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Effect of acute heat stress on the protein expression in pituitary gland of female Taiwan country chickens
|關鍵字:||急性熱緊迫;腦下腺;蛋白質體;台灣土雞;acute heat stress;pituitary gland;proteome;Taiwan country chickens||引用:||王思涵。2015。台灣土雞公雞睪丸對急性熱緊迫反應之分子機制討論。博士論文。國立中興大學。台中。 白火城、吳雨新、林仁壽。2003。家畜內分泌學。藝軒圖書出版社。台北。 李淵百。1992。台灣的土雞。國立中興大學。台中。 季培元。1979。家禽生理學。台灣商務印書館發行。台北。 邱詩堯。2013。急性熱緊迫對L2品系台灣土雞肝臟蛋白質表現之影響。碩士論文。國立中興大學。台中。 陳玉如。2003。質譜技術與蛋白質體學。醫藥基因生物技術教學資源中心。台北。 陳頌方、黎培鈺、沈蕙如。2007。找尋生物指標的利器: 差異蛋白質體學分析平台之簡介。化學 65:211-224。 謝宇揚。2014。峨嵋雞、北京油雞、絲羽烏骨雞與興大選育土雞於育成期及產蛋期行為表現之比較。碩士論文。國立中興大學。台中。 戴佩珊。2004。台灣土雞產蛋性狀遺傳變異之研究。碩士論文。國立中興大學。台中。 顏正儒。2004。台灣商用土雞、選育土雞與單冠白色來航雞在濾泡生長速度與產蛋間隔時間之比較。碩士論文。國立中興大學。台中。 Abdul, K. M., K. Terada, T. Gotoh, R. M. Hafizur, and M. Mori. 2002. Characterization and functional analysis of a heart-enriched DnaJ/ Hsp40 homolog dj4/DjA4. Cell Stress Chaperones 7:156-166. Abele, D., and S. Puntarulo. 2004. Formation of reactive species and induction of antioxidant defence systems in polar and temperate marine invertebrates and fish. Comp. Biochem. Physiol. A Mol. Integr. Physiol. 138:405-415. Akiba, Y., K. Takahashi, M. 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夏季熱緊迫會造成動物體內代謝紊亂並使其生產性能下降，熱緊迫下觸發體內多個內分泌軸線反應，而腦下腺為這些軸線上游，可調控諸多生理反應，使動物耐過惡劣的環境。目前尚缺乏熱緊迫對禽類腦下腺蛋白質影響之研究，因此，本研究之目的為探討急性熱緊迫對台灣土雞腦下腺蛋白質表現之影響，以作為改善雞隻耐熱力之基礎。試驗使用30週齡國立中興大學育成之L2品系及B品系台灣土雞母雞各12隻，所有母雞先在25˚C下適應至少兩週，再接受急性熱處理。急性熱緊迫處理組L2品系接受36˚C 4小時，B品系則是38˚C 2小時之熱緊迫處理，而後於25˚C下恢復0、2及6小時，並於每個時間點犧牲採腦下腺供蛋白質分析，對照組母雞則維持於25˚C下直到採樣。二維差異膠體電泳分析結果顯示；急性熱緊迫處理組L2及B品系母雞之腦下腺分別有143及112個蛋白質表現量具有顯著差異，蛋白質胜肽質量指紋分析鑑定後可知此等差異表現蛋白質分別屬於68及54個不同蛋白質，其中分別有64及47個蛋白質點在熱緊迫處理後表現增加和68及54個蛋白質點表現降低。此外，各有11個蛋白質點表現量隨時間而波動。基因功能分析結果顯示，差異表現蛋白質主要位於細胞膜(L2: 22%，B: 26%)、細胞質(L2: 15%，B: 20%)及細胞核(L2: 18%，B: 17%)，主要與蛋白質結合(L2: 10%，B: 26%)、離子結合(L2: 10%，B: 13%)、核苷結合(L2: 12%，B: 13%)及氧化還原活性(L2: 15%，B: 7%)等分子功能相關，並參與生物過程(L2: 16%，B: 24%)、代謝過程(L2: 19%，B: 26%)、緊迫反應(L2: 15%，B: 11%)及氧化還原(L2: 13%，B: 9%)等生物過程；急性熱緊迫後L2品系母雞腦下腺差異表現蛋白質功能分析結果顯示與內分泌相關之蛋白質(CHGA、GH、GRP、POMC、PRL及TTR)、熱緊迫蛋白質(DnaJA4、HSP70、GRP78、HSP90及HSP108)、抗氧化(AKR1E2、MNSOD、PPIA、TXNDC12及TXNDC17)及細胞骨架(LOC396224、GFAP、P02552、TUBB2B、KRT19及VIM)相關之蛋白質表現量改變。急性熱緊迫後B品系母雞腦下腺蛋白質功能分析結果顯示與內分泌相關之蛋白質(CHGA、GH、GRP、POMC、PRL及TTR)、熱緊迫蛋白質(HSP70)、抗氧化(AKR1E2、PPIA、PRDX1、TXNDC12及TXNDC17)及細胞骨架(P02552、KRT19及VIM)相關之蛋白質表現量改變。由本研究結果可知，急性熱緊迫影響腦下腺內分泌功能以調節雞隻生理狀態，同時造成雞隻氧化傷害，而熱緊迫蛋白質、抗氧化蛋白質及細胞骨架蛋白質表現量增加，可保護雞隻免於受到細胞凋亡及氧化之損傷並保護雞隻耐過熱緊迫。
Heat stress causes physiological disturbances and poor productivity to animals, which are associated with multiple axis responses in the endocrine system. The pituitary gland acts at the top of endocrine axis for the thermoregulation to environment changes. However, few studies have been tried to explore the effect of heat stress on the protein expression in pituitary gland of avian species. Hence, the purpose of this study aimed to examine the effect of acute heat stress on the protein expression in pituitary glands in Taiwan country chickens (TCCs) as a base for understanding the mechanism of thermotolerance. A total of twelve 30-wk-old hens of both L2 strain and B strain TCC hens selected for over 30 years in National Chung Hsing University were used. All hens were kept in a climate chamber at 25˚C for more than two weeks as an adaption period. L2 strain and B strain hens were subject to acute heat stress of 36˚C for 4 hr and 38˚C for 2 hr, respectively. After heat exposure, hens were recovered for 0, 2, and 6 hr at 25˚C. Pituitary glands were collected for protein analysis at the end of each time point. The control group was kept at 25˚C until sample collection. Results of two-dimensional difference gel electrophoresis showed that 143 and 112 protein spots were differentially affected in the pituitary glands after acute heat stress. Peptide mass finger printing analysis classified differentially expressed protein spots into 68 and 54 distinct proteins. There were 68 and 54 protein spots upregulated, and 64 and 47 protein spots downregulated after acute heat stress, respectively. There were 11 protein spots were fluctuated at differential time points. Gene ontology analysis revealed that most of the differentially expressed proteins located in cytoplasm (L2: 22%, B: 26%), cytosol (L2: 15%, B: 20%) and nucleus (L2: 18%, B: 17%). Most of these proteins are associated with molecular functions of protein binding (L2: 10%, B: 26%), ion binding (L2: 10%, B: 13%), nucleoside binding (L2: 12%, B: 13%), and oxidoreductase activity (L2: 15%, B: 7%) and involved in biological processes of regulation of biological process (L2: 16%, B: 24%), metabolic process (L2: 19%, B: 26%), response to stimulus (L2: 15%, B: 11%), and oxidation-reduction process (L2: 13%, B: 9%). Results of functional analysis showed that the differential proteins related to endocrine hormone (CHGA, GH, GRP, POMC, PRL, and TTR), heat shock proteins (DnaJA4, HSP70, GRP78, HSP90, and HSP108), antioxidation proteins (AKR1E2, MNSOD, PPIA, TXNDC12, and TXNDC17), and cytoskeleton proteins (LOC396224, GFAP, P02552, TUBB2B, KRT19, and VIM) were changed in L2 strain hens. Results of functional analysis showed that proteins related to endocrine hormone (CHGA, GH, GRP, POMC, PRL, and TTR), heat shock protein (HSP70), antioxidation proteins (AKR1E2, PPIA, PRDX1, TXNDC12, and TXNDC17), and cytoskeleton proteins (P02552, KRT19, and VIM) were changed in B strain hens. In conclusion, acute heat stress affected the endocrine functions for regulating the physiological state and induced oxidative damage in hens. The upregulated of heat shock proteins, antioxidation proteins, and cytoskeleton proteins protect chickens from apoptosis and oxidative damage and may thus protect chickens from the injury by high temperature.
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