A rat model of rheumatoid arthritis: TDZD-8 is associated with the protection against the induction of the synovium knee joint IL-17A/GSK3beta/ROS/alpha-SMA axis of fibrosis / Un modelo de rata de artritis reumatoide: TDZD-8 está asociado con la protección contra la inducción del eje de fibrosis de la articulación sinovial de la rodilla IL-17A/GSK3ß/ROS/α-SMA

SUMMARY: Rheumatoid arthritis (RA) that affects the synovial knee joint causes swelling of the synovial membrane and tissue damage. Interleukin-17A (IL-17A) and the enzyme glycogen synthase kinase-3β (GSK3β) are involved in the pathogenesis of RA. The link between IL-17A, GSK3β, the oxidative stress, and the profibrogenic marker alpha-smooth muscle actin (α-SMA) with and without TDZD-8, GSK3β inhibitor has not been studied before. Consequently, active immunization of rats was performed to induce RA after three weeks using collagen type II (COII) injections. The treated group received daily injection of 1 mg/kg TDZD-8 for 21 days following the immunization protocol (COII+TDZD-8). Blood and synovium tissue samples were harvested at the end of the experiment. RA development was confirmed as corroborated by a substantial increase in blood levels of the highly specific autoantibody for RA, anti-citrullinated protein antibody as well as augmentation of reactive oxidative species (ROS) levels measured as lipid peroxidation. RA induction also increased synovium tissue levels of IL-17A and the profibrogenic marker, α-SMA. All these parameters seemed to be significantly (p<0.0001) ameliorated by TDZD-8. Additionally, a significant correlation between IL-17A, ROS, and α-SMA and biomarkers of RA was observed. Thus, knee joint synovium RA induction augmented IL-17A/GSK3β/ROS/α-SMA axis mediated arthritis in a rat model of RA, which was inhibited by TDZD-8.

La artritis reumatoide (AR) que afecta la articulación sinovial de la rodilla provoca inflamación de la membrana sinovial y daño tisular. La interleucina-17A (IL-17A) y la enzima glucógeno sintasa quinasa-3β (GSK3β) están involucradas en la patogenia de la AR. No se ha estudiadol vínculo entre IL-17A, GSK3β, el estrés oxidativo y el marcador profibrogénico actina de músculo liso alfa (α-SMA) con y sin inhibidor de TDZD-8, GSK3β. En consecuencia, se realizó una inmunización activa de ratas para inducir la AR después de tres semanas usando inyecciones de colágeno tipo II (COII). El grupo tratado recibió una inyección diaria de 1 µg/ kg de TDZD-8 durante 21 días siguiendo el protocolo de inmunización (COII+TDZD-8). Se recogieron muestras de sangre y tejido sinovial al final del experimento. El desarrollo de AR se confirmó como lo corroboró el aumento sustancial en los niveles sanguíneos del autoanticuerpo altamente específico para AR, el anticuerpo antiproteína citrulinada, así como el aumento de los niveles de especies oxidativas reactivas (ROS) medidos como peroxidación lipídica. La inducción de AR también aumentó los niveles de tejido sinovial de IL-17A y el marcador profibrogénico, α-SMA. Todos estos parámetros parecían mejorar significativamente (p<0,0001) con TDZD-8. Además, se observó una correlación significativa entre IL- 17A, ROS y α-SMA y biomarcadores de AR. Por lo tanto, la inducción de AR en la sinovial de la articulación de la rodilla aumentó la artritis mediada por el eje IL-17A/GSK3β/ROS/α-SMA en un modelo de rata de AR, que fue inhibida por TDZD-8.

The effect of TNF-α inhibitor treatment on microRNAs and endothelial function in collagen induced arthritis.

Chronic inflammation causes dysregulated expression of microRNAs. Aberrant microRNA expression is associated with endothelial dysfunction. In this study we determined whether TNF-α inhibition impacted the expression of miRNA-146a-5p and miRNA-155-5p, and whether changes in the expression of these miRNAs were related to inflammation-induced changes in endothelial function in collagen-induced arthritis (CIA). Sixty-four Sprague-Dawley rats were divided into control (n = 24), CIA (n = 24) and CIA+etanercept (n = 16) groups. CIA and CIA+etanercept groups were immunized with bovine type-II collagen, emulsified in incomplete Freund's adjuvant. Upon signs of arthritis, the CIA+etanercept group received 10mg/kg of etanercept intraperitoneally, every three days. After six weeks of treatment, mesenteric artery vascular reactivity was assessed using wire-myography. Serum concentrations of TNF-α, C-reactive protein, interleukin-6, vascular adhesion molecule-1 (VCAM-1) and pentraxin-3 (PTX-3) were measured by ELISA. Relative expression of circulating miRNA-146a-5p and miRNA-155-5p were determined using RT-qPCR. Compared to controls, circulating miRNA-155-5p, VCAM-1 and PTX-3 concentrations were increased, and vessel relaxation was impaired in the CIA (all p<0.05), but not in the CIA+etanercept (all p<0.05) groups. The CIA group had greater miRNA-146a-5p expression compared to the CIA+etanercept group (p = 0.005). Independent of blood pressure, miRNA-146a-5p expression was associated with increased PTX-3 concentrations (p = 0.03), while miRNA-155-5p expression was associated with impaired vessel relaxation (p = 0.01). In conclusion, blocking circulating TNF-α impacted systemic inflammation-induced increased expression of miRNA-146a-5p and miRNA-155-5p, which were associated with endothelial inflammation and impaired endothelial dependent vasorelaxation, respectively.

Suppression of nitrosative stress and inflammation of the knee joint synovium in collagen type ii-induced rheumatoid arthritis by the inhibition of glycogen synthase kinase-3ß / Supresión del estrés nitrosativo e inflamación de la sinovial de la articulación de la rodilla en la artritis reumatoide inducida por colágeno tipo ii mediante la inhibición de la glucógeno sintasa quinasa-3 ß

SUMMARY: Rheumatoid arthritis (RA), an inflammatory autoimmune disease that causes cartilage degradation and tissue destruction, can affect synovial joints such as the knee joint. The link between the nitrosative stress enzyme inducible nitric oxide synthase (iNOS) and the cytokine interleukin-1 (IL-1β) in RA-induced knee joint synovial membrane damage with and without the incorporation of the GSK3β inhibitor TDZD-8 has never been studied. As a result, we used active immunization method with collagen type II (COII) for twenty one days to induce RA in rats. TDZD-8 (1 mg/kg; i.p.) was given daily into matched immunized rats for three weeks after day 21 (COII+TDZD-8). Blood and tissue samples were taken 42 days after immunization. A dramatic increase in rheumatoid factor (RF) blood levels, as well as considerable synovial tissue damage and inflammatory cell infiltration of the synovial membrane, were used to validate the onset of RA following COII immunization. COII immunization increased tissue levels of iNOS protein and IL- 1β mRNA and protein expression, which TDZD-8 suppressed considerably (p<0.0001). Furthermore, there was a significantly (p<0.001) positive correlation between iNOS, inflammatory biomarkers, and RF. We concluded that TDZD-8 reduced RA-induced IL-1β -iNOS axis-mediated arthritis in the rat knee joint synovium.

RESUMEN: La artritis reumatoide (AR), es una enfermedad autoinmune inflamatoria que causa la degradación del cartílago y la destrucción del tejido, pudiendo afectar las articulaciones sinoviales, como la articulación de la rodilla. No se ha estudiado el vínculo entre la óxido nítrico sintasa inducible por la enzima del estrés nitrosativo (iNOS) y la citocina interleucina-1 (IL-1β) en el daño de la membrana sinovial de la articulación de la rodilla provocado por AR con y sin la incorporación del inhibidor de GSK3β TDZD-8. Utilizamos el método de inmunización activa con colágeno tipo II (COII) durante veintiún días para inducir AR en ratas. Se administró TDZD-8 (1 mg/kg; i.p.) diariamente a ratas inmunizadas emparejadas durante tres semanas después del día 21 (COII+TDZD- 8). Se tomaron muestras de sangre y tejido 42 días después de la inmunización. Se observó un gran aumento de los niveles sanguíneos del factor reumatoideo (FR), así como un daño considerable del tejido sinovial e infiltración de células inflamatorias en la membrana sinovial, para validar la aparición de la AR después de la inmunización con COII. La inmunización con COII aumentó los niveles tisulares de la proteína iNOS y la expresión de proteína y ARNm de IL-1β, que TDZD-8 suprimió considerablemente (p<0,0001). Además, hubo una correlación positiva significativa (p<0,001) entre iNOS, biomarcadores inflamatorios y FR. Concluimos que TDZD- 8 redujo la artritis mediada por el eje IL-1β-iNOS inducida por la AR en la sinovial de la articulación de la rodilla de rata.

Different protective efficacies of a novel antigen-specific DNA vaccine encoding chicken type â ¡ collagen via intramuscular, subcutaneous, and intravenous vaccination against experimental rheumatoid arthritis.

Tolerizing DNA vaccines encoding key autoantigens are one of emerging strategies for the treatment of rheumatoid arthritis (RA). Among these vaccines, the most representative is pcDNA-CCOL2A1, an antigen-specific DNA vaccine encoding chicken type Ⅱ collagen (CCⅡ) with significant therapeutic and prophylactic efficacy in collagen-induced arthritis (CIA) rat models. We compared the in situ expression levels of CCOL2A1-mRNA and CCⅡ protein and the protective efficacies against CIA after a single dose (300 µg/kg) of this vaccine via intramuscular (IM), subcutaneous (SC) and intravenous (IV) vaccinations. The IM vaccination routes resulted in good protective efficacies in terms of decreasing CIA incidence and severity and significantly improved radiographic and histopathologic findings and scores of joints. Furthermore, IM, SC, and IV vaccinations markedly decreased serum levels of anti-type Ⅱ collagen (CⅡ) IgG antibodies, but only IM vaccination significantly reduced serum levels of rheumatoid factor (RF) and anti-cyclic citrullinated peptide (anti-CCP) antibody. The vaccine exhibited a continuous CCOL2A1-mRNA expression in the tail and abdominal subcutaneous tissue injection sites, but no CCOL2A1-mRNA signal was observed in muscle. Strikingly, CCⅡ protein expression levels at the three injection sites were comparable with minimal variation. IM administration may be considered the preferred route for RA treatment in clinical practice.

A Double-Blind, Randomized, Placebo-Controlled Trial to Evaluate the Efficacy of a Hydrolyzed Chicken Collagen Type II Supplement in Alleviating Joint Discomfort.

Joint pain and disease affects more than one in four adults in the United States. We conducted a double-blind, randomized, placebo-controlled trial to investigate the efficacy of a hydrolyzed chicken collagen type II (HCII) supplement in reducing joint-related discomfort such as pain and stiffness, and in improving mobility. We enrolled adults aged 40-65 (65.5% were women) who had joint discomfort, but had no co-morbidities, and who were not taking pain medications. The participants were randomized to receive either the HCII supplement (n = 47) or a placebo (n = 43) for eight weeks. At the baseline, and at week 4 and week 8, we administered the Western Ontario and McMaster Universities Arthritis Index (WOMAC) survey with three additional wrist-related questions and the Visual Analog Scale for assessments of joint-related symptoms. In the WOMAC stiffness and physical activity domains and in the overall WOMAC score, the HCII group had a significant reduction in joint-related discomforts compared with the placebo group. For example, at week 4, the HCII group had a 36.9% reduction in the overall WOMAC score, compared with a 14.3% reduction in the placebo group (p = 0.027). This HCII product is effective in reducing joint pain and stiffness and in improving joint function among otherwise healthy adults.

Niacinamide and undenatured type II collagen modulates the inflammatory response in rats with monoiodoacetate-induced osteoarthritis.

The current work aimed to examine the properties of oral supplementation of niacinamide and undenatured type II collagen (UCII) on the inflammation and joint pain behavior of rats with osteoarthritis (OA). Forty-nine Wistar rats were allocated into seven groups; control (no MIA), MIA as a non-supplemental group with monosodium iodoacetate (MIA)-induced knee osteoarthritis, MIA + undenatured type II collagen (UCII) at 4 mg/kg BW, MIA + Niacinamide at 40 mg/kg BW (NA40), MIA + Niacinamide at 200 mg/kg BW (NA200), MIA + UCII + NA40 and MIA + UCII + NA200. Serum IL-1ß, IL-6, TNF-α, COMP, and CRP increased in rats with OA and decreased in UCII and NA groups (p < 0.05). Rats with osteoarthritis had greater serum MDA and knee joint MMP-3, NF-κB, and TGß protein levels and decreased in treated groups with UCII and NA (p < 0.05). The rats with OA also bore elevated joint diameters with joint pain behavior measured as decreased the stride lengths, the paw areas, and the paw widths, and increased the Kellgren-Lawrence and the Mankin scores (p < 0.05) and decreased in UCII treated groups. These results suggest the combinations with the UCII + NA supplementation as being most effective and reduce the inflammation responses for most OA symptoms in rats.

Undenatured type II collagen prevents and treats osteoarthritis and motor function degradation in T2DM patients and db/db mice.

Osteoarthritis (OA) has been scarcely researched among patients with diabetes mellitus. This study aims to confirm the preventive and therapeutic effects of undenatured type II collagen (UC II) on OA in aging db/db mice and in patients with T2DM. Firstly, aging db/db mice were randomly assigned to three groups: the UC II intervention (UC II) group, old model (OM) group and positive control group. Meanwhile db/m mice and young db/db mice were used as the normal control and young control groups, respectively. Secondly, fifty-five T2DM patients diagnosed with knee OA were randomly assigned to two groups: UC-II and placebo control groups. After a three-month intervention in both mice and T2DM patients, the subjects' gait and physical activities were assessed and the serum biomarkers including inflammatory cytokines, oxidative stress factors and matrix metalloproteinases (MMPs) were measured. Compared with the OM group mice, those in the UC II group showed a significantly greater superiority in terms of motor functions including the movement trajectories area (163.25 ± 20.3 vs. 78.52 ± 20.14 cm2), the tremor index (0.42 vs. 1.23), standing time (left hind: 0.089 ± 0.03 vs. 0.136 ± 0.04 s), swing (right front: 0.12 ± 0.02 vs. 0.216 ± 0.02 s), stride length (right hind: 7.2 ± 0.9 vs. 5.7 ± 1.1 cm), step cycle (right hind: 0.252 ± 0.05 vs. 0.478 ± 0.11 s) and cadence (14.12 ± 2.7 vs. 7.35 ± 4.4 steps per s). In addition, the levels of IL-4, IL-10, CTX- II and TGF-ß in the UC II group were 1.74, 2.23, 1.67 and 1.84 times higher than those in the OM group, respectively, while the levels of MMP-3 and MMP-13 in the UC II group were half those in the OM group. Correspondingly, UC II intervention significantly decreased the scores of pain, stiffness and physical function (p < 0.05), whereas the 6 MWT and total MET distances in the UC II group increased remarkably (p < 0.05). After a three-month period of intervention, the varus angle significantly decreased from 4.6 ± 2.0° to 3.0 ± 1.4° and the knee flexion range obviously increased from 57.9 ± 14.0° to 66.9 ± 10.4°. Importantly, the declining trend in the levels of hs-CRP and MDA and the incremental trend in the SOD level were consistent in the db/db mice and OA patients following UC II administration.

Infrared radiation from cage bedding moderates rat inflammatory and autoimmune responses in collagen-induced arthritis.

The development of collagen type II (CII)-induced arthritis (CIA), a model of rheumatoid arthritis, in rats housed in cages with bedding composed of Celliant fibres containing ceramic particles, which absorb body heat and re-emit the energy back to the body in the form of infrared radiation (+IRF rats), and those housed in cages with standard wooden shaving bedding (-IRF control rats) was examined. The appearance of the first signs of CIA was postponed, while the disease was milder (judging by the arthritic score, paw volume, and burrowing behaviour) in +IRF compared with -IRF rats. This correlated with a lower magnitude of serum anti-CII IgG antibody levels in +IRF rats, and lower production level of IL-17, the Th17 signature cytokine, in cultures of their paws. This could be partly ascribed to impaired migration of antigen-loaded CD11b + dendritic cells and their positioning within lymph nodes in +IRF rats reflecting diminished lymph node expression of CCL19 /CCL21. Additionally, as confirmed in rats with carrageenan-induced paw inflammation (CIPI), the infrared radiation from Celliant fibres, independently from immunomodulatory effects, exerted anti-inflammatory effects (judging by a shift in pro-inflammatory mediator to anti-inflammatory/immunoregulatory mediator ratio towards the latter in paw cultures) and ameliorated burrowing behaviour in CIA rats.

Effect of an oral preparation containing hyaluronic acid, chondroitin sulfate, hydrolyzed collagen type II and hydrolyzed keratin on synovial fluid features and clinical indices in knee osteoarthritis. A pilot study.

The aim of this study was to evaluate the effect of an oral preparation containing a naturally occurring matrix of hydrolyzed collagen type II, chondroitin sulfate (CS), and hyaluronic acid (HA), and bioactive oligopeptides of natural hydrolyzed keratin (K) in patients affected by knee OA through the evaluation of synovial fluid (SF) and clinical changes before and after treatment. Thirty patients with knee OA and swollen joint were included in the study and submitted to arthrocentesis. Patients were randomized in two groups: 1) the treatment group (N.15) took a dietary supplement containing 120 mg HA, 240 mg CS and 300 mg K once a day for 4 weeks; 2) the control group (N.15) was only submitted to arthrocentesis. Patient symptoms were evaluated at the beginning and at the end of the study by the WOMAC self-assessment questionnaire, the Lequesne algofunctional index, and the VAS forms. SF changes were evaluated by measuring local inflammatory indices, cytokines IL-1ß, IL-8, IL-6, IL-10 and GM-CSF. The group of patients treated with the oral supplement showed an improvement in the clinical indices WOMAC (p<0.01), Lequesne (p=0.014) and VAS pain (p<0.01). On the contrary, no significant changes were found in the control group. The SF collected from the treated group showed a reduction of IL-8 (p=0.015), IL-6 and IL-10 levels, while no changes in cytokines were observed in the control group. This pilot study suggests that an oral administration of a preparation containing a combination of HA, CS and K can improve some clinical parameters and affect cytokine concentrations in SF in patients with knee OA.

Comparative study between human mesenchymal stem cells and etanercept as immunomodulatory agents in rat model of rheumatoid arthritis.

To compare human adipose tissue mesenchymal stem cells (AT-MSCs) and etanercept as immunomodulatory agents for collagen-induced arthritis (CIA). CIA was induced by rats' immunization with collagen type II (CII) in complete Freund's adjuvant in days 0 and 7. Before the onset of CIA, prevention group received five doses of AT-MSCS intraperitoneally. After establishment of arthritis, rats received either five doses of AT-MSCs or phosphate-buffered saline (PBS) intraperitoneally or six doses of etanercept subcutaneously. Clinical and histopathological evaluation were performed in all groups; serum levels of tumor necrosis factor-α (TNF-α), interleukin-10 (IL-10), and anti-collagen II were assessed by enzyme-linked immunosorbent assay (ELISA). A total percent of autoreactive T and regulatory T (Treg) cells were quantified using spleen immune histochemical analysis. AT-MSCs were able to delay the onset of CIA, suppress the ongoing clinical and histopathological signs, decrease serum levels of TNF-α and anti-collagen type II, and downregulate the autoreactive T cells as etanercept. AT-MSCs were more potent in Treg cells upregulation, producing high serum levels of IL10. AT-MSCs might have a therapeutic effect in CIA via their potency in immune cell education, representing an effective new promising approach in rheumatoid arthritis in human.

Bioactive fractions from Securidaca inappendiculata alleviated collagen-induced arthritis in rats by regulating metabolism-related signaling.

Securidaca inappendiculata is a xanthone rich medicinal plant that has been used in the treatment of inflammation and autoimmune diseases like rheumatoid arthritis (RA) for centuries; however, the material base and mechanism of action responsible for its anti-arthritis effect still remains elusive. The objective of this study is to evaluate the therapeutic effects of xanthone-enriched extract of the plant against collagen-induced arthritis (CIA) in rats and explore the underlying mechanisms. The xanthone-deprived fraction (XDF) and xanthone-rich fraction (XRF) were obtained by using a resin adsorption coupled with acid-base treatment method, and their chemical composition difference was characterized by UPLC-MS/MS analysis. Effects of the two on CIA were analyzed using radiographic, histological, and immunohistochemical analyses. The results indicated that XRF alleviated joint structures destructions with the higher efficacy than XDF, and decreased levels of TNF-α, IL-6, and anti-cyclic citrullinated peptide antibody in CIA rats significantly. Furthermore, XRF inhibited nicotinamide phosphoribosyl transferase (NAMPT) mediated fat biosynthesis and utilization indicated by clinical evidences and metabonomics analysis, which thereby disrupted energy-metabolism feedback. In addition, Toll-like Receptor 4 and High Mobility Group Protein 1 expressions were downregulated in XRF-treated CIA rats. Collective evidences suggest NAMPT could be an ideal target for RA treatments and reveal a novel antirheumatic mechanism of S. inappendiculata by regulating NAMPT controlled fat metabolism.

Intra-articular injection of indomethacin-methotrexate in situ hydrogel for the synergistic treatment of rheumatoid arthritis.

Rheumatoid arthritis is a chronic systemic autoimmune disease that causes joint swelling and cartilage damage. The objective of the present work was to develop a temperature-sensitive hydrogel (D-NGel) containing nanoparticles (D-NPs), which could simultaneously deliver combination indomethacin and methotrexate. D-NPs were formed by multiple non-covalent interactions between PEI-SS and the carboxyl-containing hydrophobic small molecule drugs IND and MTX, which were then loaded into a temperature-sensitive hydrogel matrix. The Tsol/gel of the temperature-sensitive hydrogel matrix composed of 27% F127 and 10% F68 was 33 °C and the gelation time was less than 15 s. The resultant D-NGel was injected into the articular cavity of collagen-induced arthritis rats and quickly transformed in situ into gels which slowly released drug in the joint fluid for up to 72 h. The D-NGel effectively reduced joint swelling, bone erosion and expression of inflammatory cytokines in the ankle fluid and knee joint fluid. In addition, liver and kidney function tests and histopathological examination indicated there was a good biological safety for D-NGel. In conclusion, this work has demonstrated the great potential of the D-NGel for sustained co-delivery of IND and MTX for the synergistic treatment of rheumatoid arthritis, treating both the symptoms and the root causes of rheumatoid arthritis.

Lipid accumulation and mitochondrial abnormalities are associated with fiber atrophy in the skeletal muscle of rats with collagen-induced arthritis.

Rheumatoid arthritis (RA) has a negative impact on muscle mass, and reduces patient's mobility and autonomy. Furthermore, RA is associated with metabolic comorbidities, notably in lipid homeostasis by unknown mechanisms. To understand the links between the loss in muscle mass and the metabolic abnormalities, arthritis was induced in male Sprague Dawley rats (n = 11) using the collagen-induced arthritis model. Rats immunized with bovine type II collagen were compared to a control group of animals (n = 11) injected with acetic acid and complete Freund's adjuvant. The clinical severity of the ensuing arthritis was evaluated weekly by a semi-quantitative score. Skeletal muscles from the hind limb were used for the histological analysis and exploration of mitochondrial activity, lipid accumulation, metabolism and regenerative capacities. A significant atrophy in tibialis anterior muscle fibers was observed in the arthritic rats despite a non-significant decrease in the weight of the muscles. Despite moderate inflammation, accumulation of triglycerides (P < 0.05), reduced mitochondrial DNA copy number (P < 0.05) and non-significant dysfunction in mitochondrial cytochrome c oxidase activity were found in the gastrocnemius muscle. Concomitantly, our results suggested an activation of the muscle specific E3 ubiquitin ligases MuRF-1 and MAFbx. Finally, the adipose tissue from the arthritic rats exhibited decreased PPARγ mRNA suggesting reduced adipogenic capacities. In conclusion, the reduced adipose tissue adipogenic capacity and skeletal muscle mitochondrial capacity are probably involved in the activation of protein catabolism, inhibition of myogenesis, accumulation of lipids and fiber atrophy in the skeletal muscle during RA.

Effectiveness of Huai Qi Huang Granules on Juvenile Collagen-induced Arthritis and Its Influence on Pyroptosis Pathway in Synovial Tissue.

Huai Qi Huang (HQH) exerts great effects in clinic, such as anti-inflammation, immune-regulation, anti-cancer, and so on. However, the mechanism by which HQH protects juvenile idiopathic arthritis (JIA) is obscure. Thus, we explored deeply the protective mechanisms in juvenile collagen-induced arthritis (CIA) rat model. Pyroptosis is Gasdermin D (GSDMD)-dependent programmed cell death, involved in many diseases, such as sepsis. We investigated whether GSDMD-induced pyroptosis take part in mechanisms of juvenile CIA arthritis. Juvenile Wistar rats (3-4 weeks) were injected intradermally with fully emulsified bovine type II collagen and complete Freund's adjuvant to establish CIA rat models. Later, the CIA rats received oral administration of HQH (4.16 g/kg) once a day from the day 21 of modeling, with the treatment lasting for 28 days. Varieties of indicators were measured for evaluation of anti-inflammation effect of HQH, including hind paw swelling, arthritis scores, micro CT, and histopathological changes and the level of pro-inflammatory cytokines in the serum, including tumor necrosis factor alpha (TNF-±) and interleukin-18 (IL-18). The expression of GSDMD and caspase-1 in the joint synovial tissues was detected. The results demonstrated that the expression of the pyroptotic protein GSDMD and its upstream caspase-1 was significantly increased in the synovial tissues of CIA rats. The treatment of HQH ameliorated the symptoms in CIA rats, reduced levels of pro-inflammatory cytokines and hind paw swelling, down-regulated the expression of GDSMD and caspase-1. GSDMD-induced pyroptosis participated in the pathogenesis of CIA rats. The study supported that HQH can effectively improve joints inflammation of juvenile collagen-induced arthritis rats by inhibiting pyroptosis pathway in the joint synovial tissues.

Effect of H2 treatment in a mouse model of rheumatoid arthritis-associated interstitial lung disease.

Rheumatoid arthritis (RA)-associated interstitial lung disease (ILD), a primary cause of mortality in patients with RA, has limited treatment options. A previously established RA model in D1CC transgenic mice aberrantly expressed major histocompatibility complex class II genes in joints, developing collagen II-induced polyarthritis and anti-cyclic citrullinated peptide antibodies and interstitial pneumonitis, similar to those in humans. Molecular hydrogen (H2 ) is an efficient antioxidant that permeates cell membranes and alleviates the reactive oxygen species-induced injury implicated in RA pathogenesis. We used D1CC mice to analyse chronic lung fibrosis development and evaluate H2 treatment effects. We injected D1CC mice with type II collagen and supplied them with H2 -rich or control water until analysis. Increased serum surfactant protein D values and lung densities images were observed 10 months after injection. Inflammation was patchy within the perilymphatic stromal area, with increased 8-hydroxy-2'-deoxyguanosine-positive cell numbers and tumour necrosis factor-α, BAX, transforming growth factor-ß, interleukin-6 and soluble collagen levels in the lungs. Inflammatory and fibrotic changes developed diffusely within the perilymphatic stromal area, as observed in humans. H2 treatment decreased these effects in the lungs. Thus, this model is valuable for studying the effects of H2 treatment and chronic interstitial pneumonia pathophysiology in humans. H2 appears to protect against RA-ILD by alleviating oxidative stress.

Novel Hydrolyzed Chicken Sternal Cartilage Extract Improves Facial Epidermis and Connective Tissue in Healthy Adult Females: A Randomized, Double-Blind, Placebo-Controlled Trial.

CONTEXT: Dietary supplement manufacturers claim cutaneous anti-aging properties for their products; however, research supporting these claims remains sparse. OBJECTIVES: The study intended to determine if a correlation existed between the effects of a collagen dietary supplement and changes associated with skin aging. DESIGN: The study was a 12-week, double-blind, placebo-controlled trial. SETTING: The study took place at a clinical facility specializing in dermatological testing that could perform biophysical, instrumental analysis on the effects of proprietary supplement on human skin. PARTICIPANTS: Participants were 128 females, aged 39-59 (50.57 ± 5.55). INTERVENTION: Participants were randomly assigned to an intervention or a placebo. The intervention consisted of twice daily oral administration of a supplement containing 500 mg BioCell Collagen, a chicken sternal cartilage derived dietary ingredient composed of a naturally-occurring matrix of hydrolyzed collagen type-II (≥300 mg), chondroitin sulfate (≥100 mg), hyaluronic acid (≥50 mg). OUTCOME MEASURES: The primary parameters included transepidermal water loss, viscoelasticity, hydration, (indirect) collagen content, chromophore (melanin) content and hemoglobin level, and photographic analysis. An expert visually graded participants' skin to determine the intervention's efficacy, measuring facial lines and wrinkles, crow's feet lines and wrinkles, skin texture and smoothness, and skin tone. The presence of erythema and/or dryness determined tolerance. Secondary outcome measures were tolerance and incidence of adverse events, and the participant's perception of the supplement's value. RESULTS: For the 113 participants completing the study, the dietary supplementation compared to a placebo: (1) significantly reduced facial lines and wrinkles (P = .019) and crow's feet lines and wrinkles (P = .05), (2) increased skin elasticity (P = .008) and cutaneous collagen content (P < .001) by 12%, (3) improved indicators associated with a more youthful skin appearance based on visual grading and wrinkle width (P = .046), and (4) decreased skin dryness and erythema. No difference existed between the supplement and the placebo for skin-surface water content or retention. The supplement was well tolerated, with no reported adverse reactions. CONCLUSIONS: Dietary supplementation with chicken, sternal cartilage extract supports the accumulation of types-I/III collagen in skin to promote increased elasticity and reduced skin wrinkling.

Resveratrol Alleviates Rheumatoid Arthritis via Reducing ROS and Inflammation, Inhibiting MAPK Signaling Pathways, and Suppressing Angiogenesis.

Rheumatoid arthritis (RA) is a systemic autoimmune disease primarily affecting joints and is featured by chronic synovial inflammation and angiogenesis. We employed a bovine type-II collagen (BIIC)-induced Sprague-Dawley rat arthritis model and an in vitro RA model based on interleukin (IL)-1ß-stimulated rat synovial cells (RSC-364) to explore the preventive effect of resveratrol on RA and the underlying mechanisms. We found that resveratrol ameliorated BIIC-elicited synovitis and RA-related pathological hallmarks such as inflammatory cell infiltration and angiogenesis in the synovial tissue. Also, BIIC-stimulated rats displayed increased serum levels of proinflammatory cytokines and reactive oxygen species (ROS), as manifested by elevated serum malonaldehyde contents combined with reduced superoxide dismutase activity. It is noteworthy that resveratrol abolished BIIC-induced ROS and inflammation, confirming the antioxidative and anti-inflammatory actions of resveratrol in the context of RA. Furthermore, immunoblotting indicated that resveratrol downregulated the increase in the levels of hypoxia-inducible factor-1α (HIF-1α) and that of the activated phosphorylation of p38 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase in IL-1ß-stimulated RSC-364 cells. Moreover, we observed that resveratrol-treated RSC-364 cells displayed both G0/G1 cell-cycle arrest and enhanced levels of apoptosis. Altogether, the present evidence established the preventive role of resveratrol in RA progression. Mechanistically, resveratrol inhibits MAPK signaling pathways, likely by reducing ROS accumulation, to suppress the inflammatory response and cell proliferation and to provoke cell apoptosis in the synovial tissue, along with mitigation of HIF-1α-mediated angiogenesis. Thus resveratrol appears to hold great potential for clinical translation as a novel RA therapeutic.

miR-145-5p Increases Osteoclast Numbers In Vitro and Aggravates Bone Erosion in Collagen-Induced Arthritis by Targeting Osteoprotegerin.

BACKGROUND Osteoprotegerin (OPG) inhibits bone resorption and binds with strong affinity to receptor activator of NF κB ligand (RANKL), thereby preventing RANKL from binding to its receptor RANK. Osteoclasts have documented effects on bone erosion of rheumatoid arthritis (RA). The aim of this study was to examine the role of miR-145-5p in the regulation of RA osteoclast differentiation and bone erosion. MATERIAL AND METHODS Expression of microRNA-145-5p in human peripheral blood mononuclear cells (PBMC) and synovial tissue was assayed by real-time polymerase chain reaction (RT-PCR). OPG, RANK, and RANKL expression in RAW-264.7 cells was examined by RT-PCR and Western blot analysis. Osteoclast formation was detected by tartrate-resistant acid phosphatase (TRAP) staining. The effect of miR-145-5p on predicted target mRNAs was examined by luciferase reporter assays. Collagen-induced arthritis (CIA) was induced by injecting DBA/1 mice with bovine type II collagen (CII), and miR-145-5p agomir was administered by intravenous injection. Morphological changes in the CIA joint were assessed by micro-computed tomography (CT) and histopathology. RESULTS miR-145-5p levels significantly increased in RA PBMC and synovial tissue compared with normal PBMC and osteoarthritis (OA) tissue. After transfection of RAW-264.7 cells with miR-145-5p, RANK and RANKL expression increased significantly, while OPG expression decreased significantly. TRAP staining results showed osteoclast numbers increased. Micro-CT analysis of the arthritic joints showed that the miR-145-5p agomir caused bone erosion in mice, and histopathological analysis revealed that miR-145-5p agomir aggravates cartilage erosion. CONCLUSIONS Our findings indicate that administration of miR-145-5p aggravates joint erosion in CIA mice. This suggests that miR-145-5p is a potential target for the treatment of RA.

Allylpyrocatechol attenuates methotrexate-induced hepatotoxicity in a collagen-induced model of arthritis.

The cornerstone of treatment for rheumatoid arthritis is low dose methotrexate (MTX), but its use is limited by concerns regarding its potential for hepatotoxicity. Allylpyrocatechol (APC), a phytoconstituent sourced from leaves of Piper betle demonstrated antioxidant, anti-inflammatory, and antiarthritic properties. The present study aimed to evaluate the combined effect of APC and MTX on limiting progression of lipopolysaccharide accelerated collagen-induced arthritis, along with reduction of MTX-induced hepatic damage. A collagen-induced arthritis (CIA) model was established by immunising Sprague-Dawley rats with bovine collagen type II (CII) and lipopolysaccharide, followed by a booster dose of CII on day 15. Rats from days 11-27 were administered APC (20 mg/kg), methotrexate (1.5 mg/kg), or a combination of MTX and APC. The combinatorial therapy of APC and MTX significantly improved the parameters of arthritis as evident from the reduction in paw oedema and arthritic score and was endorsed by radiological and histopathological changes. This combination prevented the rise in levels of proinflammatory cytokines, tumour necrosis factor (TNF-α), and interleukin 6 (IL-6). Furthermore, unlike MTX-monotherapy, the APC-MTX combination decreased the associated cachexia, splenomegaly, and oxidative stress. Importantly, the hepatic damage mediated by MTX monotherapy was effectively attenuated by the inclusion of APC. Taken together, antioxidants such as APC when combined with MTX not only potentiated the antiarthritic effect but importantly alleviated the MTX-induced hepatic damage, thus endorsing its effectiveness in preventing progression of articular diseases such as rheumatoid arthritis.

Oral administration of undenatured native chicken type II collagen (UC-II) diminished deterioration of articular cartilage in a rat model of osteoarthritis (OA).

OBJECTIVE: The aim of this study was to determine the ability of undenatured native chicken type II collagen (UC-II) to prevent excessive articular cartilage deterioration in a rat model of osteoarthritis (OA). METHODS: Twenty male rats were subjected to partial medial meniscectomy tear (PMMT) surgery to induce OA. Immediately after the surgery 10 rats received vehicle and another 10 rats oral daily dose of UC-II at 0.66 mg/kg for a period of 8 weeks. In addition 10 naïve rats were used as an intact control and another 10 rats received sham surgery. Study endpoints included a weight-bearing capacity of front and hind legs, serum biomarkers of bone and cartilage metabolism, analyses of subchondral and cancellous bone at the tibial epiphysis and metaphysis, and cartilage pathology at the medial tibial plateau using histological methods. RESULTS: PMMT surgery produced moderate OA at the medial tibial plateau. Specifically, the deterioration of articular cartilage negatively impacted the weight bearing capacity of the operated limb. Immediate treatment with the UC-II preserved the weight-bearing capacity of the injured leg, preserved integrity of the cancellous bone at tibial metaphysis and limited the excessive osteophyte formation and deterioration of articular cartilage. CONCLUSION: Study results demonstrate that a clinically relevant daily dose of UC-II when applied immediately after injury can improve the mechanical function of the injured knee and prevent excessive deterioration of articular cartilage.