Melatonin Delays Arthritis Inflammation and Reduces Cartilage Matrix Degradation through the SIRT1-Mediated NF-κB/Nrf2/TGF-ß/BMPs Pathway.

Cartilage, a flexible and smooth connective tissue that envelops the surfaces of synovial joints, relies on chondrocytes for extracellular matrix (ECM) production and the maintenance of its structural and functional integrity. Melatonin (MT), renowned for its anti-inflammatory and antioxidant properties, holds the potential to modulate cartilage regeneration and degradation. Therefore, the present study was devoted to elucidating the mechanism of MT on chondrocytes. The in vivo experiment consisted of three groups: Sham (only the skin tissue was incised), Model (using the anterior cruciate ligament transection (ACLT) method), and MT (30 mg/kg), with sample extraction following 12 weeks of administration. Pathological alterations in articular cartilage, synovium, and subchondral bone were evaluated using Safranin O-fast green staining. Immunohistochemistry (ICH) analysis was employed to assess the expression of matrix degradation-related markers. The levels of serum cytokines were quantified via Enzyme-linked immunosorbent assay (ELISA) assays. In in vitro experiments, primary chondrocytes were divided into Control, Model, MT, negative control, and inhibitor groups. Western blotting (WB) and Quantitative RT-PCR (q-PCR) were used to detect Silent information regulator transcript-1 (SIRT1)/Nuclear factor kappa-B (NF-κB)/Nuclear factor erythroid-2-related factor 2 (Nrf2)/Transforming growth factor-beta (TGF-ß)/Bone morphogenetic proteins (BMPs)-related indicators. Immunofluorescence (IF) analysis was employed to examine the status of type II collagen (COL2A1), SIRT1, phosphorylated NF-κB p65 (p-p65), and phosphorylated mothers against decapentaplegic homolog 2 (p-Smad2). In vivo results revealed that the MT group exhibited a relatively smooth cartilage surface, modest chondrocyte loss, mild synovial hyperplasia, and increased subchondral bone thickness. ICH results showed that MT downregulated the expression of components related to matrix degradation. ELISA results showed that MT reduced serum inflammatory cytokine levels. In vitro experiments confirmed that MT upregulated the expression of SIRT1/Nrf2/TGF-ß/BMPs while inhibiting the NF-κB pathway and matrix degradation-related components. The introduction of the SIRT1 inhibitor Selisistat (EX527) reversed the effects of MT. Together, these findings suggest that MT has the potential to ameliorate inflammation, inhibit the release of matrix-degrading enzymes, and improve the cartilage condition. This study provides a new theoretical basis for understanding the role of MT in decelerating cartilage degradation and promoting chondrocyte repair in in vivo and in vitro cultured chondrocytes.

Effect of Composite Probiotics on Antioxidant Capacity, Gut Barrier Functions, and Fecal Microbiome of Weaned Piglets and Sows.

This study investigated the efficacy of a composite probiotics composed of lactobacillus plantarum, lactobacillus reuteri, and bifidobacterium longum in alleviating oxidative stress in weaned piglets and pregnant sows. Evaluations of growth, oxidative stress, inflammation, intestinal barrier, and fecal microbiota were conducted. Results showed that the composite probiotic significantly promoted average daily gain in piglets (p < 0.05). It effectively attenuated inflammatory responses (p < 0.05) and oxidative stress (p < 0.05) while enhancing intestinal barrier function in piglets (p < 0.01). Fecal microbiota analysis revealed an increase in the abundance of beneficial bacteria such as faecalibacterium, parabacteroides, clostridium, blautia, and phascolarctobacterium in piglet feces and lactobacillus, parabacteroides, fibrobacter, and phascolarctobacterium in sow feces, with a decrease in harmful bacteria such as bacteroides and desulfovibrio in sow feces upon probiotic supplementation. Correlation analysis indicated significant negative associations of blautia with inflammation and oxidative stress in piglet feces, while treponema and coprococcus showed significant positive associations. In sow feces, lactobacillus, prevotella, treponema, and CF231 exhibited significant negative associations, while turicibacter showed a significant positive association. Therefore, the composite probiotic alleviated oxidative stress in weaned piglets and pregnant sows by modulating fecal microbiota composition.

Bifidobacterium longum GL001 alleviates rat intestinal ischemia-reperfusion injury by modulating gut microbiota composition and intestinal tissue metabolism.

Intestinal ischemia-reperfusion (IIR) injury leads to inflammation and oxidative stress, resulting in intestinal barrier damage. Probiotics, due to their anti-inflammatory and antioxidant properties, are considered for potential intervention to protect the intestinal barrier during IIR injury. Bifidobacterium longum, a recognized probiotic, has targeted effects on IIR injury, but its mechanisms of action are not yet understood. To investigate the mechanism of Bifidobacterium longum intervention in IIR injury, we conducted a study using a rat IIR injury model. The results showed that Bifidobacterium longum could alleviate inflammation and oxidative stress induced by IIR injury by suppressing the NF-κB inflammatory pathway and activating the Keap1/Nrf2 signaling pathway. Bifidobacterium longum GL001 also increased the abundance of the gut microbiota such as Oscillospira, Ouminococcus, Corynebacterium, Lactobacillus, and Akkermansia, while decreasing the abundance of Allobaculum, [Prevotella], Bacteroidaceae, Bacteroides, Shigella, and Helicobacter. In addition, Bifidobacterium longum GL001 reversed the changes in amino acids and bile acids induced by IIR injury and reduced the levels of DL-cysteine, an oxidative stress marker, in intestinal tissue. Spearman correlation analysis showed that L-cystine was positively correlated with Lactobacillus and negatively correlated with Shigella, while DL-proline was positively correlated with Akkermansia. Moreover, bile acids, cholic acid and lithocholic acid, were negatively correlated with Lactobacillus and positively correlated with Shigella. Therefore, Bifidobacterium longum GL001 may alleviate IIR injury by regulating the gut microbiota to modulate intestinal lipid peroxidation and bile acid metabolism.

Lactiplantibacillus plantarum GL001 alleviates jejunal oxidative damage induced by intestinal ischemia-reperfusion injury by influencing jejunal tissue metabolism through the improvement of jejunal microbial composition.

Intestinal ischemia-reperfusion (IIR) injury is associated with inflammation and oxidative stress, yet its precise mechanisms remain not fully understood. IIR injury is closely linked to the gut microbiota and its metabolites. The anti-inflammatory and antioxidant effects of Lactiplantibacillus plantarum are specific to IIR. In our study, we conducted a 30-day pre-treatment of SD rats with both a standard strain of Lactiplantibacillus plantarum and Lactiplantibacillus plantarum GL001. After a 7-day cessation of treatment, we induced an IIR injury model to investigate the mechanisms by which Lactiplantibacillus plantarum alleviates IIR damage. The results demonstrate that Lactiplantibacillus plantarum effectively mitigates the inflammatory and oxidative stress damage induced by IIR. Lactiplantibacillus plantarum GL001 can improve the gut microbiota by reducing the abundance of harmful bacteria and increasing the abundance of beneficial bacteria. In IIR intestinal tissue, the levels of secondary bile acids are elevated. The content of the bacterial metabolite Calcimycin increases. Annotations of metabolic pathways suggest that Lactiplantibacillus plantarum GL001 can alleviate IIR damage by modulating calcium-phosphorus homeostasis through the regulation of parathyroid hormone synthesis, secretion, and action. Microbiota-metabolite correlation analysis reveals a significant negative correlation between calcimycin and Lactonacillus and a significant positive correlation between calcimycin and Shigella. There is also a significant positive correlation between calcimycin and secondary bile acids. Lactiplantibacillus plantarum GL001 can alleviate oxidative damage induced by IIR through improvements in gut microbiota and intestinal tissue metabolism.

Curcumin alleviates spleen immunotoxicity induced by decabrominated diphenyl ethers (BDE-209) by improving immune function and inhibiting inflammation and apoptosis in broilers.

This study was conducted to assess the mitigating effects of curcumin (Cur) on immunotoxicity in the spleen of broilers induced by the polybrominated diphenyl ether BDE-209. Eighty one-day-old broilers were allocated to the following four groups: control group, BDE-209 (0.4 g/kg) group, BDE-209 (0.4 g/kg) + Cur (0.3 mg/kg) group, and Cur (0.3 mg/kg) group. Growth performance, immunological function, inflammation, and apoptosis were assessed after 42 days of treatment. The findings demonstrate that firstly, Cur restored spleen damage caused by BDE-209 by increasing body weight, decreasing feed-to-gain ratio, correcting the spleen index, and improving the histopathological structure of the spleen. Secondly, Cur relieved BDE-209-induced immunosuppression by increasing the levels of the immunoglobulins IgG, IgM, and IgA in the serum, as well as the levels of white blood cells and lymphocytes. The levels at which GATA binding protein 3, T-box expressed in T cells, interferon-γ, and interleukin (IL)- 4 are expressed were controlled. The ratio of T helper (Th) type 1 (Th1) to Th2 cells in the spleen of broilers was also controlled. Thirdly, Cur reduced the expression of Toll like receptor (TLR) 2, TLR4, nuclear factor (NF)-κB, IL-8, IL-6, and IL-1ß, which alleviated BDE-209-induced inflammation in broilers. Cur reduced BDE-209-induced apoptosis by increasing the expression of the bcl-2 protein, decreasing the expression of cleaved caspase-3 and bax proteins, decreasing the bax/bcl-2 protein ratio, and decreasing the mean optical density of TUNEL. These results suggest that Cur protects broiler spleens from BDE-209-induced immunotoxicity via modulating humoral immunity, the equilibrium between Th1 and Th2 cells, the TLRs/NF-κB inflammatory pathway, and the apoptotic pathway.

Dexmedetomidine attenuates hepatic ischemia-reperfusion injury-induced apoptosis via reducing oxidative stress and endoplasmic reticulum stress.

Dexmedetomidine (DEX) affords a hepatoprotective effect during ischemia-reperfusion (IR) injury (IRI); however, the underlying mechanism remains elusive. In this work, using a rat liver IR model and a BRL-3A cell hypoxia-reoxygenation (HR) model, we explored whether DEX protects the liver against IRI by decreasing oxidative stress (OS), endoplasmic reticulum stress (ERS), and apoptotic pathways. We found that DEX significantly increased SOD and GSH activity while decreasing ROS and MDA levels in BRL-3A cells, successfully preventing HR-induced OS damage. DEX administration reduced JNK, ERK, and P38 phosphorylation and blocked HR-induced MAPK signaling pathway activation. Additionally, DEX administration reduced the expression of GRP78, IRE1α, XBP1, TRAF2, and CHOP, which reduced HR-induced ERS. NAC prevented the MAPK pathway from being activated and inhibited the ERS pathway. Further research showed that DEX significantly reduced HR-induced apoptosis by suppressing the expression of Bax/Bcl-2 and cleaved caspase-3. Similarly, animal studies demonstrated DEX exerted a protective effect of the liver by alleviating histopathological injury and enhancing liver function, mechanically DEX reduced cell apoptosis in liver tissue by reducing oxidative stress and ERS. In conclusion, DEX mitigates OS and ERS during IR, thereby suppressing cell apoptosis, thus providing protection to the liver.

Oral administration of live combined Bacillus subtilis and Enterococcus faecium alleviates colonic oxidative stress and inflammation in osteoarthritic rats by improving fecal microbiome metabolism and enhancing the colonic barrier.

Osteoarthritis (OA) causes intestinal damage. The protective effect of probiotics on the intestine is indeed effective; however, the mechanism of protection against intestinal damage in OA is not clear. In this study, we used meniscal/ligamentous injury (MLI) to mimic OA in rats and explored the colonic protective effects of Bacillus subtilis and Enterococcus faecium on OA. Our study showed that treatment with B. subtilis and E. faecium attenuated colonic injury and reduced inflammatory and oxidative stress factors in the serum of osteoarthritic rats. α- and ß diversity of the fecal flora were not different among groups; no significant differences were observed in the abundances of taxa at the phylum and genus levels. We observed the presence of the depression-related genera Alistipes and Paraprevotella. Analysis of fecal untargeted metabolism revealed that histamine level was significantly reduced in the colon of OA rats, affecting intestinal function. Compared to that in the control group, the enriched metabolic pathways in the OA group were primarily for energy metabolisms, such as pantothenate and CoA biosynthesis, and beta-alanine metabolism. The treatment group had enriched linoleic acid metabolism, fatty acid biosynthesis, and primary bile acid biosynthesis, which were different from those in the control group. The differences in the metabolic pathways between the treatment and OA groups were more evident, primarily in symptom-related metabolic pathways such as Huntington's disease, spinocerebellar ataxia, energy-related central carbon metabolism in cancer, pantothenate and CoA biosynthesis metabolic pathways, as well as some neurotransmission and amino acid transport, and uptake- and synthesis-related metabolic pathways. On further investigation, we found that B. subtilis and E. faecium treatment enhanced the colonic barrier of OA rats, with elevated expressions of tight junction proteins occludin and Zonula occludens 1 and MUC2 mRNA. Intestinal permeability was reduced, and serum LPS levels were downregulated in the treatment group. B. subtilis and E. faecium also regulated the oxidative stress pathway Keap1/Nrf2, promoted the expression of the downstream protective proteins HO-1 and Gpx4, and reduced intestinal apoptosis. Hence, B. subtilis and E. faecium alleviate colonic oxidative stress and inflammation in OA rats by improving fecal metabolism and enhancing the colonic barrier.

Melatonin Prevents Chondrocyte Matrix Degradation in Rats with Experimentally Induced Osteoarthritis by Inhibiting Nuclear Factor-κB via SIRT1.

Osteoarthritis (OA) is a common degenerative joint disease characterized by an imbalance of cartilage extracellular matrix (ECM) breakdown and anabolism. Melatonin (MT) is one of the hormones secreted by the pineal gland of the brain and has anti-inflammatory, antioxidant, and anti-aging functions. To explore the role of MT in rats, we established an OA model in rats by anterior cruciate ligament transection (ACLT). Safranin O-fast green staining showed that intraperitoneal injection of MT (30 mg/kg) could alleviate the degeneration of articular cartilage in ACLT rats. Immunohistochemical (IHC) analysis found that MT could up-regulate the expression levels of collagen type II and Aggrecan and inhibit the expression levels of matrix metalloproteinase-3 (MMP-3), matrix metalloproteinase-13 (MMP-13), and ADAM metallopeptidase with thrombospondin type 1 motif 4 (ADAMTS-4) in ACLT rats. To elucidate the mechanism of MT in protecting the ECM in inflammatory factor-induced rat chondrocytes, we conducted in vitro experiments by co-culturing MT with a culture medium. Western blot (WB) showed that MT could promote the expression levels of transforming growth factor-beta 1 (TGF-ß1)/SMAD family member 2 (Smad2) and sirtuin 2-related enzyme 1 (SIRT1) and inhibit the expression of levels of phosphorylated nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibi-tor (p-p65) and phosphorylated IκB kinase-α (p-IκBα). In addition, WB and real-time PCR (qRT-PCR) results showed that MT could inhibit the expression levels of MMP-3, MMP-13, ADAMTS-4, inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2) in chondrocytes induced by interleukin-1ß (IL-1ß), and up-regulate the expression of chondroprotective protein type II collagen. We found that in vivo, MT treatment protected articular cartilage in the rat ACLT model. In IL-1ß-induced rat chondrocytes, MT could reduce chondrocyte matrix degradation by up-regulating nuclear factor-kB (NF-κB) signaling pathway-dependent expression of SIRT1 and protecting chondrocyte by activating the TGF-ß1/Smad2 pathway.

Effects of Lactiplantibacillus plantarum 19-2 on immunomodulatory function and gut microbiota in mice.

This study aims to evaluate the effects of Lactiplantibacillus plantarum 19-2 (L. plantarum 19-2) on mice treated with the alkylating agent cyclophosphamide (CTX). Our findings show that L. plantarum 19-2 restored the spleen and thymus index and the number of white blood cells and lymphocytes% in CTX treated mice. Serum immunoglobulin levels in CTX-treated mice were increased by L. plantarum 19-2. In addition, as compared to the model group, L. plantarum 19-2 upregulated the content of SIgA, while L. plantarum 19-2 regulates the mRNA and protein expression levels of GATA-3, T-bet, IFN-γ, and IL-4 in small intestinal tissues, which adjusted mucosal barriers, structural status, and the balance of Helper T-cell 1 and Helper T-cell 2. Lactiplantibacillus plantarum 19-2 regulated the distribution of intestinal flora in mice, promoting the growth of Bacteroides and Proteobacteria. In addition, L. plantarum 19-2 inhibited the growth of several harmful bacteria, including Actinobacteria and Firmicutes.

Dark-light cycle disrupts bone metabolism and suppresses joint deterioration in osteoarthritic rats.

BACKGROUND: Light alteration affects the internal environment and metabolic homeostasis of the body through circadian rhythm disorders (CRD). CRD is one of the factors that induce and accelerate osteoarthritis (OA). Therefore, the aim of this study was to evaluate the effects of continuous dark-light (DL) cycle on joint inflammation, bone structure, and metabolism in normal and OA Sprague-Dawley (SD) rats. METHODS: Interleukin (IL)-1ß, IL-6, inducible nitric oxide synthase (iNOS), and tumor necrosis factor (TNF)-α were used to evaluate the systemic inflammation in rats. The pathological changes and inflammatory reactions of the cartilage and synovium of the knee joint in rats were evaluated by Safranin O-fast green and immunological staining. Bone turnover was assessed by histomorphometry and µCT scanning, as well as bone metabolism markers and proteins. The expression changes of clock proteins BMAL1, NR1D1, PER3, and CRY1 in representative tissues were detected by western blotting. RESULTS: DL cycle significantly inhibited body weight gain in normal and OA rats. The levels of proinflammatory factors in the peripheral blood circulation and degradation enzymes in the cartilage were significantly decreased in OA+DL rats. DL cycle significantly destroyed the structure of subchondral bone in hindlimbs of OA rats and reduced trabecular bone numbers. The decrease of bone mineral density (BMD), percent bone volume with respect to total bone volume (BV/TV), trabecular number (TB.N), osteoclast number, and mineralization could also be found. The ratio of the receptor activator of nuclear factor-kappa B ligand/osteoprotegerin (RANKL/OPG) in the bone marrow of OA rats was markedly increased under DL, along with the activation of the mononuclear/phagocyte system. The expression of representative clock proteins and genes BMAL1, PER3, and CRY1 were markedly changed in the tissues of OA+DL rats. CONCLUSIONS: These results suggested that DL cycle dampened the arthritis and promoted bone resorption and bone mass loss. DL cycle affects bone turnover by regulating osteoclast production in osteoarthritic rats.

Intra-articular Injection of Baicalein Inhibits Cartilage Catabolism and NLRP3 Inflammasome Signaling in a Posttraumatic OA Model.

Baicalein has been shown to have chondroprotective potential in vitro. However, its effect on disease modification in osteoarthritis (OA) is largely unknown. The present study is aimed at determining whether baicalein could slow the progression of OA and inhibit OA-related inflammation in a rat model of destabilization of the medial meniscus (DMM) and the underlying mechanisms. The rats subjected to DMM surgery were treated with baicalein (0.8, 1.6, and 3.2 µg/L, 50 µL, once a week) by intra-articular injection for 6 weeks. Dexamethasone (0.4 mg/mL, 50 µL, once a week) was used as a positive control. Histologic grading of cartilage degeneration was performed using the Osteoarthritis Research Society International (OARSI) recommended grading system (on a scale of 0-6). The expression levels of molecules associated with cartilage homeostasis and inflammatory cytokines were analyzed; moreover, the NLRP3 inflammasome activation and cartilage oxidative stress-associated molecules were determined. Baicalein treatment reduced the OARSI score and slowed OA disease progression in a dose-dependent manner within a certain range. Compared with DMM rats, intra-articular injection of baicalein led to (1) reduced levels of inflammatory mediates such as IL-1ß and TNF-α, (2) reduced immunochemical staining of MMP-13 and ADAMTS-5, (3) suppressed immunochemical staining loss of type II collagen, (4) reduced expression of cartilage degradation markers including CTX-II and COMP in urine, and (5) inhibited NLRP3 inflammasome activation rather than regulated expression of SOD, GSH, and MDA. In contrast to the administration of baicalein, dexamethasone injection showed similar effects to slow OA progression, while dexamethasone inhibited NLRP3 inflammasome partly through decreasing levels of SOD, GSH, and MDA. This study indicated that baicalein may have the potential for OA prevention and exerts anti-inflammatory effects partly via suppressing NLRP3 inflammasome activation without affecting oxidative stress-associated molecules, and inhibition of cartilage catabolism enzymes in an OA rat model.