本文關(guān)鍵詞： 低頻低能量超聲 微泡造影劑 前列腺癌 體外實(shí)驗(yàn) 化療 機(jī)制　出處：《上海交通大學(xué)》2015年博士論文　論文類型：學(xué)位論文

【摘要】：目的:研究低頻低能量超聲聯(lián)合微泡對(duì)兩種不同的人雄激素非依賴型前列腺癌細(xì)胞的化療增效作用并對(duì)其機(jī)制進(jìn)行初步探索。方法:1.流式細(xì)胞儀檢測(cè)Calcein(分子量622.53)、FD4(分子量3,000-5,000)以及FD70(分子量59,000-77,000)三種不同分子量熒光物質(zhì)進(jìn)入細(xì)胞的陽(yáng)性率;2.實(shí)驗(yàn)分4組:對(duì)照組、超聲聯(lián)合微泡組,米托蒽醌組,超聲聯(lián)合微泡+米托蒽醌組。不同分組的DU-145細(xì)胞干預(yù)后培養(yǎng)24小時(shí),使用MTT和克隆實(shí)驗(yàn)檢測(cè)細(xì)胞的增殖情況,使用Transwell小室測(cè)定細(xì)胞的遷移能力,使用透射電鏡觀察超聲聯(lián)合微泡對(duì)細(xì)胞亞顯微結(jié)構(gòu)的影響;3.實(shí)驗(yàn)分4組:對(duì)照組、超聲聯(lián)合微泡組,米托蒽醌組,超聲聯(lián)合微泡+米托蒽醌組。不同分組的PC-3細(xì)胞干預(yù)后培養(yǎng)24小時(shí),使用MTT和克隆實(shí)驗(yàn)進(jìn)行細(xì)胞增殖的檢測(cè),并與DU-145細(xì)胞結(jié)果進(jìn)行比較,使用流式細(xì)胞儀,Hoechst染色檢測(cè)細(xì)胞的凋亡情況,使用透射電鏡觀察超聲聯(lián)合微泡對(duì)細(xì)胞凋亡以及亞顯微結(jié)構(gòu)的影響,最后使用流式細(xì)胞儀檢測(cè)Calcein(分子量622.53)進(jìn)入細(xì)胞的陽(yáng)性率。結(jié)果:1.該參數(shù)的低頻低能量超聲聯(lián)合微泡可以增加三種不同分子量物質(zhì)Calcein、FD4以及FD70進(jìn)入細(xì)胞的百分比,空白對(duì)照組與超聲聯(lián)合微泡組的Calcein、FD4以及FD70熒光陽(yáng)性百分比分別為:9.74±2.55%vs 31.26±3.34%;6.15±0.93%vs 20.57±2.89%;2.80±1.68%vs 12.92±3.54%(P0.05);2.超聲聯(lián)合微泡+米托蒽醌組的DU-145細(xì)胞增殖率為50.68%±5.85%,明顯小于米托蒽醌組72.28%±4.61%(P0.05);超聲聯(lián)合微泡+米托蒽醌組的DU-145細(xì)胞集落數(shù)量為18.7±5.5,明顯小于米托蒽醌組的45.0±6.0(P0.05);超聲聯(lián)合微泡+米托蒽醌組的DU-145細(xì)胞遷移數(shù)為2.7±2.5,明顯小于米托蒽醌組的DU-145細(xì)胞遷移數(shù)17.7±7.1(P0.05);透射電鏡結(jié)果顯示低頻低能量超聲聯(lián)合微泡組的DU-145細(xì)胞的細(xì)胞質(zhì)中出現(xiàn)的自噬泡比空白對(duì)照組的多。3.超聲聯(lián)合微泡治療組的DU-145細(xì)胞的吸光度為0.7078±0.0604,與空白對(duì)照組的0.7533±0.1275無(wú)統(tǒng)計(jì)學(xué)差異(P0.05);超聲聯(lián)合微泡治療組的PC-3細(xì)胞的吸光度為0.6337±0.0577,明顯小于空白對(duì)照組的0.7413±0.1252(P0.05);超聲聯(lián)合微泡+米托蒽醌組的PC-3細(xì)胞的吸光度為0.3663±0.0493,明顯小于米托蒽醌組的0.5295±0.0668(P0.05);超聲聯(lián)合微泡+米托蒽醌組的PC-3細(xì)胞的細(xì)胞集落數(shù)量為11.0±5.0,明顯小于米托蒽醌組的41.3±4.5(P0.05);流式細(xì)胞儀檢測(cè)超聲聯(lián)合微泡組的PC-3細(xì)胞的凋亡率為9.30%±0.72%,明顯高于空白對(duì)照組的2.37%±0.71%(P0.05);Hoechst染色檢測(cè)檢測(cè)超聲聯(lián)合微泡組的PC-3細(xì)胞的凋亡率為8.17±2.77%,明顯高于空白對(duì)照組的1.80±1.30%(P0.05)。透射電鏡結(jié)果顯示超聲聯(lián)合微泡治療組的PC-3細(xì)胞出現(xiàn)凋亡細(xì)胞,同時(shí)發(fā)現(xiàn)超聲聯(lián)合微泡組的PC-3細(xì)胞的細(xì)胞質(zhì)中出現(xiàn)的自噬泡比空白對(duì)照組的多�？瞻讓�(duì)照組與超聲聯(lián)合微泡組的Calcein熒光陽(yáng)性百分比分別為:7.27±3.20%vs 28.30±4.67%(P0.05)。結(jié)論:本實(shí)驗(yàn)條件下的低頻低能量超聲聯(lián)合微泡可以增加DU-145細(xì)胞對(duì)Calcein、FD4以及FD70這三種不同分子量物質(zhì)的通透性,并且可以增加人前列腺癌DU-145細(xì)胞以及PC-3細(xì)胞的化療療效,但二者的機(jī)制是不同的,對(duì)于DU-145細(xì)胞主要是通過(guò)增加通透性實(shí)現(xiàn)的,對(duì)于PC-3細(xì)胞是通過(guò)增加通透性及誘導(dǎo)凋亡實(shí)現(xiàn)的。
[Abstract]:Objective: To study the low-frequency and low-energy ultrasound combined with microbubbles on two different human androgen independent prostate cancer cells to chemotherapy synergistic effect and preliminary exploration on its mechanism. Methods: 1. flow cytometry Calcein (molecular weight 622.53), FD4 (molecular weight 3000-5000) and FD70 (molecular weight 59000-77000). The rate of three different molecular weight fluorescent substances into cells; 2. were divided into 4 groups: control group, ultrasound and microbubble group, mitoxantrone group, ultrasound and microbubble + mitoxantrone group. The prognosis of 24 hours of training in different groups of DU-145 stem cells, proliferation of MTT and cloning assay, migration cells were measured using Transwell cell, using transmission electron microscope to observe the effect of ultrasound mediated microbubble destruction on cell ultrastructure; 3. were divided into 4 groups: control group, ultrasound and microbubble group, mitoxantrone group, ultrasound and microbubble + mitoxantrone Anthraquinone group. Prognosis of 24 hours of training in different groups of PC-3 stem cells, detected by MTT and cloning experiments of cell proliferation, and compared with the results of DU-145 cells, flow cytometry was used to detect the apoptosis of Hoechst cells were examined by transmission electron microscopy, ultrasound combined microbubble on apoptosis and ultrastructure finally, using flow cytometry Calcein (molecular weight 622.53) positive rate into cells. Results: the low frequency ultrasonic energy 1. the parameters of the microbubbles can increase three different molecular weight substances Calcein, FD4 and FD70 into the cell percentage of Calcein control group and ultrasound combined with microbubbles group FD4, FD70 and fluorescence positive percentage were 9.74 + 2.55%vs 31.26 + 3.34%; 6.15 + 0.93%vs 20.57 + 2.89%; 2.80 + 1.68%vs 12.92 + 3.54% (P0.05); 2. ultrasound combined with microbubbles + mitoxantrone group DU-145 The cell proliferation rate was 50.68% + 5.85%, significantly less than the mitoxantrone group 72.28% + 4.61% (P0.05); ultrasound combined with microbubbles + mitoxantrone group DU-145 cell colony number was 18.7 + 5.5, significantly less than the mitoxantrone group 45 + 6 (P0.05); ultrasound combined with microbubbles + DU-145 cell migration mitoxantrone group is 2.7 + 2.5, significantly less than the mitoxantrone group DU-145 cell migration number 17.7 + 7.1 (P0.05); transmission electron microscopy results showed that the absorbance of low frequency and low energy ultrasound combined with microbubbles group in the cytoplasm of DU-145 cells appeared more autophagic vacuoles than the control group.3. ultrasound combined microbubble treatment group of DU-145 cells 0.7078 + 0.0604 and 0.7533 + 0.1275 in the control group had no significant difference (P0.05); the absorbance of ultrasound combined with microbubble treatment group of PC-3 cells was 0.6337 + 0.0577, significantly less than the control group 0.7413 + 0.1252 (P0.05); ultrasound combined with microbubbles + The absorbance of the mitoxantrone group of PC-3 cells was 0.3663 + 0.0493, significantly less than the mitoxantrone group 0.5295 + 0.0668 (P0.05); ultrasound combined with microbubbles + mitoxantrone group PC-3 cell colony number was 11 + 5, significantly less than the mitoxantrone group 41.3 + 4.5 (P0.05); flow cytometry detection of ultrasound combined with micro bubble group apoptosis rate of PC-3 cells was 9.30% + 0.72%, significantly higher than the control group of 2.37% + 0.71% (P0.05); Hoechst staining detected by ultrasound micro bubble group apoptosis rate of PC-3 cells was 8.17 + 2.77%, significantly higher than the control group of 1.80 + 1.30% (P0.05) transmission. The results of SEM showed that ultrasound combined with microbubbles group PC-3 cells apoptosis, also found that ultrasound combined with microbubbles group in the cytoplasm of PC-3 cells and the control group of autophagosomes than the blank. The blank control group and ultrasound combined with microbubbles group Calcein fluorescence The positive percentage was 7.27 + 3.20%vs 28.30 + 4.67% (P0.05). Conclusion: low frequency ultrasound energy under the experimental conditions the microbubbles can increase DU-145 cells to Calcein, FD4 and FD70 of the three different molecular weight substances and can increase the permeability of human prostate cancer DU-145 cells and PC-3 cells in response to chemotherapy. But the different mechanisms of the two, for the DU-145 cell is mainly by increasing the permeability to achieve, by increasing the permeability and inducing apoptosis for PC-3 cells.

【學(xué)位授予單位】：上海交通大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2015
【分類號(hào)】：R737.25

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