The extracellular domain of human delta-like-1 expressed and purified from CHO cells promotes expansion of hematopoietic progenitor cells / 中国实验血液学杂志
Journal of Experimental Hematology
; (6): 222-226, 2003.
Artigo
em Chinês
| WPRIM (Pacífico Ocidental)
| ID: wpr-355678
Biblioteca responsável:
WPRO
ABSTRACT
Notch signal path plays important roles in the regulation of proliferation and differentiation of hematopoietic stem cells. An extracellular domain of human Delta-like-1 (hDll-1(ext)), one of Notch ligands, was cloned and expressed in CHO cells, and the effect of hDll-1(ext) on expansion of hematopoietic stem/progenitor cells was investigated in this study. Total RNA was isolated from human marrow mononuclear cells. hDll-1(ext) was amplified by RT-PCR and cloned to T vector, then the gene was sequenced and subcloned to pcDNA3.1/Myc-His(+)A expression vector. The constructed plasmid was transfected into CHO cells with lipofectin and the expression of secreted hDll-1(ext) in G418-resistant clones was assayed by Western blot. hDll-1(ext) high-expressed clone was cultured to collect supernatant. Fusion protein hDll-1(ext) was purified from the supernatant by immobilized metal affinity chromatography (IMAC). The results showed that expression of Notch-1 receptor was detected in cord blood-derived CD34(+) cells by RT-PCR. Human umbilical blood CD34(+) cells were cultured in serum-free medium containing SCF, IL-3, VEGF, and with or without purified hDll-1(ext) for 4 or 8 days. Effect of hDll-1(ext) on the expansion of progenitor cells was analyzed then by clonogenic assays. The number of CFU-Mix and HPP-CFC generated from the culture system containing hDll-1(ext) was 1.5 times of that from the control. In conclusion, the recombinant hDll-1(ext) promotes the expansion of primitive hematopoietic progenitors.
Texto completo:
Disponível
Base de dados:
WPRIM (Pacífico Ocidental)
Assunto principal:
Farmacologia
/
Fisiologia
/
Fatores de Transcrição
/
Sítios de Ligação
/
Proteínas Recombinantes
/
RNA
/
Células-Tronco Hematopoéticas
/
Glicoproteínas
/
Transfecção
/
Expressão Gênica
Limite:
Animais
/
Humanos
Idioma:
Chinês
Revista:
Journal of Experimental Hematology
Ano de publicação:
2003
Tipo de documento:
Artigo