[Expression of coagulation factor IX in human CD34 + hematopoietic stem cells by adeno-associated virus 2].
OBJECTIVE
To investigate the expression of human coagulation factor IX (hFIX) gene in human umbilical cord blood (CB) CD34+ cells which was transfected with recombinant adeno-associated virus 2 (rAAV-2).


METHODS
The CD34+ cells were transfected with rAAV-2/hFIX and cultured for 21 days for inducing differentiation into myeloid, erythroid and megakaryocytes, respectively. The expression of hFIX was studied at mRNA, protein and biological activity levels. The cytotoxicity of rAAV-2 to CD34+ cells was evaluated by cell proliferation, cell vitality, CD antigen expressions and CFU yields.


RESULTS
The hFIX mRNA was expressed in the cultured cells which was verified by RT-PCR and DNA sequencing. An elevated level of hFIX expression and biological activities were detected with a maximum amount of 14.10 ng/10(6) cell x 24 h. During the period of 21 day culture, the cell vitality, cell proliferation, CD antigen expression and CFU yields between the transfected and un-transfected groups had no difference(P > 0.05).


CONCLUSIONS
The human CB CD34+ cells are able to produce functional hFIX after transduction by rAAV-2/hFIX. The cell proliferation and differentiation capacities of the host CD34+ cells were not affected by rAAV-2.
Department of Hematology, Xiangya Hospital of Central South University, Changsha, China.
Yan
Y
Chen
Fang-ping
FP
Chen
Jian-qiang
JQ
Peng
Xiao-bing
XB
Wu
Guang-ping
GP
Wang
Zai-fu
ZF
Jian
Hong-ya
HY
Xin
Xin-hua
XH
Wu
D018952:Antigens, CD34; D002454:Cell Differentiation; D049109:Cell Proliferation; D002478:Cells, Cultured; D000229:Dependovirus / Q000235:genetics*; D005164:Factor IX / Q000235:genetics* / Q000378:metabolism; D005312:Fetal Blood / Q000166:cytology; D015870:Gene Expression; D005822:Genetic Vectors; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D012333:RNA, Messenger / Q000235:genetics; D014162:Transfection
D018952:Antigens, CD34; D012333:RNA, Messenger; D005164:Factor IX