Characterization of apoptosis induced by grouper iridovirus in two newly established cell lines from Barramundi (Lates calcarifer)

• Main Authors: Yen-Chun Chen, 陳讌君
• Other Authors: Yu-Shen Lai
• Format: Others
• Language: zh-TW
• Published: 2008
• Online Access: http://ndltd.ncl.edu.tw/handle/01425744912026635231

碩士 === 國立宜蘭大學 === 生物技術研究所碩士班 === 96 === This study demonstares the induction of apoptosis in BM (barramundi muscle) and BSB (barramundi swim bladder) cell lines, by grouper iridovirus (GIV) infection. The apoptosis was detected and confirmed through various methods. At MOI of 10, cytopathic effect (CPE) characterized as cell shrinkage and rounding was first observed in both GIV-infected BM and BSB cells at as early as 45 min pi (post-infection). GIV infection appeared to progress more rapidly in BM cells than in BSB cell as suggested by the faster development of CPE in the infected BM cells. Cell viability assay also showed a stronger inhibitory effect on the BM (26.6%) and BSB (54.8%) cells by GIV at 4 hpi. The DNA fragmentation, Annexin V and Hoechst 33258 assays were performed at 45 min pi. The chromatin nick could be observed by TUNEL assay at 2 hpi. The above results demonstrate that BM and BSB cell lines can be triggered apoptosis by GIV. The heat-inactivated GIV and UV-inactivated GIV were used to infect cells. The results showed only UV-inactivated can induce apoptosis. Caspase-3, -8, and -9 activities in the BM- and BSB-infected cells were early activated at 30 min pi., and as the infection goes along, caspase-3, -9 had the maxima activities at 45 min pi. However, caspase-8 had the maxima activity in comparison to the mock-infected cells at 1 hpi. GIV-induce apoptosis was inhibited by a pan-caspase inhibitor, z-VAD-fmk, indicating a caspase-activation pathway. The above results showed that GIV can induce BM and BSB cells apoptosis and the apoptosis pathway is caspase-activation dependent pathway.