Nicotine alleviates the liver inflammation of non-alcoholic steatohepatitis induced by high-fat and high-fructose in mice / 北京大学学报(医学版)

Objective:To investigate the anti-inflammation effects by activation of the cholinergic anti-inflammatory pathway and its mechanisms in non-alcoholic steatohepatitis (NASH)model mice.Me-thods:6-week-old male C57BL/6J (B6)mice were randomly divided into four groups:the first group was normal mice,injected with saline;the second group was normal mice,injected with nicotine;the third group was NASH model mice,injected with saline;the fourth group was NASH model mice,injec-ted with nicotine.The experimental mice were fed with either standard chow (SC)or high-fat and high-fructose (HFHF)for 17 weeks to generate an NASH model mice.The mice received injection once daily for 3 weeks [nicotine dose,400 μg/kg].Then,their pathological characteristics and function of the liver were assessed.The expressions of interleukin-6 (IL-6)and tumor necrosis factor-α(TNF-α)in se-rum were analyzed by enzyme linked immunosorbent assay (ELISA).The expressions of alpha 7 nicotinic acetylcholine receptors (α7nAChR),Toll-like receptors-4 (TLR-4)and nuclear factor κB of phosphory-lation (p-NF-κB)in Kupffer cells were determined by Western blot and immunofluorescence assays.Re-sults:We successfully generated NASH model mice by imitating the high-fat and high-fructose dietary style of NASH patients.The results of our investigation demonstrated that nicotine could reduce signifi-cantly the levels of IL-6,and TNF-αin serum (P <0.05).The expression of p-NF-κB protein in the group which was NASH model mice injected with nicotine declined significantly as compared with the group which was NASH model mice injected with saline (P <0.05).And the expression of α7nAChR protein elevated significantly conversely (P <0.05 ).Conclusion:Activation of the cholinergic anti-inflammatory pathway could inhibit the release of inflammatory factors as TNF-αand IL-6 in NASH model mice,and the mechanism for the inhibition of inflammatory was mediated by NF-κB pathway.

Effects and mechanisms of the inflammatory reaction related to NASH and induced by activation of the cholinergic anti-inflammatory pathway / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the effects and mechanisms of the inflammatory reaction related to nonalcoholic steatohepatitis (NASH) and induced by activation of the cholinergic anti-inflammatory pathway.</p><p><b>METHODS</b>A mouse model of NASH was established by feeding a high-fat and high-sugar diet.Activation of the cholinergic anti-inflammatory pathway was achieved by nicotine administration to the NASH modeled mice and normal controls. Liver biopsies were taken and the concentrations of cytokines were measured. Isolated liver primary Kupffer cells and RAw264.7 cells were cultured, pre-treated or not with lipopolysaccharide (LPS) and exposed to nicotine, after which the supernatant concentrations of IL-6 and TNFa were determined by ELISA. The protein expression levels of phosphorylated (p)-NF-kB and I k B were detected in primary cultured Kupffer cells by western blotting.</p><p><b>RESULTS</b>The mouse model of NASH was successfully established, as evidenced by findings from liver biopsy and serum liver function tests. The degree of liver inflammation in the NASH mice decreased after nicotine administration, and the level of serum TNFa also significantly decreased. The levels of serum TNFa were 21.95+/-0.8 pg/mL in nicotine-treated mice and 38.07+/-1.7 pg/mL in the non-nicotine-treated NASH mice (P less than 0.05). The nicotine treatment also significantly reduced the concentration of TNFa in the culture supernatants of Kupffer cells after LPS stimulation; moreover, the supernatant level of TNFa decreased significantly after the nicotine treatment (Pless than 0.05). LPS stimulation of the RAw264.7 cells led to an increased level ofp-NF-kB and a reduced level ofI-kB, suggesting that the NF-kB pathway had been activated; different doses of nicotine pre-treatment led to down-regulation of the p-NF-kB level and up-regulation of the I-kB level, both in dose-dependent manners.</p><p><b>CONCLUSION</b>Activating the cholinergic anti-inflammatory pathway inhibits the NASH-related inflammatory reaction, and the mechanism for this inhibition involves the NF-kB signaling pathway.</p>

GLP-1 receptor activation effects the p38MAPK signal pathway in hepatic stellate cells / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the effects of activation of the GLP-1 receptor on the p38MAPK signaling pathway in hepatic stellate cells (HSCs).</p><p><b>METHODS</b>HSCs were isolated and identified according to morphological features; the levels of GLP-1R protein were determined by western blotting.The HSCs were randomly divided into a control grouP (normal saline treatment) and experimental grouP(liraglutide treatment); after 120 hours, the expression of p38MAPK mRNA was examined by RT-PCR and of phosphorylated (p)-p38MAPK protein was detected by western blotting.</p><p><b>RESULTS</b>GLP-1R proteins were detected in the HSCs. Compared with the control group, the experimental group showed significantly decreased p38MAPK mRNA and p-p38MAPK protein (both P < 0.01).</p><p><b>CONCLUSION</b>The p38MAPK signaling pathway could be down-regulated when GLP-1R is activated in HSCs.</p>

Effect and mechanism of rat macrophage α7-acetylcholine receptor mediated anti-inflammatory ;pathway on endotoxin-induced inflammation reaction / 中华临床感染病杂志

Objective To investigate the effect of rat macrophage α7-acetylcholine receptor (α7-AChR )-mediated cholinergic anti-inflammatory pathway on endotoxin-induced inflammation reaction . Methods Density gradient centrifugation method was used to isolate rat primary macrophages and flow cytometry was used to identify the cell purity .α7-AChR in macrophages was detected by fluorescence confocal microscopy and Western blot .After 1ipopolysaccharides ( LPS) was added to the culture media of primary culture of macrophages , the concentration of tumor necrosis factor ( TNF)-αin the supernatant was determined by enzyme linked immunosorbent assay (ELISA).The expression of p-NF-κB protein in primary cultured macrophages was detected by Western blot .ANOVA was used to analyze TNF-αlevels after adding different concentrations of nicotine .Results α7-AChR was observed by fluorescence confocal microscope in primary macrophages .Nicotine could significantly reduce the concentration of TNF-αin culture supernatants of macrophages after LPS stimulation .When the concentrations of nicotine were 0, 1, 10, 100μmol/L, the concentrations of TNF-αwere (2 123 ±86), (1 486 ±80), (1 316 ±83) and (1 090 ±77)pg/mL, respectively (t=16.33, 20.18 and 26.83, P<0.05).The level of p-NF-κB in macrophages was also reduced when nicotine added .Conclusion Activation of macrophage α7-AChR can inhibit the endotoxin-induced release of inflammatory factor TNF-α, which may be through NF-κB signal pathway .

Differential expressions of α-defensin between chronic hepatitis B and asymptomatic HBV carriers / 中华临床感染病杂志

Objective To explore different pathogenesis of chronic hepatitis B(CHB)and asymptomatic H BV carriers(ASCs)by identifying differentially expressed genes.Methods Subtracted library was constructed by suppression subtraetive hybridization(SSH),and α-defensin was identified by dot blot hybridization.Peripheral blood was collected from 46 CHB patients and 11 ASCs.and the expressions of α-defensin mRNA in peripheral blood mononuclear cell and protein in plasma were determined by the real time RT-PCR and enzyme-linked immunosorbent assay(ELISA).Results Real time RT-PCR showed that the expression of α-defensin mRNA in blood samples of CHB was 1.4-fold higher than that of ASCs.As shown by ELISA,the plasma level of α-defensin in CHB was higher than that of ASCs [(216.40±81.25)μg/L vs.(156.00±57.26)μg/L,t=2.23,P<0.05].Conclusion α-defensin may involve in the pathogenesis of CHB,for it iS over-expressed in CHB patients.