【摘要】：研究目的： 1、通過研究硫化鋅包被硒化鎘量子點（CdSe/ZnS QDs）基本物理及光學特征，驗證其是否為一種適用于生物醫(yī)學的熒光染料。 2、通過檢測CdSe/ZnS QDs及氯化鎘（CdCl2）對Hela細胞活性及凋亡率的影響，探究CdSe/ZnS QDs是否具有細胞毒性作用。 3、通過比較空白對照組、CdSe/ZnS QDs組及CdCl2組Hela細胞內(nèi)ROS產(chǎn)量，探究CdSe/ZnS QDs誘導Hela細胞凋亡可能的機制。 4、通過對凋亡相關蛋白的檢測，進一步闡述CdSe/ZnS QDs誘導細胞凋亡的機制。 方法： 本實驗采用透射電子顯微鏡（Transmission electron microscope，TEM）、紫外分光光度計（Ultraviolet spectrophotometer）及熒光光譜儀（Fluorescencespectrometer）對CdSe/ZnS QDs進行表征。透射電子顯微鏡用于了解CdSe/ZnSQDs的大小、形狀、外觀均一程度，紫外分光光度計用于檢測CdSe/ZnS QDs的吸收光波長范圍，熒光光度計用于檢測CdSe/ZnS QDs所發(fā)射熒光波長范圍。 在建立穩(wěn)定Hela傳代細胞株，，通過MTT實驗比較不同濃度CdSe/ZnS QDs及CdCl2孵育Hela細胞24h后細胞活性的改變，再通過流式細胞術比較不同濃度CdSe/ZnS QDs及CdCl2對Hela細胞凋亡率的影響。通過對比空白對照組、CdSe/ZnS QDs組及CdCl2組Hela細胞活性及凋亡率差異，初步判斷CdSeCdSe/ZnS QDs是否具有細胞毒性。 通過DCFH-DA熒光探針檢測不同濃度CdSe/ZnS QDs及CdCl2孵育Hela細胞24h后細胞內(nèi)ROS量，并比較不同組之間ROS量的差異，探究CdSe/ZnSQDs對Hela細胞內(nèi)ROS量的影響。 最后通過Western blot檢測不同濃度CdSe/ZnS QDs及CdCl2孵育Hela細胞24h后細胞內(nèi)凋亡相關蛋白Bcl-2與Bax表達量，對比不同組間Bcl-2與Bax蛋白表達量差異分析CdSe/ZnS QDs對細胞內(nèi)Bcl-2與Bax蛋白表達的影響并分析其可能的原因。 結果： 1、本實驗所用CdSe/ZnS QDs顆粒均勻呈球形，直徑小，具有良好的單分散性和晶體結構，激發(fā)光譜寬，發(fā)射光譜連續(xù)且對稱。 2、CdSe/ZnS QDs對Hela細胞的毒性作用遠強于CdCl2，且其細胞毒性作用與給藥濃度呈正比。 3、 CdSe/ZnS QDs較CdCl2誘導Hela內(nèi)產(chǎn)生更多的ROS，且CdSeQDs/ZnS對ROS的誘導效應與給藥濃度成正比。 4、CdSe/ZnS QDs較CdCl2誘導Hela細胞凋亡效應更強，且其誘導凋亡效應與給藥濃度成正比。 5、CdSe/ZnS QDs處理組細胞內(nèi)Bcl-2蛋白表達量較空白對照組及相應CdCl2處理組少，而Bax蛋白表達較空白對照組及相應CdCl2處理組多。 結論： 1、CdSe/ZnS QDs直徑小，光學特性優(yōu)，適用于生物醫(yī)學研究。 2、CdSe/ZnS QDs具有細胞毒性作用，其毒性作用強于CdCl2，且毒性作用與給藥濃度成正比。 3、CdSe/ZnS QDs能通過促進細胞內(nèi)ROS生成而誘導Hela細胞凋亡，且CdSe/ZnS QDs能通過誘導Bax表達增加、Bcl-2表達減少而誘導Hela細胞凋亡。
[Abstract]:Objective: 1. By studying the basic physical and optical properties of zinc sulfide coated cadmium selenide quantum dots (CdSe/ZnS QDs), it is proved that it is a kind of fluorescent dye suitable for biomedicine. 2. By examining the effects of CdSe/ZnS QDs and cadmium chloride (CdCl2) on the activity and apoptosis rate of Hela cells, we investigated the cytotoxicity of CdSe/ZnS QDs. 3. By comparing ROS production in Hela cells of blank control group, CdSe/ZnS QDs group and CdCl2 group, the possible mechanism of Hela cell apoptosis induced by CdSe/ZnS QDs was explored. 4. Through the detection of apoptosis-related proteins, the mechanism of apoptosis induced by CdSe/ZnS QDs was further explained. Methods: CdSe/ZnS QDs was characterized by transmission electron microscope (Transmission electron microscope,TEM), ultraviolet spectrophotometer (Ultraviolet spectrophotometer) and fluorescence spectrometer (Fluorescencespectrometer). Transmission electron microscopy (TEM) is used to understand the size, shape and appearance of CdSe/ZnSQDs. Ultraviolet spectrophotometer is used to detect the absorption wavelength range of CdSe/ZnS QDs, and fluorescence photometer is used to detect the range of fluorescence wavelength emitted by CdSe/ZnS QDs. After establishing stable Hela cell lines, the changes of cell activity of Hela cells incubated with different concentrations of CdSe/ZnS QDs and CdCl2 for 24 hours were compared by MTT assay, and the effects of different concentrations of CdSe/ZnS QDs and CdCl2 on the apoptosis rate of Hela cells were compared by flow cytometry. By comparing the difference of Hela cell activity and apoptosis rate among control group, CdSe/ZnS QDs group and CdCl2 group, the cytotoxicity of CdSeCdSe/ZnS QDs was preliminarily determined. The amount of ROS in Hela cells incubated with different concentrations of CdSe/ZnS QDs and CdCl2 for 24 hours was detected by DCFH-DA fluorescence probe, and the difference of ROS content among different groups was compared to explore the effect of CdSe/ZnSQDs on the quantity of ROS in Hela cells. Finally, Western blot was used to detect the expression of Bcl-2 and Bax in Hela cells incubated with different concentrations of CdSe/ZnS QDs and CdCl2 for 24 hours. The effect of CdSe/ZnS QDs on the expression of Bcl-2 and Bax in different groups was analyzed and the possible causes were analyzed. Results: 1. The CdSe/ZnS QDs particles used in this experiment are spherical and small in diameter, with good monodispersity and crystal structure, wide excitation spectrum, continuous and symmetrical emission spectrum. 2 the cytotoxic effect of CDS / ZnS QDs on Hela cells was much stronger than that on CdCl2, cells, and its cytotoxicity was proportional to the drug concentration. 3. More ROS, was produced in Hela induced by CdSe/ZnS QDs than that induced by CdCl2, and the effect of CdSeQDs/ZnS on ROS was directly proportional to the drug concentration. 4CdSee / ZnS QDs was more effective than CdCl2 in inducing apoptosis of Hela cells, and the apoptotic effect was in direct proportion to the drug concentration. 5The expression of Bcl-2 protein in CDSe-ZnS QDs treated group was lower than that in control group and CdCl2 treatment group, but the expression of Bax protein was higher than that in blank control group and CdCl2 treatment group. Conclusion: 1 CdSeR / ZnS QDs has small diameter and excellent optical properties, which is suitable for biomedical research. 2CdSe- / ZnS QDs has cytotoxic effect, which is stronger than CdCl2, and is directly proportional to the concentration of the drug. 3CdSee / ZnS QDs could induce apoptosis of Hela cells by promoting the production of intracellular ROS, and CdSe/ZnS QDs could induce apoptosis of Hela cells by increasing the expression of Bax and decreasing the expression of Bcl-2.
【學位授予單位】：南昌大學
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R737.3

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