| Metabolic syndromes are occurring at epidemic rates.It appears to have emerged largely from changes in our diet and reduced physical activity.An important dietary change has been the substantial increase in the amount of dietary fructose consumption from high intake of sucrose and high fructose corn syrup,a common sweetener used in the food industry.Emerging evidence from epidemiological and biochemical studies suggests that the high dietary intake of fructose has become an important causative factor in the development of the metabolic syndrome.A high flux of fructose to the liver,the main organ capable of metabolizing this sugar,perturbs glucose metabolism and glucose uptake pathways,and leads to a significantly enhanced rate of de novo lipogenesis and triglycerides(TG)synthesis,driven by the high flux of molecules from fructose catabolism.These metabolic disturbances appear to underlie the induction of insulin resistance(IR)commonly observed with high fructose feeding in both humans and animal models.The aim of the present study was to relate fructose to metabolism disorders,and established a new HepG2 cell model with glucose,fructose and high levels of free fatty acids.The mechanism of the disorders of this cell model was elucidated and the effects of functional compounds were evaluated.Finally,how would the fructose affect fatty liver cell models were studied as well.The effect of adding fructose to two levels of glucose was compared with monosaccharides.Based on the prior experiment,the fatty acids(OPA)which consisted of oleic acid(OA)and palmitic acid(PA)(molar ratio was 2:1),were added and investigated.The supertanant TG and fructosamine,intracellular TG and tatol cholesterol(TC),intracellular and extracellular malondialdehyde(MDA),and insulin stimulated glucose uptake and glycogen level were accessed by corresponding kits.The ROS level was determined by fluorescent staining.Cell cycle analysis was carried out on flowcytometry.Intracellular and extracellular free fatty acids were quantified using gas chromatography,and the levels of glyoxal,methylglyoxal and glyceraldehyde were tested by gas chromatography mass spectrometry.The uric acid inside of the cells was analyzed by high performance liquid chromatography.To determined the level of secreted proteins and cytokines,including apolipoprotein B,C-reaction protein,tumor necrosis factor-? and interleukin-6,Elisa kits were used.Gene expression of metabolism and synthesis of lipids were examined using RT-PCR.After confirmation of the impact of fructose towards cells,the cell model were established by chosen treatments.Moreover,the intervension of some drugs,oryzanol and polyphenols on established model cells were evaluated.Furthermore,fatty liver cell model was induced by combination of glucose and OPA to preliminary evaluate the impact of fructose on fatty liver cells.The present study was the first to provide the insight into the action by which combined glucose,fructose and fatty acids cause metabolic disorders.(1)The data showed that fructose could synergistically increase the levels of ROS,extracellular TG,MDA,fructosamine and fatty acids.Additionally,the level of extracellular TG was more elevated when OPA was added,and IR level,intracellular TG and TC were synergistically increased by fructose and glucose.(2)In the cell model,PA worked synergistically with monosaccharides and mainly accounted for the induction of IR,ROS and extracellular TG release,while OA mainly associated with intracellular TG formation.(3)When the concentrations of glucose and fructose used in the model were chosen as 10 mM and 15 mM,respectively,the cell cycle for model cells showed no significant difference from normally cultured cells.(4)The levels of metabolites and cytokines did not changed by fructose.The results indicated that fructose induced ROS might result from the increase in PA or palmitoleic acid level.(5)The alterations of apolipoprotein B or MTTP gene expression were not noted.Only C/EBP? mRNA was significantly upregulated by fructose,and HMGCR mRNA was increased by fructose when OPA was applied in the meantime.Furthermore,metformin,lovastatin,aminoguanidine,oryzanol and polyphenols were evaluated in the established cell model.Fructose-induced metabolic disorder was alleviated by oryzanol and most polyphenols,such as flavones,flavonols,flavanones,isoflavones and phenolic acids.Relative gene expression of cells treated with rutin,kaempferol,genistein,protocatechuic acid,oryzanol and malvidin-3-glucoside were investigated,and beyond which the most active compounds were malvidin-3-glucoside,rutin and oryzanol.The present study also discovered that fructose could affect fatty liver cells.Interestingly,the medium contained fructose could enhance the intracellular TG on fatty liver cell models and the degree for steatohepatitis cells was higher than that of the steatosis cells.In addition,elevated concentrations of fructose would significantly increase the ROS of fatty liver cell models. |
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