The Role Of Txnip In The Differentiation Of Porcine Preadipocytes Regulated By Glucose And Vitamin D

The main factors affecting the flavor and carcass quality of meat include the fat deposition and distribution of adipose tissue in the body.Glucose is the main substrate for de nove lipogenesis,also promoted the lipogenesis by regulating the transcription factor carbohydrate response element binding protein?ChREBP?expression through its intermediate metabolites.Vitamin D is mainly stored in adipose tissue and may play an important role in adipogenesis.Thioredoxin interacting protein?Txnip?,a cellular reduction/oxidation balance regulated protein,the binding of Txnip inactivation thioredoxin?Trx?,and then regulate many fundamental cellular processes,including proliferation,differentiation,apoptosis and metabolism.In this thesis,porcine subcutaneous preadipocytes were isolated and cultured.And Txnip expression of the cells were silenced by Txnip-siRNA or over-expressed by LV5-Txnip.At the same time,glucose and vitamin D were added in the medium in order to study the role of Txnip in the differentiation of porcine preadipocytes regulated by them.These results provides theoretical basis for elucidating the mechanism of porcine adipogenesis process.The main results of the study are as follows:1.Construction of Txnip siRNA vector and overexpression vectorThe siRNA lentiviral expression plasmid targeting porcine Txnip gene was successfully constructed with the silencing rate of Txnip expression in porcine preadipocyte cells as 75%after siRNA transfeciton.The porcine Txnip overexpression lentiviral expression plasmid was constructed with the overexpression rate of Txnip in porcine preadipocytes significantly increased about 85 times after overexpression.2.Effect of vitamin D on porcine preadipocytesWith the concentration of 1×10^-99 mol/L and 1×10^-1010 mol/L,1,25?OH?₂VD₃promoted the proliferation of porcine preadipocyts with the differentiation abilities inhibited.With the concentration of 1×10^-66 mol/L,1,25?OH?₂VD₃ inhibited the proliferation of porcine preadipocytes,meanwhile,the differentiation and proliferation abilities of porcine preadipocytes were not significantly promoted or inhibited with the concentration of 1,25?OH?₂VD₃ at 1×10^-77 mol/L and 1×10^-88 mol/L.3.Effect of Txnip on differentiation of porcine preadipocytesThe silencing of Txnip expression promoted the differentiation abilities and PPAR?expression levels in porcine preadipocytes?P<0.05?,while the overexpression of Txnip inhibited the differentiation abilities and PPAR?expression levels?P<0.05?,indicating that the expression of Txnip could inhibited the differentiation abilities of porcine preadipocytes by inhibiting the expression of PPAR?.4.The roles of Txnip in the differentiation of porcine preadipocytes inhibited by1,25?OH?₂VD₃In this part,it was found that low concentration of 1,25?OH?₂VD₃(1×10^-10-1×10^-99 mol/L)promoted the proliferation of porcine preadipocytes and inhibited the cell differentiation abilities,while high concentration(1×10^-8-1×10^-7mol/L)inhibited the proliferation of porcine preadipocytes with promoted differentiation abilities.And low concentration of 1,25?OH?₂VD₃ inhibits the differentiation of porcine preadipocytes by up-regulating Txnip gene expression.5.The role of Txnip in the differentiation of porcine preadipocytes induced by glucoseIt was found that glucose at a concentration of 15 mmol/L promoted differentiation of porcine preadipocytes?P<0.05?and silencing of Txnip expression promoted differentiation of porcine preadipocyts?P<0.05?,which was dependent on the presence of glucose.Txnip overexpression had no effect on the differentiation of porcine preadipocytes.When treated with glucose,Txnip overexpression could attenuate the promotion of glucose on porcine preadipocyte differentiation.In conclusion,the promotion of glucose on the differentiation of porcine preadipocytes was inhibited by Txnip expression.In conclusion,low concentration of 1,25?OH?₂VD₃ promoted the proliferation porcine preadipocytes and inhibited the differentiation of the cells,and high concentration of 1,25?OH?₂VD₃ stimilated the differentiation and inhibited the proliferation of the porcine preadipocytes,which indicated that 1,25?OH?₂VD₃ had significantly bidirectional effects on proliferation and differentiation of porcine preadipocytes.Txnip inhibited the differentiation of porcine preadipocytes through reducing PPAR?expression,which depends on the presence of glucose.Low concentration of 1,25?OH?₂VD₃ might inhibit the differentiation of porcine preadipocytes by inducing the Txnip expression which upregulated the expression of PPAR?.Glucose promoted the differentiation of porcine preadipocytes,but Txnip attenuated this stimulation of glucose on adipocyte differentiation.