Relationship Between The Embryonic Reactive Oxygen Species Level And Embryo Apoptosis Pathways In Mouse

In mammals,the embryonic blocking phenomenon often occurs in the early stages of development,affecting the quality of embryos,even lead to apoptosis.Reactive oxygen species(H2O2)is one of the cause cell apoptosis.Now inducing apoptosis pathways major include death receptor signaling pathway,mitochondria pathways and endoplasmic reticulum pathways,but because of Caspase-12 activate the apoptosis induced by endoplasmic reticulum stress,but the process of active oxygen stress caused whether the presence of Caspase-12 also need to study.On the other hand,reactive oxygen species induced mouse early embryonic apoptosis which apoptotic pathways plays a main role is unknown.So the aim of this study is to investigate the effects of Caspase-12 participate in the endoplasmic reticulum pathways apoptosis from ROS on the molecular level,determine the reactive oxygen species mainly perform in the process of early embryo in mice induced apoptosis pathways.In this study,we investigated the effect of in vivo,in vitro culture and treated with hydrogen peroxide(H2O2)three groups,comparing the embryos using DCFH-DA reactive oxygen content in the determination of mouse embryos(2-cell,4-cell and blastocysts),and by using TUNEL staining method to detect DNA fragmentation of blastocyst.Next analyse the expression of mRNA level of caspase-12,confirmed caspase-12 participate in apoptosis which the endoplasmic reticulum pathway induced.Then analyse the expression of mRNA level of the death receptor pathway of caspase-8,the mitochondrial apoptotic pathways of caspase-9 and casapse-3.Determine the different apoptosis pathways in the process of embryo apoptosis induced by ROS of progression.We observed a higher H202 level in vitro in 2-cell,4-cell and in blastocysts.And H2O2,treatment groups significant difference with other groups,verify the correctness of groups.And observed presence of H2O2 treatment groups decreased the total cell number and increase the apoptotic index,where as in vitro ignificantly(P<0.05).A higher apoptotic index and a lower total cell number were found in H2O2,group versus the vivo group(P<0.05),Further validated the increase of ROS level affects not only influence the apoptosis index,thus affecting the quality of the embryo,provides the basis for subsequent better study.Next the expression of Caspase-12 in embryos,confirmed that the Caspase-12 participate in the active oxygen stress caused by the endoplasmic reticulum pathway induced.In addition,the death receptor pathway of Caspase-8 gene expression level significant increased in 2-cell,4-cell stage H2O2 groups Caspase-8 gene mRNA expression quantity highest;and in blastocysts the higer expression of genes associated with a higher H2O2 level.Under the same level of reactive oxygen species,the highest expression of genes caspase-8 in vivo,in vitro and H2O2 groups.In conclusion,our data demonstrated that Caspase-12 involved in the active oxygen caused by endoplasmic reticulum stress mediated apoptosis;umder the same reactive oxygen level focus on the death receptor pathway.The death receptor pathway induced embryo apoptosis was the main pathway in 2-cell and 4-cell stage;The blastocyst stage three apoptotic pathways in no primary and secondary.