Garlic-derived Exosomes-like Nanoparticles Improve Macrophage-mediated Inflammatory Response And Abnormal Lipid Metabolism In Hepatocytes

Non-alcoholic fatty liver disease(NAFLD)is a common chronic liver disease,and mainly characterized by diffuse hepatic steatosis and triglyceride accumulation.With the improvement of people’s living standards,the prevalence of NAFLD is gradually increasing,and NAFLD is becoming an emerging public health problem worldwide.During the formation and development of NAFLD,macrophages,one kind of key cells in the innate immune system,increase the accumulation of lipids and induce the inflammatory response in hepatocytes by releasing proinflammatory cytokines and the crosstalk between macrophages and hepatocytes.The increasing evidence indicates that,similar with exosomes secreted by mammalian cells,edible plant-derived exosomes-like nanoparticles(ELNs)can act as extracellular messengers to mediate intercellular communication and improve certain pathologic and physiological features in some animal models.Our previous works demonstrated that garlicderived exosomes-like nanoparticles(GELNs)showed an interesting characteristic to inhibit the inflammatory response in lipopolysaccharide(LPS)-treated macrophages.Additionally,GELNs also improved the inflammatory response in the adipose tissue of high-fat diet(HFD)-fed mice.But what is the roles of GELNs in hepatocytes and associated molecular mechanisms remains unclear.At present,we firstly evaluated the effects of GELNs on the inflammatory response induced by LPS in Kupffer cells and differentiated THP-1 macrophages.Data revealed that GELNs significantly attenuated the inflammatory response in LPS-treated differentiated THP-1 macrophages,but not in Kupffer cells.To explore whether GELNs mediated the inflammatory response in hepatocytes via macrophage-hepatocyte crosstalk,we co-cultured LO2 human normal liver cells with conditional media from LPS-treated THP-1 macrophages.The q RT-PCR assay revealed that conditional media from LPS treatment induced a significant increase of TNFα,IL-1β,and IL-6 in LO2 hepatocytes,but once the differentiated THP-1 macrophages were preincubated with GELNs in the presence of LPS,the role of conditional media in the m RNA levels of TNFα,IL-1β,and IL-6 was dramatically attenuated.At the same time,we investigated the effect of the conditional media on the accumulation of lipid droplets and the expression of genes associated with lipogenesis in oleic acid(OA)-incubated LO2 hepatocytes.Oil red O staining demonstrated that conditional media of THP-1 macrophages accelerated the accumulation of lipid droplets in OA-treated LO2 hepatocytes,but preincubation with GELNs distinctively inhibited the accumulation of lipid droplets.Similarly,data from q RT-PCR assay indicated that treatment with conditional media from LPS-treated THP-1 macrophages significantly increased the expression of FAS and ACC1,the two key genes for the biosynthesis of fat acids in the presence of OA,but the conditional media from GELNs-treated THP-1 macrophages remarkably inhibited the expression of FAS and ACC1 in OA-treated LO2 hepatocytes.Furthermore,beside with decreasing the liver weight,administration of GELNs for continued 4 weeks(i.g.once every other day)markedly suppressed the inflammatory response in the liver tissues of HFD-fed mice by decreasing the m RNA levels of TNFα and IL-1β,but not IL-6.At the same time,although GELNs supplementation had no significant effects on triglyceride(TG)and low-density lipoprotein cholesterol(LDL-C),but distinctively increased the content of high-density lipoprotein cholesterol(HDL-C)and decreased the level of total cholesterol(TC)in the serum of HFD-fed mice.Additionally,administration of GELNs evidently improved the dysfunction of liver in HFD-fed mice by decreasing aspartate aminotransferase(AST),and alanine aminotransferase(ALT).H&E staining of liver specimens showed that there were a lot of vacuoles in the liver of HFD-fed mice,administration of GELNs markedly prevented this phenomenon of the vacuole.And,GELNs supplementation significantly decreased the accumulation of lipid droplets in the livers of HFD-fed mice.To elucidate the mechanisms of GELNs preventing the accumulation of lipid droplets,we determined the expression of genes associated with fatty acid biosynthesis.The results demonstrated that administration of GELNs decreased the m RNA and protein levels of FAS and ACC1 in the livers of HFD-fed mice.Taken together,the findings of this study demonstrated that GELNs showed antiinflammatory properties in the differentiated THP-1 macrophages,and ameliorated the inflammatory response and dysfunction of hepatocytes via the crosstalk between macrophages and hepatocytes,hinted that GELNs might be a promising component to manage obesity because of its anti-inflammatory activity and it is of interest for future studies to investigate the potential application of GELNs in improving the metabolic inflammation of obesity and NAFLD.