The Effect Of Preconceptional High Protein Diet On The Homeostasis Of Offspring’s Hepatic Glucose And Lipids And The Mechanism Study In Mouse

Systemic glucose and lipids homeostasis are important for health.As the major organ of glucose and lipid metabolism,liver plays critical role in preserving systemic metabolism healthy.Changes in dietary nutrient levels is one of the factors which affect glucose and lipid metabolism.It has been reported that unbalanced maternal dietary nutrient alters the epigenetic inheritance of offspring,which subsequently results in the changes of offspring’s glucose and lipid homeostasis.However,most of these studies were focused on the nutrient changes during maternal gestation and lactation periods,few studies were focused on that of preconception period.The daily protein intake level is increasing in human.Therefore,it is important to study the effect of preconceptional high protein diet on offspring’s hepatic glucose and lipid metabolism,and the epigenetic mechanisms.In the current study,C57BL/6J female mice were fed with high-protein diet or normal level protein diet during their before mating.The aim is to investigate the effect of preconceptional high protein diet on the offspring’s hepatic glucose and lipids metabolism and the DNA methylation level of offspring’s liver.The experimental study is as follows:Trial 1 Effect of preconceptional high protein on the hepatic glucose and lipid metabolism of offspringTwenty 28-day-old C57BL/6J female mice with similar body weight were divided into two groups,one group were fed with high protein diet(HP,50% Casein in diet)and the other group were fed with normal protein diet(NP,20% Casein in diet)for 12 weeks.Mice were mated with normal male mouse at the age of 16 weeks,and then were fed with normal protein diet.The male offspring were weaned at 21-day-old,and were then randomly divided into two groups for each treatment respectively.One group were fed with normal chow diet and the other with high-fat diet(HFD,60% energy from fat).The four groups(Chow-NP,Chow-HP,HFD-NP and HFD-HP)were measured with body weight biweekly.The offspring were performed with insulin tolerance test at age of 20 weeks,and with glucose tolerance test at age of 22 weeks.At age of 24 weeks,mice were performed with insulin signaling study(n=3-4),and harvested for blood and liver sample collection.The results are showed as follows:(1)The mice fed with the high-protein diet(HP)took in as much as 2.2 folds crude protein than the mice fed with normal-protein diet(NP)did during the study period,though the food intake was slightly decreased in the HP group.The overall body weight was also slightly lower for the HP group as compared to the NP group.(2)As compared to the offspring of NP group,the offspring of HP group had higher overall body weight during the age of 4-16 weeks,both under normal chow diet fed condition(Chow)and under high fat diet fed condition(HFD).The high fat diet in the offspring significantly increased their body as compared to the chow diet.(3)At the age of 24 weeks,the offspring of HP group had similar tissue weight index of liver,peri-renal fat,subcutaneous fat and brown fat,as compared to the offspring of NP group.The high fat diet in the offspring significantly increased their tissue weight index as compared to the chow diet.However,no interact effect was observed between the maternal high-protein diet and the offspring high fat diet.(4)As compared with the NP group,maternal high protein diet had no effect on offspring’s blood glucose level,but serum insulin level trended to be higher in the HFDHP group than that in the HFD-NP group.Furthermore,the high fat diet in the offspring significantly increased their serum blood glucose level and serum insulin concentration as compared to the chow diet.(5)As compared to the Chow-NP offspring,the Chow-HP offspring showed impaired insulin tolerance test and unchanged glucose tolerance test.Both ITT and GTT were unchanged in HFD-HP group as compared to the HFD-NP group.(6)The result of insulin signaling test showed that,as compared with the control treatment,maternal high protein diet trended to increase the hepatic insulin sensitivity in the HFD fed offspring(P=0.057).However,for their offspring fed on chow diet,there is no significant effect.(7)Compared with the control treatment,maternal high protein diet significantly reduced the liver TAG content of their offspring(P<0.001),both in the Chow diet and HFD fed offspring.The high fat diet in the offspring significantly increased their liver TAG content as compared to the chow diet(P<0.001).Besides,there was an interaction effect between the maternal high-protein diet and the offspring high fat diet(P< 0.001).(8)Compared with the control treatment,maternal high protein diet significantly increased serum levels of TAG,TC,HDL and LDL in the HFD fed offspring(P<0.05).However,only serum TAG level was increased by maternal high protein diet in the chow diet fed offspring(P<0.05).(9)Compared with control diet,maternal high protein diet significantly decreased the expression of fat acid synthesis related genes,such as Acc1,Srebp1,Fabp1 and lipoprotein metabolic genes,such as Apoe,Cyp8b1 and Lipa in the liver of HFD fed offspring.Trial 2 Effect of maternal high protein on the DNA methylation level of offspring’s liverLiver samples of the HFD fed offspring from the trial 1 were collected for analysis of the DNA methylation level.The results showed as follows:(1)In the gene of mice liver,the most m C located in the CG,which accounted for more than 97% of total m C.Besides,the highest methylation region is between the first intron and the last exon,while the lowest methylation region is the first exon.(2)Compared with the control treatment,maternal high protein diet increased the number of DNA methylation sites in their offspring.(3)The enriched pathways which related to glucose and lipids metabolism included type II diabetes,AMPK signaling pathway,nonalcoholic fatty liver,insulin resistance,PI3K/AKT and PPAR signaling.The genes related lipids and lipoprotein metabolism included Srebp1,Apoa5,Pparα,Ampkγ2,Irs1 and Cyp8b1.In summary,maternal high protein diet might alter the homeostasis of offspring’s lipids and lipoprotein metabolism,by changing the DNA methylation level of the related genes in the offspring’s liver.This study provides a reference for the rational protein intake of preconceptional female.