Role of CD95/Fas and its ligand in the regulation of the growth of human CD34(++)CD38(-) fetal liver cells.
The functional significance of CD95/Fas expressed by candidate hematopoietic stem cells (HSCs) from human fetal liver was studied by testing the effect of agonistic anti-CD95 monoclonal antibody (mAb) CH-11 and soluble CD95 ligand (sCD95L) on the growth of CD34(++)CD38(-)lineage cells in vitro. Candidate fetal HSCs exhibited a dose-dependent proliferative response to CH-11 as well as to sCD95L when combined with kit ligand (KL) + interleukin 3 (IL-3) under serum-deprived culture conditions. CH-11 mAb increased, in a synergistic fashion, the number of myeloid colony-forming unit culture (CFU-C) generated by candidate HSCs in liquid cultures with the cytokine combinations KL + IL-3, KL + granulocytemacrophage colony-stimulating factor, and KL + IL-6. CH-11 mAb and sCD95L also enhanced erythropoiesis supported by KL + IL-3 + erythropoietin (Epo). Furthermore, sCD95L was able to increase the number of megakaryocytes, granulocytes, and CD34- cells generated in the presence of KL + IL-3 + Epo + thrombopoietin. An analysis performed using Western blotting revealed that the membrane-bound CD95L (mCD95L) was expressed by both immature (total CD34+/++) and mature (CD34-) hematopoietic lin(-) FL cells. Among the CD34(++)lin(-)cells, both the freshly isolated CD38+ and CD38 subsets as well as CD95+ and CD95- cells constitutively expressed mCD95L, demonstrating that the CD95/CD95L system represents a paracrine and potentially autocrine regulator of early hematopoiesis. To study the role of the endogenously produced CD95L, we determined the effects of a neutralizing anti-CD95L NOK-1 on the growth of candidate HSCs. By blocking the endogenous CD95L with NOK-1 mAb, we observed an increase in CFU-C generated by candidate HSCs. We conclude that the endogenous CD95L has an inhibitory effect on fetal candidate HSCs, which can be blocked by sCD95L and CH-11 mAb.
Bárcena
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The Fetal Treatment Center Research Laboratory, University of California, San Francisco 94143-0570, USA. barcena@itsa.ucsf.edu
Muench
M O
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Song
K S
KS
Ohkubo
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Harrison
M R
MR
D036541:ADP-ribosyl Cyclase; D051997:ADP-ribosyl Cyclase 1; D000328:Adult; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D015703:Antigens, CD; D018952:Antigens, CD34 / Q000032:analysis; D000943:Antigens, Differentiation / Q000032:analysis; D001773:Blood Cells / Q000166:cytology; D001854:Bone Marrow Cells / Q000166:cytology; D019070:Cell Lineage; D004306:Dose-Response Relationship, Immunologic; D004357:Drug Synergism; D053222:Fas Ligand Protein; D005312:Fetal Blood / Q000166:cytology; D006411:Hematopoiesis, Extramedullary / Q000187:drug effects / Q000502:physiology*; D016298:Hematopoietic Cell Growth Factors / Q000494:pharmacology; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006413:Hematopoietic System / Q000166:cytology / Q000196:embryology* / Q000254:growth & development; D006801:Humans; D007231:Infant, Newborn; D008099:Liver / Q000166:cytology* / Q000196:embryology; D008562:Membrane Glycoproteins / Q000502:physiology*; D009244:NAD+ Nucleosidase / Q000032:analysis; D009928:Organ Specificity; D011993:Recombinant Fusion Proteins / Q000494:pharmacology; D019014:fas Receptor / Q000502:physiology*
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D018952:Antigens, CD34; D000943:Antigens, Differentiation; C505661:FASLG protein, human; D053222:Fas Ligand Protein; D016298:Hematopoietic Cell Growth Factors; D008562:Membrane Glycoproteins; D011993:Recombinant Fusion Proteins; D019014:fas Receptor; D036541:ADP-ribosyl Cyclase; C498154:CD38 protein, human; D009244:NAD+ Nucleosidase; D051997:ADP-ribosyl Cyclase 1