A pathological joint-liver axis mediated by matrikine-activated CD4+ T cells.

The knee joint has long been considered a closed system. The pathological effects of joint diseases on distant organs have not been investigated. Herein, our clinical data showed that post-traumatic joint damage, combined with joint bleeding (hemarthrosis), exhibits a worse liver function compared with healthy control. With mouse model, hemarthrosis induces both cartilage degeneration and remote liver damage. Next, we found that hemarthrosis induces the upregulation in ratio and differentiation towards Th17 cells of CD4+ T cells in peripheral blood and spleen. Deletion of CD4+ T cells reverses hemarthrosis-induced liver damage. Degeneration of cartilage matrix induced by hemarthrosis upregulates serological type II collagen (COL II), which activates CD4+ T cells. Systemic application of a COL II antibody blocks the activation. Furthermore, bulk RNAseq and single-cell qPCR analysis revealed that the cartilage Akt pathway is inhibited by blood treatment. Intra-articular application of Akt activator blocks the cartilage degeneration and thus protects against the liver impairment in mouse and pig models. Taken together, our study revealed a pathological joint-liver axis mediated by matrikine-activated CD4+ T cells, which refreshes the organ-crosstalk axis and provides a new treatment target for hemarthrosis-related disease. Intra-articular bleeding induces cartilage degradation through down-reulation of cartilage Akt pathway. During this process, the soluble COL II released from the damaged cartilage can activate peripheral CD4+ T cells, differention into Th17 cells and secretion of IL-17, which consequently induces liver impairment. Intra-articular application of sc79 (inhibitor of Akt pathway) can prevent the cartilage damage as well as its peripheral influences.

Fishmeal Protein Replacement by Defatted and Full-Fat Black Soldier Fly Larvae Meal in Juvenile Turbot Diet: Effects on the Growth Performance and Intestinal Microbiota.

A 12-week feeding trial was conducted to investigate the effect of the same fishmeal protein level replaced by black soldier fly larvae (Hermetia illucens) meal (BSFL) with different lipid contents on the growth performance and intestinal health of juvenile turbot (Scophthalmus maximus L.) (initial body weight 12.64 g). Three isonitrogenous and isolipidic diets were formulated: fish meal-based diet (FM), diets DF and FF, in which 14% fish meal protein of the FM diet was replaced by defatted and full-fat BSFL, respectively. There were no significant differences in growth performance, intestinal morphology, and mucosal barrier function between the DF and the FM group. However, diet FF markedly reduced the growth performance, intestinal perimeter ratio, and the gene expression of anti-inflammatory cytokine TGF-ß (P < 0.05). Compared to group FF, the communities of intestinal microbiota in group DF were more similar to group FM. Moreover, diet DF decreased the abundance of some potential pathogenic bacteria and enriched the potential probiotics, such as Bacillus. Diet FF obviously altered the composition of intestinal microbiota and increased the abundance of some potential pathogenic bacteria. These results suggested that the application of defatted BSFL showed more positive effects on fish growth and intestinal health than the full-fat BSFL, and the intestinal microbiota was closely involved in these effects.

Vitamin D3 deficiency induced intestinal inflammatory response of turbot through nuclear factor-κB/inflammasome pathway, accompanied by the mutually exclusive apoptosis and autophagy.

Vitamin D3 (VD3) participated widely in the nuclear factor-κB (NF-κB)-mediated inflammation, apoptosis, and autophagy through the vitamin D receptor (VDR). However, the molecular mechanisms remain not understood in teleost. The present study investigated the functions of VD3/VDR on intestinal inflammation, autophagy, and apoptosis of turbot in vivo and in vitro. Triple replicates of 30 fish were fed with each of three diets with graded levels of 32.0 (D0), 1012.6 (D1), and 3978.2 (D2) IU/kg VD3. Obvious intestinal enteritis was observed in the D0 group and followed with dysfunction of intestinal mucosal barriers. The intestinal inflammatory response induced by VD3 deficiency was regulated by the NF-κB/inflammasome signalling. The promotion of intestinal apoptosis and suppression of intestinal autophagy were also observed in the D0 group. Similarly, VD3 deficiency in vitro induced more intense inflammation regulated by NF-κB/inflammasome signalling. The mutually exclusive apoptosis and autophagy were also observed in the group without 1,25(OH)2D3 in vitro, accompanied by similar changes in apoptosis and autophagy increased apoptosis. The gene expression of VDRs was significantly increased with the increasing VD3 supplementation both in vivo and in vitro. Moreover, VDR knockdown in turbot resulted in intestinal inflammation, and this process relied on the activation of inflammasome mediated by NF-κB signalling. Simultaneously, intestinal apoptosis was promoted, whereas intestinal autophagy was inhibited. In conclusion, VD3 deficiency could induce intestinal inflammation via activation of the NF-κB/inflammasome pathway, intestinal apoptosis, and autophagy formed a mutually exclusive relation in teleost. And VDR is the critical molecule in those processes.

The alleviation of skin wound-induced intestinal barrier dysfunction via modulation of TLR signalling using arginine in gilthead seabream (Sparus aurata L).

The present study sought to investigate the effect of arginine on the involvement of toll-like receptors (TLRs) in skin wound-induced intestinal barrier dysfunction in gilthead seabream (Sparus aurata L.). Two replicates of fish (n = 8) were fed a commercial diet (CON, total 2.75% arginine), CON diet enriched with 1% arginine (ARG1, total 3.65% arginine) and 2% arginine (ARG2, total 4.53% arginine) for 30 days. Half of the fish were sampled, whereas the others were injured and sampled 7 days post-wounding. The intestinal histology results showed that a more intense infiltration of mixed leucocytes was evident in the wounded fish, which was remarkably reduced in fish that were fed the ARG1 diet. Serum IgM levels were significantly higher in the ARG1 group than levels in the CON group at 7 days post-wounding. Compared with the fish in the CON group after wounding, dietary administration of 1% arginine markedly downregulated the gene expression of TLRs (TLR2 and TLR5), MyD88, and proinflammatory cytokines (CSF1R, IL-1ß, and TNFα), but significantly enhanced the gene expression of IκBα, the anti-inflammatory cytokine TGF-ß1, and tight junction proteins (tricellulin and occludin) in wounded fish. Furthermore, the ARG2 diet demonstrated no additional benefits on intestinal cells, compared to both the ARG1 and the CON diets, and it even appeared to induce negative effects. In summary, dietary administration of 1% arginine significantly inhibited intestinal inflammatory response and tight junction disruption in skin-wounded gilthead seabream by modulating TLR signalling in the intestine.

Dietary administration of the probiotic Shewanella putrefaciens to experimentally wounded gilthead seabream (Sparus aurata L.) facilitates the skin wound healing.

The effect of the probiotic Shewanella putrefaciens Pdp11 (SpPdp11) was studied on the skin healing of experimentally wounded gilthead seabream (Sparus aurata L.). Two replicates (n = 12) of fish were fed CON diet or SP diet for 30 days. Half of the fish were sampled while the others were injured and sampled 7 days post-wounding. Results by image analysis of wound areas showed that SpPdp11 inclusion facilitated wound closure. Compared with the CON group, fish in SP group sampled 7 days post-wounding had a significantly decreased serum AST and increased ALB/GLOB ratio. Furthermore, protease and peroxidase activities were significantly increased in skin mucus from fish in SP group sampled 7 days post-wounding, compared with those fed CON diet. Additionally, SP diet up-regulated the gene expression of antioxidant enzymes, anti-inflammatory cytokines, and re-epithelialization related genes in the fish skin. Furthermore, significant decreases in pro-inflammatory cytokines expression were detected in fish from SP group, respect to control ones. Overall, SpPdp11 inclusion facilitated the wound healing and the re-epithelialization of the damaged skin, alleviated the inflammatory response in the wound area through intensifying the antioxidant system, and enhancing the neo-vascularization and the synthesis of matrix proteins in the skin wound sites of fish.

Skin wound healing in gilthead seabream (Sparus aurata L.) fed diets supplemented with arginine.

Dietary administration of arginine on the wound healing process of gilthead seabream was studied. Two replicates of fish (n = 8) were fed with either a commercial diet [control diet (CON), no arginine added] and the CON diet supplemented with 1% arginine (ARG1) or with 2% arginine (ARG2) for 30 days. Afterward, half of the fish were sampled while the other half were injured and continued to be fed the same diet for an extra week. Results by image analysis showed that the wound closure rate was significantly improved in fish that were fed the ARG1 diet, compared with those in the CON group. After seven days of wound healing, the aminotransferase and creatine kinase levels in the serum and the protease and peroxidase activities in the skin mucus were down-regulated, while the immunoglobulin M level in the skin mucus was up-regulated in the ARG1 group after wounding and in the CON group before wounding. Compared with the CON diet, the ARG1 diet remarkedly depressed the gene expression of mpo, il-8, and tnf-α, and enhanced the gene expression of tgf-ß1, igf-1, pcna, krt2, mmp9, fn1α, and colIα and the antioxidant enzyme cat in the skin tissues after wounding. Furthermore, compared with both the ARG1 and the CON groups, negative effects of the ARG2 diet on wound healing were demonstrated. In conclusion, a 1% arginine supplementation facilitates skin wound healing and prevents a systemic inflammation reaction by alleviating the inflammatory response and enhancing the re-epithelialization and ECM biosynthesis in skin wound sites.

Influence of skin wounds on the intestinal inflammatory response and barrier function: Protective role of dietary Shewanella putrefaciens SpPdp11 administration to gilthead seabream (Sparus aurata L.).

The effects of skin wounds on the intestinal barrier function and the beneficial effects of the dietary administration of Shewanella putrefaciens (known as SpPdp11) in gilthead seabream (Sparus aurata L.) were studied. Two replicates of fish were fed a commercial diet (control, CON) or CON diet enriched with 109 cfu g-1 SpPdp11 (SP diet) for 30 days. After this time, half of the fish were sampled, while the others were injured below the lateral line (wounded fish, W) and fed the same diets for an extra week before sampling (CON + W and SP + W groups). The intestinal histology and gene expression of different genes relevant for the intestinal barrier function were studied. The results showed that injured fish had a disordered enterocyte nucleus disposition, a more intense infiltration of mixed leucocytes and a thicker lamina propria in the intestine compared to the control fish. However, the fish in the SP + W group did not present these pathological symptoms in the intestine. No significant variations in the number of goblet cells were detected among the different experimental groups. Pro-inflammatory cytokines (colony-stimulating factor receptor 1, CSF1R, myeloperoxidase, MPO and interleukin-1ß, IL-1ß), mucins (intestinal mucin, IMUC and mucin 2, MUC2), and immunoglobulin T heavy chain (IGHT) were up-regulated, while tight junction protein occludin was down-regulated in the intestine from fish of the CON + W group. Similarly, the dietary administration of SpPdp11 markedly depressed the gene expression of pro-inflammatory cytokines, MUC2 and IGHT, but increased the gene expression of anti-inflammatory cytokine transforming growth factor-ß1 (TGF-ß1) and the tight junction proteins tricellulin and occluding after wounding. In brief, the skin wounds provoked an intestinal inflammatory response that included changes in the mucus layer and tight junction disruptions. Besides this, preventive administration of SpPdp11 alleviated the intestinal dysfunctions caused by skin wounds in gilthead seabream.

Citric acid mitigates soybean meal induced inflammatory response and tight junction disruption by altering TLR signal transduction in the intestine of turbot, Scophthalmus maximus L.

A 12-week feeding trial was conducted to investigate the effect of citric acid on the involvement of TLRs in the soybean meal induced inflammatory response and tight junction disruption in the distal intestine of juvenile turbot (Scophthalmus maximus L.). Four isonitrogenous and isolipidic practical diets were formulated: fish meal-based diet (FM); 40% fish meal protein in FM replaced with soybean meal protein (SBM); SBM + 1.5% citric acid and SBM + 3% citric acid. Compared to the FM, diet SBM significantly increased the gene expression of TLRs (TLR2, TLR3, TLR5b, TLR9, TLR21, TLR22) and MyD88, as well as TLR related molecules (NF-κB, IRF-3, p38 and JNK), which were remarkably reduced by dietary citric acid. Similarly, citric acid supplementation in SBM markedly depressed gene expression of pro-inflammatory cytokines (TNF-α and IFN-γ) and pore-forming tight junction protein Claudin-7, and enhanced gene expression of the anti-inflammatory cytokine TGF-ß1 and TJ proteins related to the decrease in paracellular permeability (Claudin-3, Claudin-4, Occludin, Tricellulin and ZO-1). Compared to the SBM, the concentration of IgM and C4 in serum was significantly reduced by dietary citric acid. In brief, dietary citric acid could synchronously inhibit TLRs-dependent inflammatory response regulated by NF-κB and IRF3, as well as cause TLRs-dependent tight junction disruption modulated by p38 and JNK. Therefore, citric acid could function on mitigating soybean meal induced enteropathy in the distal intestine of juvenile turbot.

Sodium butyrate supplementation in high-soybean meal diets for turbot (Scophthalmus maximus L.): Effects on inflammatory status, mucosal barriers and microbiota in the intestine.

A 12-week feeding trial was conducted to evaluate the effects of dietary sodium butyrate (NaBT) on the intestinal health of juvenile turbot (Scophthalmus maximus L.), in terms of inflammatory status, mucosal barriers and microbiota. Three isonitrogenous and isolipidic practical diets were used: (1) fish meal based group (FM); (2) soybean meal group (SBM), soy protein replacing 40% fish meal protein in FM; (3) NaBT group, 0.2% NaBT supplemented in SBM. Each diet was fed to triplicate tanks (30 fish in each tank). The current results showed that 0.2% dietary NaBT improved the growth performance of fish and alleviated the enteropathy, increasing the absorptive surface and mitigating the infiltration of mixed leukocytes in lamina propria. Fish fed the NaBT diet presented increased activities of intestinal brush border enzyme and similar nutrient digestibility with the FM group. Compared to SBM, the inclusion of 0.2% NaBT in diet significantly up-regulated the intestinal gene expression of tight junction proteins and down-regulated the gene expression of TNF-α and NF-κB. The gut microbial communities of the NaBT group were closer to the FM group than to the SBM group, in terms of PCoA, UPGMA and Heatmap analyses based on weighted Unifrac distance. The relative abundance of several dominant bacteria at the phylum (Proteobacteria, Bacteroidetes, Deinococcus-Thermus and Actinobacteria) and genus level (Thermus, Acinetobacter, Bacteroides and Silanimonas) were altered by dietary NaBT. In conclusion, dietary NaBT had positive roles in protecting the intestinal health of turbot from the impairment of soybean meal.