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Glucogallin Attenuates the LPS-Induced Signaling in Macrophages and Protects Mice against Sepsis.

ABSTRACT: The anti-oxidant and anti-inflammatory effect of beta-glucogallin (BGG), a plant-derived natural product, was evaluated in both in vitro and in vivo studies. For the in vitro study, the ability of BGG pre-treatment to quench LPS-induced effects compared to LPS alone in macrophages was investigated. It was found that BGG pre-treatment showed a significant decrease in ROS, NO, superoxide, and pro-inflammatory cytokines (TNF-alpha, IL-4, IL-17, IL-1β, and IL-6) and increased reduced glutathione coupled with the restoration of mitochondrial membrane potential. Gene profiling and further validation by qPCR showed that BGG pre-treatment downregulated the LPS-induced expression of c-Fos, Fas, MMP-9, iNOS, COX-2, MyD88, TRIF, TRAF6, TRAM, c-JUN, and NF-κB. We observed that BGG pre-treatment reduced nuclear translocation of LPS-activated NF-κB and thus reduced the subsequent expressions of NLRP3 and IL-1β, indicating the ability of BGG to inhibit inflammasome formation. Molecular docking studies showed that BGG could bind at the active site of TLR4. Finally, in the LPS-driven sepsis mouse model, we showed that pre-treatment with BGG sustained toxic shock, as evident from their 100% survival. Our study clearly showed the therapeutic potential of BGG in toxic shock syndrome.

SUBMITTER: Singh R

PROVIDER: S-EPMC9569803 | biostudies-literature | 2022 Sep

REPOSITORIES: biostudies-literature

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Glucogallin Attenuates the LPS-Induced Signaling in Macrophages and Protects Mice against Sepsis.

Singh Rajveer R Chandel Shivani S Ghosh Arijit A Matta Tushar T Gautam Anupam A Bhattacharya Arka A Babu Srivalliputturu Sarath SS Sukla Soumi S Nag Debasish D Ravichandiran Velayutham V Roy Syamal S Ghosh Dipanjan D

International journal of molecular sciences 20220924 19

The anti-oxidant and anti-inflammatory effect of beta-glucogallin (BGG), a plant-derived natural product, was evaluated in both in vitro and in vivo studies. For the in vitro study, the ability of BGG pre-treatment to quench LPS-induced effects compared to LPS alone in macrophages was investigated. It was found that BGG pre-treatment showed a significant decrease in ROS, NO, superoxide, and pro-inflammatory cytokines (TNF-alpha, IL-4, IL-17, IL-1β, and IL-6) and increased reduced glutathione cou ...[more]

PMID: 36232563

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Project description:BackgroundSepsis is a high-mortality disease without effective therapeutic options. The hyperactivation of the monocyte-macrophage system, especially M1 macrophages, triggers the onset of septic shock. Gentiopicroside (GENT), the main active component in the traditional Chinese medicinal herb Radix Gentianae, has been shown to have anti-inflammatory properties. Nevertheless, this anti-inflammatory effect has not been fully elucidated.MethodsIn vitro, we stimulated primary bone marrow-derived macrophages (BMMs) or peritoneal elucidated macrophages (PEMs) by lipopolysaccharide (LPS) and interferon (IFN)-γ and pre-treated with GENT and we tested the cytokines such as interleukin (IL)-1β, IL-6 and tumor necrosis factor (TNF) α production by enzyme linked immunosorbent assay (ELISA) or real-time quantitative PCR (qPCR). Further, we determined the NF-κB-mediated inflammatory pathway such as IKKα/β and p65 phosphorylation by Western blot. Then we detected the p65 nuclear localization by immunofluorescent staining. Moreover, NF-κB inhibitor and p65-targeted siRNAs were further used to validate the anti-inflammatory mechanism of GENT. In vivo, GENT (50 mg/kg) was administered intragastrically before and after LPS (40 mg/kg) injection. The death time were recorded and the serum levels of IL-1β, IL-6 and TNFα were tested by ELISA, and the IL-1β, IL-6 and TNFα mRNA expression in the lung were test by qPCR and the M1 infiltration in the lung were determined by F4/80 and INOS immunofluorescent staining.ResultsIn vitro, we observed that GENT reduced the inflammatory cytokine production of BMMs stimulated by (LPS)/IFN-γ and ameliorated the phosphorylation of IKKα/β and p65, the degradation of IκBα, and the translocation of p65 into the nucleus. We did not find GENT has any effect on MAPK signaling under LPS/IFN-γ stimulation. NF-κB inhibitor and p65 siRNAs eliminated the inhibition effect of GENT. In vivo, we observed GENT prevented mice from dying in the LPS-induced shock model and decreased the serum levels of IL-1β and IL-6, the mRNA expression of IL-1β, IL-6 and TNFα in lung tissue, and the amount of M1 macrophage infiltration in the lung.ConclusionsGENT prevented LPS/IFN-γ-induced inflammatory cytokine production by macrophages through the NF-κB signaling pathway in vitro and protected against the endotoxin shock induced by LPS in vivo.

Project description:BackgroundGut-resident macrophages (gMacs) supplemented by monocytes-to-gMacs differentiation play a critical role in maintaining intestinal homeostasis. Activating transcription factor 4 (ATF4) is involved in immune cell differentiation. We therefore set out to investigate the role of ATF4-regulated monocytes-to-gMacs differentiation in sepsis-induced intestinal injury.MethodsSepsis was induced in C57BL/6 wild type (WT) mice and Atf4- knockdown ( Atf4+/ - ) mice by cecal ligation and puncture or administration of lipopolysaccharide (LPS). Colon, peripheral blood mononuclear cells, sera, lung, liver, and mesenteric lymph nodes were collected for flow cytometry, hematoxylin and eosin staining, immunohistochemistry, quantitative reverse transcription polymerase chain reaction, and enzyme-linked immunosorbent assay, respectively.ResultsCD64, CD11b, Ly6C, major histocompatibility complex-II (MHC-II), CX3CR1, Ly6G, and SSC were identified as optimal primary markers for detecting the process of monocytes-to-gMacs differentiation in the colon of WT mice. Monocytes-to-gMacs differentiation was impaired in the colon during sepsis and was associated with decreased expression of ATF4 in P1 (Ly6C hi monocytes), the precursor cells of gMacs. Atf4 knockdown exacerbated the impairment of monocytes-to-gMacs differentiation in response to LPS, resulting in a significant reduction of gMacs in the colon. Furthermore, compared with WT mice, Atf4+/- mice exhibited higher pathology scores, increased expression of inflammatory factor genes ( TNF-α, IL-1β ), suppressed expression of CD31 and vascular endothelial-cadherin in the colon, and increased translocation of intestinal bacteria to lymph nodes and lungs following exposure to LPS. However, the aggravation of sepsis-induced intestinal injury resulting from Atf4 knockdown was not caused by the enhanced inflammatory effect of Ly6C hi monocytes and gMacs.ConclusionATF4, as a novel regulator of monocytes-to-gMacs differentiation, plays a critical role in protecting mice against sepsis-induced intestinal injury, suggesting that ATF4 might be a potential therapeutic target for sepsis treatment.

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Dataset Information

Glucogallin Attenuates the LPS-Induced Signaling in Macrophages and Protects Mice against Sepsis.

Publications

Glucogallin Attenuates the LPS-Induced Signaling in Macrophages and Protects Mice against Sepsis.

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