg-1 body weight for 28 days exhibited a strong anti-arthritis effect and could improve all the testing parameters at the same efficacy as those of the reference drug (Mobic 0.2 mg.kg-1 body weight). These findings suggest a potential application of the ethanol extract of C. aspera for arthritis treatment.

Evaluation of the anti-arthritic activity of Capparis spinosa L. roots extracts incomplete Freund’s adjuvant-induced arthritis in mice.

This study aimed to estimate the ant-arthritic activity of Capparis spinosa L. roots extract in complete Freund’s adjuvant (CFA)-induced arthritis mice by assessment of paw thickness, serological detection of inflammatory markers [rheumatoid factor (RF) and tumor necrosis factor-alpha (TNF-)] using ELISA, and histopathology. In total, 30 male mice were selected, prepared, and divided equally into five groups, including 2 control (NC and PC)and 3 experimental: EG1 (arthritis mice treated total effective dose of extract), EG2(arthritis mice treated total effective dose of Diclofenac Sodium), and EG3 (arthritis mice treated a half effective dose of each the extract and Diclofenac). Clinically, the extract administration lowered the paw thickness from day 7th onwards, the 21st day of study, while the extract administration lowered the paw thickness in EG1 from day 7th onwards compared to EG2 and EG3. For hematology, significant decreases were reported in RBCs, HCT, Hb, WBCs and neutrophils due to arthritis, and significant amelioration was seen obviously in mice of EG1 as a result of therapy. Significant increases in platelets, lymphocytes and monocytes were observed in PC and experimentally groups, significantly improving the values of treated groups due to therapy, in particular, EG1. All study groups ' values of MCV, MCH, MCHC, basophils and eosinophils have differed insignificantly. Concerning RF, the highest significant value was reported in PC while the lowest in NC, but without significant differences between EG1, EG2 and EG3 values. For TNF-, elevation had been recorded in PC mice, whereas reduction in NC, EG1 and EG3. Also, the value of EG2 was significantly higher than recorded in mice of EG1 and EG3. Concerning histology, the findings of EG1 revealed a significant amelioration compared to other groups. In conclusion, C. spinosa L. root extract demonstrated anti-inflammatory and anti-arthritic activity. Furthermore, studies to detect the therapeutic effects of the root extract on other systemic or local diseases are needed. Keywords: Paw thickness; Rheumatoid factor; Inflammatory markers; Diclofenac Sodium; Iraq

Activities of ethanol-extract of red ginger (Zingiber officinale var. Rubrum) on Completed Freund's Adjuvant-induced arthritis in mice

Background: Red ginger has been known to have anti-inflammatory and analgesic potency. Objective: This study aimed to analyse the ethanolic extract of red ginger (EERG) in mice's arthritis model induced by Completed Freund’s Adjuvant (CFA). Method: Red ginger was extracted using ethanol 96%. Arthritis mice (n = 25) were injected with CFA i.p., and sham mice (n = 5) were injected with normal saline. On day seven, mice were divided into six groups: sham, CFA, gabapentin 100 mg/kg BW, and EERG (dose 200; 400; 600 mg/kg BW). Treatments were administered orally once a day for seven days. Latency time and plantar thickness were measured on days zero, one, three, five, seven, eight, ten, twelve, and fourteen. On day 15, the mice were sacrificed, and the blood and spinal cords were collected. The haematology profiles were determined. Result: The EERG significantly prolonged the latency time towards thermal stimulus and decreased plantar thickness in arthritis mice, the same as gabapentin which served as the control. The EERG also reduced the number of leukocytes, lymphocytes, and neutrophils, and improved the morphology of the spinal cord's dorsal horn of arthritis mice. Conclusion: The EERG of 400 mg/kg BW significantly affects arthritis-induced hyperalgesia.

Tripterygium hypoglaucum extract ameliorates adjuvant-induced arthritis in mice through the gut microbiota.

Traditionally, Tripterygium hypoglaucum (Levl.) Hutch (THH) are widely used in Chinese folk to treat rheumatoid arthritis (RA). This study aimed to investigate whether the anti-RA effect of THH is related with the gut microbiota. The main components of prepared THH extract were identified by HPLC-MS. C57BL/6 mice with adjuvant-induced arthritis (AIA) were treated with THH extract by gavage for one month. THH extract significantly alleviated swollen ankle, joint cavity exudation, and articular cartilage destruction in AIA mice. The mRNA and protein levels of inflammatory mediators in muscles and plasma indicated that THH extract attenuated inflammatory responses in the joint by blocking TLR4/MyD88/MAPK signaling pathways. THH extract remarkably restored the dysbiosis of the gut microbiota in AIA mice, featuring the increases of Bifidobacterium, Akkermansia, and Lactobacillus and the decreases of Butyricimonas, Parabacteroides, and Anaeroplasma. Furthermore, the altered bacteria were closely correlated with physiological indices and drove metabolic changes of the intestinal microbiota. In addition, antibiotic-induced pseudo germ-free mice were employed to verify the role of the intestinal flora. Strikingly, THH treatment failed to ameliorate the arthritis symptoms and signaling pathways in pseudo germ-free mice, which validates the indispensable role of the intestinal flora. For the first time, we demonstrated that THH extract protects joint inflammation by manipulating the intestinal flora and regulating the TLR4/MyD88/MAPK signaling pathway. Therefore, THH extract may serve as a microbial modulator to recover RA in clincial practice.ver RA in clincial practice.

Sinomenine ameliorates adjuvant-induced arthritis by inhibiting the autophagy/NETosis/inflammation axis

Studies have found that neutrophil extracellular traps (NETs) which are the specific dying form of neutrophil upon activation have fundamental role in the rheumatoid arthritis onset and progression. The purpose of this study was to explore the therapeutic effect of Sinomenine on adjuvant-induced arthritis in mice, and the neutrophil activities regulated by Sinomenine. The rheumatoid arthritis model was established by local injection of adjuvant and the Sinomenine treatment was administered orally for 30 days, during which, arthritic scores were evaluated and the joint diameter was measured to determine disease progression. The joint tissues and serum were acquired for further tests after sacrifice. Cytometric beads assay was performed to measure the concentration of cytokines. For paraffin-embedded ankle tissues, hematoxylin and erosin staining and Safranin O-fast staining were adopted to monitor the tissue changes of joint. In order to analyze the inflammation, NETs and autophagy of neutrophils in vivo, immunohistochemistry assays were applied to detect the protein expression levels in the local joints. To describe the effect brought by Sinomenine on inflammation, autophagy and NETs in vitro, the western blotting and the immunofluorescence assays were performed. The joint symptoms of the adjuvant induced arthritis were alleviated by the Sinomenine treatment significantly in terms of the ankle diameter and scores. The improvement of local histopathology changes and decrease of inflammatory cytokines in the serum also confirmed the efficacy. The expression levels of interleukin-6, P65 and p-P65 in the ankle areas of mice were remarkably reduced by Sinomenine. Compared with the model group, the decreased expression levels of lymphocyte antigen 6 complex and myeloperoxidase in the Sinomenine treating group showed the inhibitory effect of Sinomenine on the neutrophil migration. The expression of protein arginine deiminase type 4 (PAD4), ctrullinated histone H3 (CitH3) and microtubule-associated protein 1 light chain 3B (LC3B) had the similar tendency. Upon activation of lipopolysaccharide (LPS) in vitro, Sinomenine suppressed the phosphorylation of P65, extracellular signal-regulated kinase (ERK) and P38 of neutrophil. Meanwhile, Sinomenine inhibited NETs formation induced by phorbol 12-myristate 13-acetate (PMA), which were demonstrated by the decreased expression of neutrophil elastase (NE), PAD4 and CitH3. Sinomenine also inhibited PMA-induced autophagy in vitro based on the changes of Beclin-1 and LC3B. Sinomenine has good efficacy in treating adjuvant induced arthritis via regulating neutrophil activities. Apart from inhibiting activation of nuclear factor kappa-B (NF-κB) and mitogen-activated protein kinase (MAPK) pathways, the mechanism includes suppression of NETs formation via autophagy inhibition.

Consumption of Saturated Fatty Acids-Rich Lard Benefits Recovery of Experimental Arthritis by Activating PPAR-γ.

This study investigates the impacts of lard and related fatty acids intake on rheumatoid arthritis (RA) animal models. Collagen-induced arthritis (CIA) and adjuvant-induced arthritis (AIA) are induced in SD rats and C57 BL/6 mice respectively, which are fed by lard-rich diet (LRD) for 42 days with intake restriction or not. AIA SD rats are treated by representative fatty acids for 30 days. Body weight, arthritis score, and metabolic profile are periodically recorded. Monocyte distribution, cytokine/metabolites levels, gene expression, and tissue damages are investigated by flow cytometry, ELISA, colorimetry, PCR, and histological methods. After being treated by fatty acids in vitro, THP-1 monocytes and the corresponding medium are collected for ELISA, PCR, immunoblotting, and reporter gene assays. Irrespective of intake amounts, LRD decreases inflammatory cytokines and inhibits glycolysis in all rheumatic rodents. Furthermore, it alters monocyte distribution and promotes PPAR-γ expression in AIA mice. Overall evidences show that both saturated (SF) and unsaturated fatty acids (USF) from lard can attenuate inflammation by activating PPAR-γ. Silencing PPAR-γ abrogates their anti-inflammatory effects in vitro. Besides, SF can stimulate TLR4/NF-κB pathway. Lard consumption is beneficial for active inflammatory arthritis recovery. Even SF can activate PPAR-γ and consequently attenuate inflammation.

Phytochemical screening and determination of treatment dose of Capparis spinosa L. roots extract in adjuvant induced arthritis in mice

This study was aimed to investigate phytochemical properties of Capparis spinosa roots extract and to evaluate anti-arthritic activity of this extract in adjuvant-induced arthritis in mice which initiated using the complete Freund's adjuvant. For these purposes, the roots of C. spinosa were collected from different rural areas in Wasit province, and the extract was prepared and analyzed phytochemically. Also, a total of 42 male mice were prepared, divided equally to 7 groups involved 2 controls [negative (NC) and positive (PC)] as well as 5 treated (TG1, TG2, TG3, TG4 and TG5) received respectively 100, 150, 200, 250 and 300 mg/kg B.W of the extract for 21 days. Phytochemical analysis of C. spinosa L. roots extract showed a significant elevation in alkaloids and lacking of steroid, with existence of tannins, saponins, polysaccharides carbohydrates and glycosides, proteins and amino acid, coumarins, flavonoids, and polyphenolic compounds. The clinical findings of the right paw thickness for mice of different study groups were revealed a significant variation in their values. Comparing to NC, values of PC were increased significantly; while, the findings of treated groups with different concentrations of C. spinosa L. roots extract were variable significantly.

Novel anti-arthritic mechanisms of trans-cinnamaldehyde against complete Freund’s adjuvant-induced arthritis in mice: involvement of NF-кB/TNF-α and IL-6/IL-23/ IL-17 pathways in the immuno-inflammatory responses

Trans-cinnamaldehyde (TCA), a natural cinnamaldehyde derivative of cinnamon oil, is known for anti-inflammatory, anti-bacterial, anti-fungal, anti-diabetic, and anti-cancer activities. However, no study has examined the protective mechanisms of TCA on complete Freund’s adjuvant (CFA)-induced arthritis. Chronic arthritis was induced in mice by triple dose injection of 0.1 ml CFA in the first two days, then a treatment with TCA (100 mg/kg, i.p.) and the anti-arthritic drug; methotrexate (MTX, 0.75 mg/kg, i.p., 3 times/week) started from day 10 after CFA and continued till day 35.TCA ameliorated the CFA-induced arthritis features, indicated by the decrease in serum rheumatoid factor, paw swelling, arthritis index and the arthritis changes in limb histology. Additionally, TCA treatment showed anti-inflammatory actions through downregulation of TNF-α, NF-κB and COX-2 expressions and marked reduction in IL-1β, IL-6, IL-23 and IL-17 levels in inflamed paw tissues.Consequently, TCA can decrease arthritis progression and inhibit the immune/inflammatory responses initiated by TNF-α/IL-1β/IL-6/IL-23/IL-17 signals, via NF-κB modulation, almost to the same extent accomplished by MTX. Therefore, TCA could be a promising anti-arthritic drug.

Ameliorative effect of tuna elastin peptides on AIA mice by regulating the composition of intestinal microorganisms and SCFAs

• Tuna elastin peptides relieved adjuvant-induced arthritis in mice. • Tuna elastin peptides modulated gut microbiota and SCFAs metabolite profile. • Tuna elastin peptides induced a dose-dependent pattern in the AIA model. • The relieving arthritis of tuna elastin peptides were transmissible by FMT. Elastin peptides display a wide range of biological activities. This study aimed to investigate the ameliorative effect of tuna elastin peptides on adjuvant-induced arthritis (AIA). The daily administration of tuna elastin peptides markedly downregulated the levels of proinflammatory cytokines and upregulated the immunosuppressive cytokines in the serum, bone, and colon of the AIA model. 16S rRNA sequencing showed that tuna elastin peptide-treated mice exhibited reversed gut microbiota dysbiosis and increased abundance of Lactobacillus and Clostridium . Additionally, metabolomics indicated that short-chain fatty acid (SCFAs) production was increased in tuna elastin peptide-treated mice. The ameliorating arthritis effects of tuna elastin peptides were transmitted by transplanting the fecal microbiota from mice treated with high levels of elastin peptides. These results indicate the potential of tuna elastin peptides as beneficial functional food that ameliorates rheumatoid arthritis.

Pinocembrin Reduces Arthritic Symptoms in Mouse Model via Targeting Sox4 Signaling Molecules.

Rheumatoid arthritis (RA) is a chronic autoimmune, multifactorial, inflammatory disorder characterized by hyperplasia and infiltration of inflammatory cells at the synovial lining leading to destruction of cartilage and bone tissues. Pinocembrin (PCB) is a natural flavonoid extracted as a pure molecule from honey, propolis, and some plants. In this study, we evaluated the antiarthritic effect of PCB in adjuvant induced arthritis (AIA) mice. Treating the AIA mouse model with PCB reduced the arthritis symptoms/score, including edema size, extent of hind paw redness, abnormal movement, and holding inability. At the pathological level, PCB significantly decreased the joint erosion and percentages of infiltrated inflammatory cells. Biochemically, PCB interacts with the transcription factor, SRY-related HMG-box 4 (Sox4), and then modulates its dysregulated expression and the expression of Sox4/Stat3 signaling molecules in AIA mice. These molecules include tumor necrosis factor-α, nuclear transcription factor kappaB, and cyclooxygenase-2, besides the microRNAs; miR-132, miR-202-5p, and miR-7235, which are dysregulated in adjuvant-induced arthritis model relative to the control mice. The possible PCB interaction with Sox4 transcriptional protein was confirmed through molecular docking where three hydrogen bonds were formed at ARG and LYS residues at a stable binding energy of -4.72. Taken together, our data demonstrate that PCB could serve as a therapeutic drug in treatment of RA.

Targeted and Combined TPCA-1-Gold Nanocage Therapy for In Vivo Treatment of Inflammatory Arthritis.

Rheumatoid arthritis (RA) is an autoimmune disease that is currently incurable. Inhibition of inflammation can prevent the deterioration of RA. 2-[(Aminocarbonyl)amino]-5-(4-fluorophenyl)-3-thiophenecarboxamide (TPCA-1) suppresses inflammation via the inhibition of nuclear factor-κ (NF-κB) signaling pathway. Gold-based therapies have been used to treat inflammatory arthritis since the 1940s. Hyaluronic acid (HA) is a targeting ligand for CD44 receptors overexpressed on activated macrophages. Therefore, a combined therapy based on TPCA-1, gold, and HA was explored for the treatment of RA in this study. We used gold nanocages (AuNCs) to load TPCA-1 and modified the TPCA-1 (T) loaded AuNCs with HA and peptides (P) to construct an anti-inflammatory nanoparticle (HA-AuNCs/T/P). An adjuvant-induced arthritis (AIA) mice model was used to investigate the in vivo anti-inflammatory efficacy of HA-AuNCs/T/P. In vivo distribution results showed that HA-AuNCs/T/P had increased and prolonged accumulation at the inflamed paws of AIA mice. Treatment by the HA-AuNCs/T/P suppressed joint swelling and alleviated cartilage and bone damage. By loading to HA-AuNCs/T/P, the effective concentration of TPCA-1 was greatly reduced from 20 to 0.016mg/kg mice. This study demonstrated that HA-AuNCs/T/P could effectively suppress inflammation and alleviate the symptoms of AIA mice, suggesting a great potential of HA-AuNCs/T/P for the treatment of RA.

Targeted and MMP-2/9 responsive peptides for the treatment of rheumatoid arthritis.

Bioactive peptides are attractive candidates for drug development. QAW is a tripeptide that is obtained from an anti-inflammatory protein-Annexin A1 (ANXA1). Previous studies showed that QAW alleviated inflammation in experimental colitis and inflammatory bowel disease via NF-κB inhibition. This study establishes adjuvant-induced arthritis (AIA) mouse models and explores the anti-inflammatory efficacy of QAW in AIA mice. To enhance the targeting, responsiveness, and efficacy of QAW to inflammation, QAW (Q) is modified with a cell penetrating peptide (T), a matrix metalloproteases-2/9 (MMP-2/9) digestive peptide (M), and an inflammation targeting peptide-RGD (R). The designed RMTQ demonstrates enhanced delivery to cytoplasm, higher reduction of pro-inflammatory factors, and better efficacy than QAW. The anti-inflammatory efficacy of RMTQ is similar to that of DEX in this study whereas RMTQ treatment shows a higher safety than that of DEX. In sum, this study demonstrates that RMTQ can be a potential therapeutic for inflammatory arthritis.

A Novel Phytochemical, DIM, Inhibits Proliferation, Migration, Invasion and TNF-α Induced Inflammatory Cytokine Production of Synovial Fibroblasts From Rheumatoid Arthritis Patients by Targeting MAPK and AKT/mTOR Signal Pathway.

In rheumatoid arthritis(RA) pathogenesis, activated RA fibroblast-like synoviocytes (RA-FLSs) exhibit similar proliferative features as tumor cells and subsequent erosion to cartilage will eventually lead to joint destruction. Therefore, it is imperative to search for compounds, which can effectively inhibit the abnormal activation of RA-FLSs, and retard RA progression.3′3-Diindolylmethane (DIM), the major product of the acid-catalyzed oligomerization of indole-3-carbinol from cruciferous vegetables, has been reported to be functionally relevant to inhibition of migration, invasion and carcinogenesis in some solid tumors. In this study, we explored the anti-proliferation, anti-metastasis and anti-inflammation effects of DIM on RA-FLSs as well as the underlying molecular mechanisms. To do this, primary RA-FLSs were isolated from RA patients and an animal model. Cell proliferation, migration and invasion were measured using CCK-8, scratch, and Transwell assays, respectively. The effects of DIM on Matrix metalloproteinases (MMPs) and some inflammatory factors mRNA and key molecules such as some inflammatory factors and those involved in aberrantly-activated signaling pathway in response to tumor necrosis factor α(TNF-α), a typical characteristic mediator in RA-FLS, were quantitatively measured by real-time PCR and western blotting. Moreover, the effect of DIM on adjuvant induced arthritis(AIA) models was evaluated with C57BL/6 mice in vivo. The results showed that DIM inhibited proliferation, migration and invasion of RA-FLS in vitro. Meanwhile, DIM dramatically suppressed TNF-α–induced increases in the mRNA levels of MMP-2, MMP-3, MMP-8, and MMP-9; as well as the proinflammatory factors IL-6, IL-8, and IL-1β. Mechanistic studies revealed that DIM is able to suppress phosphorylated activation not only of p38, JNK in MAPK pathway but of AKT, mTOR and downstream molecules in the AKT/mTOR pathway. Moreover, DIM treatment decreased expression levels of proinflammatory cytokines in the serum and alleviated arthritis severity in the knee joints of AIA mice. Taken together, our findings demonstrate that DIM could inhibit proliferation, migration and invasion of RA-FLSs and reduce proinflammatory factors induced by TNF-α in vitro by blocking MAPK and AKT/mTOR pathway and prevent inflammation and knee joint destruction in vivo, which suggests that DIM might have therapeutic potential for RA.

Mannan-Binding Lectin Attenuates Inflammatory Arthritis Through the Suppression of Osteoclastogenesis.

Mannan-binding lectin (MBL) is a vital element in the host innate immune system, which is primarily produced by the liver and secreted into the circulation. Low serum level of MBL is reported to be associated with an increased risk of arthritis. However, the underlying mechanism by which MBL contributes to the pathogenesis of arthritis is poorly understood. In this study, we investigated the precise role of MBL on the course of experimental murine adjuvant-induced arthritis (AIA). MBL-deficient (MBL−/−) AIA mice showed significantly increased inflammatory responses compared with wild-type C57BL/6 AIA mice, including exacerbated cartilage damage, enhanced histopathological features and high level of tartrate-resistant acid phosphatase (TRAP)-positive cells. MBL protein markedly inhibited the osteoclast formation from human blood monocytes induced by receptor activator of nuclear factor-κB ligand (RANKL) and macrophage colony-stimulating factor (M-CSF) in vitro. Mechanistic studies established that MBL inhibited osteoclast differentiation via down-regulation of p38 signaling pathway and subsequent nuclear translocation of c-fos as well as activation of nuclear factor of activated T-cells c1 (NFATc1) pathway. Importantly, we have provided the evidence that concentrations of MBL correlated negatively with the serum levels of amino-terminal propeptide of type I procollagen (PINP) and C-terminal telopeptide of type I collagen (β-CTX), serum markers of bone turnover, in patients with arthritis. Our study revealed an unexpected function of MBL in osteoclastogenesis, thus providing new insight into inflammatory arthritis and other bone-related diseases in patients with MBL deficiency.

Microneedle-Assisted Transdermal Delivery of Etanercept for Rheumatoid Arthritis Treatment.

Rheumatoid arthritis (RA) is a complicated autoimmune disease. The clinical applications of etanercept (EN), a TNF-α inhibitor, can efficiently halt the development of RA. EN is mainly administrated by subcutaneous injection, which may cause low compliance, side effects, and infection risk. In this study, a hyaluronic acid crosslinked microneedle system (MN) was constructed as the transdermal alternative to deliver EN. We describe the formulation, fabrication, characterization, and transdermal insertion study of MN. In vitro bioactivity of EN was conducted and analyzed by dynamic light scattering and circular dichroism spectrum. In vivo evaluation of MN was studied on adjuvant-induced arthritis mice. The MN possessed sufficient mechanical strength, good biocompatibility, little influence on the bioactivity of EN, and high anti-inflammatory efficacy. This work represents a successful example of delivering macromolecule therapeutic treatment by MN for RA treatment. The transdermal delivery of EN by MN offers a new treatment option for RA patients.

Hesperidin inhibits synovial cell inflammation and macrophage polarization through suppression of the PI3K/AKT pathway in complete Freund's adjuvant-induced arthritis in mice

Rheumatoid arthritis (RA) is an autoimmune disease characterized by synovitis. Synovitis can cause joint injury by releasing inflammatory factors and metalloproteinases (MMPs). Therefore, it is necessary to find drugs that can control synovitis in the process of RA. Herein, we investigate the anti-inflammatory effect of Hesperidin (HSN) on fibroblast-like synovial (FLS) cells induced by lipopolysaccharide (LPS) and the protective action of M1 polarization level of synovial macrophages on antigen-induced arthritis (AIA) in order to elucidate the reduction of inflammatory cytokines and MMPs and the inhibition of macrophage activation.The functional effect of HSN on LPS-induced mRNA and protein expressions of inflammatory cytokines and MMPs in FLS cells as well as on LPS-induced macrophage M1 and M2 polarization markers was determined by quantitative real-time PCR (qPCR) or Western blot analyses, respectively. AIA in 2-month-old mice was generated using intraperitoneal injection with HSN (20 mg/kg/day) or LY294002 (20 mg/kg/day). The results show HSN significantly inhibited the LPS-induced gene expression of the inflammatory mediators. Furthermore, treatment with HSN relieved the antigen-induced arthritis and reduced the protein levels of MMP3, MMP9, and MMP13 in FLS and inhibited the polarization of macrophages to M1. Based on the results of our analyses, we concluded that HSN has significant anti-inflammatory activities and reduces the potential of MMPs in rheumatoid arthritis and the degree of polarization of macrophages to M1. Through the study of signaling pathways, we established that the inhibition of the PI3K/AKT signaling pathway by HSN may show therapeutic effects in the progression of rheumatoid arthritis.

Interferon-γ facilitated adjuvant-induced arthritis at early stage.

Interferon-γ (IFN-γ) is a versatile cytokine which broadly involves in the inflammatory diseases, mediating both immune activation and tolerance. Here, we aimed to investigate the role of IFN-γ in the initiation of adjuvant-induced arthritis (AIA). In an AIA mice model, increasing IFN-γ mRNA was observed at day 3 and peaked on day 7. At day 3, the majority of IFN-γ-producing cells were located around vessels observed by immunofluorescent staining. Recombinant IFN-γ or anti-IFN-γ antibody was injected into the AIA paw on day 2 to study the outcome of AIA. The recipients of IFN-γ showed increased synovial inflammation, whereas anti-IFN-γ antibody injection repressed the expansion of inflammatory cells. As the percentages of blood monocytes were approximately equivalent, we hypothesized that IFN-γ might impact the access of innate leucocytes from blood to expand local inflammation at this stage. Analysis of tissue CD31 and vascular cell adhesion molecule-1 (VCAM-1) expressions suggested a positive effect of these factors in the development of inflammation, and IFN-γ affected the VCAM-1 expression. To further verify this idea, mice regionally injected with IFN-γ were systematically administrated with anti-VCAM-1 antibody during AIA induction. The IFN-γ expression was inhibited, and the development of AIA was partly abolished in these mice regardless of regional IFN-γ injection. These data suggested that IFN-γ might be critical for the expansion of AIA at early stage through helping inflammatory cell access.

Fatty oil from Securidaca inappendiculata exerted therapeutic effects on adjuvant-induced arthritis in mice via suppression on fibroblast-like synoviocyte

Securidaca inappendiculata Hassk. (SI) is a medicinal plant used to treat rheumatoid arthritis (RA) in South China. A substantial amount of fatty oil was isolated from SI (SIF), however little knowledge about its chemical composition and medicinal potentials was obtained. In this study, we analyzed its chemical composition with methyl esterification based GC–MS method, and investigated the therapeutic potentials on adjuvant-induced arthritis (AA) in mice. MTT and western-blot methods were employed to investigate its effects on proliferation rate and protein expressions in MH7A cells, respectively. It was revealed SIF was mainly comprised of saturated and monosaturated fatty acids, and the two predominant compounds were palmitic acid (36.89%) and oleic acid (31.12%). Treatment with SIF at 100 mg/kg resulted in significant alleviation of AA severity in mice, together with reduced synovial hyperplasia and inflammatory infiltration in joints, and decreased levels of sialic acid, malondialdehyde and alkaline phosphatase in serum. Results from immunohistochemical assays hinted the protective effects of SIF on joints were associated to the inhibition on production of some pathological factors in synovium, including IL-1β, TNF-α and MMP-9. SIF inhibited the proliferation of MH7A cells in a concentration dependent manner, and abrogated phosphorylation of p65 in vitro. These evidences collectively suggested SIF could suppress the pathological functions of fibroblast-like synoviocyte, and protect joints from destruction under AA conditions.