Effect And Mechanism Of Gestational Obesity On Glucose And Lipid Metabolism In Offspring

Part IObjective With the improvement of people’s material living standards,the incidence of obesity has been increasing year by year.At the same time,the incidence of gestational obesity has also increased significantly.Gestational obesity not only harms the health of the pregnant mother,leading to short-term complications such as gestational diabetes and pre-eclampsia,but also has long-term effects on the obesity susceptibility and insulin sensitivity of the offspring,increasing the risk of metabolic diseases in adulthood.Gestational obesity can be divided into pre-pregnancy obesity and excessive gestational weight gain(EGWG)according to the period of weight gain.Studies indicate that EGWG is an independent risk factor for adverse health effects in offspring.At present,there are few studies on the effect of EGWG on hepatic lipid metabolism of offspring,and there is also a lack of general animal model of EGWG.In order to investigate the effect of EGWG on offspring metabolism,a rat model of EGWG was constructed and the effect of EGWG on fetal hepatic lipid metabolism and its possible mechanism was explored.Methods Healthy Sprague-Dawley(SD)rats(16 females and 8 males)were caged together with 2 females and 1 male and tested for pregnancy after one week of adaptive feeding and the sperm observed by microscope was considered to be pregnant for 0.5 day.Pregnant rats were randomly divided into normal diet(ND)group and high-fat diet(HFD)group.ND group was given normal diet and HFD group was given high-fat diet.The body weight during pregnancy of the pregnant rats was recorded.Cesarean section was performed at day 21.5 of gestation and the birth weight of the fetal rats was recorded.Hematoxylin-eosin(HE)staining and oil red O staining were used to observe hepatic lipid deposition of the pregnant rats and the fetal rats.Glycerol phosphate oxidase-peroxidase(GPO-PAP)method was used to detect triglyceride(TG)and total cholesterol(TC)levels in livers and serum of the pregnant rats and the fetal rats.Real-time polymerase chain reaction(RT-PCR)and Western blot were used to measure the m RNA expression levels and protein expression levels of key genes in hepatic lipid metabolism FASN,SREBP1 c,SREBP2 and CPT1 a of the fetal rats.Results There was no difference in pre-pregnancy body weight between the HFD group and the ND group,and the difference in the weight and the weight gain during pregnancy were gradually obvious between the two groups.At day 21.5 of gestation,the weight of the pregnant rats,the weight gain during pregnancy of the pregnant rats and the birth weight of the fetal rats of the HFD group [(467.75±22.05)g,(181.50±9.68)g,(5.51±0.17)g] were significantly higher than those of the ND group [(430.88±18.80)g,(148.50±10.86)g,(4.85±0.35)g](P<0.05).Both HE staining and oil red O staining presented increased hepatic lipid deposition in the pregnant rats and the fetal rats of the HFD group.The hepatic levels of TG and TC in the pregnant rats of the HFD group were3.09 times and 2.08 times higher than those of the ND group,and the serum levels of TG and TC were 3.44 times and 1.23 times higher than those of the ND group,respectively.The hepatic levels of TG and TC in the fetal rats of the HFD group were 2.98 times and1.52 times higher than those of the ND group and the serum levels of TG and TC were1.52 times and 1.44 times higher than those of the ND group,respectively(P<0.05).RT-PCR and Western blot showed that the m RNA and protein expression levels of key genes in hepatic lipid metabolism FASN,SREBP1 c,SREBP2 in the fetal rats of the HFD group were significantly higher than those of the ND group,the m RNA and protein expression levels of CPT1 a in the fetal rats of the HFD group were significantly lower than those of the ND group(P<0.05).Conclusions HFD during pregnancy for 21 days can successfully construct the animal model of EGWG.EGWG can lead to hepatic lipid deposition in the fetal rats,the mechanism of which may be related to the expression changes of key genes in hepatic lipid metabolism FASN,SREBP1 c,SREBP2 and CPT1 a in the fetal rats.Part IIObjective In metabolic studies,different timing of intervention can lead to different phenotypes in offspring.Studies have shown that maternal obesity has independent programming effects on offspring obesity and metabolism at different developmental Windows such as early pregnancy and late pregnancy.Through the intervention of high-fat diet in maternal rats at different periods,the levels of glucose and lipid metabolism of offspring rats in different periods were detected,and the specific effects of high-fat diet in different periods on offspring were clarified,in order to provide a basis for the study of maternal obesity,which has important practical significance.In order to clarify the differences in the effects of high-fat diet intervention before and during pregnancy on glucose and lipid metabolism of offspring,this study established the animal model of gestational obesity by giving SD rats high-fat diet before pregnancy and/or during pregnancy and lactation,and the differences in glucose and lipid metabolism of offspring were observed and the possible mechanism was explored.Methods Healthy SD rats(36 females and 18 males)were randomly divided into four groups according to different diets before pregnancy and during pregnancy and lactation: CC group(normal diet before pregnancy and during pregnancy and lactation),HC group(high-fat diet before pregnancy and normal diet during pregnancy and lactation),CH group(normal diet before pregnancy and high-fat diet during pregnancy and lactation),HH group(high-fat diet before pregnancy and during pregnancy and lactation),and all offspring were given normal diet after weaning.The body weight of maternal rats before pregnancy and during pregnancy and the body weight of offspring rats were recorded.Male offspring rats from each group at birth(0w),juvenile stage(3w)and adult stage(12w)were selected,fasting glucose(GLU)levels were detected by glucometer,fasting insulin(INS)levels were detected by enzyme-linked immunosorbent assay(ELISA)kit,and the index of homeostasis model assessment of insulin resistance(HOMA-IR)was calculated.Glucose tolerance and insulin sensitivity were detected by glucose tolerance test(GTT)and insulin tolerance test(ITT),and the area under the curve(AUC)of GTT and ITT were calculated.Hepatic TG and TC levels were detected by GPO-PAP method,hepatic lipid deposition were observed by HE staining and oil red O staining.The m RNA expression levels and protein expression levels of hepatic key genes in glucose metabolism pathway IR,IRS,AKT,lipid metabolism FASN,SCD1,SREBP1 c and Wnt signaling pathway LRP6,β-catenin,TCF4 were detected by RT-PCR and Western blot.Results The pre-pregnancy weight of maternal rats in HC group [(320.00±12.39)g] and HH group [(315.56±8.60)g] which were given a high-fat diet before pregnancy was significantly higher than those in CC group [(287.11±8.65)g] and CH group [(281.56±7.76)g] which were given a normal diet before pregnancy,and the difference was statistically significant(P<0.05).The weight gain during pregnancy of maternal rats in HC group [(176.00±11.69)g],CH group [(178.22±10.41)g] and HH group [(188.67±6.06)g] which were given a high-fat diet before and/or during pregnancy was significantly higher than those in CC group [(160.33±8.86)g] which were given a normal diet before and during pregnancy(P<0.05).At birth,the birth weight of neonatal rats in HC group,CH group and HH group were significantly higher than that in CC group(P<0.05),and the birth weight of neonatal rats in HH group was the largest.The hepatic TG and TC levels and the m RNA and protein expression levels of FASN and SREBP1 c and the protein expression levels of SCD1 of neonatal rats in CH group and HH group were significantly higher than those in CC group(P<0.05),and hepatic lipid deposition was aggravated.The hepatic TG levels and the protein expression levels of FASN and SCD1 in HH group were significantly higher than those in the other three groups(P<0.05).There was no significant difference in the levels of fasting GLU,INS,HOMA-IR indexes and the m RNA and protein expression levels of IR,IRS,AKT,LRP6,β-catenin,TCF4 in the four groups(P>0.05).At three weeks of age,the body weight,hepatic TG and TC levels,and the m RNA and protein expression levels of FASN,SCD1,SREBP1 c of offspring rats in HC group,CH group,and HH group were significantly higher than those in CC group(P<0.05),and hepatic lipid deposition was aggravated,and the HH group of the three groups increased the most.The fasting GLU and INS levels,HOMA-IR index,GTT-AUC,ITT-AUC and the protein expression level of p-IRS in HH group were significantly higher than those in CC group.The m RNA and protein expression levels of IR,IRS,LRP6,β-catenin,TCF4 in HH group were significantly lower than those in CC group(P<0.05).At twelve weeks of age,the body weight,fasting GLU and INS levels and the HOMA-IR index,GTT-AUC,ITT-AUC,hepatic TG and TC levels,the protein expression level of p-IRS,and the m RNA and protein expression levels of FASN,SCD1,SREBP1 c of offspring rats in HC group,CH group,and HH group were significantly higher than those in CC group(P<0.05),and hepatic lipid deposition was aggravated.The m RNA expression levels of IR,IRS,AKT,LRP6,β-catenin,TCF4 and the protein expression levels of IR,IRS,p-AKT,LRP6,β-catenin,TCF4 were significantly lower in HC group,CH group and HH group than those in CC group,and the protein expression levels of p-IRS in HC group,CH group and HH group was significantly higher than those in CC group(P<0.05).Conclusions High-fat diet before pregnancy and/or during pregnancy and lactation can successfully construct animal models of gestational obesity.High-fat diet before pregnancy and during pregnancy and lactation has the greatest effect on glucose and lipid metabolism in offspring.Compared with rats fed high-fat diet before pregnancy,high-fat diet during pregnancy had a more significant effect on lipid metabolism of neonatal offspring,and there was no significant difference in the effects of the two intervention methods on juvenile and adult offspring.Maternal high-fat diet before pregnancy has adverse effect on offspring even if the maternal rats change to normal diet during pregnancy and lactation.The offspring born to maternal rats of high-fat diet showed abnormal lipid metabolism first and then abnormal glucose metabolism.The effect of high-fat diet on glucose metabolism in offspring was considered to be related to the expression changes of genes in glucose metabolism and Wnt signaling pathway,and the effect of high-fat diet on lipid metabolism in offspring was considered to be related to the expression changes of key genes in lipid metabolism.