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Zi Qi Decoction Alleviates Liver Fibrosis by Inhibiting the Toll-Like Receptor 4 (TLR4)-Related Nuclear Factor kappa b (NF-κB) and Mitogen-Activated Protein Kinase (MAPK) Signaling Pathways.

ABSTRACT: BACKGROUND Hepatic stellate cells (HSCs) play a vital role in hepatic fibrogenesis. Our recent clinical study indicated that the Zi Qi decoction, a Traditional Chinese Medicine formula, exhibited good efficacy in alleviating liver fibrosis, but the underlying mechanism remains elusive. MATERIAL AND METHODS Rats repeatedly injected with CCl₄ and cells stimulated with lipopolysaccharide were used as in vivo and in vitro models for liver fibrosis, respectively. The viability of LX-2 cells was evaluated with MTT assay. Relative messenger RNA (mRNA) expression of representative extracellular matrix (ECM) components was detected with real-time quantitative polymerase chain reaction (RT-qPCR). Moreover, total and phosphorylation levels of ECM proteins and pathway-related proteins were detected with western blotting. Immunofluorescent staining was used to show the nuclear translocation of nuclear factor kappa b (NF-kappaB) p65. Hematoxylin & eosin (H&E) and Masson trichrome staining and immunohistochemistry were performed to evaluate the extent of liver fibrosis. The levels of alanine transaminase (ALT), aspartate transaminase (AST), gamma-glutamyl transpeptidase (GGT), Hyp, tumor necrosis factor-alpha (TNF-alpha), and interleukin-6 (IL-6) were tested with an enzyme-linked immunosorbent assay. In addition, 7.0T micro-magnetic resonance imaging (micro-MRI) was used to evaluate the severity of hepatic damage. RESULTS The Zi Qi decoction inhibited lipopolysaccharide-mediated upregulation of mRNA and protein levels of representative ECM proteins both in vivo and in vitro. The Zi Qi decoction also suppressed activation of the Toll-like receptor 4 (TLR4)-related NF-kappaB signaling pathway and subsequently inhibited the nuclear translocation of activated NF-kappaB. Moreover, another TLR4 downstream pathway, mitogen-activated protein kinase (MAPK), was simultaneously restrained. The results of liver pathology and MRI in rat models also suggested the efficacy of the Zi Qi decoction in attenuating liver damage. CONCLUSIONS The Zi Qi decoction inhibited liver fibrosis by inhibiting the TLR4-related NF-kB and MAPK signaling pathways and preventing activation of HSCs.

SUBMITTER: Zhou J

PROVIDER: S-EPMC8054620 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundAcute pancreatitis (AP) is a common acute abdomen inflammation, characterized by the dysregulation of digestive enzyme production and secretion. Many studies have shown that Da Cheng Qi Decoction (DCQD) is a secure, effective prescription on AP. In this study, cerulein-stimulated AR42J cells damage model was established to further explore the feasibility and underlying mechanism of DCQD as a potential inhibitor of JAK2/STAT3 pathway for the treatment of AP.MethodsCell viability of DCQD was measured using a cell counting Kit-8 assay. Pancreatic biochemical markers such as amylase, lipase, and C-reactive protein production were measured by assay kits, respectively. Cytokines (TNF-α, IL-6, IL-10, and IL-1β) were assayed by ELISA. Protein location and protein expression were detected by immunofluorescence staining and Western blotting, respectively. Gene expression was assessed by real-time PCR. For mechanistic analysis of the effect of DCQD on JAK2/STAT3 signaling pathway, selective JAK2 inhibitor (Fedratinib) and STAT3 inhibitor (Stattic) as well as STAT3 activator (Garcinone D) were used.ResultsDCQD protected cells by regulating cerulein-induced inflammation and reducing the secretion of pancreatic biochemical markers. Moreover, DCQD could not only inhibit the nuclear translocation of p-STAT3, but also decrease the mRNA expression of JAK2 and STAT3 as well as the ratio of p-JAK2/JAK2 and p-STAT3/STAT3 in protein level. Additionally, DCQD could regulate the mRNA and protein expression of JAK2/STAT3 downstream effectors, Bax and Bcl-XL. The activated effect of cerulein on JAK2/STAT3 pathway was also reversed by JAK2 inhibitor Fedratinib or STAT3 inhibitor Stattic. And the overexpression of JAK2/STAT3 pathway, via STAT3 activator Garcinone D, did exert damage on cells, which bore a resemblance to cerulein.ConclusionThe activation of JAK2/STAT3 pathway may play a key role in the pathogenesis of cerulein-stimulated AR42J pancreatic acinar cell injury. DCQD could improve inflammatory cytokines and cell injury, which might be mediated by suppressing the activation of JAK2/STAT3 signaling pathway.

Project description:BackgroundJianpi Yangxue Qufeng Compound (JPYXQFC) is a Chinese medicine widely used in the clinical treatment of atopic dermatitis (AD) and has a significantly therapeutic effect. However, the mechanism of JPYXQFC in AD has been not understood clearly.ObjectiveThis study aimed to explore the effect of JPYXQFC on AD model cells and rats by regulating TLR4/MyD88/NF-κB signaling pathway.MethodsThe rats (n > 5) were given JPYXQFC decoction orally twice a day for three days, and their abdominal aortic blood was collected. HaCaT cell proliferation rate was tested by cell counting kit-8 (CCK-8) assays. We induced AD rat model through 2, 4-dinitrofluorobenzene (DNFB). AD rats were given oral JPYXQFC decoction and cetirizine (positive control). HaCaT cells were pretreated with JPYXQFC drug serum or cetirizine for 0.5 h and then stimulated with TNF-α/IFN-γ for 1 h. The mRNA levels of TLR4, MyD88, NF-κB, IL-4, IL-13, MCP1, TNF-α and TSLP were detected by quantitative real-time PCR (Q-RT-PCR), and TLR4/MyD88/NF-κB pathway protein expression was tested by Western blot. The total serum levels of immunoglobulin E (IgE), thymus and activation regulated chemokine/chemokine (C-C motif) ligand 17 (TARC/CCL17) were detected by enzyme-linked immunosorbent assay (ELISA). The epidermal thickness was detected by hematoxylin and eosin (HE) staining. The dermatitis area and score were measured by a ruler and a four-point scoring method, respectively.ResultsJPYXQFC significantly inhibited mRNA and protein expression of the TLR4/MyD88/NF-κB pathway and Histone H3 in TNF-α/IFN-γ-induced HaCaT cells and DNFB-induced rats, decreased the mRNA of IL-4, IL-13, MCP1, CCL22, TSLP and the level of AD-related genes IgE and TAEC/CCL17 of TNF-α/IFN-γ-induced HaCaT cells. Meanwhile, JPYXQFC significantly reduced the dermatitis area and dermatitis score in DNFB-induced rats, inhibited the levels of pro-inflammatory cytokines IL-6 and TNF-α, and upregulated FLG, as well as inhibited the levels of IgE and TARC/CCL17 in the serum of AD rats.ConclusionJPYXQFC alleviates AD by inhibiting the activation of TLR4/MyD88/NF-κB pathway.

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Zi Qi Decoction Alleviates Liver Fibrosis by Inhibiting the Toll-Like Receptor 4 (TLR4)-Related Nuclear Factor kappa b (NF-κB) and Mitogen-Activated Protein Kinase (MAPK) Signaling Pathways.

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