Effect of ChREBP on carbohydrate feeding regulation in mandarin fish Siniperca chuatsi

Carnivorous fish have poor tolerance to carbohydrate in feed and low utilization rate of carbohydrate.Therefore,the balance of carbohydrate and lipids in the nutrient metabolism of carnivorous fish,the ef fective conversion and utilization of carbohydrate and lipids,and the feedback regulation of feeding are the key links for the e fficient utilization of carnivorous fish feed.Carbohydrate response element binding protein(ChREBP)is a new transcription factor found in recent years in the glucose signaling pathway,and can also participate in feeding regulation.We performed in-vivo and in-vitro experiments to reveal the role of ChREBP in the glucose metabolism and feeding in mandarin fish.The mRNA expression of ChREBP and appetite regulatory factors were measured after intraperitoneal injection of glucose in mandarin fish Siniperca chuatsi and cotransfection with glucose and glucose+siRNA in the hypothalamic cells in mandarin fish.The results reveal that at hour 2 and 4 post intraperitoneal injection with 1 mg/g BW glucose,the blood glucose level of the mandarin fish increased significantly,but food intake decreased significantly,and it also displayed a significantly increased ChREBP mRNA expression levels in liver.At hour 4 post injection,hypothalamic ChREBP mRNA level was significantly increased,whereas the mRNA expression levels of appetite genes neuropeptide Y(npy)and agouti-related peptide(AgRP)were decreased significantly.When the glucose concentration was 40 mmol/L,the expression level of ChREBP mRNA in mandarin fish hypothalamic cells was significantly up-regulated,but the expression level of appetite gene npy mRNA was down-regulated.When siRNA and glucose were co-transfected into mandarin fish brain cells,the expression level of chrebp mRNA was significantly decreased,and the appetite gene npy mRNA was significantly increased.The results indicated that glucose regulated food intake through the modulation of appetite gene npy by ChREBP.

AMPK-ChREBP axis mediates de novo milk fatty acid synthesis promoted by glucose in the mammary gland of lactating goats

To investigate the role of glucose in regulating milk fatty acid synthesis,6 lactating Guanzhong dairy goats were infused with 0,60,or 100 g/d glucose via the external pubic artery in a 33 repeated Latin square experiment.A concomitant in vitro experiment was conducted to investigate possible mechanisms whereby glucose regulates milk fatty acid synthesis.RNA sequencing was used for cellular transcriptome analysis.Drugs,MK-2206,rapamycin,and dorsomorphin were used to block cellular mammalian AMP-activated protein kinase(AMPK),AKT serine/threonine kinase 1,and mechanistic target of rapamycin kinase signaling pathways,respectively.Carbohydrate response element binding protein(ChREBP)was knockdown and overexpressed to investigate its role in regulating milk fatty acid synthesis in mammary epithelial cells.Glucose infusion linearly elevated the concentration of C8:0(P=0.039)and C10:0(P=0.041)in milk fat while it linearly decreased(P=0.049)that of C16:0.This result was in agreement with the upregulation of genes related to de novo synthesis of fatty acids and lipid droplet formation,including adipose differentiation-related protein,butyrophilin subfamily 1 member A1,fatty acid synthase(FASN)and ChREBP.Their expression increased(P<0.05)linearly in the lactating goat mammary gland.In vitro,glucose linearly stimulated the expression of genes related to de novo synthesis of fatty acids and cellular triacylglycerol in cultured mammary epithelial cells.RNA sequencing and inhibition studies revealed that glucose induced transcriptomic changes increasing lipogenic pathways,with AMPK responding to glucose by controlling ChREBP and FASN.Knockdown and overexpression of ChREBP highlighted its essential role in lipogenesis.The knockdown and overexpression of ChREBP protein also revealed an essential role in regulating the de novo synthesis of fatty acids.Collectively,our data highlight that glucose supplementation promotes de novo fatty acid synthesis via the AMPK-ChREBP axis,hence increasing milk fat yield in the goat mammary gland.Results from the current study provide possible strategies to manipulate the fatty acid composition as well as improve ruminant milk quality.