Identification of CD34+ subsets after glycoprotease selection: Engraftment of CD34+Thy-1+Lin- stem cells in fetal sheep

D. R. Sutherland, E. L. Yeo, Alexander Keith Stewart, R. Nayar, R. DiGiusto, E. Zanjani, R. Hoffman, L. J. Murray

Research output: Contribution to journalArticle

55 Citations (Scopus)

Abstract

Epitopes on the CD34 molecule detected by some CD34 antibodies can be cleaved by a unique glycoprotease from Pasteurella haemolytica, which cleaves only glycoproteins rich in O-linked glycans. A method to isolate CD34+ cells from adult bone marrow was developed subsequently, in which CD34+ cells were isolated in high purity and yield following immunomagnetic bead selection and detachment with the glycoprotease. Using a variety of other cell-surface markers shown here to be insensitive to glycoprotease, committed progenitors of T lymphoid, B lymphoid, monomyeloid, megakaryoblastic, or erythroid lineages could be identified. Significantly candidate hematopoietic stem cells (HSC) that are contained within a CD34+Lin- (CD2-, CD14-, CD15-, CD16-, CD19- (or CD34+CD38-) subset expressing the Thy-1 antigen (CDw90), c-kit receptor (CD117), and CDw109 but lacking expression of CD71 and HLA-DR antigens also were detected. Functionally distinct subsets of glycoprotease-selected CD34+ cells were identified and subfractionated using flow cytometry and fluorescence-activated cell sorting (FAGS). These subsets included candidate HSCs expressing the CD34+Thy-1+Lin- phenotype, which were sorted from a CD34+ fraction of a mobilized peripheral blood (MPB) sample. In a fetal sheep model, when CD34+Thy-1+Lin- cells were injected intraperitoneally, they were capable of homing to the marrow where they generated long-term multilineage hematopoiesis and maintained human CD34+ cells, indicating that candidate HSC subsets of CD34+ cells selected with this highly specific enzyme were capable of engraftment in vivo. The ability to identify and purify virtually any phenotypically defined subset of glycoprotease-selected CD34+ stem/progenitor cells should facilitate the study of hematopoiesis in vitro and in animal models in vivo as well as the development of novel genetic techniques for the correction of specific blood cell disorders in humans.

Original languageEnglish (US)
Pages (from-to)795-806
Number of pages12
JournalExperimental Hematology
Volume24
Issue number7
StatePublished - 1996
Externally publishedYes

Fingerprint

Fetal Stem Cells
Sheep
Hematopoiesis
Hematopoietic Stem Cells
Flow Cytometry
Stem Cells
Thy-1 Antigens
Mannheimia haemolytica
Proto-Oncogene Proteins c-kit
Genetic Techniques
HLA-DR Antigens
Bone Marrow Cells
Polysaccharides
Epitopes
Blood Cells
Glycoproteins
Animal Models
Bone Marrow
Phenotype
Antibodies

Keywords

  • CD34thy-1Lin
  • Engraftment
  • Flow cytometry
  • Glycoprotease selection

ASJC Scopus subject areas

  • Cancer Research
  • Cell Biology
  • Genetics
  • Hematology
  • Oncology
  • Transplantation

Cite this

Sutherland, D. R., Yeo, E. L., Stewart, A. K., Nayar, R., DiGiusto, R., Zanjani, E., ... Murray, L. J. (1996). Identification of CD34+ subsets after glycoprotease selection: Engraftment of CD34+Thy-1+Lin- stem cells in fetal sheep. Experimental Hematology, 24(7), 795-806.

Identification of CD34+ subsets after glycoprotease selection : Engraftment of CD34+Thy-1+Lin- stem cells in fetal sheep. / Sutherland, D. R.; Yeo, E. L.; Stewart, Alexander Keith; Nayar, R.; DiGiusto, R.; Zanjani, E.; Hoffman, R.; Murray, L. J.

In: Experimental Hematology, Vol. 24, No. 7, 1996, p. 795-806.

Research output: Contribution to journalArticle

Sutherland, DR, Yeo, EL, Stewart, AK, Nayar, R, DiGiusto, R, Zanjani, E, Hoffman, R & Murray, LJ 1996, 'Identification of CD34+ subsets after glycoprotease selection: Engraftment of CD34+Thy-1+Lin- stem cells in fetal sheep', Experimental Hematology, vol. 24, no. 7, pp. 795-806.
Sutherland, D. R. ; Yeo, E. L. ; Stewart, Alexander Keith ; Nayar, R. ; DiGiusto, R. ; Zanjani, E. ; Hoffman, R. ; Murray, L. J. / Identification of CD34+ subsets after glycoprotease selection : Engraftment of CD34+Thy-1+Lin- stem cells in fetal sheep. In: Experimental Hematology. 1996 ; Vol. 24, No. 7. pp. 795-806.
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