本文選題：人支氣管上皮細(xì)胞　切入點(diǎn)：納米級(jí)石英　出處：《昆明醫(yī)科大學(xué)》2012年碩士論文　論文類型：學(xué)位論文

【摘要】：[目的]研究納米級(jí)石英與苯并芘聯(lián)合作用對(duì)人支氣管上皮細(xì)胞(BEAS-2B)的影響。 [方法]本實(shí)驗(yàn)選用人支氣管上皮細(xì)胞為靶細(xì)胞,選用粒徑為30nm的納米級(jí)石英和3、4-苯并芘為研究對(duì)象,設(shè)置納米級(jí)石英的濃度分別為25mg/L、50mg/L、100mg/L、300mg/L,3、4-苯并芘的濃度為5μmol/L,進(jìn)行各物質(zhì)的單獨(dú)染毒和聯(lián)合染毒。并設(shè)置正常達(dá)爾伯克必需基本培養(yǎng)基(Dulbecco's Minimum Essential Medium, DMEM)為空白對(duì)照組。細(xì)胞培養(yǎng)6、12、24、48、72小時(shí)后,用WST-1法進(jìn)行細(xì)胞毒性試驗(yàn)。細(xì)胞培養(yǎng)6、12、24小時(shí)后用倒置相差顯微鏡對(duì)細(xì)胞形態(tài)生長(zhǎng)情況進(jìn)行觀察；用流式細(xì)胞技術(shù)測(cè)定細(xì)胞內(nèi)活性氧代謝產(chǎn)物(Reactive Oxygen Species, ROS)的表達(dá)情況；細(xì)胞培養(yǎng)6、24、48小時(shí)后用流式細(xì)胞技術(shù)測(cè)定細(xì)胞周期改變。結(jié)果應(yīng)用SPSS17.0統(tǒng)計(jì)軟件進(jìn)行統(tǒng)計(jì)學(xué)處理。 [結(jié)果]1.細(xì)胞存活率的變化納米級(jí)石英顆粒和苯并芘分別作用于人支氣管上皮細(xì)胞后,隨作用劑量增加、作用時(shí)間延長(zhǎng),細(xì)胞存活率呈下降趨勢(shì)。納米級(jí)石英和苯并芘聯(lián)合作用組與納米級(jí)石英單獨(dú)作用組相比,細(xì)胞存活率下降更快。各實(shí)驗(yàn)組與對(duì)照組比較,細(xì)胞存活率差異均具有統(tǒng)計(jì)學(xué)意義(P0.05)。2.細(xì)胞形態(tài)學(xué)的變化空白對(duì)照組細(xì)胞形態(tài)學(xué)無(wú)明顯變化。納米級(jí)石英顆粒和苯并芘分別作用于人支氣管上皮細(xì)胞后,隨作用劑量增加、作用時(shí)間延長(zhǎng),細(xì)胞體積增大。當(dāng)納米級(jí)石英與苯并芘聯(lián)合作用時(shí),細(xì)胞形態(tài)學(xué)的變化明顯增強(qiáng)。3.細(xì)胞內(nèi)ROS的表達(dá)程度納米級(jí)石英顆粒和苯并芘分別作用于人支氣管上皮細(xì)胞后,隨作用劑量增加、作用時(shí)間延長(zhǎng),與空白對(duì)照組比較,細(xì)胞內(nèi)ROS表達(dá)的平均熒光強(qiáng)度均逐漸升高；當(dāng)納米級(jí)石英與苯并芘聯(lián)合作用時(shí),細(xì)胞內(nèi)ROS表達(dá)的平均熒光強(qiáng)度明顯升高(P0.01)�？瞻讓�(duì)照組細(xì)胞內(nèi)ROS表達(dá)的平均熒光強(qiáng)度無(wú)明顯變化(P0.05)。4.細(xì)胞周期的變化培養(yǎng)6小時(shí)后聯(lián)合組細(xì)胞中DNA合成前期(G1期)細(xì)胞所占的比例為65.8±6.5%,與納米級(jí)石英組和空白對(duì)照組相比無(wú)統(tǒng)計(jì)學(xué)差異(P0.05),隨培養(yǎng)時(shí)間延長(zhǎng),聯(lián)合組與納米級(jí)石英組G1期細(xì)胞所占的比例逐漸下降,但聯(lián)合組下降趨勢(shì)更為明顯(P0.01),空白對(duì)照組G1期細(xì)胞所占的比例無(wú)明顯變化。培養(yǎng)6小時(shí)后聯(lián)合組細(xì)胞中DNA合成期(S期)細(xì)胞所占的比例為21.9±2.1%,與納米級(jí)石英組和空白對(duì)照組相比差異有統(tǒng)計(jì)學(xué)意義(P0.05),隨培養(yǎng)時(shí)間延長(zhǎng),聯(lián)合組、納米級(jí)石英組S期細(xì)胞所占的比例均有上升,但聯(lián)合組趨勢(shì)更為明顯(P0.01),空白對(duì)照組S期細(xì)胞所占的比例無(wú)明顯變化(P0.05)。 [結(jié)論]受納米級(jí)石英與苯并芘刺激后,人支氣管上皮細(xì)胞喪失原有形態(tài),細(xì)胞形態(tài)發(fā)生改變；細(xì)胞內(nèi)活性氧代謝物過(guò)度表達(dá)：細(xì)胞周期由G1向S期過(guò)渡。其作用表現(xiàn)出時(shí)間-效應(yīng)關(guān)系和劑量-效應(yīng)關(guān)系,作用時(shí)間越長(zhǎng)、作用劑量越大,其刺激作用越強(qiáng)。當(dāng)二者聯(lián)合作用時(shí),對(duì)人支氣管上皮細(xì)胞表現(xiàn)出更強(qiáng)的細(xì)胞毒性作用。
[Abstract]:[objective] to study the effect of combined action of nanometer quartz and benzopyrene on human bronchial epithelial cells (BEAS-2B). [methods] Human bronchial epithelial cells were selected as target cells, and 30nm nano-quartz and 3o 4-benzopyrene were selected as the research objects. We set the concentration of nanoscale quartz to 25 mg / L 50 mg / L 10 mg / L 10 mg / L 3 mg / L 3 mg / L 3 渭 mol / L benzopyrene 5 渭 mol / L for each substance separately and in combination, and set Dulbeccos Minimum Essential Media (DMEM), the basic medium required for normal Dalbek, as a blank control group. After cell culture, 612, 24, 4872 hours later, The cytotoxicity test was carried out by WST-1 method. The morphological growth of the cells was observed by inverted phase contrast microscope after 24 hours of cell culture, and the expression of reactive Oxygen species (ROS) was detected by flow cytometry. Cell cycle changes were measured by flow cytometry after 48 hours of cell culture. Results SPSS17.0 statistical software was used to analyze the changes of cell cycle. [results] 1. The changes of cell survival rate after treated with silica nanoparticles and benzopyrene, respectively, with the increase of the dose of action, the time of action was prolonged. The cell survival rate of the group treated with nano-quartz and benzopyrene decreased more rapidly than that of the group treated with nano-quartz alone, and the survival rate of each experimental group was higher than that of the control group. There was no significant change in cell morphology in blank control group. After treated with nanometer quartz particles and benzopyrene, the cell morphology increased with the increase of the dose. When the action time was prolonged, the cell volume increased, and when the nanoscale quartz was combined with benzopyrene, (3) the expression of ROS in cells was increased significantly. The effect of nano-quartz particles and benzopyrene on human bronchial epithelial cells increased with the increase of the dose, and the time of action was prolonged, compared with the blank control group. The average fluorescence intensity of ROS expression in cells increased gradually, and when combined with benzopyrene, The average fluorescence intensity of ROS expression in cells was significantly higher than that in control group (P 0.01). There was no significant change in the average fluorescence intensity of ROS expression in the blank control group. The cell cycle was changed after 6 hours of cell cycle change. After 6 hours of culture, the cells in the combined group were in the early stage of DNA synthesis and G1 phase. The proportion of cells was 65.8 鹵6.5. There was no significant difference in the proportion of cells between the nano-quartz group and the blank control group (P 0.05), but with the increase of culture time, there was no significant difference between the two groups. The proportion of G1 phase cells in combination group and nanometer quartz group decreased gradually. However, the decreasing trend of the combined group was more obvious than that of the control group, but the percentage of G1 phase cells in the blank control group had no significant change. After 6 hours of culture, the proportion of the cells in the DNA synthesis phase of the combined group was 21.9 鹵2.1, which was similar to that of the nano-quartz group and the blank group. The difference between the white control group and the control group was statistically significant (P 0.05). In the combined group, the percentage of S phase cells in the nano quartz group was increased, but the trend of the combined group was more obvious than that of the control group (P 0.01), but the percentage of S phase cells in the blank control group had no significant change. [conclusion] after being stimulated by nanometer quartz and benzopyrene, human bronchial epithelial cells lose their original morphology and change in morphology. Overexpression of Ros metabolites in cells: the cell cycle transitioned from G1 to S phase. The cell cycle showed a time-effect relationship and a dose-effect relationship. The longer the action time, the greater the dose. The stronger the stimulative effect is, the stronger the cytotoxic effect on human bronchial epithelial cells is when combined with them.
【學(xué)位授予單位】：昆明醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類號(hào)】：R363

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