碩士 / 國立彰化師範大學 / 生物技術研究所 / 101 / Thioacetamide (TAA) is a hepatotoxin that can cause hepatocellular carcinoma in mice and zebrafish, but little is known about its toxic effect on embryonic development. In the previous study, we have showed TAA can enhanced ROS generation and induced the edema in pericardial sac (eps) formation, but the cellular mechanism was still unclear. In this study, we try to identify the cells which involved in TAA induced eps formation, and overexpression of manganese superoxide dismutase (Mn-SOD) to reduce eps formation in TAA treatment. First, we test that the toxicological mechanism of TAA on eps formation, and we treated with 32mM TAA of zebrafish embryos between 14 –36 hpf. The result show the heart beat rate 50-60%, and a narrow elongated heart malformations, these symptoms similar to heart failure, and also observed eps formation. We use of transgenic technology to build real-time in vivo dynamic analysis of ROS in liver specificity at zebrafish model, the results confirm the embryo can promote superoxide and H2O2 generated after 32 mM TAA treatment, and 32 mM TAA treatment zebrafish embryos can induction of TNF-α highly expressed and the SOD activity was inhibited. Overexpress Mn-SOD transgenic fish by Tol2 transposon system were generated and confirmed by RT-PCR, Western blotting and enzyme activity assay. The result obviously show the Mn-SOD mRNA expression increased approximately 2 fold, protein expression increased approximately 3 fold, and the activity increased approximately 1.3-1.5 fold. The transgenic F1 transgenic fish embryos were treated with TAA, the results showed that TNF-α expression is reduced to 1.5 fold, and the activity can rescue of SOD. We use antibody of anti-human MC tryptase to Immunohistochemistry, we found the mast cells were congregate and degranulate at the pericardial sac, and overexpression of Mn-SOD decrease amount of the mast cells in the pericardial sac in the number and protein oxidative damage is reduced by approximately 4 fold, the eps formation rate was reduced to about 10-20%. Our results showed TAA induced eps/heart failure-like syndrome are due to enhanced ROS generation, and induced mast cells activation during embryonic development. To overexpress Mn-SOD can reduce intracellular ROS, Mast cells numbers, and oxidative damages, and can partially rescue TAA induced embryo toxicity.
| Identifer | oai:union.ndltd.org:TW/101NCUE5108110 |
| Date | January 2013 |
| Creators | Ya-Ging Liu, 劉雅菁 |
| Contributors | Chuian - Fu Ken, 耿全福 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | zh-TW |
| Detected Language | English |
| Type | 學位論文 ; thesis |
| Format | 111 |
Page generated in 0.1209 seconds