【作者】 孙咏梅；

【导师】 刘韶平；

【作者基本信息】 山东大学 ， 影像医学与核医学， 2009， 博士

【摘要】 第一部分彩色多普勒超声诊断甲状腺癌的多因素分析背景甲状腺癌是头颈部比较常见的恶性肿瘤,近年来发病率有上升趋势,早期甲状腺癌经手术切除后,几乎可以根治,而晚期甲状腺癌则会发生淋巴结、肺、骨等远处转移,早期诊断、早期治疗至关重要。目前,临床上对甲状腺癌的诊断主要依靠触诊及影像学检查(高频超声、核素、CT、MRI)及免疫学测定等,其中,彩色多普勒超声是目前甲状腺及颈部疾病最重要的检查手段,随着多种超声显像技术的不断改进和应用,超声能够观察到2～3mm的微小病灶,能够较清晰地显示病灶内部结构、有无包膜和微钙化等细微结构的改变以及病灶血流分布和血流频谱的变化,为进一步提高甲状腺疾病的诊断率提供了有利条件。然而,由于甲状腺结节病理类型繁多,生物学特性差异大,且具有多发性及多源性的特点,使得超声对部分不典型结节的诊断存在困难,在超声检查中还存在一定比例的漏诊和误诊,对于部分良恶性结节的鉴别尚有争议。目的通过观察145例经手术和病理证实的甲状腺结节的彩色多普勒超声表现,将声像图表现与病理结果结合进行单因素和Logistic多因素分析,以获得超声诊断甲状腺癌的最相关的声像图指标,为方便、快捷地鉴别甲状腺良、恶性结节提供理论依据。方法选择2006年2月至2008年8月因甲状腺结节在我院普外科收治的患者145例,所有患者术前行彩色多普勒超声检查,观察并记录病灶的二维及彩色多普勒图像表现,包括病灶数目、大小、边界、内部回声、砂粒样强回声、声晕、后方衰减、血流分级及血流阻力指数、颈部肿大淋巴结等。术后与病理对照。将甲状腺良、恶性结节超声表现结合病理结果进行单因素和Logistic多因素回归分析,筛选甲状腺癌相关声像图指标。结果145例甲状腺结节包括乳头状癌48例,滤泡样癌6例,髓样癌2例,转移癌1例;结节性甲状腺肿69例,甲状腺腺瘤19例。57例甲状腺癌中,边界模糊者39例(68.4%);低回声46例(80.7%),有砂粒样强回声35例(61.4%),有后方衰减21例(36.8%),血流阻力指数＞0.7者19例(33.3%),有淋巴结转移23例(40.4%)。颈部肿大淋巴结位于中颈部者占76%(19/25)。转移淋巴结表现为增大、变圆,皮质增厚,髓质偏心或消失,内部回声增强,不均匀,可见砂粒样强回声,可呈不同程度囊性改变,实性淋巴结内血流增多、紊乱。88例甲状腺良性结节中,边界模糊者8例(9.1%),低回声26例(29.5%),有砂粒样强回声4例(4.5%),有后方衰减6例(6.8%),血流阻力指数＞0.7者5例(5.7%)。经单因素分析,对诊断甲状腺癌有意义的声像图表现有单发病灶、边界模糊、低回声、砂粒样强回声、后方衰减及高血流阻力。经Logistic多因素回归分析,甲状腺癌相关声像图特征为:砂粒样强回声、低回声、边界模糊及单发病灶。结论1.经Logistic多因素相关回归分析显示,单发病灶、砂粒样强回声(微钙化)、低回声、边界模糊,为诊断甲状腺癌的主要依据。2.甲状腺癌的淋巴结转移常为多分区转移,Ⅲ区(中颈部)为最常见的转移部位;主要声像图特点有:淋巴结增大、变圆,皮髓质分界不清,淋巴门结构破坏或消失;淋巴结内部回声偏强,有时可出现较多细小点状钙化;部分淋巴结内部出现囊性改变。3.彩色多普勒超声对甲状腺良恶性结节的鉴别诊断具有重要意义,能为临床诊断及治疗提供准确信息。第二部分乳头状甲状腺癌的超声表现与骨桥蛋白表达的关系研究背景甲状腺癌是头颈部最常见的恶性肿瘤之一,近年来发病率有增长趋势,乳头状甲状腺癌占所有甲状腺恶性肿瘤的75%以上,其发病机理及生物学过程至今不明。骨桥蛋白(osteopontin,OPN)是一种含有特异RGD序列的分泌型磷酸化糖蛋白,具有促进细胞趋化、黏附和迁移的功能,近年来,OPN与肿瘤的发生发展,特别是它与肿瘤转移的关系日益引起人们的关注。目前,已有文献报道OPN在乳腺癌、结肠癌、胃癌、肝癌、卵巢癌等多种恶性肿瘤中呈高表达,在头颈部癌(包括甲状腺癌)的报道较少见。本文通过研究乳头状甲状腺癌组织中的OPN表达情况及探讨其与超声表现之间的关系,旨在研究乳头状甲状腺癌的分子生物学机制,以期为乳头状甲状腺癌的生物学行为和预后进行评估。目的分别从mRNA和蛋白质水平检测骨桥蛋白(OPN)在乳头状甲状腺癌中的表达,探讨OPN的表达与乳头状甲状腺癌超声表现特征之间的关系。方法1.彩色多普勒超声检查对48例乳头状甲状腺癌患者术前行甲状腺彩色多普勒超声检查,分析其声像图特征。2.实时荧光定量PCR(FQ-PCR)法检测OPNmRNA的表达收集48例乳头状甲状腺癌及对照组(30例甲状腺良性病变及20例正常甲状腺组织)的术后标本,分别从标本组织中提取总RNA,经逆转录反应得到cDNA,以GAPDH基因作为参照,通过实时定量PCR技术检测OPNmRNA的表达。分析OPNmRNA的表达与各项超声表现特征之间的关系。3.免疫组织化学检测取组织切片,用免疫组织化学法行OPN的组织学检测,观察乳头状甲状腺癌及对照组的OPN蛋白表达情况。结果1.乳头状甲状腺癌中OPNmRNA的表达水平为56.82±49.16,高于甲状腺良性病变(19.72±28.56)及正常甲状腺组织(13.76±20.65)(P＜0.05)。2.免疫组织化学结果表明,OPN主要定位于癌细胞的细胞质,乳头状甲状腺癌中OPN蛋白阳性表达高于甲状腺良性病变及正常甲状腺组织(P＜0.05)。3.本组病例超声显示微钙化的乳头状甲状腺癌组织中OPNmRNA的表达水平(68.89±67.96)明显高于超声未显示微钙化者(20.62±15.90),伴淋巴结转移的癌组织OPNmRNA的表达(81.26±67.08)高于无淋巴结转移者(38.83±34.69)(P＜0.05)。超声显示的肿块大小、肿块边界及血流分级与OPN表达无明显相关。结论1.OPN在乳头状甲状腺癌组织中高表达,OPN在乳头状甲状腺癌的发生、发展中起到一定作用。2.乳头状甲状腺癌OPNmPNA的表达量与超声显示的微钙化及淋巴结转移存在相关性,OPN可能参与了微钙化的形成,超声表现在一定程度上反映了肿瘤组织中OPN的表达水平,对评估乳头状甲状腺癌的生物学行为和预后具有一定价值。第三部分乳头状甲状腺癌的超声表现与微血管密度测定的相关研究背景肿瘤新生血管形成在其生长、侵袭转移中起着重要作用,研究肿瘤新生血管一直是目前肿瘤研究的热点。测量微血管密度(MVD)是目前常用的评价肿瘤血管生成情况的方法,MVD能较为精确地反映肿瘤内的血管状态,已被公认为是预测肿瘤预后的一个独立指标。目前认为多数实体肿瘤MVD明显增高,并与其侵袭性有关。CD34是一种新型的微血管标记物,其显示血管内皮细胞时,更具敏感性和特异性,易于观察,用抗CD34抗体作微血管染色较其它标记物能提供更多有用而可靠的信息。目前随着彩色多普勒超声的广泛使用,超声检查已成为临床诊断甲状腺疾病的主要检查手段之一,对良恶性结节的鉴别诊断已积累了一定的经验,但在部分结节的鉴别诊断方面仍有争议。近年来微血管密度测定技术的发展为甲状腺结节性疾病的诊断提供了一个新的方向,MVD可精确标识肿瘤组织内微血管,而超声血流显像则是对肿瘤内血管分布的宏观测估,本研究拟探讨二者的内在联系,以为临床提供更多诊断及评估愈后的信息。目的通过对48例乳头状甲状腺癌及对照组的彩色多普勒超声检查、微血管密度测定,并与病理结果对照,分析声像图表现与微血管密度之间的关系,评价彩色多普勒超声、微血管密度测定在乳头状甲状腺癌中的诊断价值。方法1.彩色多普勒超声检查采用ATL HDI-5000及GE LOGIQ 9彩色超声诊断仪,观查并记录48例乳头状甲状腺癌的声像图表现。二维超声观察甲状腺结节的部位、大小、数目、边界、内部回声及钙化情况、颈部有无肿大淋巴结等。用彩色多普勒血流显像(CDFI)及彩色多普勒能量显像(PDI)技术检测每个结节内部血流情况,将结节内部血流丰富程度按Alder半定量法分为0～Ⅲ级。2.微血管密度测定用抗CD34单克隆抗体作免疫组化染色,检测48例乳头状甲状腺癌、30例甲状腺良性病变及20例正常甲状腺组织中的微血管密度,在200倍视野下(0.739mm~2)由两名计数者以双盲法对每一例标本作微血管计数,取二者的均数为该例的MVD值。结果1.乳头状甲状腺癌的超声表现本组乳头状甲状腺癌最常见的声像图表现为不规则形低回声结节,边界模糊和结节内回声不均质及簇状“砂粒样”强回声。常伴有淋巴结转移。结节内部血流分级:0级9例,Ⅰ级13例,Ⅱ级16例,Ⅲ级10例。2.甲状腺良恶性结节微血管密度值乳头状甲状腺癌组织微血管密度值(126.8±42.1)高于甲状腺良性病变(90.2±38.5)和正常甲状腺组织(73.7±27.6)(P＜0.05),甲状腺良性病变与正常甲状腺组织间差异无统计学意义(P＞0.05)。3.乳头状甲状腺癌超声血流分级与微血管密度不同血流分级微血管密度均值分别为:91.3±35.4(0级),105.5±30.8(Ⅰ级),137.5±43.0(Ⅱ级),169.3±37.2(Ⅲ级)。经方差分析,各级间数值差异有统计学意义(P＜0.05),Ⅱ级显著高于0级和Ⅰ级,Ⅲ级显著高于Ⅱ级。随血流分级的增高,微血管密度显著增高。4.乳头状甲状腺癌声像图特征与微血管密度的相关性体积较大的结节(≥2cm)22例,微血管密度值为140.1±37.8,体积较小的结节(＜2cm)26例,微血管密度值为115.5±43.3,两组间差异有统计学意义(P＜0.05),表明体积较大结节的微血管密度高于较小结节。淋巴结转移阳性组微血管密度值为145.3±44.5,淋巴结转移阴性组为114.7±36.7,经t检验,两组间差别有统计学意义(P＜0.05),表明乳头状甲状腺癌淋巴结转移阳性组微血管密度值高于阴性组。结节边界和砂粒样钙化与微血管密度均无明显相关(P＞0.05)。结论1.乳头状甲状腺癌的微血管密度高于甲状腺良性病变及正常甲状腺组织。2.乳头状甲状腺癌的二维声像图特征中,结节大小及淋巴结转移与微血管密度相关;彩色多普勒超声显示的血流分级能反映微血管密度。3.彩色多普勒超声与微血管密度结合使用,将能更准确地反映肿瘤内的血管生成情况,能为乳头状甲状腺癌的临床诊断及预后评估提供更多信息。更多还原

【Abstract】 BackgroundThyroid cancer is common head and neck malignancies, in recent years, the disease incidence rate had the trend of escalation, the early-time thyroid cancer excised after the surgery, nearly may permanently cure, but the later-period thyroid cancer will have distant place shifts , such as lymph nodes, lungs, bone and other distant metastasis, early diagnosis, early treatment is essential. At present, the clinical diagnosis of thyroid cancer mainly depends on palpation and imaging (high-frequency ultrasound, radionuclide, CT, MRI) and immunoassay, etc, color Doppler ultrasound is currently the most important means of inspection for the thyroid and neck disease, with the development of ultrasound imaging and the application of new technologies, ultrasound can observe the minimal disease of 2-3mm ,and more clearly show the internal structure of nodules, with or without capsule and small structures such as micro-calcification changes, as well as color Doppler signals and Doppler changes in the spectrum, have provided favorable conditions to improve the diagnostic rate of thyroid disease. However, due to different types of pathological thyroid nodules, large differences in biological characteristics, and with multiple and multi-derived characteristics, ultrasonic performance of some nodules is not typical, there are difficulties in diagnosis, and there is a certain proportion of the missed diagnosis and misdiagnosis in the ultrasound examination, differential diagnosis of some benign and malignant nodules still controversial.ObjectiveBy observing the color Doppler ultrasonography performance of 145 cases of thyroid nodules confirmed by surgery and pathology , carried on the single factor and the multi-factor analysis, to obtain the most relevant ultrasonographic features for the diagnosis of thyroid cancer ,to provide a theoretical basis for identifying benign and malignant thyroid nodules easily.MethodsSelected 145 cases of thyroid nodules admitting to our hospital general surgical department from June 2006 to August 2008. all patients were preoperatively with color Doppler ultrasonography, careful observation and recording of two-dimensional and color Doppler ultrasonic performance of lesions was made, including lesion number, size, border, internal echo, sand-like strong echoes, halo sound, rear attenuation, flow classification , flow resistance index,and neck lymph node swelling. After operation and pathology comparison, carried on the single factor and the Logistic multi-factor regression analysis, screened the thyroid cancer related ultrasonic characteristics.Results145 cases of thyroid nodules, including 48 cases of papillary carcinoma, 6 cases of follicular carcinoma, 2 cases of medullary carcinoma, 1 cases of metastatic carcinoma; 69 cases of nodular goiter, 19 cases of thyroid adenoma.88 cases of benign thyroid nodules, 8 cases (9.1%)with fuzzy borders, low-echo 26cases (29.5%), 4 cases with sand-like strong echo (4.5%), 6 cases with rear attenuation (6.8%),5 cases with blood flow RI> 0.7 (5.7%); 57 cases of thyroid cancer , with fuzzy boundaries 39(68.4%); hypoechoic cases 46(80.7%), with sand-like strong echo 35cases (61.4%),21 cases with rear attenuation (36.8%),19cases with flow RI> 0.7 (33.3% ).23 cases with lymph node metastasis (40.4%). Swollen lymph nodes in the middle neck were 76% (19/25). Metastatic lymph nodes showed increasing and round, cortical thickening and medullary eccentric or disappear, internal echogenic heterogeneous, sand-like strong echoes, varying degrees of cystic change, blood flow increasing and disorder.By single factor analysis, to diagnosis the thyroid cancer meaningful ultrasonic characteristics including single lesion. fuzzy border, low-echo, sand-like strong echoes, rear attenuation, high blood flow resistance and neck lymph node swelling.By Logistic regression analysis, thyroid cancer associated sonographic characteristics including: single lesion ,sand-like strong echoes, hypoechoic, and boundary ambiguity. Conclusions1. Multi-factor Logistic regression analysis showed that, the related factors for diagnosis of thyroid cancer includes: single lesion ,sand-like strong echoes (microcalcification), low echo, and boundary ambiguity.2. Lymph node metastasis of thyroid cancer often was multi-division transfer, III District (in the middle neck) as the most common metastatic sites; the main ultrasonic features included: lymph nodes increased, round, corticomedullary demarcation, lymph gate structure destroyed or disappeared; lymph internal echo heterogeneous, and sometimes may appear sand-like strong echoes; some cystic changes occur within lymph nodes.3. Color Doppler ultrasound is of great significance in the differential diagnosis of benign and malignant thyroid nodules , can provide accurate information for the clinical diagnosis and treatment.PART TWO Expression of osteopontin in thyroid papillary carcinoma and its relationship with ultrasonographic features.BackgroundThyroid cancer is common head and neck malignancies, in recent years, the disease incidence rate had the trend of escalation, of all thyroid malignancies papillary thyroid carcinoma accounts for more than 75 percent, and its pathogenesis and biological processes are not yet completely clear. Osteopontin (OPN) is a calcium-binding secreted phosphorylated glycoprotein containing specific RGD- sequence, with the promotion of cell chemotaxis, adhesion and migration functions. At present, OPN has been reported in the breast, colon, stomach, liver, ovarian cancer and other malignant tumors with high expression,but in head and neck cancer (including thyroid cancer) reported rare. In this paper, by studying expression of OPN in papillary thyroid carcinoma and investigating the relationship between expression of OPN and ultrasonic performance of papillary thyroid carcinoma, to study the molecular mechanisms of papillary thyroid cancer ,and to evaluate the biological behavior and prognosis of the tumor.ObjectiveTo investigate the expression of osteopontin(OPN) in human thyroid papillary carcinoma and its relationship with ultrasonographic features.MethodsUltrasonographic scanning was performed in 48 patients with thyroid papillary carcinoma before operation, ultrasonographic features were analyzed. The 48 specimens were collected, the expressions of OPN mRNA and protein were analyzed by using FQ - PCR and immunohistochemical staining respectively. 30 samples of thyroid benign lesions and 20 samples of normal thyroid tissues were selected as control group. Results of OPN expressions were compared with ultrasonographic features.ResultsThe expression levels of OPN mRNA and protein in thyroid papillary carcinoma were higher than those of control. Expression of OPN was significant related with the presence of microcalcification in ultrasonography,the expression of OPN in microcalcification group was higher than that in non-microcalcification group. The expression of OPN in the group with lymph nodes metastasis was higher than that in the group without lymph nodes metastasis. No significant relevance was found between the expression of OPN and other ultrasonographic features(include tumor size, margin and blood flow).ConclusionsThe expression levels of OPN in thyroid papillary carcinoma were higher than that in benign lesions and normal thyroid tissues. OPN may play an important role in the development of thyroid papillary carcinoma ; The expression levels of OPN mRNA was related with the presence of microcalcification and lymph nodes metastasis in ultrasonography. OPN may take part in the formation of the microcalcification, the ultrasonographic features can reflect the expression of OPN, so the biological behavior and prognosis of thyroid papillary carcinoma can be assessed by ultrasonographic features to some extent. PART THREE Microvessel Density Determination in Thyroid Papillary Carcinoma and its Relationship with Ultrasonographic Features.BackgroundTumor angiogenesis plays an important role in its growth, invasion and metastasis, studying tumor angiogenesis in cancer research has been hot, measuring microvessel density (MVD) is now commonly used in evaluation of tumor angiogenesis, MVD can precisely reflect the angiogenesis in the tumor, has been regarded as an independent indicator for prediction of tumor prognosis. At present,in the majority of solid tumors , MVD was significantly increased and related to their invasiveness. CD34 is a new type of microvascular markers, which shows the vascular endothelial cells, the more sensitivity and specificity, ease of observation, with anti-CD34 antibody for microvessel staining can provide more useful and reliable information than the other markers.At present, with the extensive use of ultrasonography ,color Doppler ultrasound has become one of the main inspection means for clinical diagnosis of thyroid disease, on the differential diagnosis of benign and malignant nodules has accumulated certain experience, but in some aspects of differential diagnosis of nodules is still controversial. In recent years , determination of MVD provides a new direction for the development of technology in thyroid nodular disease diagnosis, MVD accurately identifies tumor microvessel, and ultrasonic blood flow classification is the distribution of tumor blood vessels within the macro-test assessment, This study was to investigate the intrinsic link between the two ,and to provide more useful information for clinical diagnosis and assessment.ObjectiveThrough the observation of two-dimensional and color Doppler ultrasonography of 48 cases of papillary thyroid carcinoma and the control group, determination of MVD, compared with pathological results, evaluated color Doppler ultrasound and determination of MVD in the diagnosis of papillary thyroid carcinoma.MethodsColor Doppler ultrasonic examination Using ATL HDI-5000 and GE LOGIQ 9 color ultrasound diagnostic apparatus , observed and recorded the ultrasonography of 48 cases of papillary thyroid carcinoma. Two-dimensional ultrasonic observation of thyroid nodules included location, size, number, boundaries (fuzzy or clear), internal echo ,calcification (with particular attention to the existence of microcalcifications), neck lymph node swelling, and so on. Using color Doppler flow imaging (CDFI) and power Doppler imaging(PDI) to detect blood flow in each nodule, the nodule internal flow was divided into 4 levels(0-III) according to the Alder semi-quantitative methods.Determination of MVDWith anti-CD34 monoclonal antibodies for immunohistochemical staining to detect the MVD in 48 cases of papillary thyroid carcinoma, 30 cases of benign thyroid lesions and 20 cases of normal thyroid tissue, made the capillaries counting in 200 time of fields (0.739mm~2) by two counters for each specimen for microvessel count using double-blind method, took the two’s mean value for one specimen.ResultsUltrasonic performance of Papillary thyroid carcinoma Papillary thyroid carcinoma in this group the most common sonographic performance showed irregular hypoechoic nodules, and nodules with fuzzy boundary , non-homogeneous and cluster "sand-like" strong echo, more than one strip flow points in the majority of nodules, often accompanied by lymph node metastasis. The flow in the nodules is divided into 4 levels according to the Alder semi-quantitative method, grade 0 9 cases , grade I 13 cases , grade II -III 26 cases.MVD in benign and malignant thyroid nodulesMVD of papillary thyroid cancer tissue (126.8±42.1) is higher than that of benign thyroid lesions (95.2±38.5) and normal thyroid tissue (83.7±27.6)(p<0.05). The difference of MVD between benign thyroid lesions and normal thyroid tissue has no significances.Papillary thyroid carcinoma power Doppler grading vs MVDMean MVD were: 97.3±35.4 (grade 0), 104.6±30.8 (grade I), 127.5±43.0 (grade II), 141.3±37.2 (grade III). By analysis of variance at all levels ,the numerical differences were significant (p<0.05),grade II was significantly higher than grade 0 and I, grade III were significantly higher than grade II, with the flow classification increased MVD was significantly higher.Correlation between MVD and lymph node metastasisMVD in lymph node metastasis positive group was 140.3±44.5, lymph node negative group was 116.0±36.7,by t-test, the difference between the two groups was statistically significant (p<0.05),showed that the MVD in papillary thyroid carcinoma lymph node metastasis positive group was higher than that of the negative group.Correlation between papillary thyroid carcinoma sonographic features and MVDMVD of 22 Larger nodules (≥2cm) was 139.4±37.8, 26 smaller nodules (<2cm) was 117.5±43.3, difference between the two groups was statistically significant (P<0.05), showed that larger nodules had higher MVD than smaller nodules. Nodular borders and gravel-like calcification were not significantly associated with MVD.Conclusions1. MVD of papillary thyroid carcinoma was higher than that of benign lesions and normal thyroid tissue.2. Of ultrasonic features , the nodule size and lymph node metastasis were related to MVD significantly. The application of color Doppler Imaging accurately reflect the tumor microvessel situation.3. Color Doppler ultrasound combined with the MVD determination will be able to more accurately reflect the tumor blood supply situation, and provide more information for clinical diagnosis and assessment.更多还原