本文選題：硫酸銦 + DNA損傷��； 參考：《蘇州大學(xué)》2012年碩士論文

【摘要】：目的：銦（indium,In）是一種稀散金屬，在信息宇航能源軍事工業(yè)及醫(yī)藥衛(wèi)生領(lǐng)域均有著很廣泛的應(yīng)用。隨著銦及其化合物的使用增多，，接觸人群也在不斷擴(kuò)大。通過(guò)各種途徑接觸吸收的銦通過(guò)血液轉(zhuǎn)運(yùn)到軟組織及骨骼，并隨尿及糞便排出體外。本研究旨在了解硫酸銦的細(xì)胞毒性DNA損傷及細(xì)胞毒性機(jī)制，試圖探討硫酸銦可能的細(xì)胞損傷機(jī)制，為提出有效保護(hù)職業(yè)暴露人群健康的措施提供基礎(chǔ)依據(jù)。 方法：(1)以小鼠成纖維細(xì)胞（L929）作為實(shí)驗(yàn)系統(tǒng)，采用四甲基偶氮唑鹽法（MTT比色法）檢測(cè)不同濃度硫酸銦對(duì)L929細(xì)胞作用不同時(shí)間段后，細(xì)胞存活率的變化，了解濃度-時(shí)間-毒性效應(yīng)的關(guān)系；（2）采用單細(xì)胞凝膠電泳實(shí)驗(yàn)（SCGE彗星實(shí)驗(yàn)）研究不同濃度硫酸銦對(duì)L929細(xì)胞DNA單鏈斷裂損傷的程度；（3）采用免疫熒光技術(shù)研究不同濃度硫酸銦對(duì)L929細(xì)胞DNA雙鏈斷裂損傷的程度；（4）用流式細(xì)胞技術(shù)DCFH-DA法檢測(cè)不同濃度硫酸銦處理L929細(xì)胞后活性氧（ROS）含量的變化；（5）流式細(xì)胞術(shù)用JC-1檢測(cè)不同濃度硫酸銦作用下對(duì)L929細(xì)胞線(xiàn)粒體膜電位的改變。 結(jié)果：（1）MTT法顯示，在24h及48h處理過(guò)程中，當(dāng)硫酸銦終濃度1mmol/L及以上，各染毒組吸光度值與陰性對(duì)照組相比較具有統(tǒng)計(jì)學(xué)差異（P 0.05）。隨染毒濃度增加，細(xì)胞存活率也呈濃度依賴(lài)性降低。（2）單細(xì)胞凝膠電泳結(jié)果顯示，在1～4mmol/L硫酸銦作用2h及12h后，彗星細(xì)胞拖尾率隨著濃度的增加而增加，且有統(tǒng)計(jì)學(xué)意義（P 0.05）。其他分析指標(biāo)彗星尾長(zhǎng)、Olive尾矩和彗尾DAN含量也明顯高于陰性對(duì)照組(P 0.05)。（3）免疫熒光實(shí)驗(yàn)顯示：在1～6mmol/L硫酸銦作用2h后，細(xì)胞核內(nèi)γH2AX熒光強(qiáng)度明顯增強(qiáng)，焦點(diǎn)數(shù)增加，與陰性對(duì)照組比較差異具有統(tǒng)計(jì)學(xué)意義（P 0.05）。（4）流式細(xì)胞儀檢測(cè)結(jié)果顯示，隨硫酸銦染毒濃度升高，活性氧（ROS）含量相應(yīng)升高并呈現(xiàn)線(xiàn)性相關(guān)關(guān)系。（5）經(jīng)流式細(xì)胞儀檢測(cè)，在硫酸銦染毒12h條件下，隨染毒濃度增加線(xiàn)粒體膜電位水平降低，且差異具有統(tǒng)計(jì)學(xué)意義(P 0.05)。 結(jié)論：（1）在體外培養(yǎng)的條件下，硫酸銦能明顯抑制L929細(xì)胞的增殖。在各個(gè)染毒時(shí)間段內(nèi)細(xì)胞存活率隨染毒濃度增加而降低，并呈濃度依賴(lài)關(guān)系。（2）1mmol/L硫酸銦染毒2h可引起細(xì)胞DNA單鏈斷裂。（3）在2h染毒條件下硫酸銦可引起細(xì)胞DNA雙鏈斷裂。（4）硫酸銦可引起L929細(xì)胞內(nèi)活性氧含量增加。（5）硫酸銦可引起L929細(xì)胞線(xiàn)粒體膜電位降低。本次研究結(jié)果顯示，硫酸銦可引起體外培養(yǎng)的L929細(xì)胞活性氧含量增加，造成DNA損傷，同時(shí)引起膜電位降低，影響線(xiàn)粒體穩(wěn)定性。
[Abstract]:Objective: indium Inis is a dilute metal in information? Space? energy Military industry and medical and health fields have a very wide range of applications. As the use of indium and its compounds increases, the number of people exposed to it is expanding. Indium absorbed through various ways is transported through blood to soft tissues and bones and discharged with urine and faeces. The purpose of this study was to understand the cytotoxic DNA damage and cytotoxic mechanism of indium sulfate, and to explore the possible mechanism of cell damage in indium sulfate, and to provide the basis for effective measures to protect the health of occupational exposed population. Methods Mouse fibroblasts (L929) were used as experimental system. MTT colorimetric assay was used to detect the survival rate of L929 cells treated with different concentrations of indium sulfate for different time periods. To understand the relationship between concentration, time and toxicity, the single cell gel electrophoresis (SCGE) assay and comet assay were used to study the damage degree of DNA single strand break in L929 cells with different concentrations of indium sulfate. (3) Immunofluorescence technique was used to study the degree of DNA double strand break damage in L929 cells treated with different concentrations of indium sulfate. Flow cytometry (DCFH-DA) was used to detect the changes of reactive oxygen species (Ros) content in L929 cells treated with different concentrations of indium sulfate. Flow cytometry was used to detect the mitochondrial membrane potential of L929 cells treated with different concentrations of indium sulfate. Results in 24 h and 48 h treatment, when the final concentration of indium sulfate was 1 mmol / L or above, the absorbance value of each group was significantly different from that of negative control group (P 0.05). The survival rate of comet cells decreased in a concentration-dependent manner with the increase of concentration. The results of single cell gel electrophoresis showed that the trailing rate of comet cells increased with the increase of concentration after 2 h and 12 h exposure to 1 mol / L indium sulfate, and there was statistical significance (P 0.05). Olive tail moment and DAN content in comet tail were also significantly higher than those in negative control group (P 0.05). The results of immunofluorescence assay showed that the fluorescence intensity of 緯 H 2AX and the number of focal points in the nucleus were significantly increased after 2 h exposure to 1 渭 mol / L indium sulfate. The results of flow cytometry showed that the content of reactive oxygen species rose with the increase of indium sulfate concentration and showed a linear correlation with flow cytometry. When treated with indium sulfate for 12 h, the level of mitochondrial membrane potential decreased with the increase of concentration of indium sulfate, and the difference was statistically significant (P 0.05). Conclusion in vitro culture, indium sulfate can significantly inhibit the proliferation of L929 cells. The cell survival rate decreased with the increase of the exposure concentration during each exposure period. In a concentration-dependent manner, 1 mmol / L indium sulfate could induce single strand breaks of DNA in L929 cells for 2 h.) under the condition of 2 h exposure, indium sulfate could induce double strand breaks of DNA. 4) indium sulfate could increase the content of reactive oxygen species in L929 cells. Mitochondrial membrane potential of L 929 cells decreased. The results showed that indium sulfate could increase the content of reactive oxygen species in cultured L929 cells, cause DNA damage and decrease the membrane potential and affect the stability of mitochondria.
【學(xué)位授予單位】：蘇州大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類(lèi)號(hào)】：R114

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