Preparation method and application of human pluripotent stem cell senescence model

A technology of human pluripotent stem cells and cell aging, applied in the field of biomedicine, can solve the problems of lack of cell models of Chinese mutant HGPS patients, etc., and achieve the effects of easy sampling, high safety, and simple cell separation methods

Pending Publication Date: 2019-05-21
深伦生物科技(深圳)有限公司
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Problems solved by technology

[0006] The first object of the present invention is to provide a method for preparing a human pluripotent stem cell aging model carrying a novel gene mutation, so as to solve the problem of the lack of cell models of Chinese mutant HGPS patients, and to study the pathological conditions of Chinese mutant HGPS, Molecular pathogenic mechanisms and simulated in vitro pathology provide cellular models

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  • Preparation method and application of human pluripotent stem cell senescence model
  • Preparation method and application of human pluripotent stem cell senescence model
  • Preparation method and application of human pluripotent stem cell senescence model

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Embodiment Construction

[0032] The present invention will be described in detail below in conjunction with specific embodiments.

[0033] The initial subjects of this experiment included 3 normal healthy people (control group) and 3 Chinese HGPS patients carrying the c.1579C>T (p.R527C) mutation (experimental group). Basic conditions such as gender and age of normal people and patients are matched with each other. Part of the patient information is as follows: patient E-1: female, 3 years old; patient E-2: male, 5 years old; patient E-3: male, 7 years old. The rest of the information is kept confidential. The main implementation steps include:

[0034] 1) Peripheral blood collection from normal people and Chinese HGPS patients, isolation of peripheral blood mononuclear cells (PBMC), culture and expansion of primary PBMC.

[0035] 1.1) Alcohol disinfection, use heparin sodium anticoagulant blood collection tube to collect 2ml of blood from the head vein of normal people and HGPS patients, shake the...

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Abstract

The invention relates to the technical field of biomedicine, and provides a preparation method and application of a human pluripotent stem cell senescence model. The preparation method of the human pluripotent stem cell senescence model comprises the steps that peripheral blood monocytes from Chinese mutant HGPS patients are reprogrammed into Chinese HGPS-iPSC; c.1579C)T(p. R527C) mutation of LMNAis repaired by using a single nucleotide mutation repair technology mediated by CRISPR/Cas9 and ssODNs; normal iPSC, Chinese HGPS-iPSC and mutation correction iPSC are differentiated into functionalMSCs respectively in vitro, and the expression, cell proliferation, apoptosis, cell cycle, cell senescence and the like of LMNA protein in three iPSC-MSCs are detected, thereby disclosing the effect of the c.1579C)T(p. R527C) mutation on an expression function of cell-related passage signaling molecules, and studying the molecular mechanism and pathogenic mechanism of the Chinese HGPS which are different from Caucasian population mutant HGPS.

Description

technical field [0001] The invention relates to the technical field of biomedicine, in particular to a preparation method and application of a human pluripotent stem cell aging model. Background technique [0002] Progeria, also known as Hutchinson-Gilford Progeria Syndrome (HGPS), is an extremely rare congenital genetic disease, and its patients experience rapid physical aging during infancy, with an incidence of about one in a million First, patients generally show symptoms such as skin sclerosis at about 1 year old, and obvious aging phenotypes can be seen at about 3 years old, and aging faces, subcutaneous fat loss, and osteolysis can also appear. The patient's aging rate is 5-10 times that of normal children, and the premature death of cardiovascular and cerebrovascular diseases is generally caused by accelerated cardiovascular atherosclerosis before the teens. At present, it has been preliminarily recognized that the cells of HGPS patients have obvious defects: includ...

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/10C12N5/0735C12N5/074C12N5/0775C12N15/85C12N15/90C12Q1/02
Inventor 周焱莫翠萍舒伟廖进齐方崇州韩倩刘佳周光前
Owner 深伦生物科技(深圳)有限公司
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