| Budget Amount *help |
¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 2006: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 2005: ¥2,600,000 (Direct Cost: ¥2,600,000)
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| Research Abstract |
1. GFP expression in vitro
Lentivirus vector with GFP transgene was transduced to bladder cancer cell line, KU-7, and prostate cancer cell line, PC-3, to establish human bladder and prostate cancer cell lines that continuously express the GFP. KU-7-GFP and PC-3-GFP were established and GFP expression was detected by the western blotting.
2. GFP expression in vivo
(1) Subcutaneous model : KU-7, KU-7-GFP, PC-3, or PC-3-GFP was inoculated to nude mouse subcutaneously, and GFP expression was observed by Macro-Imaging system. All mice that inoculated KU-7-GFP or PC-3-GFP were recognized GFP after 1-2 weeks.
(2) Bladder orthotopic model : 5×106 of KU-7 or KU-7-GFP was injected to the bladder of nude mouse through urethra and GFP expression was observed. In all six mice, GFP expression was not detected until eight weeks. On the other hand, when KU-7-GFP was transplanted from subcutaneous near bladder, GFP expression was detected.
(3) Bone metastasis model : GFP expression was detected by imaging system, when PC-3-GFP was transplanted into the tibia of nude mice.
(4)Long term GFP expression : KU-7-GFP was cultured until forty passage in vitro, and was inoculated to subcutaneous in nude mice. GFP expression was detected at one week.
In conclusion, GFP expression of bladder and prostate cancer cell lines was able to detect in vivo and both orthotopic and metastatic model were useful for the experiment without sacrificing mice. Because the sensitivity of detection for the expression of GFP is not very high, replication competent retrovirus vector would not be detected in this system. Therefore improvement such as is light cable and camera are needed to detect the replication competent retrovirus vector.
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