Mobilized peripheral blood CD34+ cells express more amphotropic retrovirus receptor than bone marrow CD34+ cells.
OBJECTIVE
The increased susceptibility to gene transfer by amphotropic retroviral vectors of mobilized peripheral blood (PB) CD34+ cells compared to their bone marrow (BM) counterparts may depend, among other factors, on the level of expression of the amphotropic receptor on the progenitor cell. Using a previously described flow cytometry strategy, we have studied retrovirus binding to mobilized CD+ cells, derived from cancer patients treated with high-dose chemotherapy and growth factor(s), that are efficiently transduced by N2 retrovirus vector.


METHODS
We measured the binding of the retrovirus to the cells using a rat monoclonal antibody reactive with the gp70 envelope glycoprotein, common to all replication-defective amphotropic retroviruses. Antibody-virus-cell complexes were indirectly labeled and analyzed by flow cytometry. We compared the binding of PA317-N2 vector to CD34+ cells derived from steady-state BM, steady-state PB and mobilized PB from cancer patients treated with high-dose chemotherapy and cytokine.


RESULTS
The fluorescence intensity of mobilized CD34+ cells was approximately one log higher than that of steady-state BM or PB CD34+ cells, indicating that the expression of the amphotropic receptor was increased. Moreover, the virus binding was proportional to the gene transfer rate, as assessed by G418 resistance into mobilized PB-derived CFU-GM. The increase in fluorescence intensity appeared to be restricted to CD34+ cell subset, neither CD2+ nor CD14+ cells bound the virus in an appreciable amount.


CONCLUSIONS
Virus binding, as assessed by indirect immunofluorescence assay, is increased in mobilized CD34+ cells. The increased binding may contribute to their high susceptibility to retrovirus vector infection.
Bregni
M
M
Cristina Gandini Transplantation Unit, Istituto Nazionale Tumori, Milan, Italy.
Di Nicola
M
M
Siena
S
S
Belli
N
N
Milanesi
M
M
Shammah
S
S
Ravagnani
F
F
Gianni
A M
AM
D018952:Antigens, CD34 / Q000276:immunology*; D001854:Bone Marrow Cells / Q000276:immunology* / Q000378:metabolism*; D002478:Cells, Cultured / Q000821:virology; D005434:Flow Cytometry; D005822:Genetic Vectors; D019650:Hematopoietic Stem Cell Mobilization; D006801:Humans; D007937:Leukapheresis; D007962:Leukocytes / Q000276:immunology* / Q000378:metabolism*; D011991:Receptors, Virus / Q000096:biosynthesis* / Q000235:genetics; D012190:Retroviridae / Q000235:genetics / Q000502:physiology*
D018952:Antigens, CD34; D011991:Receptors, Virus