本文選題：微泡 + 空化　； 參考：《第三軍醫(yī)大學(xué)》2014年碩士論文

【摘要】：背景： 血管生成是腫瘤生長(zhǎng)、轉(zhuǎn)移和侵襲的重要條件。已知直徑超過1～2mm的實(shí)體腫瘤生長(zhǎng)和轉(zhuǎn)移離不開腫瘤血管生成。與正常血管相比，腫瘤新生血管發(fā)育不成熟，其結(jié)構(gòu)上存在諸多缺陷：如血管內(nèi)皮連接不緊密、基底膜不完整、缺少血管平滑肌層、周細(xì)胞連接疏松等�？鼓[瘤血管生成靶向治療主要利用腫瘤新生血管壁存在的特異性生物靶點(diǎn)，通過生物或者化學(xué)方法攻擊和破壞腫瘤新生血管，以此達(dá)到治療腫瘤的目的。然而，現(xiàn)有的靶向抗腫瘤新生血管的治療方法，如血管生成抑制劑等，未取得突出的治療效果，同時(shí)也會(huì)產(chǎn)生一些副作用影響到機(jī)體正常的生理過程（如傷口愈合、組織修復(fù)等），也無法避免長(zhǎng)期使用產(chǎn)生的耐藥性。 超聲激勵(lì)的血管腔內(nèi)微泡空化可以物理破壞腫瘤微血管，阻斷腫瘤血流灌注，其機(jī)制主要是空化效應(yīng)產(chǎn)生的沖擊波和微射流等機(jī)械效應(yīng)對(duì)腫瘤血管壁造成的可逆性或不可逆性機(jī)械損傷。前期實(shí)驗(yàn)發(fā)現(xiàn)，采用峰值負(fù)壓為4.8MPa的脈沖式聚焦超聲激勵(lì)微泡可以產(chǎn)生腫瘤血管毀損性損傷以及長(zhǎng)達(dá)24h的微循環(huán)阻斷，而當(dāng)峰值負(fù)壓降為2.6MPa時(shí)僅能使腫瘤血流阻斷一小時(shí)。采用微泡超聲空化治療腫瘤具有安全、高效、操作簡(jiǎn)單、重復(fù)性好等優(yōu)點(diǎn)，具有良好的發(fā)展前景。為了進(jìn)一步了解超聲空化強(qiáng)度與腫瘤微血管損傷的關(guān)系，我們采用三個(gè)水平峰值負(fù)壓超聲聯(lián)合微泡治療兔VX2腫瘤，并分析腫瘤超聲空化治療前后超聲造影血流灌注情況及空化強(qiáng)度對(duì)微血管密度的影響。 目的： 1.通過觀察不同水平峰值負(fù)壓超聲聯(lián)合微泡空化對(duì)腫瘤血流灌注的影響，初步分析超聲峰值負(fù)壓對(duì)腫瘤微循環(huán)阻斷程度的影響作用。 2.通過比較各水平超聲峰值負(fù)壓組與對(duì)照組腫瘤的微血管密度，探討微泡超聲空化治療的超聲峰值負(fù)壓與微血管密度的量效關(guān)系。 材料方法： 1.實(shí)驗(yàn)材料 ⑴儀器設(shè)備：①DCT-700型新型數(shù)字化超聲空化治療儀，深圳市威爾德醫(yī)療電子有限公司生產(chǎn)。超聲發(fā)射頻率為1.0MHz，脈沖重復(fù)頻率為100Hz，占空比為1.5%，峰值負(fù)壓多檔可調(diào)。②超聲診斷儀，S2000型彩色多普勒超聲診斷儀（西門子公司生產(chǎn)），配備有CPS超聲造影成像模式，配有9L4高頻線陣探頭，發(fā)射頻率為7～9MHz，中心頻率8MHz。 ⑵實(shí)驗(yàn)動(dòng)物：由第三軍醫(yī)大學(xué)附屬新橋醫(yī)院的實(shí)驗(yàn)動(dòng)物中心提供，共46只雄性健康的新西蘭大白兔，體質(zhì)量在1.5～2.0kg之間。 ⑶實(shí)驗(yàn)試劑：①本實(shí)驗(yàn)中用于超聲空化治療與超聲造影所用的微泡為第三軍醫(yī)大學(xué)附屬新橋醫(yī)院超聲科所制的超聲造影劑-脂氟顯，，其中心氣體為全氟丙烷，微泡濃度為（4～9）×109/mL，微泡直徑為2～8μm，平均2μm。②小鼠抗人CD31單克隆抗體（即血小板內(nèi)皮細(xì)胞黏附分子-1抗體），購自Abcam公司，保存于4°冰箱。 2.實(shí)驗(yàn)方法 ⑴不同水平峰值負(fù)壓超聲對(duì)兔VX2肌肉移植瘤空化治療實(shí)驗(yàn)： 16只腿部肌肉種植VX2腫瘤荷瘤種兔，共30個(gè)腫瘤，隨機(jī)分成3個(gè)不同水平超聲峰值負(fù)壓幅度處理組，分別為2184kPa組、1785kPa組、1019kPa組。各峰值負(fù)壓組采用相應(yīng)峰值負(fù)壓參數(shù)進(jìn)行超聲輻照聯(lián)合經(jīng)靜脈注射微泡（0.2ml）治療腫瘤，并于治療前、后進(jìn)行超聲造影檢查，分別對(duì)超聲造影峰值強(qiáng)度和曲線下面積值進(jìn)行分析，即刻處死實(shí)驗(yàn)兔于治療后，獲取腫瘤組織標(biāo)本進(jìn)行HE（hematoxylin-eosin staining）染色，并觀察腫瘤組織病理改變。 ⑵超聲峰值負(fù)壓與兔VX2肌肉移植瘤微血管密度量效關(guān)系研究實(shí)驗(yàn)： 30只腿部肌肉種植VX2腫瘤荷瘤兔隨機(jī)分成4組：分別為上述三個(gè)水平峰值負(fù)壓組（n=8）及對(duì)照組（n=6），對(duì)照組不予以任何處理。另三組給予超聲激勵(lì)微泡空化治療腫瘤。治療后即刻取出腫瘤組織，固定，包埋，切片，行免疫組化染色，對(duì)微血管密度（MVD）進(jìn)行計(jì)數(shù)，并比較各組腫瘤微血管密度值。 結(jié)果： 1.不同水平峰值負(fù)壓超聲對(duì)兔VX2肌肉移植瘤空化治療實(shí)驗(yàn)：峰值負(fù)壓2184kPa組腫瘤治療后造影血流灌注完全消失，其PI值由治療前的（40.59.9）%降至治療后的（11.67.8）%，AUC由（1299.1512.6）%s降至（280.1186.1）%s (P0.05)；1785kPa組治療后造影腫瘤血流灌注明顯下降，PI由（42.57.8）%降至（24.214.8）%，AUC由（1378.1494.6）%s降至（549.4463.4）%s (P0.05)；1019kPa組腫瘤治療后造影與治療前比較腫瘤血流灌注輕度增強(qiáng)，但治療前后PI與AUC比較差異無統(tǒng)計(jì)學(xué)意義(P0.05)。病理檢查各組腫瘤治療后均可見部分腫瘤微血管斷裂，紅細(xì)胞溢出，腫瘤微血管出血，紅細(xì)胞滲入組織間隙，腫瘤組織水腫，但2184kPa組腫瘤細(xì)胞較其它組水腫明顯。 2.峰值負(fù)壓為2184kPa、1785kPa、1019kPa的三組腫瘤MVD值分別為（157.15±124.37）、（141.12±60.45）、（311.64±73.93），均低于對(duì)照組腫瘤MVD值（619.73±282.78），差異有統(tǒng)計(jì)學(xué)意義(P0.05)，余各組兩兩比較差異無統(tǒng)計(jì)學(xué)意義(P＞0.05)。 結(jié)論： 1.超聲峰值負(fù)壓在1785kPa和2184kPa水平時(shí)超聲空化可阻斷VX2腫瘤微循環(huán)，且峰值負(fù)壓越高，阻斷效果越顯著。峰值負(fù)壓為1019kPa水平時(shí)不僅不能阻斷腫瘤微循環(huán)，還可能造成腫瘤血流灌注輕度增強(qiáng)。 2.經(jīng)靜脈微泡聯(lián)合超聲空化可以減少腫瘤微血管數(shù)，可能與超聲空化毀損性破壞腫瘤微血管有關(guān)；且峰值負(fù)壓在2184kPa與1785kPa水平減少腫瘤微血管數(shù)要多于峰值負(fù)壓在1019kPa水平。
[Abstract]:Background:
Angiogenesis is an important condition for tumor growth, metastasis and invasion. The growth and metastasis of solid tumors with known diameter of more than 1 to 2mm are inseparable from tumor angiogenesis. Compared with normal blood vessels, the neovascularization of the tumor is immature and there are many defects in its structure, such as the incompleteness of the vascular endothelium, incomplete basement membrane, and lack of smooth blood vessels. The targeting of tumor angiogenesis is mainly used to attack and destroy neovascularization through biological or chemical methods to achieve the purpose of treating tumor. However, the existing methods of targeting neovascularization, such as angiogenesis, have been used. As a inhibitor, there is no outstanding therapeutic effect, and some side effects will also affect the normal physiological processes of the body (such as wound healing, tissue repair, etc.), and the resistance to long-term use can not be avoided.
The intravascular microbubble cavitation stimulated by ultrasound can physically destroy the tumor microvessels and block the tumor blood flow. The mechanism is the reversible or irreversible mechanical damage caused by the mechanical effects such as the cavitation effect and the micro jets on the tumor vessel wall. In the previous experiment, the pulse type polymerization with the peak negative pressure of 4.8MPa was found. Jiao Chaosheng microbubbles can produce tumor vascular damage and 24h microcirculation blocking, but when the peak pressure drop is 2.6MPa, the tumor blood flow can only be blocked for an hour. Microbubble ultrasound cavitation is a safe, efficient, simple and reproducible, and has a good prospect. To understand the relationship between ultrasonic cavitation intensity and tumor microvascular injury, we used three horizontal peak negative pressure ultrasound combined with microbubbles in the treatment of rabbit VX2 tumor, and analyzed the effect of ultrasound contrast perfusion and cavitation intensity on microvascular density before and after ultrasound cavitation therapy.
Objective:
1. by observing the effect of different levels of peak negative pressure ultrasound combined with microbubble cavitation on the blood perfusion of tumor, the effect of peak negative pressure on the blocking degree of tumor microcirculation was preliminarily analyzed.
2. the quantitative relationship between the peak negative pressure and the microvascular density of the microbubble ultrasound cavitation was investigated by comparing the microvascular density of the different levels of the ultrasonic peak negative pressure group and the control group.
Material methods:
1. experimental materials
(1) equipment: (1) a new type of DCT-700 digital ultrasonic cavitation treatment instrument, produced by Shenzhen Wilder Medical Electronics Co., Ltd., the ultrasonic emission frequency is 1.0MHz, the pulse repetition rate is 100Hz, the duty ratio is 1.5%, the peak negative pressure is multiple adjustable. 2. Ultrasonic diagnostic instrument, S2000 color color color ultrasonic diagnostic instrument (SIEMENS company production). The CPS ultrasound contrast imaging mode is equipped with 9L4 high frequency linear array probe. The transmitting frequency is 7 ~ 9MHz, and the central frequency is 8MHz.
(2) experimental animals: a total of 46 healthy New Zealand white rabbits were provided from the experimental animal center affiliated to the new bridge hospital affiliated to Third Military Medical University. The body mass of the rabbits was from 1.5 to 2.0kg.
(3) experimental reagents: (1) the microbubbles used in ultrasonic cavitation therapy and ultrasound contrast in this experiment are the ultrasound contrast agents made by the ultrasound department of the affiliated Xinqiao Hospital of Third Military Medical University. The central gas is perfluoropropane, the concentration of microbubbles is (4~9) * 109/mL, the diameter of microbubbles is 2~8 u m, and the average of 2 mu m. in mice against human CD31 monoclonal anti Body (that is, platelet endothelial cell adhesion molecule -1 antibody), purchased from Abcam company, is stored in the 4 degree refrigerator.
2. experimental method
(1) cavitation treatment of rabbit VX2 muscle xenograft with different level of peak negative pressure ultrasound:
16 leg muscles were planted with VX2 tumor bearing rabbits and 30 tumors were randomly divided into 3 different levels of ultrasonic peak negative pressure amplitude treatment group, group 2184kPa, group 1785kPa and group 1019kPa. The peak negative pressure groups were treated with corresponding peak negative pressure parameters for ultrasound irradiation combined with intravenous microbubbles (0.2ml) for the treatment of tumor before and after treatment. The peak intensity of contrast-enhanced ultrasound and the area under the curve were analyzed by ultrasound contrast examination. After the treatment, the rabbit tissue specimens were stained with HE (hematoxylin-eosin staining), and the pathological changes of the tumor were observed.
2. Study on the dose effect relationship between peak negative pressure and microvessel density of rabbit VX2 muscle transplantation tumor:
30 VX2 tumor bearing rabbits were randomly divided into 4 groups: the above three level negative pressure group (n=8) and the control group (n=6). The control group was not treated with any treatment. The other three groups were treated with ultrasonic stimulation microbubble cavitation for the treatment of tumor. Density (MVD) was counted, and tumor microvessel density values were compared.
Result錛

本文編號(hào)：2024520