Reyanning mixture inhibits M1 macrophage polarization through the glycogen synthesis pathway to improve lipopolysaccharide-induced acute lung injury.

ETHNOPHARMACOLOGICAL RELEVANCE: Reyanning (RYN) mixture is a traditional Chinese medicine composed of Taraxacum, Polygonum cuspidatum, Scutellariae Barbatae and Patrinia villosa and is used for the treatment of acute respiratory system diseases with significant clinical efficacy. AIM OF THE STUDY: Acute lung injury (ALI) is a common clinical disease characterized by acute respiratory failure. This study was conducted to evaluate the therapeutic effects of RYN on ALI and to explore its mechanism of action. MATERIALS AND METHODS: Ultra-high-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was used to analyze the chemical components of RYN. 7.5 mg/kg LPS was administered to induce ALI in rats. RYN was administered by gavage at doses of 2 ml/kg, 4 ml/kg or 8 ml/kg every 8 h for a total of 6 doses. Observations included lung histomorphology, lung wet/dry (W/D) weight ratio, lung permeability index (LPI), HE staining, Wright-Giemsa staining. ELISA was performed to detect the levels of TNF-α, IL-6, IL-10, Arg-1,UDPG. Immunohistochemical staining detected IL-6, F4/80 expression. ROS, MDA, SOD, GSH/GSSG were detected in liver tissues. Multiple omics techniques were used to predict the potential mechanism of action of RYN, which was verified by in vivo closure experiments. Immunofluorescence staining detected the co-expression of CD86 and CD206, CD86 and P2Y14, CD86 and UGP2 in liver tissues. qRT-PCR detected the mRNA levels of UGP2, P2Y14 and STAT1, and immunoblotting detected the protein expression of UGP2, P2Y14, STAT1, p-STAT1. RESULTS: RYN was detected to contain 1366 metabolites, some of the metabolites with high levels have anti-inflammatory, antibacterial, antiviral and antioxidant properties. RYN (2, 4, and 8 ml/kg) exerted dose-dependent therapeutic effects on the ALI rats, by reducing inflammatory cell infiltration and oxidative stress damage, inhibiting CD86 expression, decreasing TNF-α and IL-6 levels, and increasing IL-10 and Arg-1 levels. Transcriptomics and proteomics showed that glucose metabolism provided the pathway for the anti-ALI properties of RYN and that RYN inhibited lung glycogen production and distribution. Immunofluorescence co-staining showed that RYN inhibited CD86 and UGP2 expressions. In vivo blocking experiments revealed that blocking glycogen synthesis reduced UDPG content, inhibited P2Y14 and CD86 expressions, decreased P2Y14 and STAT1 mRNA and protein expressions, reduced STAT1 protein phosphorylation expression, and had the same therapeutic effect as RYN. CONCLUSION: RYN inhibits M1 macrophage polarization to alleviate ALI. Blocking glycogen synthesis and inhibiting the UDPG/P2Y14/STAT1 signaling pathway may be its molecular mechanism.

Nrf2-Dependent Protective Effect of Paeoniflorin on α-Naphthalene Isothiocyanate-Induced Hepatic Injury.

The pathological mechanism of cholestatic hepatic injury is associated with oxidative stress, hepatocyte inflammation, and dysregulation of hepatocyte transporters. Paeonia lactiflora Pall. and its compound can improve hepatic microcirculation, dilate bile duct, and promote bile flow, which is advantageous to ameliorate liver damage. Paeoniflorin (PEA), as the main efficacy component of Paeonia lactiflora Pall., has multiple pharmacological effects. PEA improves liver injury, but it remains obscure whether the protective action on [Formula: see text]-naphthalene isothiocyanate (ANIT)-induced cholestatic liver injury is dependent on the NF-E2 p45-related Factor 2 (Nrf2) signaling pathway. In this study, C57BL/6 mice were administrated with 80 mg⋅kg[Formula: see text]⋅d[Formula: see text] ANIT followed by PEA (75, 150, and 300 mg⋅kg[Formula: see text]⋅d[Formula: see text]) orally for 10 days, respectively. Tissue histology and liver function were detected, including serum enzymes, gallbladder (GB) weight, phenobarbital-induced sleeping time (PEN-induced ST), hepatic uridine di-phosphoglucuronosyltransferase (UDPG-T), malondialdehyde (MDA), and glutathione (GSH). The expressions of protein Nrf2, sodium taurocholate cotransporting polypeptide (Ntcp), and NADPH oxidase 4 (Nox4) were evaluated. Nrf2 plasmid or siRNA-Nrf2 transfection on LO2 cells and Nrf2-/- mice were used to explore the liver protective mechanism of PEA. Compared to ANIT-treated mice, PEA decreased serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), total bilirubin (TBIL), direct bilirubin (DBIL), total bile acid (TBA), and phenobarbital-induced sleeping time. The bile secretion, hepatic UDPG-T, MDA, GSH, and liver histology were improved. The expressions of protein Nrf2 and Ntcp in liver tissues increased, but Nox4 decreased. After Nrf2 plasmid or small interfering RNA (siRNA)-Nrf2 transfection, the protective effects of PEA on LO2 cells were, respectively, strengthened or weakened. Moreover, PEA had no significant effects on ANIT-treated Nrf2-/- mice. Our results suggest that Nrf2 is essential for PEA protective effects on ANIT-induced liver injury.