RESUMO
BACKGROUND: Qingchang Wenzhong Decoction (QCWZD), a chinese herbal prescription, is widely used for ulcerative colitis (UC). Nevertheless, the active ingredients and mechanism of QCWZD in UC have not yet been explained clearly. PURPOSE: This research focuses on the identification of the effective ingredients of QCWZD and the prediction and verification of their potential targets. METHODS: The UC mice were established by adding 3.0% dextran sulfate sodium (DSS) to sterile water for one week. Concurrently, mice in the treatment group were gavage QCWZD or mesalazine. LC-MS analyzed the main components absorbed after QCWZD treatment, and network pharmacology predicted their possible targets. ELISA, qPCR, immunohistochemistry and immunofluorescence experiments were used to evaluate the colonic inflammation level and the intestinal barrier completeness. The percentage of Th17 and Treg lymphocytes was detected by flow cytometry. RESULTS: After QCWZD treatment, twenty-seven compounds were identified from the serum. In addition, QCWZD treatment significantly reduced the increased myeloperoxidase (MPO) and inflammatory cell infiltration caused by DSS in the colonic. In addition, QCWZD can reduce the secretion of inflammatory factors in serum and promote the expression of mRNAs and proteins of occludin and ZO-1. Network pharmacology analysis indicated that inhibiting IL-6-STAT3 pathway may be necessary for QCWZD to treat UC. Flow cytometry analysis showed that QCWZD can restore the normal proportion of Th17 lymphocytes in UC mice. Mechanistically, QCWZD inhibited the phosphorylation of JAK2-STAT3 pathway, reducing the transcriptional activation of RORγT and IL-17A. CONCLUSIONS: Overall, for the first time, our work revealed the components of QCWZD absorbed into blood, indicated that the effective ingredients of QCWZD may inhibit IL-6-STAT3 pathway and inhibit the differentiation of Th17 lymphocytes to reduce colon inflammation.
Assuntos
Colite Ulcerativa , Animais , Colite Ulcerativa/induzido quimicamente , Colite Ulcerativa/tratamento farmacológico , Colite Ulcerativa/metabolismo , Colo , Sulfato de Dextrana , Modelos Animais de Doenças , Inflamação/metabolismo , Interleucina-17/metabolismo , Interleucina-6/metabolismo , Mesalamina/metabolismo , Mesalamina/farmacologia , Mesalamina/uso terapêutico , Camundongos , Camundongos Endogâmicos C57BL , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/metabolismo , Ocludina/metabolismo , Peroxidase/metabolismo , Células Th17 , ÁguaRESUMO
The excessive deposition of extracellular matrix (ECM) is the main characteristic of liver fibrosis, and hepatic stellate cells (HSCs) are the main source of ECM. The removal of activated HSCs has a reversal effect on liver fibrosis. Western blot and MTT analysis indicated that curcumol could relieve hepatic fibrosis by promoting HSCs receptor-interacting protein kinase 1/3 (RIP1/RIP3)-dependent necroptosis. Importantly, autophagy flow was monitored by constructing the mRFP-GFP-LC3 plasmid, and it was found that curcumol cleared activated HSCs in a necroptosis manner that was dependent on autophagy. Our study suggested that the activation of necrosome formed by RIP1 and RIP3 depended on Atg5, and that autophagosomes were also necessary for curcumol-induced necroptosis. Furthermore, microscale thermophoresis and co-immunoprecipitation assay results proved that curcumol could target Sirt1 to regulate autophagy by reducing the acetylation level of Atg5. The HSCs-specific silencing of Sirt1 exacerbated CCl4 -induced liver fibrosis in mice. The deacetylation of Atg5 not only accelerated the accumulation of autophagosomes but also enhanced the interaction between Atg5 and RIP1/RIP3 to induce necroptosis. Overall, our study indicated that curcumol could activate Sirt1 to promote Atg5 deacetylation and enhanced its protein-protein interaction function, thereby inducing autophagy and promoting the necroptosis of HSCs to reduce liver fibrosis.