An Investigation Of The Association Between Glycolipids Metabolism And Vitamin D Status In Children/Adolescents And In Animal Modeling

Part Ⅰ. Investigation of glycolipid profiles in overweight/obese childrenand adolescents and the best cut-off value of anthropometric indice for predicting dyslipidemia Background:The prevalence of adolescents’ obesity and overweight has dramaticallyelevated in China. Obese children were more likely to be insulin resistant and dyslipidemia, which are risk factors of cardiovascular diseases. However there was no cut-off point of anthropometric values to predict the risk factors in Chinese adolescents. The present study was to investigate glycolipid metabolism status of adolescents in Shanghai and to explore the correlations between body mass index standard deviation score (BMI-SDS) and metabolic indices, and to determine the best cut-off value of BMI-SDS for predicting dyslipidemia. Methods:Fifteen schools in Shanghai’s two districts were chosen by cluster sampling andprimary screening was done in children aged 9-15 years old. After screening of bodyweight and height, overweight and obese adolescents and age-matched children with normal body weight were randomly recruited. Anthropometric measurements, biochemical measurements of glycolipid profiles were done. SPSS 19.0 was used to analyze the data. Receiver operating characteristic (ROC) curves were made and the best cut-off values of BMI-SDS to predict dyslipidemia were determined while the Youden indice were maximum. Results:Five hundred and thirty-eight adolescents were enrolled in this research, amongwhich 283 have normal bodyweight,115 were overweight and 140 were obese. No significant differences of the ages among 3 groups were found. There were significant differences of WC-SDS (p<0.001), triacylglycerol (p<0.05), high and low density lipoprotein cholesterol(p<0.01), fasting insulin(p<0.01) and C-peptide (p<0.001) among 3 groups. Significant difference of fasting glucose was only found between normal weight and overweight group. Significant difference of total cholesterol was found between obese and normal weight group. There was no significant difference of glycated hemoglobin among 3 groups. The same tendency was found in boys but not in girls. Only HDL-C reduced and TG increased while BMI elevated in girls. The best cut-off value of BMI-SDS was 1.22 to predict dyslipidemia in boys. The BMI cut-off was 21.67kg/m2 in boys. Conclusions:Overweight and obese youths had reduced insulin sensitivity and dyslipidemia. When BMI-SDS elevated up to 1.22 and BMI was higher than 21.67kg/m2 in boys, the risk of dyslipidemia increased.Part Ⅱ. Survey of the vitamin D status in children/adolescents and the associated factorsBackground:The Vitamin D endocrine system plays several important roles in human’s body, such as bone health, keratinocyte differentiation and immune system. Accumulating researches suggest vitamin D deficiency is high risk factors of obesity, diabetes and cardiovascular disease. But hypovitaminosis D is prevalent worldwide and the relationship between vitamin D deficiency and obesity or metabolic indice is still controversial. Therefore this study aims to evaluate the vitamin D status in Chinese adolescents, to find out the risk factors of hypovitaminosis D and the association between obesity and vitamin D.Methods:The research population in this part was derived from the population of the first part who completed the questionnaire of physical activities/dietary habits and had enough frozen serum specimen to have 25(OH)D tested. The questionnaire was filled out collaborated by the children and their guardians.25(OH)D levels were performed by electrochemical luminescence using Roche E411 analyzer. The data obtained was put into computer by Epidata 3.1 and was analyzed by SPSS 19.0. The prevalence of hypovitaminosis D was determined. The variations of 25(OH)D level caused by gender, seasons and BMI were analyzed. The risk factors of hypovitaminosis D were determined by multi-variation logistic regression. The association of vitamin D3 and the metabolic indice was analyzed by gender.Results:Four hundred and forty-one adolescents were enrolled in this part research, among which 284 were male and 157 were femal. Three hundred and thirty-eight participants had physical exam in winter and 103 had exammed in summer. The mean serum 25(OH)D level of all the objects was 21.93±8.11ng/ml. The mean 25(OH)D concentration in boys was 22.41±8.11ng/ml, significantly higher than that of gilrs’(18.97ng/ml (P25, P75:15.49ng/ml, 25,21 ng/ml), p<0.05). The prevalence of hypovitaminosis D was 42.40%, with 2.95% objects were severe deficiency. The prevalence of vitamin D deficiency in girls was 55.41%, significantly higher than the prevalence in boys(39.79%, p<0.01). The mean level of 25(OH)D in winter was 19.59ng/ml, significantly lower than that in summer(25.58±7.83ng/ml, p<0.001). Significant differences were found in gender(p=0.005), age(p<0.001), BMI-SDS(p=0.003), WC-SDS(p=0.005), seasons(p<0.001)and non-alcoholic fatty liver disease (NAFLD) ratios (p=0.018) among different groups of vitamin D sufficiency, insufficiency and deficiency. The logistic regression suggested that female (OR=2.451, p=0.001), older ages (OR= 1.285, p=0.011) and winter (OR=1.957, p=0.038) were risk factors of vitamin D deficiency. In male adolescents, the level of 25(OH)D was negatively associated with BMI (r=-0.304 p<0.001), waist circumstance (r=-0.320, p<0.001) and the existence of NAFLD (r=-0.185, p=0.002). HDL was positively associated with 25(OH)D concentrations (r=0.293, p<0.001). No relationship between 25(OH)D and metabolic indice in girls was found.Conclusions:The vitamin D3 status in adolescents was very worrying, with about half were vitamin D3 deficency and more severe in females. The risk factors of hypovitaminosis D were female, older ages and winter. The level of 25(OH)D was adversely associated with BMI, WC, NAFLD and positively associated with HDL in boys.Part Ⅲ. Glycolipid indice of obese SD rats and the effect of vitamin D3 supplementionBackground:The recognition of vitamin D’s importance is expanding from traditional bone health to endocrine system and glycolipids metabolism. Research findings suggest that vitamin D may improve insulin secretion and sensitivity and also can regulate lipid profiles. Dietary induced obese SpragueDawley (SD) rats were used in this research to analyze differences of metabolic indice and 25(OH)D level among obese rats, normal weight rats and diet-induced obesity resistant (DIO-R) rats. Vitamin D3 oil at different doses were administrated through oral gavage, therefore to determine the impact of different dosage on rats’ body weight, fat weight, lipids, fasting glucose and insulin.Methods:Eighty 4-week-old male SD rats were fed with high-fat, high-calorie foods for 8 weeks, while 10 other SD rats were fed with standard chow as the control group. DIO rats were judged by weight higher than 120% of the control group’s average weight and DIO-R rats were those did not met the criteria. Ten rats in each group underwent chloral hydrate anesthesia as cross-sectional survey. Blood was taken from ventricle and centrifuged to obtain serum. TC, TG, HDL, LDL and glucose were tested by automatic biochemical analyzer while insulin was tested by ELISA kit. Fat surrounding the kidneys and the testicles were collected and weighed up. Forty obese rats were allocated to 4 groups randomly and underwent 250IU/kg.d,500IU/kg.d, 1000IU/kg.d vitamin D3 and olive oil supplementation. Four weeks later, blood samples and. SPSS 19.0 was utilized to compare the body weight, Lee’s index, fat weight and glycolipid indice among DIO, DIO-R and the control groups. Paired t test was use to estimate the interested indice changes before and after vitamin D3 supplementation.Results:After 8-week high-fat feeding, the DIO group has signifacanly higher mean body weight(456.50±12.51g, p<0.001), fat weight(17.45±3.36g, p<0.001), Lee’s index(3.15±0.05, p<0.01) than the control group (374.50±30.42g,9.21±1.77g,3.02±0.95, respectively). The mean body weight (385.70±29.30g, p<0.001), fat weight (10.68±2.56g, p<0.001) and Lee’s index (3.06±0.09, p<0.05) of DIO-R group was significantly lower than those of DIO group, but comparable to the control group. The mean level of 25(OH)D of DIO and DIO-R group was 24.24±1.29ng/ml,25.29±2.03ng/ml respectively, significantly higher than the control group(12.53±1.29ng/ml, p<0.001). No statistically differences were found in TG, insulin and HDL among three groups. The mean TC level (1.44±0.14mmol/L, p<0.05), Glu (9.40±1.84 mmol/L, p<0.001) and LDL (0.74±0.04mmol/L, p<0.05) of DIO group were significantly higher than those of the control group (1.25±0.18mmol/L,4.90±0.7mmol/L,0.73±0.03 mmol/L, respectively). The indice of DIO-R group were similar to those of DIO group, but higher than the control group. The results of linear regression suggested that the glucose concentration was positively associated with body weight(r=0.551, p=0.002) and Lee’s index (r=0.392, p=0.032). In the supplementation trial, the mean body weight of 250IU/kg-d and 500IU/kg-d intervention group decreased after one week. Two weeks later, the mean weight of 500IU/kg-d intervention group was 458.10±2.16g, significantly lower than the 1000 IU/kg-d intervention group. But when the intervention ended after four weeks, there were no statistically differences of body weight among the four groups. The baselines and end-points of TC, TG, LDL, Glu,25(OH)D among the four groups were of no statistically differences.The mean HDL at the end point was significantly higher in 500IU/kg-d, 1000IU/kg-d group and the control group than in 250IU/kg-d group.The mean insulin after 500IU/kg-d vitamin D3 intervention was significantly lower than that of 250IU/kg·d intervention and had no statistically differences with the 1000IU/kg-d group and the control group. The mean 25(OH)D was significantly elevated (p<0.001) after vitamin D3 supplementation while significantly decreased (p<0.001) after olive oil intake. The results of paired t test showed that TG (p<0.05) and HDL(p<0.01) decreased and glucose(p<0.01)/ insulin(p<0.05) increased in 250IU/kg.d group; TC/TG decreased (p<0.05) and glucose elevated (p<0.01) in 500IU/kg.d group; only glucose (p<0.01) rised in 1000IU/kg.d group; TC/TG/HDL decreased (p<0.05) and glucose increased (p<0.01) in olive oil supplementation.Conclusions:High fat diet could induce male SD rats into obesity, accompanied by increased body fat, dyslipidemia and elevated glucose. When fed with high fat diet, even normal weight rats would have dyslipidemia. With enough vitamin D3 intake, obese rats could achieve optimical serum 25(OH)D level. Vitamin D3 supplementation could reverse the vitamin D deficiency and 500IU/kg.d suggested a favorable regulation of lipid profiles.Part Ⅳ. Expression of liver vitamin dhydroxylase CYP27A1 in obese SD ratsBackground:Accumulating observational studies have found the association between vitamin D deficiency and obesity. One assumption of this phenomenon is the reduction of liver hydroxylase of vitamin D. Therefore, this research investigated the expression of CYP27A1 in obese rats.Methods:Ten DIO rats were obtained by 8-week high-fat feeding. Liver specimen were get from the DIO rats and the control rats (n=10). Pathological slides were hematoxylin-eosin stained to evaluate the steatosis scoring. Immunohistochemical staining and Western blot was applied to detect the expression of CYP27A1.Results:Five of the control group’s rats were scored 1 point and another 5 rats were scored 2 points as to the hepatocyte steatosis. Nine of the DIO rats got 3 points and 1 got 1 points. It was of significant difference between the ratios (p<0.001). The immunohistochemical staining showed that CYP27A1 expressed in the centrallobular and was decreased in DIO rats. The relative expression of CYP27A1 of the control group detected by western blot was 2.08±0.37, significantly higher than that of DIO rats (1.27±0.28, p<0.001). The CYP27A1 concentration was negatively associated with the scoring of hepatocyte steatosis (r=-0.548, p=0.023).Conclusions:Obeses rats had severe hepatocyte steatosis and decreased expression of vitamin D hydroxylase CYP27A1. The expression of CYP27A1 was adversely associated with hepatocyte steatosis.