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研究生:楊致豪
研究生(外文):Chih-hao Yang
論文名稱:暴露低劑量PCB 153和PCB 118影響第三子代雄性大鼠發育和生殖功能之研究
論文名稱(外文):The transgenerational effects of low-level exposure of PCB 153 and PCB 118 on developmental and reproductive outcomes in F3 male offspring
指導教授:許昺奇許昺奇引用關係
學位類別:碩士
校院名稱:國立高雄第一科技大學
系所名稱:環境與安全衛生工程研究所
學門:工程學門
學類:環境工程學類
論文種類:學術論文
論文出版年:2011
畢業學年度:99
語文別:中文
論文頁數:58
中文關鍵詞:超代作用多氯聯苯153生殖功能
外文關鍵詞:transgenerational actionF3PCB153reproductive function
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  多氯聯苯 153(2,2’,4,4’,5,5’-hexachlorobiphenyl, PCB 153)為人體脂肪組織常見之雙鄰位多氯聯苯同源物;多氯聯苯 118(2,3’,4,4’,5-pentachlorobiphebyl, PCB 118)則是被WHO之毒性作用歸類為Group I之重要指標之一。本研究於大鼠暴露後,分析第三雄性子代影響生長發育或生殖功能之評估。
  母鼠懷孕第8天到第15天每天以胃管餵食暴露,經過第一子代(F1)交錯繁殖,產下第二子代(F2)分別為CC組(父母皆控制組)、CH組(父控制組;母暴露組)、HC組(父暴露組;母控制組)與HH組(父母皆為暴露組)。F2公鼠再與空白母鼠交配,產下第三子代(F3)。本研究F3主要探討CC組和HC組在離乳前後觀察生長發育和量測體重與尿道到肛門口距離(AGD),並於第112天進行犧牲,分析精子功能等。
研究結果發現暴露於PCB 153造成F3生長發育上開眼有較為遲緩(p<0.05);離乳前經校正後AGD暴露組有明顯較短(p<0.05);離乳後校正後AGD暴露組有明顯較長(p<0.05);在生殖功能方面,暴露組儲精囊則有較輕(p<0.05);精子數增加(p<0.05);αT、COMP αT和O2-.傷害均有明顯增高(p<0.05);精子型態則是無勾增加(p<0.05);睪丸DNA含量二倍體明顯增加(p<0.05)。
PCB 118方面,長毛暴露組有遲緩現象(p<0.05);離乳後校正後AGD暴露組明顯較短(p<0.05);生殖功能方面,暴露組MMP (mitochondrial membrane potential)則是比控制組高;暴露組睪丸DNA含量瘦長形精細胞比控制組來的高,在二倍體和四倍體則來的低。
  本研究發現母鼠懷孕時期暴露低劑量PCB 153和PCB 118,可能具有超代作用,影響雄性F3之生殖功能。未來研究應針對PCBs暴露時間與PCBs如何引起父親精子細胞遺傳性之改變而導致下一代疾病的發生機轉再進一步釐清。
2,2’,4,4’,5,5’-hexachlorobiphenyl (PCB153) is one of the two-ortho congeners of PCBs and being found more often in human adipose tissue. 2,3’,4,4’,5-pentachlorobiphebyl, (PCB 118) is classified by WHO as the toxic effects of an important indicator of the Group I. The objective is to evaluate the transgenerational effects of low-level exposure of PCB 153 on developmental and reproductive outcomes in F3 male offspring.
The female rats were gavaged per day between gestational day (GD) 8 and 15. The F1 control males were bred to F1 control females to generate the F2 control generation (CC group). F1 control males were bred to F1 treated females to generate the F2 reverse outcross generation (CH group) and F1 treated males were bred to F1 control females to generate the F2 outcross generation (HC group). F1 treated females to generate the F2 treated generation (HH group). The F2 male were bred to control females to generate the F3 offspring and F3 were sacrificed at PND 112 to evaluate testicular and sperm functions.
We found in F3 rat offspring, the time of eye opening were significantly delayed in PCB 153 exposed group. Adjusted AGD of PCB 153 exposed group was shorter than CC group before weaning, however, adjusted AGD of PCB 153 exposed group was longer than CC group after weaning. In reproductive function respect of F3, a decrease of seminal vesicle weight in PCB 153 exposed group was observed (p<0.05). The sperm count was significantly increased in PCB 153 exposed group (p<0.05). The αT, COMP αT and super oxide anion (O2-.) were significantly increased in PCB 153 exposed group (p<0.05). In morphology, flattened head was significantly increased in PCB 153 exposed group (p<0.05). The percentage of diploid cells was significantly increased in PCB 153 exposed group than in control (p<0.05).
In PCB 118 group, the time of incisor eruption occurrence was delayed in HC group (p<0.05). The length of adjusted AGD was significantly shorter than control after weaning (p<0.05). In reproductive function respect, the sperm mitochondrial membrane potential (MMP) was significantly increased in PCB 118 exposed group (p<0.05). The percentage of elongated spermatids (ES) was significantly increased in PCB 118 exposed group (p<0.05), but in diploid and tetraploid was significantly decreased in PCB 118 exposed group (p<0.05).
In summary, gestational exposure to low level of PCB 153 and PCB 118 might alter sperm quality in F3 male offspring via tansgenerational effects. Further studies are required to determine the crucial time of exposure of PCBs, and to identify factors that result in germline-transmitted adult-onset diseases and given the implications of heritable effects of paternal exposure of PCBs could be considerable.
摘要 ................................................................................................................................... I
Abstract ........................................................................................................................... III
誌謝 .................................................................................................................................. V
目錄 ................................................................................................................................ VI
表目錄 .......................................................................................................................... VIII
圖目錄 ............................................................................................................................ IX
專有名詞縮寫與中英文對照 .......................................................................................... X
一、緒論 .......................................................................................................................... 1
1.1 前言 ................................................................................................................... 1
二、文獻探討 .................................................................................................................. 3
2.1 多氯聯苯介紹 ................................................................................................... 3
2.1.1 多氯聯苯概論 ........................................................................................ 3
2.1.2 多氯聯苯PCB 153與PCB 118之危害特性 ....................................... 5
2.3 超代作用 ........................................................................................................... 6
2.4 雄性生殖系統介紹 ........................................................................................... 7
2.4.1 雄性生殖機能 ........................................................................................ 7
2.4.2 精子生成步驟與精子構造 .................................................................... 8
2.4.3 細胞週期與睪丸DNA含量關係 ....................................................... 11
2.5 反應性氧化物質 ............................................................................................. 12
2.5.1 何謂反應性氧化物質 .......................................................................... 12
2.5.2 反應性氧化物質對精子功能的影響 .................................................. 13
三、研究設計與方法 .................................................................................................... 15
3.1 研究設計 ......................................................................................................... 15
3.2 動物來源與處理 ............................................................................................. 18
3.3 雄性生殖毒理評估 ......................................................................................... 19
3.3.1 藥品與詴劑 .......................................................................................... 19
3.3.2 儀器設備 .............................................................................................. 22
3.3.3 副睪精子及睪丸單細胞懸浮液之配製 .............................................. 24
3.3.4 精子型態評估 ...................................................................................... 24
3.3.5 精子核染色質結構分析 ...................................................................... 25
3.3.6 精子粒線體功能 .................................................................................. 26
3.3.7 精子細胞內過氧化氫(H2O2) ............................................................... 27
3.4.8 精子細胞內超氧陰離子( ? -
2 O ) ............................................................ 28
3.3.9 睪丸細胞DNA 含量分析 ................................................................... 29
3.3.10 睪丸組織切片 .................................................................................... 30
3.4 流式細胞儀 ..................................................................................................... 32
3.4.1 工作原理 .............................................................................................. 32
3.4.2 分析原理 .............................................................................................. 32
3.5 化學冷光分析 ................................................................................................. 35
3.6 統計分析 ......................................................................................................... 36
四、結果 ........................................................................................................................ 37
4.1 第二母代(F2)生殖影響評估 ........................................................................... 37
4.2 第三(F3)雄性子代離乳前影響評估 ............................................................... 38
4.2.1 離乳前生長發育觀察 .......................................................................... 38
4.2.2 離乳前體重與AGD ............................................................................ 39
4.2.2 離乳後體重與AGD ............................................................................ 41
4.3 第三雄性子代(F3)生殖功能影響評估 ........................................................... 42
4.3.1 F3 體重、組織器官重量與睪固酮 ....................................................... 42
4.3.2 精子功能 .............................................................................................. 44
4.3.3 精子型態 .............................................................................................. 46
4.3.4 F3 睪丸細胞週期DNA 含量 ................................................................ 47
五、討論 ........................................................................................................................ 48
六、結論 ........................................................................................................................ 54
參考文獻 ........................................................................................................................ 55
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