本文選題：雙酚A + 調(diào)節(jié)性T細(xì)胞��； 參考：《安徽醫(yī)科大學(xué)》2013年碩士論文

【摘要】：研究背景雙酚A（bispheno1A，BPA）是生產(chǎn)碳酸聚酯、塑料制品等工業(yè)產(chǎn)品的原材料，用于奶瓶、可重復(fù)用水杯、牙密封劑、食品與飲料的包裝材料、水管、熱敏紙、卡板紙及其他產(chǎn)品的添加劑等制造。據(jù)報(bào)道，2008年BPA一年的產(chǎn)量高達(dá)80億磅，而且以每年6%~10%的速度增長(zhǎng)。BPA在高溫、聚合不完全、長(zhǎng)期使用等情況下通過(guò)酯鍵水解而釋放，導(dǎo)致人群及動(dòng)物的慢性低劑量暴露。 BPA具有擬雌激素作用，通過(guò)干擾雌激素活性等途徑對(duì)人體多個(gè)系統(tǒng)如內(nèi)分泌、生殖、發(fā)育等產(chǎn)生有害作用，而孕期及生命早期（包括嬰幼兒期、青春期等）暴露BPA，可能與發(fā)育期免疫功能異常及相關(guān)疾病有關(guān)，這引起了廣泛的關(guān)注。流行病學(xué)研究提示，生命早期暴露如BPA等內(nèi)分泌干擾物可能是過(guò)去幾十年發(fā)達(dá)國(guó)家兒童哮喘、過(guò)敏性皮炎、糖尿病、兒童腫瘤發(fā)病率上升的原因之一。動(dòng)物實(shí)驗(yàn)對(duì)圍生期或成年期BPA等內(nèi)分泌干擾物暴露誘導(dǎo)的免疫毒性的研究越趨增多，但至今BPA暴露引起的免疫發(fā)育毒性機(jī)制尚未完全闡明。Treg細(xì)胞對(duì)機(jī)體發(fā)揮維持免疫穩(wěn)態(tài)和免疫負(fù)性調(diào)控的關(guān)鍵作用，有報(bào)道稱成年期BPA暴露能抑制Treg細(xì)胞數(shù)量，使T/B細(xì)胞及巨噬細(xì)胞等應(yīng)答過(guò)度，導(dǎo)致免疫功能異常及免疫性疾病。本課題組假設(shè)母鼠孕期及哺乳期BPA暴露，誘導(dǎo)的仔鼠免疫功能的異常與失衡，可能與BPA抑制Treg細(xì)胞數(shù)量這一途徑有關(guān)。 研究目的通過(guò)檢測(cè)孕期和哺乳期經(jīng)水暴露不同劑量BPA母鼠所生仔鼠斷乳期及青春期脾臟Treg細(xì)胞數(shù)量、特征性轉(zhuǎn)錄因子Foxp3mRNA表達(dá)水平及血清中IL-10、TGF-β含量，結(jié)合仔鼠生長(zhǎng)發(fā)育狀況，，探討圍生期BPA暴露對(duì)子代Treg細(xì)胞的影響。 研究方法選用SPF級(jí)8周齡ICR雌鼠、9周齡ICR雄鼠；適應(yīng)性飼養(yǎng)1周，按照雌：雄比例2:1合籠交配，雌鼠查到陰栓確認(rèn)為妊娠，將孕鼠隨機(jī)分為空白對(duì)照組、溶劑對(duì)照組（DMSO）、BPA低中高劑量組（2.2829、22.829、228.29μg/L）。母鼠自妊娠開始日（GD0）至仔鼠斷乳（PND21）經(jīng)飲水暴露BPA，PND21與PND42處死仔鼠，收集外周血并取脾臟；流式細(xì)胞儀檢測(cè)仔鼠脾臟Treg細(xì)胞數(shù)量；熒光定量RT-PCR檢測(cè)脾臟Treg細(xì)胞特征性轉(zhuǎn)錄因子Foxp3mRNA表達(dá)水平；ELISA檢測(cè)血清中IL-10、TGF-β含量；使用SPSS15.0統(tǒng)計(jì)軟件分析實(shí)驗(yàn)結(jié)果。 結(jié)果 1一般情況母鼠孕期狀況良好，未出現(xiàn)死亡、流產(chǎn)、死胎、死產(chǎn)等情況；組間飲水量差異無(wú)統(tǒng)計(jì)學(xué)意義，約為0.12ml/（g·d）；母鼠孕期體重差異無(wú)統(tǒng)計(jì)學(xué)意義。仔鼠出生性別比及平均窩仔數(shù)差異均無(wú)統(tǒng)計(jì)學(xué)意義；僅PND42時(shí)高劑量組仔鼠體重較空白對(duì)照組有顯著增加（P0.05），其余各時(shí)點(diǎn)各組間仔鼠體重差異均無(wú)統(tǒng)計(jì)學(xué)意義。 2圍生期BPA暴露對(duì)仔鼠Treg細(xì)胞的影響 2.1對(duì)仔鼠脾臟Treg細(xì)胞數(shù)量的影響圍生期BPA暴露后仔鼠脾臟Treg細(xì)胞數(shù)量隨母鼠BPA暴露劑量的增加而呈下降趨勢(shì)。PND21時(shí)中、高劑量組Treg細(xì)胞數(shù)量與對(duì)照組相比明顯降低，差異有統(tǒng)計(jì)學(xué)意義（P0.05，P0.01），而低劑量組則無(wú)明顯差異；PND42時(shí)各組間Treg細(xì)胞數(shù)量均無(wú)明顯差異，BPA暴露組仔鼠Treg細(xì)胞數(shù)量與對(duì)照組相比則表現(xiàn)為輕微下降，但與PND21相比，BPA暴露組Treg細(xì)胞數(shù)量均所有回升。 2.2對(duì)仔鼠脾臟Foxp3mRNA表達(dá)的影響暴露組仔鼠Foxp3mRNA表達(dá)水平隨母鼠BPA暴露劑量的增加而呈下降趨勢(shì)。PND21時(shí)中、高劑量組Foxp3mRNA表達(dá)水平與對(duì)照組相比有明顯降低（P0.05，P0.01），而低劑量組則無(wú)明顯差異；PND42時(shí)各組間Foxp3mRNA表達(dá)水平相比均無(wú)明顯差異，與對(duì)照組相比，BPA暴露組表現(xiàn)為輕微下降，但與PND21時(shí)相比，BPA暴露組Foxp3mRNA表達(dá)水平均所有回升。 3對(duì)Treg細(xì)胞相關(guān)細(xì)胞因子的影響 3.1對(duì)仔鼠血清IL-10水平的影響PND21時(shí)高中低劑量組仔鼠血清IL-10含量隨劑量增加而逐漸下降，高劑量組IL-10含量與對(duì)照組相比有顯著降低（P0.05），低、中劑量組與對(duì)照組相比雖有降低但差異均無(wú)統(tǒng)計(jì)學(xué)意義；PND42時(shí)各組仔鼠血清IL-10含量均無(wú)顯著差異，暴露組與對(duì)照組相比均有減少，與PND21相比則均有所回升。 3.2對(duì)血清TGF-β水平的影響PND21時(shí)高中低劑量組仔鼠血清TGF-β含量隨劑量增加而逐漸上升，高、中劑量組TGF-β含量與對(duì)照組相比有顯著增加（P0.05，P0.01），低劑量組與對(duì)照組相比差異無(wú)統(tǒng)計(jì)學(xué)意義；PND42時(shí)低、中、高劑量組與對(duì)照組相比呈逐漸增加，但差異無(wú)統(tǒng)計(jì)學(xué)意義，與PND21相比則5組均有下降，而暴露組下降幅度更為明顯。 4仔鼠血清IL-10、TGF-β水平與Treg細(xì)胞數(shù)量的相關(guān)性相關(guān)性分析顯示仔鼠血清IL-10含量與Treg細(xì)胞數(shù)量之間存在不顯著正相關(guān)；而TGF-β含量與Treg細(xì)胞數(shù)量之間存在顯著負(fù)相關(guān)。 結(jié)論 1圍生期BPA暴露對(duì)孕鼠、斷乳前仔鼠體重?zé)o明顯影響。 2圍生期暴露BPA降低仔鼠斷乳期Treg細(xì)胞數(shù)量及Foxp3mRNA表達(dá)水平。 3圍生期暴露BPA對(duì)Treg細(xì)胞相關(guān)細(xì)胞因子有影響：降低血清中IL-10含量，增加血清中TGF-β含量。 4圍生期暴露BPA，仔鼠Treg細(xì)胞數(shù)量的減少與相關(guān)細(xì)胞因子含量存在相關(guān)。
[Abstract]:Research background bisphenol A (bispheno1A, BPA) is the raw material for the production of carbonated polyesters, plastic products and other industrial products, used in bottles, reproducible cups, dental sealants, food and beverage packaging materials, water pipes, thermosensitive paper, cardboard and other products additives. It is reported that in 2008, the output of BPA was as high as 8 billion pounds, and per year. The rate of 6%~10% increased in.BPA at high temperature, incomplete polymerization, and was released by hydrolysis of ester bonds in the case of long term use, resulting in chronic low dose exposure to people and animals.
BPA has the effect of estrogenic activity by interfering with the activity of estrogen, such as endocrine, reproduction, and development, and so on. The exposure to BPA in pregnancy and early life (including infancy, puberty, etc.) may be associated with abnormal immune function and related diseases in the development period, which has aroused widespread concern. Studies have suggested that early exposure to endocrine disruptors, such as BPA, may be one of the reasons for the increase in the incidence of childhood asthma, allergic dermatitis, diabetes, and children's tumours in developed countries for the past few decades. Animal experiments have increased the immunological toxicity induced by endocrine disruptors such as perinatal or adult BPA, but up to now, BPA The mechanism of Immunogenicity Induced by exposure has not yet fully elucidated the key role of.Treg cells in maintaining immune homeostasis and negative immunological regulation. It is reported that adult BPA exposure can inhibit the number of Treg cells and excessively respond to T/B cells and macrophages and lead to immune dysfunction and immune diseases. The abnormal and unbalanced immune function induced by BPA exposure during pregnancy and lactation may be related to the inhibition of the number of Treg cells by BPA.
Objective to investigate the effect of perinatal BPA exposure on the progeny Treg cells by detecting the weaning period and the number of Treg cells in puberty spleens, the expression level of the characteristic transcription factor Foxp3mRNA and the content of IL-10, TGF- beta in the serum, and the growth and development of the offspring of BPA mice.
The study methods were SPF 8 weeks old ICR female rats and 9 weeks old ICR male rats. The female rats were bred for 1 weeks. The female rats were copulated with the male proportion of 2:1 cage. The female rats were found to be pregnant, and the pregnant rats were randomly divided into the blank control group, the solvent control group (DMSO), the low and high dose group of BPA (2.2829,22.829228.29 mu g/L). The female rats were from the beginning of pregnancy (GD0) to the offspring. PND21 was exposed to BPA by drinking water, PND21 and PND42 were sacrificed to kill the offspring, collect the peripheral blood and take the spleen; flow cytometry was used to detect the number of Treg cells in the spleen of the offspring of the offspring; the fluorescence quantitative RT-PCR was used to detect the Foxp3mRNA expression level of the characteristic transcription factors of the spleen Treg cells; ELISA detected IL-10, TGF- beta content in serum; and the analysis of the SPSS15.0 statistical software was used. Experimental results.
Result
1 normal female mice were in good condition during pregnancy. There was no death, abortion, stillbirth, and stillbirth. There was no statistical difference between the group and 0.12ml/ (G. D). There was no statistical significance in the weight difference between the female rats during pregnancy and the difference of the sex ratio and the average litter size of the offspring. The weight of the high dose group at PND42 was higher than that of the high dose group. There was a significant increase in the blank control group (P0.05).
The effect of 2 perinatal exposure to BPA exposure on Treg cells in offspring rats
2.1 effect on the number of Treg cells in spleens of the offspring rats, the number of Treg cells in the spleen of the offspring was decreased with the increase of BPA exposure dose after perinatal exposure, and the number of Treg cells in the high dose group was significantly lower than that of the control group (P0.05, P0.01), but there was no significant difference between the low dose group and the low dose group, while PND42 in the low dose group was not significantly different. There was no significant difference in the number of Treg cells in each group. The number of Treg cells in the BPA exposure group was slightly lower than that in the control group, but the number of Treg cells in the BPA exposed group all increased all compared with the PND21.
The effect of 2.2 on the expression of Foxp3mRNA in the spleens of the offspring rats, the expression level of Foxp3mRNA in the exposed group was decreased with the increase of BPA exposure dose in.PND21, and the Foxp3mRNA expression level in the high dose group was significantly lower than that in the control group (P0.05, P0.01), but there was no significant difference between the low dose group and the low dose group, and the Foxp3mRNA expression level between each group at PND42. There was no significant difference between the two groups. Compared with the control group, the BPA exposure group showed a slight decrease, but compared with the PND21 group, the expression level of Foxp3mRNA in the BPA exposure group all recovered.
The effect of 3 on Treg cell related cytokines
The effect of 3.1 on the serum IL-10 level of young mice PND21 decreased with the increase of dose in the low dose group of high and low dose group, and the content of IL-10 in high dose group was significantly lower than that of the control group (P0.05). The low dose group was lower than the control group, but the difference was not statistically significant compared with the control group, and the serum IL-10 content of each group in each group at PND42. There was no significant difference between the exposed group and the control group, and all of them rose again compared with PND21.
3.2 when the serum TGF- beta level was affected by PND21, the serum level of TGF- beta in the low dose group of high and low dose group increased with the increase of dose, and the content of TGF- beta in the middle dose group was significantly higher than that of the control group (P0.05, P0.01). The low dose group had no significant difference compared with the control group; the low dose group was lower than the control group, and the high dose group was compared with the control group. Gradually increased, but the difference was not statistically significant, compared with PND21, the 5 groups decreased, while the exposure group decreased more significantly.
The correlation analysis of the serum IL-10, TGF- beta level and the number of Treg cells in the sera of 4 offspring showed that there was no significant positive correlation between the serum IL-10 content and the number of Treg cells in the offspring, but there was a significant negative correlation between the content of TGF- beta and the number of Treg cells.
conclusion
1 the BPA exposure during perinatal period had no significant effect on the weight of pregnant rats before weaning.
2 exposure to BPA during perinatal period reduced the number of Treg cells and Foxp3mRNA expression in weaned rats.
3 exposure to BPA during perinatal period has an effect on Treg cell related cytokines: decreasing IL-10 content in serum and increasing TGF- beta content in serum.
4 during perinatal exposure to BPA, the reduction of Treg cell number in offspring rats was related to the content of related cytokines.
【學(xué)位授予單位】：安徽醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2013
【分類號(hào)】：R114

【共引文獻(xiàn)】

相關(guān)期刊論文 前1條

1 時(shí)國(guó)慶;李棟;盧曉s

本文編號(hào)：2103557