| IVWith the development of economy and society and the increase of femaleeducation level,advanced maternal age gradually becomes a social problem.Especially the implementation of China’s two-child policy,it is increasingly common for women to delay childbearing in China.In addition to the adverse effects to mother,the delayed motherhood has a negative effect on the growth and development of children.However,the influence of advanced maternal age on the neural development and intelligence of children is still controversial.We have proved that the offspring born to old female rats have poorer ability of study and memory using Morris water maze test,and a set of genes are differentially expressed in hippocampus of offspring born to old female rats when compared with those in offspring born to young female rats through gene chip,q RT-PCR and Western blot.In order to further explore the influence of the delayed motherhood on the growth and development of offspring,especially on the neural development and intelligence,q RT-PCR and Western blot were used to detect the dynamic expression of Skp2 gene and its substrate,p27 and p21,in the hippocampus of offspring born to old female rats.We also detected the expression of the neural development related genes in the hippocampus of adult offspring,which showed the effect of delayed motherhood on the development of hippocampus in offspring at the molecular level.ObjectiveTo investigate the effects of delayed motherhood on the development of offspring,to detect the expression of Skp2,p21 and p27 in hippocampus of offspring during the neuronal development,to study the expression of neural development related genes in hippocampus of adult offspring,to study the possible effect of delayed motherhood on the ability of study,memory and cognitive in offspring.Methods1.Experimental animals and groups: Sprague–Dawley rats were from laboratory animal center of Zhengzhou University.Young female rats(3-month-old)and old female rats(12-month-old)were randomly selected and cohabited with male rats(3-month-old)separately,and the offspring were divided into two groups: Young mother group and old mother group.2.Reproductive ability.The pairing-to-birth days,litter size and sex of pups after parturition were recorded.3.The weight of offspring.The weight of offspring rats was recorded at 1,3,7,14,21,28,60 days after birth.4.The detection of the expression of Skp2,p21 and p27.The offspring were sacrificed at 1,3,7,14,21,28,60 days after birth,and the hippocampus tissue was separated.The m RNA expression of Skp2 was detected by q RT-PCR.The pritein expression of Skp2 and its substrate p27 and p21 were detected by Western blot at 60 days after birth.5.The detection of neural development related genes in adult hippocampus.The m RNA expression of Edn1,Hspa4,Fos,Bcl2 and Grin1 in the hippocampus was detected by q RT-PCR in the hippocampus of offspring rats at 60 days after birth.6.Statistical analysis.SPSS17.0 statistical software was used for data analysis.ANOVA was used to analyze weight change in young mother group and old mother group.The proportion of male and female,the development index and the expression of Skp2,p21,p27 at different time points during development were tested by independent samples t test.The inspection level: α=0.05.Results1.Reproductive ability.The pairing-to-birth days of old female rats were significantly longer than that of youth female rats.The litter size of old female rats is less than that of youth female rats significantly(P < 0.01).The proportion of birth sex had no significant difference between the offspring born to old mother and the offspring born to young mother(P > 0.05).2.The weight of offspring.The weight of offspring was recorded on the 1,3,7,14,21,28,60 days after birth.The weight of the two groups increased gradually during the development,and there was no significantly difference between old mother group and young mother group(P > 0.05).3.The expression of Skp2,p27 and p21 during the development.The m RNA expression of Skp2 in the hippocampus decreased with the development time at old mother group and young mother group.The m RNA expression of Skp2 at 7 days after birth in old mother group was less than that in young mother group significantly(P < 0.05)and There was no significantly difference between two groups at other time point(P > 0.05).The protein expression of Skp2 in the hippocampus decreased with the development time at old mother group and young mother group.From 1 days to 14 days after birth the protein expression of Skp2 in two groups was similar,and during 21 days to 60 days after birth the protein expression of Skp2 in old mother group is higher than that in young mother group significantly(P < 0.05).The protein expression of p27 decreased during the development and there was no significantly difference between two groups(P > 0.05).The protein expression of p21 increased during the development and it was higher at 28 and 60 days after birth in old mother group than that in young mother group significantly(P < 0.05).4.Detection of neural development related genes.At 60 days after birth,the m RNA expression of Edn1 and Hspa4 in old mother group was less than that in young mother group(Edn1 P < 0.05,Hspa4 P < 0.01),and there was no significantly difference in m RNA expression of Fos,Bcl2,and Grin1 between two groups(P >0.05).Conclusion1.The expression of Skp2 and p21 may be related to the change of neural function and the decline of learning and memory function in offspring born to old mother,but Skp2-p27/p21 may be not the important mechanisms when delayed motherhood changes offspring neural development.2.Edn1 and Hspa4 are less expressed in adult offspring born to old mother than that in young group,suggesting that these two genes may be involved in the neural function change in offspring by the delayed motherhood. |
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