A method for constructing an immunodeficiency mouse transplantation model of human stem cells

A construction method and technology of stem cells, applied in medical science, veterinary instruments, veterinary surgery, etc., can solve the problems that are not suitable for clinical and scientific research experiments, hinder the implantation and survival of human hematopoietic stem cells, and the microenvironment of human hematopoietic stem cells. Clear and other issues, to achieve the effect of simple and easy implementation, low price, and less trauma

Active Publication Date: 2016-04-20
INST OF HEMATOLOGY & BLOOD DISEASES HOSPITAL CHINESE ACADEMY OF MEDICAL SCI & PEKING UNION MEDICAL COLLEGE
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Problems solved by technology

However, NOD / SCID mice also have many disadvantages: (1) there are still low levels of NK cell activity, different major histocompatibility sites, and complete fusion cell functions such as the functional expression of NK cell perforin molecules, less than 10 % of NOD / SCID mice will experience immune escape and produce B lymphocytes and T lymphocytes. These remaining immune responses will not be conducive to the implantation of human cells; (2) Microenvironment of human hematopoietic stem cell transplantation in NOD / SCID mice It is unclear, mostly of mouse origin, that human stem cells cannot utilize mouse hematopoietic cytokines due to lack of cross-reactivity to mouse cytokines, and lack of interaction with appropriate adhesion molecules will continue to hamper human Engraftment and survival of hematopoietic stem cells
However, in addition to the relatively large trauma to mice, these methods use relatively expensive antibodies and cytokines, which can only be limited to research in some scientific research laboratories, and are not suitable for wide application in clinical and scientific research experiments.

Method used

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  • A method for constructing an immunodeficiency mouse transplantation model of human stem cells
  • A method for constructing an immunodeficiency mouse transplantation model of human stem cells
  • A method for constructing an immunodeficiency mouse transplantation model of human stem cells

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Embodiment 1

[0026] A method for constructing an immunodeficiency mouse (NOD / SCID) transplantation model of human stem cells (the method for improving the implantation rate of human stem cells in an immunodeficiency mouse transplantation model), comprising the steps of:

[0027] 1) Take 8-week-old NOD / SCID mice, female, and raise them in an SPF-grade sterile room. The feed, drinking water, and litter are all autoclaved, and the antioxidant N-acetylcysteine ​​(100mg / kg) the mice in the experimental group were injected intraperitoneally, once every other day, and N-acetylcysteine ​​was added to the drinking water of the mice in the experimental group at the same time, and the concentration of N-acetylcysteine ​​was 1 mg / ml to reduce the Reactive oxygen species levels in the mouse bone marrow, the recipient mice 137 Cs source sublethal dose irradiation, dose 2.0Gy, dose rate 0.7Gy / min, the collected human CD34+ cells were transplanted through the tail vein of the mice within 24 hours after t...

Embodiment 2

[0034] A method for constructing an immunodeficiency mouse (NOD / SCID) transplantation model of human stem cells (the method for improving the implantation rate of human stem cells in an immunodeficiency mouse transplantation model), comprising the steps of:

[0035] 1) Take 8-week-old NOD / SCID mice, female, and raise them in an SPF-grade sterile room. The feed, drinking water, and litter are all autoclaved, and the antioxidant N-acetylcysteine ​​(100mg / kg) the mice in the experimental group were injected intraperitoneally, once every other day, and N-acetylcysteine ​​was added to the drinking water of the mice in the experimental group at the same time, and the concentration of N-acetylcysteine ​​was 1 mg / ml to reduce the Reactive oxygen species levels in the mouse bone marrow, the recipient mice 137 Cs source sublethal dose irradiation, dose 2.0Gy, dose rate 0.7Gy / min, the mice were anesthetized by intraperitoneal injection of pentobarbital sodium (50mg / kg) within 24 hours a...

Embodiment 3

[0044] Using the method of limiting dilution, the number of transplanted cells in Example 2 and Comparative Example 2 was changed, and other conditions remained unchanged, and the number of transplanted cells was: 100, 50, 20, and 10 cells / group. Such as Figure 4 As shown, according to the principle of Poisson distribution, after treatment with the antioxidant N-acetylcysteine, NOD / SCID mice were injected with tibial bone marrow (IT), contralateral bone marrow (BM) and spleen (SP). The probabilities of human SCID-repopulating cells (SCID-repopulating cells, SRC) were: 1 / 36, 1 / 50, 1 / 57; the control groups were: 1 / 108, 1 / 203, 1 / 448. It shows that NOD / SCID mice treated with antioxidants are 3.0, 4.1 and 7.9 times more sensitive for detecting human SCID replantation cells, respectively.

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Abstract

The invention discloses a method for constructing an immunodeficiency mouse transplant model of human stem cells. The method comprises the following steps of: performing intraperitoneal injection on a receptor mouse (NOD / SCID) by adopting an N-acetylcysteine (NAC) antioxidant before transplanting, supplying drinking water containing NAC to the receptor mouse, continuously supplying the drinking water containing the NAC to the receptor mouse after the human stem cells are transplanted, and thus obtaining the immunodeficiency mouse transplant model of the human stem cells. The optimal model of NOD / SCID mouse transplant is successful established; and by the method, no matter in caudal vein or medullary cavity transplant, the implantation rate of the human stem cells is effectively improved, and the mouse is low in death rate and good in status in the transplant process and can survive for a long term. The method is simple, novel, easy to implement and low in wound to the mouse; and the NAC antioxidant used by the method is low in price.

Description

technical field [0001] The invention relates to a biotechnology method, in particular to a method for constructing an immunodeficiency mouse transplantation model of human stem cells (HSCs) for improving the implantation rate of human stem cells (HSCs) in the immunodeficiency mouse transplantation model. Background technique [0002] NOD / SCID (Non-obesediabeticseverecombinedimmunodeficiency) is a non-obese diabetic / severe combined immunodeficiency mouse, which is an immunodeficiency mouse produced by backcrossing SCID (severe combined immunodeficiency) mice and NOD (non-obese diabetic) mice. It is the most commonly used immunodeficiency mouse model at present. The mouse not only has the characteristics of SCID mice, lacks functional T and B lymphocytes, and has dual immunodeficiency of cells and humoral, but also has multiple inherent immune deficiencies of NOD mice. Including low NK cell and complement activity, limited development and function of myeloid cells, etc., so it...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61D1/00
Inventor 程涛胡林萍高瀛岱
Owner INST OF HEMATOLOGY & BLOOD DISEASES HOSPITAL CHINESE ACADEMY OF MEDICAL SCI & PEKING UNION MEDICAL COLLEGE
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