[Transfection and expression of hRI gene on human umbilical blood stem cells and gene therapy for mouse melanoma].
In order to explore the transfection and expression of hRI gene on human umbilical blood stem cells, and observe it's effect on the tumor growth. After enriching human umbilical cord blood CD34+ cells with a high-gradient magnetic cell sorting system (MACS), transfected them with supernatant of retrovirus containing human Ribonuclease inhibitor (hRI) cDNA. Hematopoietic progenitor clonogenic assay and PCR were used to evaluate transfection efficiency, and Western-blot and immune fluorescence were used to evaluate the expression quantity of hRI gene after transfection. Observe the effect of RI on the growth of melonoma in B16C57BL mice. The results showed that human umbilical blood CD34+ cells were highly purified by MACS, which made the purity of human umbilical blood CD34+ cells average 96.15%. hRI can be transfected on umbilical blood CD34+ cells, and the transfection efficiency was 35%. The positive expression of hRI gene on transfected CD34+ cells is identified by Western-blot and immune fluorescence assay. Mice injected with transfected CD34+ cells show a significant restraint of the tumor growth, a lower efficiency of tumor formation, a lower weight of the tumor and a longer incubation period of tumor formation with respect to the control groups. The results demonstrated the capacity of RI to inhibited the tumor growth by blocking the vasculature in tumor.
Deng
Yan-Jie
YJ
Department of Obstetrics and Gynecology, Second Affiliated Hospital of China Medical University, Shenyang 110004, China. yanjiedeng@yahoo.com.cn
Zhang
Shu-Lan
SL
Shang
Xi-Ying
XY
Fu
Pan-Feng
PF
Cui
Xiu-Yun
XY
D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D005312:Fetal Blood / Q000166:cytology; D015316:Genetic Therapy; D006412:Hematopoietic Stem Cells / Q000378:metabolism*; D006801:Humans; D008545:Melanoma / Q000473:pathology / Q000628:therapy*; D051379:Mice; D008810:Mice, Inbred C57BL; D009419:Nerve Tissue Proteins / Q000096:biosynthesis / Q000235:genetics*; D014162:Transfection; D014407:Tumor Cells, Cultured
D018952:Antigens, CD34; D009419:Nerve Tissue Proteins; C088463:ribonuclease inhibitor, brain