In vivo selection of transduced hematopoietic stem cells and little evidence of their conversion into hepatocytes in vivo.
OBJECTIVE
FMEV-type retroviral vector provides high transgene expression in hepatocytes and hematopoietic stem cells (HSCs). Here, we examined whether these vectors could provide a sufficient drug-resistance gene expression in HSCs and whether transduced HSCs could differentiate into hepatocytes in vivo.


METHODS
The CD45(+)/Lin(-) cells were transduced in vitro by FMEV-type vectors containing human O(6)-methylguanine-DNA methyltransferase (MGMT)/reporter genes and transferred into recipient mice. After the treatment with temozolomide and O(6)-benzylguanine (TMZ/BG) in vivo, we analyzed the transgene expression in peripheral blood cells by flow-cytometry. Immunohistochemistry was performed on the liver slices in partial hepatectomized recipient mice.


RESULTS
After TMZ/BG treatment, transduced host cells were enriched in recipient mice. In the liver, we observed the efficient transgene expression in many small cells along sinusoids. However, only few large cells in hepatic lobules expressed albumin. They also expressed both a transgene and a recipient marker gene, suggesting the fusion of donor HSCs with recipient hepatocytes.


CONCLUSIONS
This vector expressed a drug-resistance gene in HSCs highly enough to protect them from the drugs. But, the conversion of HSCs into hepatocytes in vivo might be a rare event in this model.
Yamaguchi
Kanji
K
Department of Clinical Molecular Biology, Faculty of Medicine, Kyoto University, Kyoto, Japan.
Itoh
Katsuhiko
K
Masuda
Tomoko
T
Umemura
Atsushi
A
Baum
Christopher
C
Itoh
Yoshito
Y
Okanoue
Takeshi
T
Fujita
Jun
J
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology; D002454:Cell Differentiation / Q000502:physiology; D002460:Cell Line; D003606:Dacarbazine / Q000031:analogs & derivatives / Q000494:pharmacology; D004351:Drug Resistance / Q000235:genetics*; D005822:Genetic Vectors; D006147:Guanine / Q000031:analogs & derivatives / Q000494:pharmacology; D018380:Hematopoietic Stem Cell Transplantation / Q000379:methods*; D006412:Hematopoietic Stem Cells / Q000187:drug effects / Q000502:physiology*; D022781:Hepatocytes / Q000166:cytology* / Q000187:drug effects; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D019853:O(6)-Methylguanine-DNA Methyltransferase / Q000235:genetics*; D000077204:Temozolomide; D014161:Transduction, Genetic / Q000379:methods*; D014183:Transplantation, Heterologous
D000970:Antineoplastic Agents; D018906:Antineoplastic Agents, Alkylating; C064976:O(6)-benzylguanine; D006147:Guanine; D003606:Dacarbazine; D019853:O(6)-Methylguanine-DNA Methyltransferase; D000077204:Temozolomide