| BackgroundFetal growth restriction (FGR) includes intrauterine growth retardation(IUGR) and low birth weight(LBW). FGR is the main cause of perinatal illness and death. Metabolic syndrome (MS) includes several common chronic diseases which can cause death and disability, such as adiposity, diabetes, hypertension and so on. The epidemiological investigation shows that low birth weight is a high-risk factor of adult MS. The mechanism behind this phenomenon becomes a research focus. Because the research can't be performed in human being directly, suitable animal model should be established. The theories of fetal origins hypothesis, nutritional programming and metabolic programming provide theoretic foundations for the establishment of low birth weight model.Insulin resistance (IR) is the common mechanism of MS and first occurs in skeletal muscle which is an important organ in the metabolism of glucose. The abnormity of any factor in the PI3K pathway may induce insulin resistance. But it is unclear that the importance of pathological changes of skeletal muscle and abnormity of PI3K pathway in the insulin resistance of low birth weight.Angiotensin II (Ang II) is a key factor of renin-angiotensin system (RAS). It can influence the PI3K pathway and is connected with insulin resistance. So it needs to know the expression and effects of Ang II in the development of low birth weight.Nutritional programming points out that in the key period of development the nutrition will influence the organ's function for a long time or lifetime. So in theory, nutritional intervention on low birth weight rat can prevent insulin resistance and adult MS. Arginine (Arg) is a conditionally essential amino acid. L-Arg is the active form and has many biological effects. For example, it can promote the growth of fetus and cure IUGR, decrease blood sugar and improve insulin resistance, reduce the content of Ang II in cardiovascular tissues and restrain RAS activity. But it is still unknown whether the supplement of L-Arg in the early life can prevent insulin resistance in low birth weight.Objective(1)To observe the newborn member per nest, body weight of neonatal rat, IUGR rate and perinatal death rate when the pregnant rats were fed 10% low protein diet or 21% normal protein diet; and to test whether feeding pregnant rats 10% low protein diet was an effective method to make low birth weight rat model. To observe the changes of body weight, blood sugar level, serum insulin level and insulin resistance index (IRI) in the growth of low birth weight rats.(2)To study the changes of the structure and PI3K pathway of skeletal muscle in different age of low birth weight rats.(3)To study the expression and effects of Ang II on PI3K pathway, and analyse the mechanism of insulin resistance in low birth weight rats.(4)To examine the body weight, blood sugar, serum insulin and IRI of low birth weight rats after given L-Arg, observe the changes of structure and PI3K pathway of skeletal muscle. To analyse the alteration of Ang II after L-Arg supplement and discuss the possible mechanism of preventing insulin resistance by using L-Arg.Methods(1) Establishment of low birth weight rat model: Eighteen male and eighteen female healthy three-month SD rats were bought. Male and female rats were put into a cage at the proportion of 1:1. The pregnant rats were divided into three groups: pregnant rats of model and intervention groups were fed 10% low protein diet, while normal group were fed 21% normal protein diet. 21% normal protein diet was given to the maternal rats in the lactation and the offspring after ablactation. The maternal rats of intervention group were given a supplementation of L-Arg drinking water in the lactation. At 7d, 21d, 2m and 3m, the body weight, blood sugar and serum insulin were measured, then IRI could be calculated. (2)Study of the structure, PI3K pathway and Ang II of skeletal muscle: At 7d, 21d and 2m, the structure and ultrastructure of skeletal muscle were analysed in the offspring of nomal and model groups. The protein expression of PI3K, PKB, p-PKB, GLUT4 and insulin-stimulated p-PKB, GLUT4 of plasma membrane were also detected by western-blot. Ang II was examined by radioimmunoassay. AT1R was detected by immuno-histochemical staining technique.(3)Measure of these indexes after supplyment of L-Arg: In the offspring of intervention group, the body weight, blood sugar and serum insulin were measured, then IRI could be calculated. The structure and ultrastructure of skeletal muscle were analysed. The protein expression of PI3K, PKB, p-PKB, GLUT4 and insulin-stimulated p-PKB, GLUT4 of plasma membrane were detected by western-blot. Ang II was examined by radioimmunoassay. AT1R was detected by immuno-histochemical staining technique. All data were compared among normal, model and intervention groups.These data were counted with the SPSS-11.5. P value < 0.05 was considered significant.Results(1)The offspring in model and intervention groups got low birth weight. But there were no obvious changes in the neoborn member per nest and perinatal death rate among three groups. (2)At 7d, the body weight of offspring in model and intervention groups were lower than normal group, but the intervention group heavier than model group. At 21d, the body weight of intervention group reached normal level. While at 2m he body weight of model and intervention groups were under the nomal level obviously. There was no significant difference between the nomal and model offsprings at 3m. The difference of blood sugar, serum insulin and IRI among three groups were not distinct at 7d,21d and 2m. Although the blood sugar was higher in model offspring at 3m, there was no statistic significance.(3)From 7d to 2m, the muscle fiber of low birth weight rat atrophied and arranged in disorder. The ultrastructure of skeletal muscle was also abnormal distinctly at 2m. For example the myofibril arranged in disorder, atrophied, and broke down. Mitochondria swelling, disruption of mitochondria crest and vesicle mitochondria were observed. But all these abnormality meliorated in intervention group.(4)At 7d the expression of PI3K became higher in intervention group than model group and reached normal level at 21d, but no distinct difference could find when compared intervention group with nomal or model groups at 2m. The expression of PKB wasn't different among three groups. Before giving insulin-stimulation, expression of p-PKB was almost same in three groups at 7d. However, it was visibly higher in model group than the other groups at 21d and 2m. The expression of total GLUT4 was nearly equal. After the stimulation of insulin, the increase of p-PKB became normal in intervention group at 21d, but lower than normal group at 7d and 2m. The GLUT4 in plasma membrane was normal in intervention group and higher than the model group..(5)In intervention group the content of Ang II decreased distinctly and approached normal level at each time. The expression of AT1R also lessened. The correlation analysis showed negative correlation between Ang II and the increase of p-PKB or expression of GLUT4 in plasma membrane after insulin stimulation.Conclusions(1) From 2m to 3 m, the low birth weight offspring had catch up growth and had no insulin resistance in 2m.(2) The structure of skeletal muscle was abnormal and PI3K pathway was unusual in low birth weight rat. The expression and function of Ang II was enhanced in low birth weight rat and may restrain PI3K pathway.(3) Supplement of L-Arg in the early life could promote the catch up growth of low birth weigh rat, improve the abnormality of skeletal muscle and decrease the content and inhibiting effects of Ang II to the PI3K pathway. |
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