Glucose and Adenosine-containing Molecule Stimulate Expression of Thioredoxin Interacting Protein(Txnip) mRNA through Carbohydrate Response Element Binding Protein(ChREBP) in Primary Cultured Porcine Adipocytes
Abstract：Thioredoxin interacting protein (Txnip) is a multifunctional protein involved in many cellular and physiological processes, and can regulate cellular redox state by inhibiting the activity of thioredoxin. To explore effects of glucose and adenosine-containing molecule on expression of Txnip mRNA and regulation pathway for transcription in porcine adipocytes, the preadipocytes isolated from subcutaneous adipose tissue obtained from 3~7 days old piglets(Sus scrofa) were cultured and induced to differentiate into mature adipocytes, in which carbohydrate response element binding protein gene (ChREBP) was silenced by transfection of ChREBP-RNAi expression plasmid. Non-transfected and transfected adipocytes were cultured in medium with 0, 5 or 15 mmol/L glucose with or without NAD+ for 24 h. Expression level of Txnip and ChREBP mRNA was determined by fluorescence Real-time PCR. The results showed that ChREBP mRNA expressions were up-regulated 4.57 and 6.69 times (P＜0.01) in porcine adipocytes cultured in medium with 5 mmol/L or 15 mmol/L glucose, respectively, but there was no difference (P＞0.05) in the cells cultured in same glucose concentrations of medium with or without NAD+, which indicated NAD+ alone or accompanied by glucose have no effect on the ChREBP mRNA expression. Expression of Txnip mRNA was increased by glucose stimulation (P＜0.01). Txnip mRNA expression was not elevated by NAD+ under glucose-free condition (P＞0.05), but increased significantly in the presence of glucose (P＜0.01). In ChREBP-silencing adipocytes, levels of Txnip mRNA expression had no difference (P＞0.05) and slightly increased by glucose (P＜0.05; ~1.34 times) compared with non-transfected adiocytes cultured in glucose-free medium. However, this stimulation of glucose on Txnip mRNA expression was reduced 82% and 91% in ChREBP-silencing adipocytes cultured in medium with 5 mmol/L and 15 mmol/L glucose than that in non-transfected cells cultured under the same condition, respectively. NAD+ did not further induce Txnip mRNA expression on the basis of glucose stimulation (P＞0.05), and stimulation of NAD+ on Txnip mRNA expression reduced 91% and 93% (P＜0.01) in ChREBP-silencing adipocyes than that in non-transfected cells cultured in medium with 5 mmol/L and 15 mmol/L glucose, respectively. These results suggested that NAD+ induces Txnip transcription expression dependent on glucose, and the stimulation of glucose and adenosine-containing molecule on Txnip mRNA expression were mediated by ChREBP. This study can provide useful data for research on the role of ChREBP in glucose and lipid homeostasis.
