Mononuclear cells of umbilical cord blood differentiation to granulocyte cell in vitro
10.3760/cma.j.issn.0253-2727.2017.06.013
- VernacularTitle: 脐血单个核细胞体外诱导分化为粒系细胞的研究
- Author:
Lin CHEN
1
;
Xiaoyan XIE
;
Jiqin NIE
;
Dongli CHEN
;
Anping HUANG
;
Fang FANG
;
Mingyi QU
;
Xue NAN
;
Lijuan HE
;
Zeng FAN
;
Wen YUE
;
Xuetao PEI
1
Author Information
1. Stem Cell and Regenerative Medicine lab, Beijing Institute of Transfusion Medicine, South China Research Center for Stem Cell& Regenerative Medicine, Beijing 100850, China
- Publication Type:Journal Article
- Keywords:
Cord blood;
Mononuclear cells;
Granulocyte cells
- From:
Chinese Journal of Hematology
2017;38(6):532-536
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To explore an optimal method for granulocyte cell production from umbilical cord blood mononuclear cells.
Methods:Erythrocytes were precipitated by hydroxyethyl starch. Mononuclear cells were isolated through Ficoll density gradient centrifugation. Different media, additives and cultivation model were chosen for granulocyte induction. Cell morphology was observed by microscopy, and cell phenotype was detected by flow cytometry. The CD18 expression of granulocytes was tested by immunofluorescence assay, and phagocytosis test was executed as well.
Results:Compared to fetal bovine serum (FBS) treatment group, cell viability, counts and differentiation rate of granulocytes induced by X-VIVOTM 15 combined with TPO, SCF, G-CSF but without FBS were superior. And X-VIVOTM15 medium was better than SCGM medium at effectiveness and cost. Using two-stage mode of hematopoietic stem cell expansion followed by granulocyte induction with X-VIVOTM15 combining TPO, SCF and G-CSF, cell proliferation was nearly 132 times at day 21. Flow cytometry showed that the differentiation was lagged in 2-stage mode than in direct induction mode, CD15 expression was (69.60± 1.06) % vs (97.73±0.39) %; Wright-Giemsa staining demonstrated mature granulocytes; immunofluorescence showed the expression of lysosomal proteins CD18. A strong phagocytic function of mature granulocytes was demonstrated by phagotrophic efficiency of (51.43±0.05) %. And granulocyte had chemotaxis ability under the role of chemotactic factor IL-8.
Conclusion:Optimized culture media and cultivation mode are achieved for functional granulocytes induction in vitro.
