【作者】 李权；

【导师】 王智彪； 陈文直；

【作者基本信息】 重庆医科大学 ， 妇产科学， 2013， 博士

【摘要】 目的通过对HIFU消融治疗子宫肌瘤时引起子宫内膜反应（包括阴道流血或阴道流液）的相关因素的回顾性分析，筛出可能引起子宫内膜反应的因素。同时用细胞在超声近场内的反应情况，超声消融离体子宫肌瘤时内膜的形态学变化以及超声消融子宫肌瘤时在体子宫内膜的形态学变化的研究，为HIFU消融子宫肌瘤对子宫内膜的影响提供理论和临床依据，为合理选择适应症、减少内膜损伤的发生提供依据。资料与方法1.筛选HIFU消融治疗的单发子宫肌瘤或多发子宫肌瘤仅治疗单个肌瘤患者381例，根据有无阴道流液分为两组，筛选内容包括三方面：第一部分为年龄、症状综合评分、妊娠次数、生育次数、人流次数、BMI；第二部分为MRI检查相关指标：腹直肌厚度、脂肪厚度、腹壁厚度、子宫大小、肌瘤大小、肌瘤位置、子宫内膜厚度、肌瘤分类和肌瘤浅面到皮肤的距离；第三部分为HIFU治疗相关指标：治疗时间、辐照时间、治疗强度、治疗总能量、治疗体积、焦点到内膜距离、肌瘤边缘到内膜距离和消融后肌瘤坏死边缘到内膜距离。每例病例均有放射科医师和妇产医师共同阅片，统计分析各影响因素在阴道流液和无引道流液两组之间差异情况，寻找引起阴道流液的因素。2.取因子宫肌瘤行子宫切除的子宫内膜，行原代细胞混合培养。将子宫内膜细胞按1×10~~6的密度放入特制的培养瓶内密封，将细胞悬液分别置于与物理学焦域距离为5mm、10mm、15mm和20mm处，每组分别辐照2s、4s和6s三个组，辐照后细胞继续培养，分别在24h、48h和72h用MTT法检测细胞的增殖情况。3.取因子宫肌瘤行子宫切除的者，将切除的子宫立即低温保存后送HIFU治疗实验室行超声消融治疗。将子宫置入治疗床上特制的容器中，内膜在超声近场，监控超声可见肌瘤及宫体，用200W的能量对肌瘤消融，超声监测见肌瘤消融完毕后，立即沿肌瘤最大径线纵轴剖视肌瘤，取子宫内膜分别行HE和电镜检查。4.选择因子宫肌瘤行超声消融的后壁肌瘤患者7例，术前月经量的评分，基础性激素，MRI检查肌瘤。在行超声消融治疗后患者休息2小时无反应，行MRI复查，了解肌瘤消融情况，无发热和腹痛等反应者，预约宫腔镜检查。超声消融术后3月复诊基础性激素，月经量的评分，MRI和宫腔镜。结果1.381例接受了HIFU消融术，其有133例有不同程度阴道流液或血性分泌物，持续1周消失，占34.9%。前壁肌瘤248个，其中45例患者有阴道流液，占18.1%。后壁肌瘤133个，其中88例患者有阴道流液，占66.2%。对一般资料与有无阴道排液进行相关性分析，患者年龄、BMI、月经周期的天数、妊娠次数、生育次数与阴道排液的发生均无明显相关性（p>0.05）；患者症状综合评分（UFS）与阴道排液的发生具有正相关关系（r=0.235，p=0.000）；QOL生活质量评分（QOL）与阴道排液的发生具有负相关关系（r=-0.165, p=0.001）。对肌瘤特征与有无阴道排液进行相关性分析，腹壁厚度、肌瘤体积、肌瘤浅面到腹壁皮肤距离与阴道排液的发生均无明显相关性（p>0.05）；子宫体积与阴道排液的发生具有正相关关系（r=0.164，p=0.001）；子宫内膜厚度与阴道排液的发生具有负相关关系（r=-0.225，p=0.000）；前壁肌瘤深面到内膜的距离与阴道排液的发生具有负相关关系（r=-0.445, p=0.000）；前壁肌瘤消融区深面到内膜的距离与阴道排液的发生具有负相关关系（r=-0.442，p=0.000）；后壁肌瘤浅面到内膜的距离与阴道排液的发生具有负相关关系（r=-0.622，p=0.000）；前壁肌瘤消融区浅面到内膜的距离与阴道排液的发生具有负相关关系（r=-0.532，p=0.000）。对超声消融相关参数与有无阴道排液进行相关性分析，焦点到子宫内膜距离、剂量强度、治疗时间、治疗总能量和消融体积与阴道排液的发生均无明显相关性（p>0.05）；而辐照时间与阴道排液的发生具有正相关关系（（r=-0.622，p=0.018）。将与阴道排液相关的因素症状综合评分（UFS）、生活质量评分（QOL）、子宫体积、子宫内膜厚度、肌瘤边缘到内膜的距离、超声辐照时间，经多重Logistic回归分析逐步剔除变量包括QOL、子宫体积、子宫内膜厚度和超声辐照时间（P>0.05），有意义的变量为肌瘤边缘到内膜的距离和UFS。2.子宫内膜细胞经原代混合培养后上皮细胞可以传代3-5代，而间质细胞可以多次传代，上皮细胞排列呈旋涡状，细胞呈多角形或蝌蚪形，而间质细胞在呈梭形，经细胞鉴定角蛋白CK19阳性为上皮细胞，而波形蛋白阳性为间质细胞。子宫内膜细胞在超声近场随辐照距离的减小，细胞增殖能力变弱，在相同距离下，20mm处随辐照时间增加，细胞增殖能力增强；在距离15mm处、10mm和5mm处随辐照时间增加，细胞增殖能力减弱。而增殖明显的辐照点在20mm处辐照6秒和10mm处辐照4秒；增殖相对较小的点在5mm处辐照6秒，两两比较差异均具有显著性（P<0.01）。3.在低倍镜下可见子宫内膜层变薄，腺体少，肌层与腺体均未见明显坏死组织；高倍镜下子宫内膜细胞和肌层细胞均未见坏死。电镜下内膜与肌瘤间子宫平滑肌细胞排列呈束状，胞浆丰富，细胞核部分溶解，细胞核膜完整；血管内皮细胞肿胀明显，细胞周围间隙扩大；线粒体肿胀，可见高尔基体肿胀，细胞核膜间隙增宽呈双层；子宫内膜腺体形态存在轮廓，腺上皮细胞结构破坏，相互融合，说明超声消融离体子宫肌瘤对子宫基地层组织损伤小。4.7例患者超声消融治疗前后性激素比较差异无显著意义（P<0.05）,治疗前后月经量评分比较差异无显著意义（P<0.05）。术后宫腔镜检查，宫腔无积血，近肌瘤处子宫内膜散在充血点，与宫腔内膜色泽淡红，未见出血点，行光镜观察可见内膜下出血，间质水肿出血，部分腺体断裂。术后3月MRI检查肌瘤明显缩小，复查宫腔镜见宫腔内膜淡红色，无粘连。结论HIFU消融为非侵入性治疗方法，治疗子宫肌瘤引起阴道流液或流血是其并发症之一，阴道排液相关的因素为症状综合评分、QOL生活质量评分、子宫体积、子宫内膜厚度、肌瘤边缘到内膜的距离、超声辐照时间，经多重Logistic回归分析逐步剔除后，阴道流液与肌瘤边缘到内膜的距离密切相关；体外通过子宫内膜细胞置于超声近场不同距离，发现超声近场促进细胞增殖，随距离物理学焦域越近，增殖作用减弱，辐照时间有关，辐照时间越长，增殖减弱；而离体超声消融肌瘤时，子宫内膜凝固性坏死明显，而子宫肌层组织仅有细胞器水肿，无凝固性坏死，说明子宫内膜更易受热损伤；在临床研究中显示高强度聚焦超声消融子宫肌瘤时，近肌瘤子宫内膜有散在充血点，余子宫内膜未见充血；同时超声消融肌瘤对卵巢内分泌功能无影响，月经量也无影响，提示子宫内膜局部出血在正常的性激素作用下会增生修复。高强度聚焦超声消融子宫肌瘤在目前治疗条件下对子宫内膜是安全的。更多还原

【Abstract】 ObjectiveBy retrospectively analyzing the relevant factors which caused theendometrial reaction for the HIFU ablation treatment of uterine fibroids(including vaginal bleeding or vaginal fluid) and sieving the factors whichcan cause the endometrial reaction. While using the reaction of endometrialcell in the near field of ultrasound, studying the morphological changes ofthe endometrium using ultrasound ablation in vitro and the morphologicalchanges of the endometrium in vivo, offering theoretical basis for theimpact upon the endometrium in the ultrasound ablation of uterine fibroidsHIFU ablation, and providing the basis for a reasonable choice ofindications to reduce the occurrence of endometrial injury.Methods1. Three hundred and eighty-one patients with single uterine fibroidsor multiple uterine only individual treated fibroids were screened.According to the presence or absence of vaginal fluid; patients weredivided into two groups. The screened contents include three aspects as follows: The first part includes age, symptoms, composite scores, thenumber of pregnancy,the number of births and the number of abortions,BMI; The second part includes the related indicators of the MRIexamination: such as the thickness of rectos abdominals, fat thickness,abdominal wall thickness, size of the uterus,fibroids size, the place of thefibroids, endometrial thickness, the fibroids classification and the distancefrom fibroids shallow surface to skin; The third part includes the relatedindicators of the HIFU treatment, such as treatment time, irradiation time,intensity of treatment, the total energy of the treatment, treatment volume,the distance from the focus to the endometrial, distance from fibroids edgeto endometrium, distance from ablation fibroid necrosis edge toendometrium. The MRI pieces were read by both radiologists andgynecologist. Add up and analyze each influential factor’s differencebetween the vaginal fluid liquid and no vaginal flow liquid, seeking forthe factor that causes vaginal fluid.2. Take the endometrium of the hysterectomy due to uterine fibroids.Primary endometrial cell was mixed culture. Endometrial cells were placedin specially sealed flasks by a density of1×10~~6, the sealed flasksequipped with endometrial cells were placed at distance from physic focalregion of5mm,10mm,15mm and20mm. Each group endometrial cellsrespectively irradiated2s,4s and6s three groups. Irradiated cells werecultured with MTT detect cell proliferation at24h,48h and72hrespectively.3. To take the endometrium of the hysterectomy patient of uterinefibroids, the removal of the uterus were immediately cryopreservation inice container and send to HIFU treatment laboratory for ultrasound ablationtreatment. Put the uterine fibroids into the special containers on the JC200treatment couch. The fibroids in the focal region remote, monitoring ultrasound can be screen fibroids and uterus. Uterine fibroids were ablatedwith200W energy. When ultrasound grayscale covered fibroids, the uterinefibroids ablation finished and immediately slit fibroids along thelongitudinal axis of the fibroids. Endometrium were immediately cut andplaced into10%formalin for HE, other endometrium placed into2%glutaraldehyde for electron microscopic examination.4. To select seven cases posterior wall of the uterus fibroids of HIFUablation due to uterine fibroids. Before HIFU ablation, menstrual score,basis of sex hormones and MRI check fibroids were applied. After HIFUablation fibroids two hours, patients underwent MRI review and understandfibroids necrotic area for no fever and no abdominal pain and prepared forhysteroscopy examination. After HIFU ablation uterine fibroids threemonths, menstrual score, basis of sex hormone, MRI check fibroids andhysteroscopy examination were applied again.Results1.381patients who underwent HIFU ablation,133patiets withvarying degrees of vaginal flow or blood,disappeared about1week,accounting for34.9%.Anterior fibroids248,of which45patients vaginalfliud,accounting for18.1%; Posterior wall of fibroids133,of which88patients with vaginal fluid,accounting for66.2%.General and the presenceof vaginal discharge correlation analysis,patient age,BMI,menstrual cycledays,the number of pregnancy,number of births and the incidence ofvaginal discharge,there was no significant correlayion(p>0.05),symptomscomposite score (UFS) and the occurrence of vaginal discharge has apositive correation (r=0.235, P=0.000); quality of life (QOL) score and theoccurrence of vaginal discharge has a negative correlation(r=-0.165, p=0.001).The correlation analysis of fibroids features with or without vaginaldischarge,including abdominal wall thickness, fibroids,fibroids surface to the abdominal wall skin distance and the occurrence of vaginaldischarge,there was no significant correlation(p>0.05);uterine volume andthe occurrence of vagial discharge has a positive correlation(r=0.164,p=0.001); endometrial thickness and the occurrence of vaginaldischarge has a negative correlation(r=-0.225,p=0.000);deep surface of theanterior wall of fibroids to the endometrial distance and occurrence ofvaginal discharge has a negative correlation (r=-0.445,p=0.000); theanterior fibroid ablation zone deep surface has a negative correlation toendometrial distance and the occurrence of vaginal discharge(r=-0.442,p=0.000)；occurrence of the distance of the shallow surface of theposterior wall of the fibroids to endometrial and vaginal discharge has anegative correlation (r=-0.622,p=0.000);the anterior fibroid ablation zoneshallow surface to the endometrium of the distance and occurrence ofvaginal diacharge has a negative correlation (r=-0.532,p=0.000).Ultrasound ablation parameters with or without vaginal dischargecorrelation analysis,the focus distance to the endometrium,dose intensity,duration of treatment,the treatment of the total energy and ablation volumeand the occurrence of vaginal discharge has no significant correlation(p>0.05); while the irradiation time and the occurrence of vaginal dischargefactors symptoms score(UFS), life quality score (QOL) uterinevolume,emdometrial thickness,the endometrial fibroids edge to thedistance of the ultrasonic irradiation time. Multiple Logistic regressionanalysis gradually removing the variable including QOL,uterinevolume,endometrial thickness and the ultrasonic irradiation time(p>0.05),fibroids edge to endometrial distance and UFS of significantvariable quantity.2. Primary endometrial cells after a mixed culture of epithelial cellscan be passed to3or5generations, but stromal cells can be repeatedly passed.Under microscope, epithelial cells arranged in a spiral-shaped cellwere polygonal or tadpole-shaped but stromal cells infusiform. The cellidentification keratin CK19positive in epithelial cells, vimentin positive instromal cells. In the near-field of HIFU, endometrial cells wasproliferation.The cell proliferation ability becomes weak when distancedecreases from focal field in physics. The endometrial cell proliferationenergy was increased at the same distance of20mm with the increaseirradiation time.The endometrial cell proliferation energy was decreasingat a distance of15mm,10mm, and15mm with irradiation time increasing.The proliferation strongest point were at10mm pairwise comparison(p<0.01).3. The visible layer of the endometrium were thin under the lightmicroscope. The endometrium, glands and myometrium were no obviousnecrotic tissue under low magnification. Endometrial cells and smoothmuscle cells all shows no necrosis. On the electron microscope The smoothmuscle cells arranged in bundles, abundant cytoplasm, nucleus partialdissolution of the nuclear membrane integrity, vascular endothelial cellswelling, cell cycle gap expansion,mitochondrial swelling, swelling of theGolgi apparatus, nuclear membrane gap broadening were bunk, film glandsmorphology contour glandular epithelial cells structural damage and fusion.Ultrasound ablation from uterine fibroids has little damage for uterine baselayer.4. Seven patients of HIFU ablation treatment uterine fibroids were notstatistically significant compare before HIFU hormone and menstrualscore with after three months hormone and menstrual score usingHIFU(p>0.05). Under hysteroscopy, endometrium show color pinkhemorrhage, congestive points scattered near fibroids. Endometrium werepart glands fractur, endometrial stromal edema and hemorrhage. After three months HIFU ablation, uterine fibroids were significantly reduced by MRIexamination fibroids again. Endometrium were pink and no adhesion.ConclusionsHIFU ablation is a non-invasive treatment. The treatment of theuterine fibroids causes vaginal fluid or flooding, which is one of thecomplications. Vaginal discharge factors including symptoms compositescore,the quality of life score,uterine volume,endometrial thickess,thefibroids edge to endometrial distance,ultrasonic irradiation time,multiplelogistic regression analysis,vaginal fluid factor successively removed,endometrium is closely related to the distance to the edge of the fibroids.The tissue and cells of sound channel are affected by near-field of HIFUwith physics focal field distance and irradiation time. The near-field ofHIFU mainly promote cell proliferation.With distance from the nearerphysics focal field, the proliferation energy is reduced.When theirradiation time is longer,the proliferation energy is weakened more. Usingultrasound ablation fibroids in vitro, endometrial coagulation necrosis isobvious,but myometrium only organelle edema, no coagulation necrosis,itis easier to heat endometrial damage. On HIFU ablation uterinefibroids,endometrium near fibroids scattered congestive point,the othersendometrium didn’t blood. After HIFU ablation fibroids, ovarianendocrine function and menstrual score are no longer affected. Under thenormal sex hormone,bleeding endometrium may repair again. Therefore,endometriums of HIFU ablation fibroids were safe under the currenttreatment modalities.更多还原