Effects Of Diethylstilbestrol On The Gene Expression Of Sex Determining Factors And The Learning And Memory In Sex Reversed Chicks

With the development of agriculture and industry,more and more chemical substances are having been synthesized and used in our daily life.Some of them entered into organism disturbing their endocrine system,show hormone activities.As a result, these substances are named as endocrine disrupting chemicals(EDCs) or environmental hormone.Their effects on endocrine system,nervous system and reproductive system are having attracted the attention of people.Diethylstilbestrol(DES) were synthesized to prevent miscarriage and premature delivery of pregnant women.It was also addicted into the feeding stuff as an ingredient to help growth.Recent researches suggested DES could induce reproductive organism disease,so the using of it was banned by most countries.In this paper,we choose eggs and chicks as experiment materials to study how the xenoestrogens affect the development of chicken embryo and the genes expression involved in its sex determination process;besides,we investigate the imprinting learning of chicks after they hatched.In the first part,our aim is to establish a steady and easy- going injecting method to evaluate environmental hormone and its impact.Diethylstilbestrol was injected into eggs in different developmental stages and to different parts of them.Then Polymerase chain reaction(PCR) was used to judge the genetic sex of the chicks. Anatomise the chicks and make gonad sections to investigate if the sexual phenotype were affected by injecting environmental hormone.The result shows that injecting at day O eggs to the yolk from the blunt end of the shell is more likely to get ideal experimental result;0.1mg/egg DES have changed the gonadal sex and led to sex reversal(male→female). In the second part,semi-quantitative RT-PCR was used to investigate how the expression of sex-related genes(SF1,AMH,DMRT1,WPKCI,FET1,FOXL2) fluctuated when the chicken embryos were exposed to 0.1mg/egg DES.Through this part of experiment we could get some clue of sex determining genes and the mechanism of sex determination as well as the environmental hormone effect.We find out that the expression of SF1,AMH,DMRT1,WPKCI are changed greatly under the exposure of DES.The expression of SF1 is detccted in both males and females,and is up-regulate in the genetic males injected with DES.So SF1 gene might have positive effect on the feminization.AMH are set back(after E7) in female and in males,which are exposed to DES.So we drive the conclusion that AMH is related with the development of spermery,but it is not a key factor.DMRT1 expression is down-regulated in early development stage of the sex determining process in both genetic female and male chicks.Thus the gene DMRT1 must have some closely relationship witthe sex reversal and might be the main and key sex determining gene in birds as it is presented in former studies.WPKCI is found to be highly expressed in the critical stage(E7-E11) of the ovary development in the exposed chicken embryo.The expressions of FET1 and FOXL2 are very low and both show higher expression in E11 or E13,so they may have little relationship with the sex detemining.It is known that environmental estrogen can disturb endocrine system,inducing malformation of gonad and abnormal reproductive behavior in aves;it can affect the construction of nervous system as well.In the third section,we want to find out if the imprinting learning ability of the chicks exposed to DES would make some difference from those with regular treatment.The behavior models in this experiment we used are one-trial passive avoidance learning(PAL) task and the detour learning procedure,and the result shows DES can boost the learning and enhance the memory of genetic females,but the effect in males remain blurring.