褪黑素對雙酚A致成年雄性大鼠生殖細胞DNA損傷的拮抗作用及機制研究
發(fā)布時間：2018-09-04 08:06
【摘要】：目的：環(huán)境內(nèi)分泌干擾物雙酚A（BPA）是工業(yè)生產(chǎn)環(huán)氧樹脂、聚碳酸酯和聚苯乙烯樹脂等的前體物質(zhì)。因加入BPA可以使聚碳酸酯等塑料產(chǎn)品變得無色透明、耐用和防摔，BPA被廣泛應(yīng)用于嬰幼兒奶瓶、食品包裝材料、塑料餐具和醫(yī)療器械等塑料行業(yè)。塑料制品的大量使用使得BPA在我們的生活中廣泛存在。因BPA的化學(xué)結(jié)構(gòu)與明確的人類致癌物己烯雌酚相似，BPA可能的遺傳毒性已經(jīng)引起了國內(nèi)外學(xué)者的廣泛關(guān)注。然而，關(guān)于BPA遺傳毒性的研究結(jié)果仍存在爭議，至今尚未得到統(tǒng)一的結(jié)論。目前，有關(guān)BPA對雄性大鼠生殖細胞遺傳毒性的研究報道相對較少。本研究以BPA為研究對象，通過建立BPA亞急性暴露的動物實驗?zāi)Ｐ吞剿鰾PA暴露對成年期SD雄性大鼠生殖細胞的遺傳毒性和抗氧化劑褪黑素（MT）對BPA所致毒性效應(yīng)的拮抗作用，以探討其可能的機制。 方法：SPF級8周齡健康雄性SD大鼠，隨機分為：(1)正常對照組；（2）MT處理組，，MT10mg/kg體重劑量腹腔注射；（3）BPA處理組，BPA200mg/kg體重劑量經(jīng)口灌胃染毒；（4）MT預(yù)處理+BPA組，大鼠經(jīng)10mg/kg的MT預(yù)處理30min后再灌胃200mg/kg劑量的BPA。每組10只動物，連續(xù)處理10天。末次給藥24h后，硫代巴比妥酸法測定睪丸組織中MDA含量，黃嘌呤氧化酶法測定SOD活力；分離睪丸精母細胞，堿性彗星實驗檢測精母細胞DNA損傷，染色質(zhì)擴散免疫染色γH2AX檢測粗線期精母細胞常染色體上γH2AX陽性焦點數(shù)量；流式細胞檢測睪丸細胞群中不同DNA含量亞細胞群的細胞數(shù)量；TUNEL熒光染色檢測生殖細胞凋亡；睪丸組織HE染色觀察睪丸組織病理形態(tài)學(xué)變化。 結(jié)果：①200mg/kg體重劑量的BPA暴露10天對SD大鼠生長無明顯抑制作用，大鼠體重增量、生殖器官睪丸和附睪的重量及精子數(shù)量均未出現(xiàn)明顯變化（P＞0.05）。 ②200mg/kg體重劑量的BPA處理SD大鼠10天引起了氧化損傷，睪丸組織中MDA含量升高， SOD活力降低（P＜0.01）。 ③BPA暴露引起了睪丸精母細胞DNA損傷。精母細胞彗星參數(shù)TL, Tail DNA%, TM和OTM與對照組比較均顯著增加（P＜0.01）；SD大鼠粗線期精母細胞常染色體上γH2AX陽性焦點數(shù)量與對照組比較明顯增多（P＜0.01）。 ④BPA處理顯著降低了睪丸細胞群中4C-DNA含量亞細胞群的細胞數(shù)量（P＜0.05）。尚未引起生殖細胞凋亡增多和睪丸組織病理形態(tài)學(xué)的變化。 ⑤10mg/kg體重劑量的MT對SD大鼠進行預(yù)處理后減輕了氧化損傷，和BPA處理組比較睪丸組織內(nèi)MDA含量降低，SOD活力升高（P＜0.05）。 ⑥MT預(yù)處理拮抗了BPA暴露引起的精母細胞DNA損傷。顯著降低了DNA損傷的程度（P＜0.01）和粗線期精母細胞常染色體上γH2AX陽性焦點的數(shù)量（P＜0.05）；并增加了睪丸細胞群中4C-DNA含量亞細胞群的細胞數(shù)量，與單獨的BPA處理組比較，差異有統(tǒng)計學(xué)意義（P＜0.05）。 結(jié)論：200mg/kg BPA亞急性暴露10天可引起大鼠精母細胞DNA損傷和生殖細胞比例的改變，但尚未引起細胞周期阻滯、生殖細胞凋亡和睪丸組織形態(tài)學(xué)變化等嚴重的DNA損傷不可修復(fù)的應(yīng)答事件。BPA誘發(fā)的DNA損傷與睪丸組織中MDA含量升高，SOD活力降低有關(guān)。MT10mg/kg體重劑量預(yù)處理則可拮抗BPA誘發(fā)的損傷。結(jié)果提示，氧化應(yīng)激可能是BPA導(dǎo)致生殖細胞DNA損傷的機制之一，在BPA環(huán)境和職業(yè)暴露所致遺傳毒性的防治藥物中，MT具有潛在的應(yīng)用價值。
[Abstract]:OBJECTIVE: Bisphenol A (BPA), an environmental endocrine disruptor, is a precursor in the industrial production of epoxy resin, polycarbonate and polystyrene resins. BPA is widely used in the plastics of baby bottles, food packaging materials, plastic tableware and medical instruments because BPA can make polycarbonate and other plastic products colorless, transparent, durable and fall-proof. Material industry. The extensive use of plastic products makes BPA widespread in our lives. Due to the chemical structure of BPA is similar to the clear human carcinogen diethylstilbestrol, the possible genetic toxicity of BPA has attracted wide attention of scholars at home and abroad. However, the results of the study on the genetic toxicity of BPA are still controversial and have not yet been unified. Conclusion. At present, there are relatively few reports about the genetic toxicity of BPA on male rat germ cells. This study was conducted to explore the genetic toxicity of BPA exposure to adult SD male rats germ cells and the toxic effects of antioxidant melatonin (MT) on BPA by establishing an animal model of subacute exposure to BPA. The antagonistic effect was explored to explore its possible mechanism.
METHODS: SPF grade 8-week-old healthy male SD rats were randomly divided into: (1) normal control group; (2) MT treatment group, MT 10 mg/kg body weight dose intraperitoneal injection; (3) BPA treatment group, BPA 200 mg/kg body weight dose by oral administration; (4) MT pretreatment + BPA group, rats were pretreated with 10 mg/kg MT for 30 minutes and then gavaged with 200 mg/kg BPA. Twenty-four hours after the last administration, MDA content in testicular tissue was measured by thiobarbituric acid, SOD activity was measured by xanthine oxidase; testicular spermatocytes were isolated, DNA damage in spermatocytes was detected by alkaline comet assay, and the number of gamma H2AX positive foci on pachytene spermatocytes was detected by chromatin diffuse immunostaining (CDI). TUNEL fluorescence staining was used to detect the apoptosis of germ cells and HE staining was used to observe the pathological changes of testicular tissue.
RESULTS: 1. The growth of SD rats was not significantly inhibited after exposure to BPA at 200 mg/kg body weight for 10 days. There were no significant changes in body weight gain, testicular and epididymal weight of reproductive organs and sperm number (P > 0.05).
(2) BPA treatment with 200 mg/kg body weight caused oxidative damage in SD rats for 10 days, MDA content in testicular tissue increased and SOD activity decreased (P < 0.01).
(3) BPA exposure induced DNA damage in testicular spermatocytes. The comet parameters TL, Tail DNA, TM and OTM of spermatocytes were significantly increased compared with the control group (P < 0.01), and the number of positive focal gamma H2AX on the pachytene spermatocytes of SD rats was significantly increased compared with the control group (P < 0.01).
(4) BPA treatment significantly decreased the number of 4C-DNA subpopulations in testicular cell population (P
MT at 10 mg/kg body weight reduced oxidative damage in SD rats after pretreatment. Compared with BPA treatment group, MDA content in testicular tissue decreased and SOD activity increased (P < 0.05).
_MT pretreatment antagonized DNA damage in spermatocytes induced by BPA exposure, significantly reduced the degree of DNA damage (P < 0.01) and the number of gamma H2AX positive foci on pachytene spermatocytes (P Academic significance (P < 0.05).
CONCLUSION: Subacute exposure of 200 mg/kg BPA for 10 days can induce DNA damage in rat spermatocytes and changes in the proportions of germ cells, but it has not yet caused irreparable events such as cell cycle arrest, germ cell apoptosis and histomorphological changes of testis. The results suggest that oxidative stress may be one of the mechanisms of DNA damage in germ cells induced by BPA. MT has potential application value in the prevention and treatment of genetic toxicity induced by BPA environmental and occupational exposure.
【學(xué)位授予單位】：重慶醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2013
【分類號】：R114
[Abstract]:OBJECTIVE: Bisphenol A (BPA), an environmental endocrine disruptor, is a precursor in the industrial production of epoxy resin, polycarbonate and polystyrene resins. BPA is widely used in the plastics of baby bottles, food packaging materials, plastic tableware and medical instruments because BPA can make polycarbonate and other plastic products colorless, transparent, durable and fall-proof. Material industry. The extensive use of plastic products makes BPA widespread in our lives. Due to the chemical structure of BPA is similar to the clear human carcinogen diethylstilbestrol, the possible genetic toxicity of BPA has attracted wide attention of scholars at home and abroad. However, the results of the study on the genetic toxicity of BPA are still controversial and have not yet been unified. Conclusion. At present, there are relatively few reports about the genetic toxicity of BPA on male rat germ cells. This study was conducted to explore the genetic toxicity of BPA exposure to adult SD male rats germ cells and the toxic effects of antioxidant melatonin (MT) on BPA by establishing an animal model of subacute exposure to BPA. The antagonistic effect was explored to explore its possible mechanism.
METHODS: SPF grade 8-week-old healthy male SD rats were randomly divided into: (1) normal control group; (2) MT treatment group, MT 10 mg/kg body weight dose intraperitoneal injection; (3) BPA treatment group, BPA 200 mg/kg body weight dose by oral administration; (4) MT pretreatment + BPA group, rats were pretreated with 10 mg/kg MT for 30 minutes and then gavaged with 200 mg/kg BPA. Twenty-four hours after the last administration, MDA content in testicular tissue was measured by thiobarbituric acid, SOD activity was measured by xanthine oxidase; testicular spermatocytes were isolated, DNA damage in spermatocytes was detected by alkaline comet assay, and the number of gamma H2AX positive foci on pachytene spermatocytes was detected by chromatin diffuse immunostaining (CDI). TUNEL fluorescence staining was used to detect the apoptosis of germ cells and HE staining was used to observe the pathological changes of testicular tissue.
RESULTS: 1. The growth of SD rats was not significantly inhibited after exposure to BPA at 200 mg/kg body weight for 10 days. There were no significant changes in body weight gain, testicular and epididymal weight of reproductive organs and sperm number (P > 0.05).
(2) BPA treatment with 200 mg/kg body weight caused oxidative damage in SD rats for 10 days, MDA content in testicular tissue increased and SOD activity decreased (P < 0.01).
(3) BPA exposure induced DNA damage in testicular spermatocytes. The comet parameters TL, Tail DNA, TM and OTM of spermatocytes were significantly increased compared with the control group (P < 0.01), and the number of positive focal gamma H2AX on the pachytene spermatocytes of SD rats was significantly increased compared with the control group (P < 0.01).
(4) BPA treatment significantly decreased the number of 4C-DNA subpopulations in testicular cell population (P
MT at 10 mg/kg body weight reduced oxidative damage in SD rats after pretreatment. Compared with BPA treatment group, MDA content in testicular tissue decreased and SOD activity increased (P < 0.05).
_MT pretreatment antagonized DNA damage in spermatocytes induced by BPA exposure, significantly reduced the degree of DNA damage (P < 0.01) and the number of gamma H2AX positive foci on pachytene spermatocytes (P Academic significance (P < 0.05).
CONCLUSION: Subacute exposure of 200 mg/kg BPA for 10 days can induce DNA damage in rat spermatocytes and changes in the proportions of germ cells, but it has not yet caused irreparable events such as cell cycle arrest, germ cell apoptosis and histomorphological changes of testis. The results suggest that oxidative stress may be one of the mechanisms of DNA damage in germ cells induced by BPA. MT has potential application value in the prevention and treatment of genetic toxicity induced by BPA environmental and occupational exposure.
【學(xué)位授予單位】：重慶醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2013
【分類號】：R114
【參考文獻】
相關(guān)期刊論文 前4條
1 武紅娟;陳于;趙奇;王艷;劉露;王帥;;褪黑素對丙烯酰胺致大鼠睪丸細胞DNA損傷的拮抗作用[J];上海交通大學(xué)學(xué)報(醫(yī)學(xué)版);2012年08期
2 逄兵,吳向東,任道風(fēng),金泰^
本文編號：2221497
本文鏈接：http://sikaile.net/yixuelunwen/yufangyixuelunwen/2221497.html
最近更新
教材專著
