Development Potential Of Porcine IPS Cells In Vivo

Because of the ability of self-renew and multilineage differentiation potential, Pig iPSCsare used in the medical and biological research. The two parts of this thesis is as follows:1The production of pig iPSC chimeric embryoWe use the piggyback vector, PB[Act-RFP]DS and Act-PBase, to transfect pig iPS, andsuccessfully obtain the PS24-R, a pig iPS cell line expressing RFP. We have got chimericembryos through microinjecting the PS24-R cells into the4-8cell parthenogenetic in vitro,and then collect the statistics of PS24-R in embryos, when the chimeric embryos developed tothe blastocyst stage. The result shows that the PS24-R cells could stably express RFP usingpiggyback vector to transfect the iPS. We are able to effective observe the iPS cells inchimeric embryos. We respectively microinject one, five, ten PS24-R cells and one PS24-Rclone to pig embryos to product chimeric embryos. Along with the increase of donor cells, thechimeric rate of embryos injected gradually increases, but the blastocyst rate declines. At thesame time, compared with the parthenogenetic embryos, the expression lever of the stem cellpluripotent factors, OCT4, SOX2, NANOG, had significant increased in chimeric embryos. Insummary, using the pig iPS marked by piggyback vector Can achieve chimerism in the pigearly embryonic development stage. This work lays a foundation for the production of pig iPScells chimeric animals future.2Injecting pig iPS cells into pig testicular tissue to study the development capacityof iPS cellsIn order to obtain testicular spermatogonia defect model, we used differentconcentrations of busulfan to treat different age boars. The results showed that the number ofspermatogonial cells showed a downward trend along with the gradually increasing of theconcentrations of busulfan at0,5,10,15mg/kg. The weight of testis gradually reduced and theinhibition of ponderal growth gradually strengthened. When using the same dose busulfan toprocess different age pig, the degree of inhibition of spermatogonia has slowed along with theincrease of pig age, but did not change significantly. And then we injected the fluorescentlylabeled pig iPS cells into the testicular tissue, remove the testicles after40days, isolate and culture the testicular cells. We found fluorescence expression, and tissue sections detectingalso verified above result. All of these results indicated that using busulfan to treat boarscould get testicular spermatogonia defect model, and that pig iPS cells could survive in thetesticular tissue, however their differentiation potential should be studied further more.