Expression of c-kit mRNA and protein during the differentiation of human hematopoietic progenitor cells

Y. Yamaguchi, Y. Gunji, M. Nakamura, K. Hayakawa, M. Maeda, H. Osawa, K. Nagayoshi, T. Kasahara, T. Suda

Research output: Contribution to journalArticle

32 Citations (Scopus)

Abstract

To investigate how c-kit and c-kit ligand play a role in the function of hematopoietic stem cells, we determined the expression of c-kit in sorted human hematopoietic stem cells, CD34+CD33- cells and CD34+CD33+ cells. CD34+ cells constituted approximately 1% of the population of gated bone marrow cells and contained colony-forming cells. Two-color analysis by a fluorescence-activated cell sorter (FACS) revealed that about one-third to one-half of the total CD34+ cell population were positive for the CD33 antigen. To analyze the relative accumulation of c-kit mRNA in sorted cells, we used the reverse transcription-polymerase chain reaction (RT-PCR) method, followed by Southern blot analysis. There was a linear relationship between the amount of input RNA and products amplified in the range of 103 to 105 cells. Using this procedure, we carried out an analysis of c-kit mRNA expression in CD34+CD33-, CD34+CD33+, CD34-CD33+, and CD34-CD33- cells. Enhanced expression for c-kit mRNA was observed solely in CD34+CD33- cells. In contrast, flow cytometry shows that c-kit protein was expressed most abundantly in CD34+CD33+ cells. Colony-forming cells were generated on a human stromal cell layer for 5 weeks initiated with CD34+CD33- cells but not with CD34+CD33+ cells. During co-culture with stromal cells, CD34+CD33- cells differentiated into CD34+CD33+ cells. From these findings, it is concluded that CD34+CD33+ cells are direct progenies of CD34+CD33- cells. In this differentiation pathway, the expression of c-kit mRNA decreased and the c-kit protein increased.

Original languageEnglish
Pages (from-to)1233-1238
Number of pages6
JournalExperimental Hematology
Volume21
Issue number9
Publication statusPublished - 1993 Dec 1

Keywords

  • CD34
  • FACS
  • RT-PCR
  • c-kit

ASJC Scopus subject areas

  • Molecular Biology
  • Hematology
  • Genetics
  • Cell Biology
  • Cancer Research

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    Yamaguchi, Y., Gunji, Y., Nakamura, M., Hayakawa, K., Maeda, M., Osawa, H., Nagayoshi, K., Kasahara, T., & Suda, T. (1993). Expression of c-kit mRNA and protein during the differentiation of human hematopoietic progenitor cells. Experimental Hematology, 21(9), 1233-1238.