Hasegawa Koki, Nakano Kazuaki, Nagaya Masaki, Watanabe Masahito, Uchikura Ayuko, Matsunari Hitomi, Umeyama Kazuhiro, Kobayashi Eiji, Nagashima Hiroshi
Laboratory of Developmental Engineering, Department of Life Sciences, School of Agriculture, Meiji University, 1-1-1 Higashimita, Tama-ku, Kawasaki 214-8571, Japan.
PorMedTec Co. Ltd., 2-3227 MIta, Tama-ku, Kawasaki, Kanagawa, 214-0034, Japan.
Regen Ther. 2022 Jun 12;21:62-72. doi: 10.1016/j.reth.2022.05.010. eCollection 2022 Dec.
Previously, we performed gene knockout (KO) of interleukin-2 receptor gamma () in porcine fetal fibroblasts using zinc finger nuclease-encoding mRNAs, subsequently generating KO pigs using these cells through somatic cell nuclear transfer. The KO pigs lacked a thymus and were deficient in T lymphocytes and natural killer cells, similar to human X-linked severe combined immunodeficiency (SCID) patients. The present study aimed to evaluate whether pigs can support the growth of xenografted human cells and have the potential to be an effective animal model.
The XY pigs used in this study were obtained by mating XX females with wild-type boars. This permitted the routine production of KO pigs via natural breeding without complicated somatic cell cloning procedures; therefore, a sufficient number of pigs could be prepared. We transplanted human HeLa S3 cells expressing the tandem dimer tomato into the ears and pancreas of KO pigs. Additionally, a newly developed method for the aseptic rearing of SCID pigs was used in case of necessity.
Tumors from the transplanted cells quickly developed in all pigs and were verified by histology and immunohistochemistry. We also transplanted these cells into the pancreas of designated pathogen-free pigs housed in novel biocontainment facilities, and large tumors were confirmed.
KO pigs have the potential to become useful animal models in a variety of translational biology fields.
此前，我们使用编码锌指核酸酶的信使核糖核酸在猪胎儿成纤维细胞中进行了白细胞介素-2受体γ（IL-2Rγ）基因敲除（KO），随后通过体细胞核移植利用这些细胞培育出了IL-2Rγ KO猪。这些IL-2Rγ KO猪缺乏胸腺，T淋巴细胞和自然杀伤细胞不足，类似于人类X连锁重症联合免疫缺陷（SCID）患者。本研究旨在评估猪是否能够支持异种移植的人类细胞生长，并有无成为有效动物模型的潜力。
本研究中使用的IL-2Rγ XY猪是通过将IL-2Rγ XX雌性与野生型公猪交配获得的。这使得通过自然繁殖常规生产IL-2Rγ KO猪成为可能，而无需复杂的体细胞克隆程序；因此，可以准备足够数量的猪。我们将表达串联二聚体番茄的人类HeLa S3细胞移植到IL-2Rγ KO猪的耳朵和胰腺中。此外，如有必要，还使用了一种新开发的无菌饲养SCID猪的方法。
移植细胞产生的肿瘤在所有猪中迅速发展，并通过组织学和免疫组织化学得到证实。我们还将这些细胞移植到饲养在新型生物安全设施中的无特定病原体猪的胰腺中，并且确认形成了大肿瘤。
IL-2Rγ KO猪有潜力在各种转化生物学领域成为有用的动物模型。
