本文選題：脈沖高強(qiáng)度聚焦超聲 + 微泡造影劑��； 參考：《重慶醫(yī)科大學(xué)》2007年碩士論文

【摘要】： 背景和目的高強(qiáng)度聚焦超聲(high intensity focused ultrasound,HIFU)是一種新興的非侵入性局部治療技術(shù)。HIFU治療腫瘤的機(jī)制除了熱機(jī)制外,還有機(jī)械機(jī)制和空化機(jī)制。非熱機(jī)制(機(jī)械機(jī)制和空化機(jī)制)在HIFU治療中的作用一方面認(rèn)為應(yīng)盡量對(duì)空化現(xiàn)象進(jìn)行抑制,另一方面認(rèn)為空化有助于熱損傷和監(jiān)控。目前對(duì)HIFU形成損傷的機(jī)制中仍很難確切描述熱機(jī)制、空化機(jī)制和機(jī)械機(jī)制在組織損傷中相對(duì)獨(dú)立作用。單純的非熱損傷(機(jī)械機(jī)制和空化機(jī)制)在HIFU治療中的研究和應(yīng)用較少。 實(shí)驗(yàn)首先進(jìn)行脈沖高強(qiáng)度聚焦超聲(pulsed high intensity focused ultrasound,PHIFU)輻照離體牛肝組織,探討形成非熱損傷的輻照參數(shù)。然后在此輻照參數(shù)下進(jìn)一步研究PHIFU聯(lián)合超聲微泡造影劑(ultrasound contrast agent ,UCA)非熱損傷活體腫瘤組織的可行性及非熱損傷的生物學(xué)行為,為HIFU治療腫瘤提供新方法和基礎(chǔ)理論支持,進(jìn)一步推進(jìn)HIFU技術(shù)在腫瘤治療中的應(yīng)用。 材料和方法參數(shù)設(shè)置時(shí)維持總能量恒定,按不同工作周期1%、5%、15%、25%、50%、75%和100%分成7組,在B超圖像監(jiān)控下將熱電偶測(cè)溫探針插入牛肝,并使針尖位于PHIFU焦點(diǎn),然后進(jìn)行定點(diǎn)輻照。觀(guān)察各輻照參數(shù)下焦域溫度、灰度和組織學(xué)改變。 采用腫瘤組織塊開(kāi)腹包埋接種方法,建立兔肝VX2移植瘤動(dòng)物模型。將36只荷瘤兔隨機(jī)分為假照組、PHIFU組和PHIFU+UCA組。用第一部分實(shí)驗(yàn)篩選出的輻照參數(shù)(超聲頻率0.87 MHz、焦距150 mm、聲功率150 W、脈沖重復(fù)頻率100 Hz、工作周期15%)對(duì)兔肝VX2移植瘤進(jìn)行PHIFU輻照。治療后1天取材觀(guān)察腫瘤組織學(xué)改變、超微結(jié)構(gòu)改變,應(yīng)用末端脫氧核苷酸轉(zhuǎn)移酶(TdT)介導(dǎo)脫氧核苷酸(dUTP)缺口末端標(biāo)記技術(shù)(in situ deoxynucleotityl transferase-mediated dUTP nick end labeling,TUNEL)檢測(cè)細(xì)胞凋亡和用免疫組化法檢測(cè)細(xì)胞增殖核抗原(proliferating cell nuclear antigen,PCNA),并用凋亡指數(shù)(apoptosis index, PI)和增殖指數(shù)(proliferating index, AI)比較細(xì)胞凋亡和增殖情況。 結(jié)果PHIFU輻照離體牛肝時(shí),工作周期25%、50%、75%和100%組平均溫升分別為63.4±9.2℃、65.0±11.5℃、66.6±9.9℃、79.6±10.6℃,最高溫度大于85℃且有明顯的凝固性壞死。1%、5%和15%組平均溫升分別29.2±1.9℃、30.0±2.8℃、31.0±2.4℃,最高溫度不高于56℃且無(wú)肉眼可見(jiàn)的凝固性壞死形成。在25%、50%、75%、100%組,PHIFU損傷后形成的凝固性壞死區(qū)光鏡下主要表現(xiàn)為胞漿顏色變淡,部分核仁消失、細(xì)胞核固縮、染色質(zhì)邊集且隨工作周期延長(zhǎng)而增加。15%組光鏡下細(xì)胞間裂隙增寬,肝細(xì)胞胞漿內(nèi)見(jiàn)大量大小不等空泡,而1%組、5%組無(wú)明顯改變,核仁清晰。 PHIFU輻照兔肝VX2移植瘤后,假照組、PHIFU組和PHIFU+UCA組經(jīng)2,3,5-氯化三苯基四氮唑(2,3,5-triphenyl tetrazolium chloride ,TTC)染色后,肉眼可見(jiàn)腫瘤組織被均勻紅染,表明各組均無(wú)肉眼可見(jiàn)的凝固性壞死形成。光鏡下PHIFU組見(jiàn)腫瘤細(xì)胞胞漿嗜伊紅染色淺,細(xì)胞腫脹,胞漿疏松,胞漿內(nèi)大量空泡,少量細(xì)胞核固縮、染色質(zhì)邊集;PHIFU+UCA組見(jiàn)胞漿內(nèi)大量大小不等空泡,可見(jiàn)染色質(zhì)邊集和細(xì)胞核固縮。電鏡下見(jiàn)PHIFU組瘤細(xì)胞胞漿內(nèi)大量線(xiàn)粒體腫脹和內(nèi)質(zhì)網(wǎng)擴(kuò)張,PHIFU+UCA組部分細(xì)胞核固縮和染色質(zhì)邊集,兩組腫瘤組織內(nèi)均見(jiàn)凋亡小體及細(xì)胞胞漿內(nèi)大小不等空泡。TUNEL法檢測(cè)發(fā)現(xiàn),假照組發(fā)現(xiàn)少量陽(yáng)性染色的腫瘤細(xì)胞,PHIFU組和PHIFU+UCA組見(jiàn)較多細(xì)胞核棕褐色染色的陽(yáng)性細(xì)胞。PCNA檢測(cè)結(jié)果表明,假照組靶區(qū)組織見(jiàn)大量著色部位在腫瘤細(xì)胞核的陽(yáng)性細(xì)胞,而PHIFU組和PHIFU+UCA組陽(yáng)性染色細(xì)胞少。PHIFU組和PHIFU+UCA組腫瘤組織的AI比假照組高,而PI陽(yáng)性表達(dá)率比假照組低。PHIFU+UCA組AI比PHIFU組高,但PHIFU+UCA組PI比PHIFU組低。 結(jié)論1.脈沖工作周期長(zhǎng)在PHIFU輻照時(shí)可形成熱凝固性壞死,而脈沖工作周期短可通過(guò)非熱效應(yīng)損傷組織。2.一定工作周期的PHIFU(超聲頻率0.87 MHz、焦距150 mm、聲功率150 W、脈沖重復(fù)頻率100 Hz、工作周期15%)可通過(guò)非熱效應(yīng)損傷腫瘤,表現(xiàn)為胞漿內(nèi)大小不等空泡和促進(jìn)腫瘤細(xì)胞凋亡及抑制其增殖。3.微泡造影劑可增強(qiáng)超聲對(duì)組織的非熱效應(yīng)損傷。4.可通過(guò)控制PHIFU的工作周期實(shí)現(xiàn)非熱損傷,為HIFU治療腫瘤提供了一種新的治療思路。
[Abstract]:Background and objective high intensity focused ultrasound (high intensity focused ultrasound (HIFU)) is a new noninvasive local treatment technique for the treatment of tumors. Besides the thermal machine, there are mechanical mechanisms and cavitation mechanisms. The role of non thermal mechanism (mechanical mechanism and cavitation mechanism) in the treatment of HIFU should be as far as possible. On the other hand, cavitation is helpful for thermal damage and monitoring. At present, it is difficult to describe the thermal mechanism in the mechanism of HIFU damage. The cavitation mechanism and mechanical mechanism are relatively independent in tissue damage. The research and application of simple non thermal damage (mechanical mechanism and cavitation mechanism) in the HIFU treatment is less.
In the experiment, the isolated bovine liver tissues were irradiated by pulsed high intensity focused ultrasound (PHIFU), and the irradiated parameters of non thermal damage were discussed. Then, the non thermal injured living body tumor tissues of PHIFU combined with the ultrasonic microbubble contrast agent (ultrasound contrast agent, UCA) were further studied under the irradiation parameters. The biological behavior of the feasibility and non thermal damage can provide a new method and basic theory support for the treatment of tumor by HIFU, and further promote the application of HIFU in the treatment of tumor.
When the material and method parameters are set, the total energy is kept constant, and the thermocouple probe is inserted into the bovine liver under the monitoring of B ultrasonic image under the monitoring of different working cycles 1%, 5%, 15%, 25%, 50%, 75% and 100%. The needle tip is located at the focus of PHIFU and then irradiated at fixed point. The focal temperature, gray scale and histological changes under the irradiated parameters are observed.
The animal model of rabbit liver VX2 transplanted tumor was established by the method of open tumor tissue mass, and 36 rabbits were randomly divided into sham group, PHIFU group and PHIFU+UCA group. The irradiation parameters were selected by the first part of the experiment (ultrasonic frequency 0.87 MHz, focal length 150 mm, sound power 150 W, pulse repetition rate 100 Hz, operation cycle 15%). The tumor was irradiated by PHIFU. After 1 days of treatment, the histological changes and ultrastructural changes were observed and the terminal deoxynucleotidyl transferase (TdT) mediated deoxynucleotide (dUTP) Nick nick end labeling technique (in situ deoxynucleotityl transferase-mediated dUTP nick end labeling, TUNEL) was used to detect cell apoptosis and immunohistochemical method. Proliferating cell nuclear antigen (PCNA) was detected and apoptosis and proliferation were compared with the apoptosis index (apoptosis index, PI) and proliferation index (proliferating index, AI).
Results when PHIFU was irradiated in vitro, the average temperature rise of the 25%, 50%, 75% and 100% groups was 63.4 + 9.2, 65, 11.5, 66.6 + 9.9, 79.6 + 10.6, and the highest temperature was higher than 85, and there was obvious solidification necrosis.1%. In the 25%, 50%, 75%, 100% groups, the coagulative necrosis area in the 25%, 50%, 75%, and 100% groups was mainly characterized by the pale color of the cytoplasm, the disappearance of some nucleolus, the nuclear condensation, the aggregation of chromatin and the increase of the widening of the intercellular fissure under the light microscope of the.15% group with the prolongation of the working cycle, and a large number of sizes in the cytoplasm of the liver cells. There was no difference in vacuoles, but in the 1% group, there was no significant change in the 5% groups, and the nucleolus was clear.
After PHIFU irradiated rabbit liver VX2 transplanted tumor, in group PHIFU and PHIFU+UCA group, after 2,3,5- chlorination of three phenyl tetrazoles (2,3,5-triphenyl tetrazolium chloride, TTC), the naked eye could be found to be uniform red staining, indicating that there was no visible coagulative necrosis in all groups. The PHIFU group under light microscopy showed the cytoplasm eosinophilic staining of tumor cells. Light color, cell swelling, loose cytoplasm, a large number of vacuoles in the cytoplasm, a small amount of nuclear condensation and chromatosis, PHIFU+UCA group found a large number of different vacuoles in the cytoplasm, visible chromatin edge and nuclear condensation. Under the electron microscope, a large number of mitochondria swelling and endoplasmic reticulum dilation in the cytoplasm of the tumor cells of the PHIFU group were found, and the part of the nucleus of group PHIFU+UCA was retractable and dyed. In the two groups of tumor tissues, all the two groups of tumor tissues were found to detect apoptotic bodies and cytoplasm unequal vacuoles, and a small number of positive staining cells were found in the sham group. The results of.PCNA detection in group PHIFU and PHIFU+UCA group were found in the positive cells of multicellular nuclear brown staining. The results showed that a large number of pigmentation sites were found in the target tissue of the sham group. The positive cells of the tumor nuclei were higher in group PHIFU and group PHIFU+UCA than in group.PHIFU and group PHIFU+UCA, and AI in group PHIFU+UCA was higher than that in sham group, but the positive rate of PI was higher than that of group PHIFU in the lower.PHIFU+UCA group than that in the sham group, but the PI PHIFU+UCA group was lower than that of the PHIFU group.
Conclusion the 1. pulse working cycle can form thermocoagulation necrosis with PHIFU irradiation, and the short pulse cycle can be short of PHIFU (ultrasonic frequency 0.87 MHz, focal length 150 mm, sound power 150 W, pulse repetition rate 100 Hz, operation cycle 15%) through non thermal effect injury. Unequal vacuoles in the pulp and promoting the apoptosis of tumor cells and inhibiting the proliferation of.3. microbubbles can enhance the non thermal effect of ultrasound on tissue..4. can achieve non thermal damage by controlling the working cycle of PHIFU, which provides a new therapeutic idea for the treatment of tumor in HIFU.

【學(xué)位授予單位】：重慶醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2007
【分類(lèi)號(hào)】：R312

【引證文獻(xiàn)】

相關(guān)碩士學(xué)位論文 前1條

1 史學(xué)豹;脈沖高強(qiáng)度聚焦超聲脈沖參數(shù)與焦點(diǎn)聲壓的關(guān)系研究[D];重慶醫(yī)科大學(xué);2012年

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本文編號(hào)：1812703