[Establishment of induced pluripotent stem cell model of Aicardi-Goutières Syndrome mutated in TREX1]

Abstract

To establish and identify induced pluripotent stem cells (iPSCs) derived from patients with Aicardi-Goutières syndrome (AGS) with TREX1 gene 667G>A mutation, and obtain a specific induced pluripotent stem cell model for Aicardi-Goutières syndrome (AGS-iPSCs). A 3-year-old male child with Aicardi-Goutieres syndrome was admitted to Zhongshan People's Hospital in December 2020. After obtaining the informed consent of the patient's family members, 5 ml peripheral blood samples from the patient were collected, and mononuclear cells were isolated. Then,the peripheral blood mononuclear cells(PBMCs) were transduced with OCT3/4, SOX2, c-Myc and Klf4 by using Sendai virus, and PBMCs were reprogrammed into iPSCs. The pluripotency and differentiation ability of the cells were identified by cellular morphological analysis, real-time PCR, alkaline phosphatase staining (AP), immunofluorescence, teratoma formation experiments in mice. The results showed that the induced pluripotent stem cell line of Aicardi-Goutieres syndrome was successfully constructed and showed typical embryonic stem-like morphology after stable passage, RT-PCR showed mRNA expression of stem cell markers, AP staining was positive, OCT4, SOX2, NANOG, SSEA4, TRA-1-81 and TRA-1-60 pluripotency marker proteins were strongly expressed. In vivo teratoma formation experiments showed that iPSCs differentiate into the ectoderm (neural tube like tissue), mesoderm (vascular wall tissue) and endoderm (glandular tissue). Karyotype analysis also confirmed that iPSCs still maintained the original karyotype (46, XY). In conclusion, induced pluripotent stem cell line for Aicardi-Goutières syndrome was successfully established using Sendai virus, which provided an important model platform for studying the pathogenesis of the disease and for drug screening.