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In Vitro and In Vivo Interspecies Chimera Assay Using Early Pig Embryos

机译:体外和体内interspecies使用早期猪胚胎的嵌合测定

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Chimeric pigs harboring organs derived from human stem cells are promising for patient-specific regenerative therapies. Induced pluripotent stem cells (iPSCs) can contribute to all cell types of the fetus, including germline after injection into embryos. However, ethical concerns prohibit testing human iPSCs in chimera assays. Here, we evaluated porcine embryos as hosts for an interspecies chimera assay using iPSCs from either cynomolgus monkeys (cyiPSCs) or mouse (miPSCs). To establish an in vitro culture system compatible for cyiPSCs and porcine embryos, we determined blastocyst development in eight different stem cell media. The highest developmental rates of blastocysts were achieved in Knockout Dulbecco's modified Eagle's medium with 20% knockout serum replacement. We found that cyiPSCs injected into porcine embryos survived in vitro and were mostly located in the trophectoderm (TE). Instead, when miPSCs were injected into porcine embryos, the cells rapidly proliferated. The behavior of chimeras developed in vitro was recapitulated in vivo; cyiPSCs were observed in the TE, but not in the porcine epiblast. However, when miPSCs were injected into in vivo derived porcine embryos, mouse cells were found in both, the epiblast and TE. These results demonstrate that porcine embryos could be useful for evaluating the interspecies chimera-forming ability of iPSCs from different species.
机译:嵌入来自人干细胞的器官的嵌合猪是对患者特异性的再生疗法有望。诱导多能干细胞(IPSC)可以促进胎儿的所有细胞类型,包括注射到胚胎后的种系。然而,伦理问题禁止在嵌合体测定中检测人体IPSC。在这里,我们评估了猪胚胎作为嵌入式嵌合体测定的宿主,使用来自Cynomolgus猴子(Cyipscs)或鼠标(MIPSC)的IPSCS。建立对Cyipscs和猪胚相容的体外培养系统,我们在八种不同的干细胞介质中确定了胚泡发育。胚泡发育率最高的胚泡改性Eagle媒介培养基中达到了20%敲除血清替代品。我们发现将Cyipscs注入猪胚胎体外存活,并且主要位于促进菌(TE)中。相反,当将MIPSC注入猪胚胎时,细胞快速增殖。体内嵌合体外开发的嵌合体的行为;在TE中观察到CyipsC,但不在猪外卵细胞中观察到。然而,当将MIPSC注入体内衍生的猪胚胎时,在两者中发现小鼠细胞和TE。这些结果表明,猪胚胎可用于评估IPSCS从不同物种的IPSCS形成能力。

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