Inhibit IL-8 gene expression in prostate cancer cells by transduction of recombinant baculovirus equipped with IL-8 shRNA expression cassette

Autor: Pin-Hsuan Lu, 呂品璇
Rok vydání: 2016
Druh dokumentu: 學位論文 ; thesis
Popis: 104
Prostate cancer is the sixth leading cause of cancer death in Taiwan. The androgen- independent prostate cancer (AIPC) cells constitutively express abundant amount of the Interleukin-8 (IL-8) which has been identified in term of prostate cancer developing. In this researching issue, the IL-8 expression of four types of prostate-related cells including RWPE-1, LNCaP, PC-3 and DU-145 were compared. PC-3 and DU-145 of AIPC expressed more abundant of the IL-8 than that of RWPE-1 which represents the normal prostate cells, and LNCaP which is androgen-dependent prostate cancer (ADPC) cell line. The IL-8 expressing of PC-3 was more than that of RWPE-1 by six times. Then we design five DNA fragments which could transcribe shRNA of IL-8 by the Oligoengine software. These DNA fragments were cloned into baculovirus by the platform subjected to the application of baculovirus as a mammalian gene cloning vector, the Bac-to-Bac express system, and resulted in construction of recombinant Autographa californica multiple nucleopolyhedrovirus (AcMNPV) vAcMBac-PU6-IL8-shRNA which could express short hairpin RNA (shRNA) of IL8, and vAcMBac-PU6-IL8-scRNA which could express scramble RNA (scRNA) of IL8. Since the PC-3 but not DU-145 cells received the proer transduction of baculovirus, the following studies were only conducted on PC-3 cells. The proliferation of PC-3 cells transduction with vAcMBac-PU6-IL8-shRNA(118) or vAcMBac-PU6-IL8-shRNA(338) at multiplicities of infection (m.o.i) at 750 pfu/cell were inhibited up to 40%. Western analysis showed significantly decrease of IL-8 expression in PC-3 cells infected with vAcMBac-PU6-IL8-shRNA. Both vAcMBac-PU6-IL8-shRNA(118) and vAcMBac-PU6-IL8-shRNA(338) disturbed the PC-3 cells by arresting the cell cycle at G2/M and S phase respectively. The conclusion of these studies might described as following: baculovirus was regarded as potential vector candidate for mammalian gene delivery to inhibit PC-3 cell proliferation in consequence of expressing shRNA of IL-8 to inhibit IL-8 gene activation in prostate cancer cells. This strategy could be an alternative approach of gene therapy for prostate cancer.
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