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A PGP-1-specific one/two-photon fluorogenic probe (BH1), capable of high sensitivity, super selectivity, and visual imaging of endogenous PGP-1 activity from live mammalian cells and serum/skin tissues from patients by using one/two-photon fluorescence microscopy (O/TPFM).

Sepsis is a complex inflammatory disorder in which high mortality is associated with an excessive inflammatory response. Inhibitor of growth 4 (ING4), which is a cofactor of histone acetyltransferase and histone deacetylase complexes, could negatively regulate this inflammation. However, the exact molecular signaling pathway regulated by ING4 remains uncertain. As a pivotal histone deacetylase, Sirtuin1 (SIRT1), which is widely accepted to be an anti-inflammatory molecule, has not been found to be linked to ING4. This study investigated how ING4 is involved in the regulation of inflammation by constructing lipopolysaccharide (LPS)-induced macrophage and mouse sepsis models. Our results revealed that ING4 expression decreased, whereas the levels of proinflammatory cytokines increased in LPS-stimulated cultured primary macrophages and RAW 264.7 cells. ING4 transfection was confirmed to alleviate the LPS-induced upregulation of proinflammatory cytokine expression both in vitro and in vivo. In addition, ING4-overexpressing mice were hyposensitive to an LPS challenge and displayed reduced organ injury. Furthermore, immunoprecipitation indicated a direct interaction between ING4 and the SIRT1 protein. Moreover, ING4 could block nuclear factor-kappa B (NF-κB) P65 nuclear translocation and restrict P65 acetylation at lysine 310 induced by LPS treatment. These results are the first to clarify that the anti-inflammatory role of ING4 is associated with SIRT1, through which ING4 inhibits NF-κB signaling activation. Our studies provide a novel signaling axis involving ING4/SIRT1/NF-κB in LPS-induced sepsis., (© 2020 The Authors. Immunology & Cell Biology published by John Wiley & Sons Australia, Ltd on behalf of Australian and New Zealand Society for Immunology, Inc.)

Protein kinase Cζ (PKCζ) is a member of the atypical protein kinase C family. Its roles in macrophages or skin-resident keratinocytes have not been fully evaluated. In this study, we provide evidence that PKCζ mediates lipopolysaccharide (LPS)-induced tumor necrosis factor α (TNFα) gene expression in the mouse macrophage cell line, RAW264.7. TNFα has been proven to be one of the main culprits of chronic wounds and impaired acute wounds, which are characterized by excessive inflammation, enhanced proteolysis and reduced matrix deposition. Among the multiple effects of TNFα on keratinocytes, the induction of chemokines which are indispensable factors involved in the massive infiltration of various inflammatory cells into skin lesions serves as a crucial mechanism. In the present study, we found that PKCζ inhibitor or its specific siRNA inhibited the TNFα-induced upregulation in the levels of the chemokines, interleukin (IL)-8, monocyte chemotactic protein-1 (MCP-1) and intercellular cell adhesion molecule-1 (ICAM-1) in HaCaT keratinocytes. Moreover, under a disrupted inflammatory environment, activated keratinocytes can synthesize large amounts of matrix metalloproteinases (MMP), which has a negative effect on tissue remodeling. We discovered that TNFα promoted the expression of MMP9 in a PKCζ-dependent manner. Further experiments revealed that nuclear factor-κB (NF-κB) was a key downstream molecule of PKCζ. In addition, as shown in vitro, PKCζ was not involved in the TNFα-induced decrease in HaCaT cell migration and proliferation. In vivo experiments demonstrated that TNFα-induced wound closure impairment and inflammatory disorders were significantly attenuated in the PKCζ inhibitor group. On the whole, our findings suggest that PKCζ is a crucial regulator in LPS- or TNFα-induced inflammatory responses in RAW264.7 cells and HaCaT keratinocytes, and that PKCζ/NF-κB signaling may be a potential target for interventional therapy for TNFα-induced skin inflammatory injury.

The type III histone deacetylase SIRT1 has recently emerged as a critical immune regulator by suppressing T-cell immunity and macrophage activation during inflammation, but its mechanism in regulating inflammatory response in macrophages remains unclear. Here we show that the expression of SIRT1 in macrophage cells decreased following the release of inflammation cytokines when the cells were stimulated with LPS. IRF8, an important regulator in monocyte differentiation and macrophage polarization, showed the opposite trend, with SIRT1 expression levels increasing after the cells treated with LPS. Co-immunoprecipitation and immunofluorescence experiments showed that SIRT1 could not only interact with IRF8, but also deacetylate it. LPS treatment had no effect on the expression of IRF8 in macrophage cells in which sirt1 was specifically deleted. Our results show that IRF8 may be the target of histone deacetylase SIRT1 to regulate the inflammation in the macrophage cells.

Background: Ischemia-reperfusion injury (IRI) of the intestine is associated with high morbidity and mortality in surgical and trauma patients. T cells participate in the pathogenesis of intestinal IRI, and T-cell depletion has been shown to inhibit inflammatory responses and diminish intestinal damage. However, the mechanism by which T cells contribute to intestinal IRI is not completely understood. Regulatory T cells (Tregs) are a specific subset of T cells that suppress immune responses and protect against tissue injuries. We hypothesized that Tregs might be involved in intestinal IRI., Materials and Methods: We subjected C57/Bl6 mice to 30 min of ischemia by clamping the superior mesenteric artery followed by reperfusion. Animals were pretreated with the anti-CD25 monoclonal antibody or adoptive transfer of Tregs before induction of IRI. The number of inflammatory cells, the level of inflammatory factors, and intestinal permeability were assessed., Results: Partial depletion of Tregs with an anti-CD25 monoclonal antibody potentiated intestinal permeability induced by IRI. The Treg-depleted mice showed more neutrophils and CD4(+) T cells. In addition, depletion of Tregs led to enhanced secretion of tumor necrosis factor-α, interferon-gamma, and interleukin (IL)-4 and reduced levels of IL-10. Furthermore, we performed adoptive transfer of Tregs and found that transfer of Tregs significantly inhibited the ischemia-reperfusion-induced increase in intestinal permeability., Conclusions: Our study indicated that Tregs participate in intestinal inflammatory responses induced by IRI and that targeting Tregs could be a novel therapeutic approach to intestinal IRI., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Abstract: Severe burns initiate an inflammatory cascade within the gut, which leads to intestinal mucosal injury. Although Na+/H+ exchanger 1 (NHE1) is recognised as a pivotal player in several inflammatory processes, its role in burn-induced intestinal injury is relatively unknown. We hypothesised that NHE1 might be involved in the increased intestinal permeability and barrier breakdown after severe burns. Thus, we here investigate whether the inhibition of NHE1 has a protective effect on burn-induced intestinal injury. Mice were subjected to a 30% total body surface area (TBSA) full-thickness steam burn. Cariporide was used to assess the function of NHE1 in mice with burn-induced intestinal injury by fluorescence spectrophotometry, Western blotting and enzyme linked immunosorbent assay (ELISA). We found that severe burn increased intestinal permeability, associated with the up-regulation of NHE1 and raised inflammatory cytokine levels. Mice treated with the NHE1 inhibitor cariporide had significantly attenuated burn-induced intestinal permeability and a reduced inflammatory response. NHE1 inhibition also reduced nuclear factor-κB (NF-κB) activation and attenuated p38 mitogen-activated protein kinase (MAPK) phosphorylation. Our study suggests that NHE1 plays an important role in burn-induced intestinal permeability through the regulation of the inflammatory response. Inhibition of NHE1 may be adopted as a potential therapeutic strategy for attenuating intestinal barrier breakdown. [Copyright &y& Elsevier]

Abstract: Background: Severe burns initiate an inflammatory response characterized by the upregulation of proinflammatory cytokine, which contributes to multiple organ injury. Na+/H+ exchanger 1 (NHE1) plays a significant role in several inflammatory processes. This study was designed to investigate the role of NHE1 in burn-induced inflammation and multiple organ injury. Materials and methods: Rats were subjected to a 30% total body surface area full-thickness burn. Cariporide was used to assess the function of NHE1 in burn-induced multiple organ injury by biochemical parameters, histologic changes, and inflammatory cytokine production. Results: We found that NHE1 expression was significantly increased after burn injury. Inhibition of NHE1 by cariporide attenuated burn-induced edema and tissue injury in heart, lung, kidney, and small intestine. Cariporide also inhibited plasma levels of tumor necrosis factor α, interleukin 6, and myeloperoxidase activity. Conclusions: These results indicate that NHE1 inhibition prevents burn-induced multiple organ injury. The salutary effects afforded by NHE1 inhibition, at least in part, are mediated by attenuating systemic inflammatory response. [Copyright &y& Elsevier]

Highlights: [•] We investigate the antioxidant activities of Lycium barbarum polysaccharides (LBP). [•] We examine the role of LBP in a models of intestinal ischemia–reperfusion injury. [•] The salutary effects depend on inhibiting oxidative stress and inflammation. [Copyright &y& Elsevier]

Background: The dysbiosis of gastrointestinal microbiome is an important reason for burn-induced intestinal injury. Clostridium butyricum (C.butyricum) and its production butyrate are beneficial for the homeostasis of intestinal microflora and suppression of inflammatory response.Purpose: The roles of C.butyricum and butyrate in burn-induced intestinal injury were explored. The effects of oral administration of C.butyricum on intestinal injury were observed in burned mice.Materials and Methods: The skin surface of mice was exposed to 95 °C water to induce a burn injury. Then the intestinal microbiome structure, abundance of C.butyricum and level of butyrate were respectively observed. The correction between intestinal permeability indicated by FITC dextran level and abundance of C.butyricum or level of butyrate was analyzed. C.butyricum was cultured and orally administrated to burned mice. The levels of butyrate, FITC dextran and pro-inflammatory cytokines, including interleukin 6 (IL-6) and tumor necrosis factor α (TNF-α) were respectively measured.Results: Burn injury altered the intestinal microbiome structure of mice, and especially decreased the abundance of C.butyricum and level of butyrate. Both the abundance of C.butyricum and the level of butyrate were negatively correlated with the intestinal permeability. Oral administration of C.butyricum increased the level of butyrate, decreased levels of TNF-α and IL-6, and suppressed intestinal damage in burn-injured mice.Conclusion: Oral administration of C.butyricum significantly alleviated the intestinal damage induced by burn injury. The therapeutic effects of C.butyricum and butyrate on burn injury should be further explored, which deserves further investigation. [ABSTRACT FROM AUTHOR]

Background Oxidative stress and inflammation exert central roles in burn-induced intestinal injury. Crocetin, a natural carotenoid compound from gardenia fruits and saffron, has been shown to inhibit oxidative stress and inflammatory response. However, the possibility of crocetin to be used in the treatment of intestinal injury after burn injury has not been investigated. The purpose of the present study was to investigate the effects and potential mechanisms of crocetin in burn-induced intestinal injury. Materials and methods Several free radical–generating and lipid peroxidation models were used to systematically assess the antioxidant activities of crocetin in vitro . A common burn model was used to induce the intestinal injury in rats. Changes in the levels of malondialdehyde, superoxidase dismutase, catalase, glutathione peroxidase, tumor necrosis factor α, interleukin 6, polymorphonuclear neutrophil accumulation, intestinal permeability, and intestinal histology were examined. Results In several models of antioxidant activity, crocetin exhibited marked inhibitory action against free radicals and lipid peroxidation. Crocetin increased levels of antioxidant enzymes and reduced intestinal oxidative injury in burn models. In addition, crocetin inhibited polymorphonuclear neutrophil accumulation, ameliorated tumor necrosis factor α and interleukin 6 levels, intestinal permeability, and histological changes. Conclusions Crocetin treatment may protect against burn-induced small intestinal injury, possibly by inhibiting burn-induced oxidative stress and inflammatory response. [ABSTRACT FROM AUTHOR]