Expansion system in vitro for hematopoietic stem/progenitor cell

An in vitro expansion and hematopoietic stem technology, applied in the direction of blood/immune system cells, animal cells, vertebrate cells, etc., can solve unseen problems and achieve broad clinical application prospects and good hematopoietic reconstruction ability

Inactive Publication Date: 2012-08-22
FOURTH MILITARY MEDICAL UNIVERSITY
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  • Abstract
  • Description
  • Claims
  • Application Information

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  • Expansion system in vitro for hematopoietic stem/progenitor cell
  • Expansion system in vitro for hematopoietic stem/progenitor cell
  • Expansion system in vitro for hematopoietic stem/progenitor cell

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Embodiment

[0031] (1) Experimental materials

[0032] (1) Human umbilical vein endothelial cells were cultured as follows

[0033] (2) Umbilical cord blood was obtained from the Department of Obstetrics and Gynecology, Tangdu Hospital, Fourth Military Medical University

[0034] (3) Reagents:

[0035] M199, fetal bovine serum: Gibco

[0036] StemSpan™ serum-free medium, MethoCult™ GF H4434: Stemcell

[0037] Endothelial Cell Growth Supplement (ECGS): Sigma

[0038] Bovine Serum Albumin (BSA): Sigma

[0039] Type II collagenase: Sigma

[0040] Mitomycin C: Sigma

[0041] CD34 MicroBead Kit: Miltenyi Company

[0042] Separation buffer: PBS+2mM EDTA+0.5% BSA

[0043] Lymphocyte Separation Medium: TBD Company

[0044] PBSA: PBS + 2mM EDTA

[0045] SCF (stem cell factor): recombinant human SCF, Peprotech

[0046] TPO (thrombopoietin): recombinant human TPO, Peprotech

[0047] FL (flt3 ligand): recombinant human FL, Peprotech

[0048] IL-6 (interleukin-6): recombinant human IL-6, ...

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Abstract

The invention provides an expansion system in vitro for hematopoietic stem/progenitor cells used for expanding human umbilical cord blood in vitro of soluble fusion protein Dll1-RGD (Hdll1-RGD) of notch ligand of a vascular endothelial cell target. The effective expansion system in vitro is formed by taking human umbilical vein endothelial cells as supporting cells of a cultivation system, jointly applying exogenous growth factors SCF, TOP, FL, IL-6, IL-3 and own developed hDLL1-RGD, and co-cultivating with the hematopoietic stem/progenitor cells. According to the expansion system, the hDll1-RGD target is combined to the surface of a human umbilical vein endothelial cell, activated stimulation aiming at a notch receptor is provided for the hematopoietic stem cells, and self-renewal and propagation of the hematopoietic stem cells are maintained, so that a satisfactory expansion effect and favorable implantation capacity are obtained.

Description

technical field [0001] The invention relates to an in vitro culture method of hematopoietic stem / progenitor cells, in particular to a soluble fusion protein hDll1-RGD of Notch ligand targeting vascular endothelial cells in vitro to expand human umbilical cord blood hematopoietic stem / progenitor cells in vitro. The augmentation system belongs to the field of biotechnology. Background technique [0002] At present, hematopoietic stem cell transplantation has become one of the important methods for the clinical treatment of hematological diseases and malignant tumors. Due to the rich source of cord blood, convenient collection, and weak immunogenicity, it has more advantages than bone marrow and peripheral blood transplantation. However, the content of hematopoietic stem / progenitor cells in umbilical cord blood is low, and it is difficult to meet the needs of adults and patients with large body weight. After transplantation, hematopoietic reconstruction is delayed, and it is di...

Claims

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Application Information

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IPC IPC(8): C12N5/0789
Inventor 韩骅梁英民田登梅
Owner FOURTH MILITARY MEDICAL UNIVERSITY
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