Detection of tryptase-, chymase+ cells in human CD34 bone marrow progenitors.
BACKGROUND
Mast cells (MCs) arise from haematopoietic stem cells. We have recently reported that CD34(+) progenitors derived from human bone marrow (BM) develop into tryptase+, chymase+ MCs when cultured in the presence of recombinant human stem cell factor (rhSCF) and recombinant human IL-6 (rhIL-6). In an experiment for the expression of chymase during differentiation, chymase+ cells were detected in human BM, but tryptase+ cells were not found.


OBJECTIVE
The purpose of this study was to show the appearance of chymase+ cells in CD34(+) cells with an origin different from MC differentiation.


METHODS
CD34(+) cells from human BM were sorted with anti-CD117 monoclonal antibody (mAb), and cytospins of CD34(+), CD34(+)CD117(+), or CD34(+)CD117(-) were prepared. These cells were cultured with rhSCF+rhIL-6 for 12 weeks. Some of the cells were subjected to either histological stain with Wright-Giemsa or immunocytochemistry with anti-chymase mAb. Real-time RT-PCR was also performed to compare the transcriptional level of chymase from each cell preparation.


RESULTS
Chymase was expressed in CD34(+) cells as well as human MCs by immunocytochemistry. Substantial CD34(+)CD117(-) cells, but not CD34(+)CD117(+) cells, were stained immunocytochemically with anti-chymase mAb. For 1 week culture with rhSCF+rhIL-6, no cells expressed chymase in any preparation. Real-time RT-PCR revealed positivity for chymase mRNA in CD34(+) cells, but it reduced at 1 week of culture, and increased as cells developed into MCs. Chymase mRNA in CD34(+)CD117(+) cells was negligible compared with that in CD34(+)CD117(-). Tryptase mRNA was below the detectable level in CD34(+) cells, and increased along with MC differentiation. After 12 weeks of culture, CD34(+)CD117(+) developed predominantly into MCs, whereas CD34(+)CD117(-) developed into monocytes/macrophages.


CONCLUSIONS
Our findings suggested that chymase is present not only in MCs but also in CD34(+)CD117(-) BM progenitors, but that its origin is different from the MC lineage.
Shimizu
Y
Y
Department of Medicine and Biological Science, Gunma University Graduate School of Medicine, Maebashi, Gunma, Japan. yshimizu@med.gunma-u.ac.jp
Suga
T
T
Maeno
T
T
Tsukagoshi
H
H
Kawata
T
T
Narita
T
T
Takahashi
T
T
Ishikawa
S
S
Morishita
Y
Y
Nakajima
T
T
Hara
F
F
Miura
T
T
Kurabayashi
M
M
D018952:Antigens, CD34 / Q000032:analysis*; D002454:Cell Differentiation / Q000187:drug effects / Q000235:genetics; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D053818:Chymases; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D006801:Humans; D015850:Interleukin-6 / Q000494:pharmacology; D008407:Mast Cells / Q000201:enzymology; D019009:Proto-Oncogene Proteins c-kit / Q000032:analysis; D012333:RNA, Messenger / Q000235:genetics; D011994:Recombinant Proteins / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D012697:Serine Endopeptidases / Q000235:genetics / Q000378:metabolism*; D019089:Stem Cell Factor / Q000494:pharmacology; D053802:Tryptases
D018952:Antigens, CD34; D015850:Interleukin-6; D012333:RNA, Messenger; D011994:Recombinant Proteins; D019089:Stem Cell Factor; D019009:Proto-Oncogene Proteins c-kit; D012697:Serine Endopeptidases; D053818:Chymases; D053802:Tryptases