【作者】 李曦洲；

【导师】 施俊义； 郑建明； 倪灿荣； 李莉； 胡薇；

【作者基本信息】 第二军医大学 ， 外科学， 2010， 硕士

【摘要】 实验背景及目的乳腺癌是妇女常见的恶性肿瘤之一。全球每年约有120万名妇女患乳腺癌,上海已经成为全国乳腺癌发病率最高的城市。乳腺癌的发病机制尚不明确,随着诊断、治疗手段不断完善及普查普治工作的开展,患者生存期大幅提高,但乳腺癌的治疗、预后中仍存在许多疑问。例如相同组织病理类型、相同分期、相同治疗的患者预后不尽相同,这提示,乳腺癌存在基因水平的差异,随之也诞生了乳腺癌的基因分型。近期,国外研究表明,某些蛋白因子不仅在乳腺癌的发生、发展中发挥作用,在乳腺癌的基因分型、治疗、预后中也起到十分重要的作用。因此,有必要对这些蛋白因子在乳腺癌中的作用进行研究,进而指导乳腺癌的临床治疗和预后判断。骨桥蛋白(osteopontin, OPN)是一种具有多种功能分泌型钙结合磷酸化糖蛋白,它能与骨组织中的羟磷灰石紧密结合,参与调节骨钙的沉积。其分子量为41.5kD,存在于人的尿、乳汁等体液以及胃肠道、膀胱、胰腺、肺、支气管等组织中。在胃癌、肝癌、卵巢上皮性癌与子宫内膜癌等多种恶性肿瘤组织中高表达,与肿瘤的黏附、侵袭及转移等恶性行为密切相关。本研究应用免疫组化检测99例乳腺癌石蜡包埋标本及20例正常乳腺组织活检标本,分析骨桥蛋白在乳腺癌各分子分型中的表达及骨桥蛋白与乳腺癌病理类型、临床分期、有无淋巴结转移、预后等临床诊断的关系,探讨其临床意义。实验方法采用免疫组化法检测99例乳腺癌组织以及20例正常乳腺组织中OPN的表达。分析OPN在乳腺癌组织与正常乳腺组织中、乳腺癌各分子分型中、不同病理类型间OPN表达的差异,对OPN肿瘤组织学分级、淋巴结转移情况及预后的关系进行相关性检验和分析。实验结果免疫组化检测结果：1.正常乳腺组织20例、乳腺导管内癌25例、浸润性导管癌58例、浸润性小叶癌16例,OPN阳性表达率分别是0、20.0%、43.1%、37.5%,各类型间OPN表达阳性率不完全相等,差异有统计学意义(P<0.05)。其中,乳腺导管内癌组(20.0%)与浸润性导管癌组(43.1%)比较,差异具有统计学意义(P(0.05)其余两两比较均无统计学差异。2.在浸润性导管癌中,58例浸润性导管癌中肿瘤组织学分级Ⅰ级21例、Ⅱ级19例、Ⅲ级18例。Ⅰ级组中OPN表达阳性率为14.3%；Ⅱ级组为42.1%、Ⅲ级组为77.8%,OPN的表达与浸润性导管癌中肿瘤组织分级差异有统计学意义(P<0.05)。3.58例浸润性导管癌中肿瘤cTNM分期Ⅰ期19例、Ⅱ期21例、Ⅲ+Ⅳ期18例。Ⅰ期组中OPN表达阳性率为10.5%；Ⅱ期组为47.6%、Ⅲ+Ⅳ期组为72.2%,OPN的表达与浸润性导管癌cTNM分期差异有统计学意义(P<0.05)。4.58例浸润性癌中,淋巴结转移25例。淋巴结阳性组OPN表达阳性率为66.7%；淋巴结阴性组为22.6%,OPN表达与浸润性乳腺癌中淋巴结转移情况有差异(P<0.05),OPN高表达与腋淋巴结转移有关。5.OPN在不同家族史组、年龄组、原发肿瘤大小组异常阳性表达率差异无统计学意义(P>0.05)。6.99例乳腺癌中luminal A型45例,luminal B型27例,HER-2过表达型14例,basal-like型13例, OPN阳性表达率36.4%(36／99)。OPN在luminal A型,luminal B型,HER-2过表达型和basal-like型表达率分别为22.2%,29.6%,71.4%,和61.5%。各型间OPN表达阳性率不完全相等(P=0.001)。依据ER状态,分为两组,luminal A型与luminal B型为ER阳性组、HER-2过表达型与basal-like型为ER阴性组。ER阳性组(HER-2过表达型与basal-like型)的OPN表达率高于ER阴性组(luminal A型与uminal B型),差异有显著的统计学意义(P<0.01).7.本组全部99病例,失访6例,随访率93.9%,中位随访时间约5.1年。其中OPN阳性组33例,死亡18例,生存率为45.5%；OPN阴性组60例,死亡8例,生存率96.7%。经χ2检验分析,OPN低表达的患者生存率高于OPN高表达者,两者有显著的统计学差异(P<0.01)实验结论OPN在乳腺癌及正常乳腺组织中的表达有显著差异,其可能与某类乳腺癌的发生相关。在浸润性导管癌中,OPN表达与家族史组、年龄组、原发肿瘤大小无关,与不同组织学分级、临床TNM分期、淋巴结转移状态有关。临床分期越晚,OPN的阳性表达率越高；有淋巴结转移者,OPN的阳性表达率也越高。提示OPN阳性表达的乳腺癌细胞具有更强的侵袭、转移能力。淋巴结转移是预后不良的重要指针,进一步说明OPN的表达对乳腺浸润性导管癌的病变发展和预后有重要的意义,定量测量其表达可作为乳腺癌预后的评估指标之一。此外,实验结果提示OPN表达与乳腺癌分子分型密切相关,ER阳性组(HER-2过表达型与basal-like型)的OPN表达率高于ER阴性组(luminal A型与uminal B型),其在乳腺癌的分子分型中的价值可以得到肯定。更多还原

【Abstract】 Background and ObjectivesBreast cancer is one of the primary malignant tumor, which is endangering women’s health。There are 1,200,000 new cases for the total in the world each year. Shanghai has become the city with the highest incidence of breast cancer. Although, the pathogenesis of breast cancer is not clear yet, as the development of diagnosis and treatment and the effort of general check-up and treatment, the survival period has prolonged. But, it still has a lot of problems such as the treatment and prognosis of breast cancer. For instance, the patients with the same histopathological type by the same stages under the same treatment have different prognosis. It indicates that it has difference of gene grade for breast cancer, and its gene typing forms accordingly. Recently, the overseas studies show that some protein factors not only have effects in the process of breast cancer but also play important roles in the gene typing, therapy and prognosis. Therefore, it’s necessary to study on the function of protein factors in breast cancer, which will help instruct the clinical therapy and have evaluation of prognosis.Osteopontin (osteopontin, OPN) is a multi-functional secretory calcium-binding phosphorylated glycoprotein, which in bone tissue and integrated hydroxyapatite closely, involved in the regulation of bone deposition. Its molecular weight of 41.5kD, exist in people’s urine, milk and other body fluids as well as the gastrointestinal tract, bladder, pancreas, lung, bronchial and other organizations. In gastric cancer, liver cancer, epithelial ovarian cancer and endometrial cancer, cervical cancer and other malignant tissues of high expression with tumor adhesion, invasion and metastasis, such as closely related to malignant behavior. This study is to apply immunohistochemical method to check 99 cases of paraffin-embedded breast cancer samples and 20 cases of normal biopsy specimens of breast tissue samples, analyze the expression of OPN in molecular classification and the relationship between OPN and clinical diagnosis such as pathology type of breast cancer, clinical staging, with or without lymph node metastasis and prognosis, and discuss its clinical significance.MethodApply immunohistochemical method to check the expression of OPN in 99 cases of breast cancer tissue and 20 cases of normal breast tissue. Analyze the diversity of OPN expression in breast cancer tissue, molecular classification of breast cancer and different pathology types. Check and analyze the relationship between grading of OPN tumor histology, condition of lymph node metastasis and prognosis.ResultsImmunohistochemistry results:1.20 cases of normal breast tissue, ductal carcinoma in 25 cases,58 cases of infiltrating ductal carcinoma, invasive lobular carcinoma in 16 cases, OPN expression positive rates were 0,20.0%,43.1%,37.5%, among the various types of OPN positive expression was not completely the same, the difference was statistically significant (P<0.05). Among them, breast ductal carcinoma in situ group (20.0%) and invasive ductal carcinoma group (43.1%), the difference was statistically significant (P<0.05), the remaining two groups were no statistical differences in comparison.2. In the invasive ductal carcinoma,58 cases of invasive ductal carcinoma of histological grade in 21 cases gradeⅠ,Ⅱgrade 19 cases,Ⅲgrade 18 cases.Ⅰ-level group, OPN expression positive rate of 14.3%;Ⅱ-class group,42.1%,Ⅲ-class group,77.8%, OPN expression in invasive ductal carcinoma with tumor grade, the difference was significantly (P<0.05).3.58 cases of invasive ductal carcinoma tumor cTNM period of 19 patients with stageⅠ,Ⅱperiod of 21 cases,Ⅲ+Ⅳperiod of 18 cases. PhaseⅠgroup, OPN expression positive rate of 10.5%;Ⅱphase group,47.6%,Ⅲ+Ⅳphase group 72.2%, OPN expression and invasive ductal carcinoma cTNM staging, the difference was significantly (P<0.05).4.58 cases of invasive carcinoma, lymph node metastasis in 25 cases. OPN expression in lymph node positive group was positive rate was 66.7%; lymph node-negative group was 22.6%, OPN expression and invasive breast cancer, there are differences in lymph node metastasis (P<0.05), OPN high expression of axillary lymph node metastasis.5. OPN in different family history group, age group, primary tumor positive expression rate of abnormal large group has no statistically significant difference (P> 0.05).6.99 cases of breast cancer in the luminal A type 45 cases, luminal B-type in 27 cases, HER-2 over-expression of type 14 cases, basal-like type in 13 cases, OPN-positive expression rate of 36.4%(36/99). OPN at the luminal A type, luminal B-type, HER-2 over-expression of type and basal-like type expression rates were 22.2%,29.6%,71.4% and 61.5%. Between various types of OPN, positive expression rate was not completely the same (P=0.001). Based on ER status, they were divided into two groups, luminal A-type and luminal B type of ER-positive group, HER-2 over-expression of type and basal-like model for the ER negative group. ER-positive group (HER-2 over-expression of type and basal-like type) was higher than that of OPN expression in ER-negative groups (luminal A-type and luminal B type), the difference was significant statistically significant (P <0.01).7. This group all 99 cases,6 patients lost to follow-up rate of 93.9%, median follow-up time is about 5.1 years. OPN-positive group in which 33 cases of death in 18 cases, survival rate was 45.5%; OPN-negative group,60 cases of death in 8 cases, survival rate was 96.7%. Theχ2 tests and analysis, survival rates of OPN low expression in patients were higher than the high expression of OPN, both statistically significant difference (P <0.01)ConclusionThe expression of OPN has significant difference between breast cancer and normal breast tissues, which might be associated with the certain type of breast cancer. As to infiltrating ductal carcinoma, the expression of OPN is negated with family history, age group and primary tumor grade; it positive correlate with different histological grade, staging of clinical TNM, The later the clinical staging is, the higher the positive expression rate of OPN will be. The positive expression rate of OPN is higher with those who suffer from lymph node metastasis. It shows that the breast cancer cell with positive expression rate of OPN has stronger capability for invasion and metastasis. Lymph node metastasis is a key index for poor prognosis, which further indicates that the expression of OPN plays important roles in the development of infiltrating ductal breast cancer and that of prognosis. The quantitative measurement of expression could be acted as one of the evaluation index for prognosis of breast cancer. moreover, the experiment results suggest that OPN expression is closely related to breast cancer molecular type, ER-positive group (HER-2 over-expression of type and basal-like tpye) was higher than that of OPN expression in ER negation in ER-negative group(luminal-A-type and luminal B type), its affirmation of value in the molecular classification of breast cancer.更多还原