Human II-type thrombocytopenia iPS cell line and preparation method and application thereof

Abstract

The invention relates to the technical field of biology, in particular to a human II-type thrombocytopenia iPS cell line and a preparation method and application thereof. The iPS cell line is formed by introducing a transcription factor into CD34+ cells of a patient suffering from II-type hereditary thrombocytopenia related to c.-128G>T heterozygous mutation in a 5'-UTR region of an Ankerd26 gene,and cell reprogramming is formed; the preparation method comprises the following steps that mononuclear cells are separated from an in-vitro bone marrow specimen, sorting is carried out to obtain CD34+ cells, and the CD34+ cells are subjected to cell culture; and a virus vector containing Oct4, Sox2, Klf4 and c-Myc genes is used for transfecting the cultured CD34+ cells, then centrifuging is carried out, cell culture is carried out to enable the CD34+ cells to be reprogrammed, and the successfully reprogrammed cells are screened to obtain the iPS cell line. According to the human II-type thrombocytopenia iPS cell line and the preparation method and application thereof, the CD34+ cells derived from marrow of the patient suffering from type II thrombocytopenia and having c.-128G>T heterozygous mutation in the 5'-UTR region of the Ankerd26 gene are reprogrammed into the iPS cell line, the prepared iPS cell line is long in maintenance life, and the human II-type thrombocytopenia iPS cellline can be applied to researches such as the establishment of an in-vitro disease model of the type II thrombocytopenia and drug screening.

Description

Technical field [0001] The invention relates to the field of biotechnology, in particular to a human type II thrombocytopenia iPS cell line and its preparation method and application. Background technique [0002] Inherited thrombocytopenia (IT) is a rare disease characterized by thrombocytopenia, or (and) diseases accompanied by abnormal platelet function. The clinical manifestations are mainly petechiae, ecchymosis or bleeding symptoms of varying degrees , No specificity. The etiology has a low incidence, lack of specific clinical manifestations and is often missed or misdiagnosed as idiopathic thrombocytopenic purpura. Type II thrombocytopenia is one of the most common types of IT. It belongs to an autosomal dominant inheritance model. It is characterized by a moderate reduction in the number of megakaryocytes in the bone marrow of the patient, but the size and function of the platelets are normal. The mutation in the 5'-UTR region of the ANKRD26 gene has been confirmed to b...

AI Technical Summary

Problems solved by technology

Mutations in the 5'-UTR region of the ANKRD26 gene have been confirmed to be the causative gene mutations of type Ⅱ thrombocytopenia. Mutations in this region lead to the impact of Runt-related transcription factor 1 (RUNX1) and Friend leukemia virus integration factor 1 (FLI1) on the ANKRD26 gene. Attenuated transcriptional repression, resulting in sustained overexpression of the ANKRD26 gene, which ultimately leads to impaired megakaryocyte maturation and decreased platelet count

Because type II thrombocytopenia is a rare disease, and the number of megakaryocytes in the bone marrow is rare, it is difficult to obtain megakaryocyte samples from patients, and the preparation process of primary megakaryocytes is complicated and their lifespan is short. The pathogenic mechanism and therapeutic drugs of thrombocytopenia are greatly limited

Images