Study On The Changes And Significance Of Mitochondrial Fatty Acid Synthesis Pathway ? In Different Inflammatory

Background: Inflammation is closely related to the pathogenesis of various diseases,such as cardiovascular disease,type 1 diabetes,type 2 diabetes and their complications.Diabetes is a chronic inflammatory disease characterized by hyperglycemia.Both high blood pressure and increased expression of pro-inflammatory factors can induce an inflammatory response,and the development and progression of inflammation are closely related to mitochondria.Sepsis is a systemic inflammatory disease.Lipopolysaccharide(LPS)is widely used to induce inflammation and apoptosis in cardiomyocytes as a classical model of sepsis.Pro-inflammatory factors and apoptosis produced by cardiomyocytes are critical in the pathogenesis of myocardial dysfunction.Metformin(Met)has hypoglycemic and anti-inflammatory effects,exhibits antioxidant properties and reduces lipid oxidation in a variety of tissues.In addition,metformin has been shown to act directly on mitochondria and protect cells.It is known that mitochondria can regulate the occurrence of inflammation through ROS,autophagy,mitochondrial fission and fusion.Mitochondrial fatty acid synthesis ?(mtFAS ?)is an important component of mitochondrial function,and its end-product is lipoic acid(LA),which is the main raw material for protein lipoylation.Lipoylation is required for the activation of a variety of enzymes that catalyze the catabolism of glucose,amino acids and so on.Therefore,maintenance of mtFAS ? function is necessary and sufficient for the maintenance mitochondrial function.With regard to the inflammatory state,the changes of mtFAS ? and its significance in the treatment of inflammation have not been studied.Purposes: To explore the changes of mtFAS ? in various inflammatory states and its significance in the treatment of cardiomyocyte inflammation with metformin.Contents and methods: This study mainly studied on the changes of mtFAS ? in tumor necrosis factor-alpha and high-glucose inflammatory states,as well as the changes of mtFAS ? in lipopolysaccharide-induced inflammatory states and the changes and significance of mtFAS ? after metformin treatment.To study on the changes of MCAT,OXSM,LIAS and the lipoylation level of PDCE2 in TNF-? and high-glucose-induced inflammation.(1)Inducing inflammatory state with 10 ng/ml of tumor necrosis factor-?(TNF-?),treating 3T3-L1 preadipocytes according to the corresponding time gradient,then examining the mRNA levels of Malonyl-CoenzymeA/Acyl-carrier protein transferase(MCAT),3-Oxoacyl-Acyl-carrier protein synthase(OXSM),Lipoic acid synthase(LIAS)in mtFAS ? and the protein level of OXSM;in DIO obese mice,examining the mRNA level of OXSM in inflammatory conditions of adipose tissue with high expression of TNF-?.(2)Inducing inflammatory state with 35 mM glucose(D-glucose),treating the 3T3-L1 preadipocytes according to the corresponding time gradient,examining the mRNA levels of MCAT,OXSM,LIAS and the protein level of OXSM under high glucose inflammatory state.Examing the mRNA levels of MCAT,OXSM,and LIAS in the kidney tissues of STZ diabetic model mice and db/db diabetic model mice and the protein levels of MCAT,OXSM,LIAS and the lipoylation level of PDCE2 in kidney tissues of STZ diabetic mice.To study on the changes of MCAT,OXSM,LIAS and the lipoylation level of PDCE2 in LPS-induced cardiomyocyte inflammation and their significance in the treatment with metformin(Met).H9C2 cardiomyocytes were divided into control group,LPS-induced inflammation group and three different concentrations of Met and LPS combined treatment group.The cell viability was determined by MTT assay,the levels of apoptosis and reactive oxygen species were detected by flow cytometry,the protein level of Caspase3,BCL2,BAX,phosphorylated adenylate-activated protein kinase(PAMPK),adenylate-activated protein kinase(t-AMPK),Beclin1,Parkin,MCAT,OXSM,LIAS and the lipoylation level of PDCE2 were detected by Western blot.Results:(1)In TNF-?-induced inflammatory state,the mRNA levels of MCAT and LIAS showed transient decrease,while the mRNA level of OXSM showed persistent decrease,and the protein level of OXSM also showed persistent decrease and reached the lowest value at 24 h(P < 0.001);in the adipose tissue of DIO mice with high expression of TNF-?,the mRNA level of OXSM was significantly decreased(P < 0.001).(2)In the high-glucose-induced inflammatory state,the expression of MCAT,OXSM,and LIAS decreased transiently at the mRNA level and eventually returned to normal levels.At the protein level,OXSM expression decreased.It reached the lowest level at 4h(P < 0.001),and recovered at 24 h,but still below normal.In the kidney tissues of STZ-induced diabetic mice,the mRNA levels of MCAT and LIAS decreased significantly,but the mRNA levels of OXSM did not change.In the kidney tissue of the db/db diabetic mice,the mRNA levels of MCAT,OXSM and LIAS were significantly decreased.In the kidney tissue of STZ diabetic mice,the protein level of OXSM and the lipoylation level of PDCE2 were significantly decreased(P < 0.001).(3)In the LPS-induced inflammation state,levels of poptosis and ROS were significantly elevated in the LPS-induced inflammation group(P<0.01),proteins of autophagy-related proteins,OXSM,LIAS and the lipoylation of PDCE2 were significantly decreased(P < 0.01).Levels of apoptosis and reactive oxygen species were significantly decreased,and protein expressions of autophagy-related proteins,P-AMPK,OXSM,LIAS and the lipoylation of PDCE2 were significantly increased in the Mettreated group(P < 0.01).Conclusion: Inflammation induced by TNF-?,high-glucose and LPS can inhibite the expression of the enzyme in mtFAS ? and the lipoylation level of PDCE2.Metformin can significantly improve the expression of enzymes in mtFAS ? and reduce damage to cardiomyocytes and apoptosis caused by inflammation.