【作者】 赵冀安；

【导师】 耿翠芝；

【作者基本信息】 河北医科大学 ， 外科学， 2010， 硕士

【摘要】 目的:由于社会经济的原因越来越多的妇女推迟生育孩子的年龄,妇女开始做母亲的年龄不断增大。但是随之而来,妇女乳腺癌的发病率也迅速上升。乳腺癌被认为是多种病原学共同作用的结果,其中公认的一项危险因素是生育。许多流行病学研究证实,未经产妇女同经产妇女相比,乳腺癌发病率明显增高,然而,迄今没有充足的实验数据说明上述结果的正确性。本文通过动物实验研究生育及生育早晚对二甲基苯蒽(DMBA)诱导的SD大鼠乳腺癌发生、发展的影响,探讨生育对乳腺癌发生、发展的相关性,对更新乳腺癌防治观念,降低乳腺癌的发病率起到一定的指导作用。方法:1.本试验分别选取8周龄的健康雌性SD处鼠80只,36周龄的健康雌性SD处鼠40只,12周龄的健康雄性SD大鼠10只,共计130只。将所用120只SD雌鼠随机分为试验组和对照组:①8周龄处鼠组20只(早生育对照组);②8周龄处鼠DMBA组20只(早生育试验组)③36周龄处鼠组20只(晚生育对照组);④36周龄处鼠DMBA组20只(晚生育试验组)⑤8周龄处鼠组20只(未生育对照组);⑥8周龄处鼠DMBA组20只(未生育试验组)。2.所有大鼠在适应饲养一周后开始实验。将8周龄40只和36周龄40只SD雌性处鼠均与雄鼠交配,生育一周后(未给幼仔哺乳),将其中的早生育试验组20只和晚生育试验组20只连同未生育试验组20只处鼠分别按体重分两次给予10mg/100gDMBA灌胃(两次给药间隔2周)。生育对照组和未生育对照组给予正常饮食喂养,不给予任何干预措施。3.试验组DMBA给药时间分别达到24周后连同相应对照组全部处死并解剖,进行肉眼观察,取乳腺及肿瘤组织石蜡包埋,每个蜡块切片进行常规的H-E染色和AgNOR经典染色法进行大鼠乳腺上皮细胞的核仁组成区嗜银染色计数,用免疫组化二步法检测C-erbB-2、PCNA、Ki-67、MCM2在大鼠乳腺上皮细胞中的表达。4.所有计数结果用SPSS 13.0软件进行统计分析。结果:1.发病率:至24周试验结束时,共有32只大鼠发病。未生育DMBA组存活20只,有19只发病,发病率为95.0%;早生育DMBA组存活18只,有3只发病,发病率为16.7%;晚生育DMBA组存活17只,有10只发病,发病率为58.8%。对照组均无发病。经过统计学Fisher确切概率法处理,未生育DMBA组和早生育DMBA组之间、未生育DMBA组和晚生育DMBA组之间、早生育DMBA组和晚生育DMBA组之间的发病率均具有统计学上的差异(分别为p=9.59×10-7、p=0.014、p=0.015)。2.潜伏期:以第一次给予二甲基苯蒽至触诊大鼠发现肿瘤计算潜伏期。至24周试验结束时未生育DMBA组(10.25±0.83)周、早生育DMBA组(17.25±3.20)周;晚生育DMBA组(14.50±0.96)周。经过统计学秩和检验处理,未生育DMBA组和早生育DMBA组之间、未生育DMBA组和晚生育DMBA组之间、早生育DMBA组和晚生育DMBA组之间的发病率均具有统计学上的差异(分别为p=0.001、p=1.90×10-6、p=0.014)。3.肿瘤直径:所有发病大鼠的肿瘤直径以肿瘤的长径计数。未生育DMBA组为(2.86±0.54)厘米,早生育DMBA组肿瘤直径为(2.65±1.67)厘米;晚生育DMBA组(2.41±0.50)厘米。经过统计学秩和检验处理,三组之间的肿瘤大小无统计学差异(p=1.00、p=0.54、p=0.81)。4.AgNOR计数:未生育DMBA组为7.52±1.24;早生育DMBA组为:3.52±3.00;晚生育DMBA组为:4.76±1.20。经过统计学秩和检验处理,未生育DMBA组和早生育DMBA组之间、未生育DMBA组与晚生育DMBA组之间比较具有统计学差异(P=0.009,P=0.008)。早生育DMBA组和晚生育DMBA组之间比较无统计学差异(P=0.287)。5.C-erbB-2蛋白表达:统计学处理时将“-”和“+”归为一组(阴性表达),“++”和“+++”归为一组计算(阳性表达)。未生育DMBA组阴性表达:4,阳性表达:16;早生育DMBA组阴性表达:15,阳性表达:3;晚生育DMBA组阴性表达:12,阳性表达:5。用Fisher确切概率法统计:未生育DMBA组和早生育DMBA组之间、未生育DMBA组与晚生育DMBA组之间比较具有统计学差异(p=0.00023,p=0.003)。早生育DMBA组和晚生育DMBA组之间比较无统计学差异(p=0.443)。6.PCNA的表达:统计学处理时将“-”和“+”归为一组(阴性表达),“++”和“+++”归为一组计算(阳性表达)。未生育DMBA组阴性表达:6,阳性表达:14;早生育DMBA组阴性表达:15,阳性表达:3;晚生育DMBA组阴性表达:13,阳性表达:4。用Fisher确切概率统计:未生育DMBA组和早生育DMBA组之间、未生育DMBA组与晚生育DMBA组之间比较具有统计学差异(p=0.001,p=0.008)。早生育DMBA组和晚生育DMBA组之间比较无统计学差异(p=0.691)。7.Ki-67的表达:统计学处理时将“-”和“+”归为一组,“++”和“+++”归为一组计算。未生育DMBA组阴性表达:3,阳性表达:17;早生育DMBA组阴性表达:15,阳性表达:3;晚生育DMBA组阴性表达:11,阳性表达:6。用Fisher确切概率统计:未生育DMBA组和早生育DMBA组之间、未生育DMBA组与晚生育DMBA组之间比较具有统计学差异(p=5.1×10-5,p=0.003)。早生育DMBA组和晚生育DMBA组之间比较无统计学差异(p=0.264)。8.MCM2的表达:统计学处理时将“-”和“+”归为一组,“++”和“+++”归为一组计算。未生育DMBA组阴性表达:4,阳性表达:16;早生育DMBA组阴性表达:15,阳性表达:3;晚生育DMBA组阴性表达:11,阳性表达:6.用Fisher确切概率统计:未生育DMBA组和早生育DMBA组之间、未生育DMBA组与晚生育DMBA组之间比较具有统计学差异(p=0.00023,p=0.008)。早生育DMBA组和晚生育DMBA组之间比较无统计学差异(p=0.264)。结论:1.给予SD大鼠单次剂量10mg/100g DMBA两次灌胃,可以成功制作大鼠乳腺癌的动物模型,以模拟人体乳腺肿瘤的发生、发展过程,进行有关乳腺癌发生机理的研究和各种干预试验。2.生育可以降低DMBA诱导的SD大鼠乳腺组织的致癌作用。3.未生育SD大鼠对DMBA的敏感性强于生育SD大鼠,致癌率高于生育SD大鼠。4.晚生育SD大鼠对DMBA的敏感性强于早生育SD大鼠,致癌率高于早生育SD大鼠。5.生育可以降低DMBA诱导的SD大鼠乳腺癌组织中AgNOR、C-erbB-2、PCNA、Ki-67、MCM2的表达,预示着有较好的预后。更多还原

【Abstract】 Objective: With the development of society and economy, more and more women postpone the age of the reproduction, the age of being mother is becoming older and older. Meanwhile, the incidence of breast cancer rises quickly. Breast cancer is considered as outcome of many etiology factors effect, it is well known that reproduction is one of the risk factors. Many epidemiology researches have certified that nulliparous women are at increased risk of breast cancer compared with parous women.We study the effect of reproduction and the age of reproduction to epithelial cells of mammary gland induced by DMBA in SD rat, which can help us clear the effect of occurrence and development of breast cancer, to renew the idea of prevention and cure of breast cancer, and direct clinical work to breast cancer.Methods:1.80 healthy female virgin SD rats of 8 weeks、40 healthy female virgin SD rats of 36 weeks and 10 healthy male SD rats of 12 weeks. Each category was radomedly divided into 2 groups: control group ( non DMBA) and experimental group (DMBA):①20 female virgin SD rats of 8 weeks (control group of early reproduction) ;②20 female virgin SD rats of 8 weeks (experimental group of early reproduction);③20 female virgin SD rats of 36 weeks (control group of late reproduction);④20 female virgin SD rats of 36 weeks (experimental group of late reproduction);⑤20 female virgin SD rats of 8 weeks (control group of nulliparity) ;⑥20 female virgin SD rats of 8 weeks (experimental group of nulliparity)2.40 female virgin SD rats of 8 weeks and 40 female virgin SD rats of 36 weeks are mated with male SD rats.When female virgin SD rats breeded, 20 female virgin SD rats of 8 weeks and 20 female virgin SD rats of 36 weeks are feeded with 10mg/100g DMBA twice, as well as another 20 female virgin SD rats of 8 weeks.3.All female rats were killed after 24 weeks. The morphological change was observed in macroscopic observation. Breast tissue get from rats were imbedded with paraffin then stained with H-E Silver-binding nucleoar organizer regions (AgNOR) were counted and the expressions of C-erbB-2、PCNA、Ki-67and MCM2 were detected by S-P method.4.The data was organized and processed with SPSS13.0 statistical software.Results:1.Incidence:There have 32 rats with breast cancer in 55 survival rats, 19 in experimental group of nulliparity (DMBA), 3 in experimental group of early reproduction (DMBA), 10 in experimental group of late reproduction (DMBA), 0 in all control groups. It shows statistical difference between group of nulliparity (DMBA) and group of early reproduction (DMBA)、group of nulliparity (DMBA) and group of late reproduction (DMBA)、group of early reproduction (DMBA) and group of late reproduction (DMBA). (p=9.59×10-7、p=0.014、p=0.015)2.Latency: group of nulliparity (DMBA)(10.25±0.83)weeks; group of early reproduction (DMBA)(17.25±3.20)weeks; group of late reproduction (DMBA)(14.50±0.96)weeks. It shows statistical difference between group of nulliparity (DMBA) and group of early reproduction (DMBA)、group of nulliparity (DMBA) and group of late reproduction (DMBA)、group of early reproduction (DMBA) and group of late reproduction (DMBA). (p=0.001、p=1.90×10-6、p=0.014) 3. Diameter of tumor: group of nulliparity (DMBA()2.86±0.54)cm; group of early reproduction (DMBA)(2.65±1.67)cm; group of late reproduction (DMBA)(2.41±0.50)cm.It shows no statistical difference in three groups.(p=1.00、p=0.54、p=0.81)4.The AgNOR in rat breast epithelioglandular: The AgNOR in rat breast epithelioglandular of group of nulliparity (DMBA)(7.52±1.24)higher than that of group of early reproduction (DMBA)(3.52±3.00)and group of late reproduction (DMBA)(4.76±1.20).(P=0.009,P=0.008)It shows no statistical difference between group of early reproduction (DMBA) and group of late reproduction.(p=0.287)5.The positive expression of C-erbB-2 in rat breast epithelioglandular: The positive expression of C-erbB-2 in rat breast epithelioglandular of group of nulliparity (DMBA) (16/20) higher than that of group of early reproduction (DMBA()3/18)and group of late reproduction (DMBA()5/17) (p=0.00023,p=0.003). It shows no statistical difference between group of early reproduction (DMBA) and group of late reproduction.(p=0.443)6.The positive expression of PCNA in rat breast epithelioglandular: The positive expression of PCNA in rat breast epithelioglandular of group of nulliparity (DMBA) (14/20) higher than that of group of early reproduction (DMBA)(3/18)and group of late reproduction (DMBA)(4/17)(p=0.001,p=0.008 ) . It shows no statistical difference between group of early reproduction (DMBA) and group of late reproduction.(p=0.691)7.The positive expression of Ki-67 in rat breast epithelioglandular: The positive expression of Ki-67 in rat breast epithelioglandular of group of nulliparity (DMBA) (17/20) higher than that of group of early reproduction (DMBA)(3/18)and group of late reproduction (6/17)(DMBA)(p=5.1×10-5,p=0.003). It shows no statistical difference between group of early reproduction (DMBA) and group of late reproduction.(p=0.264)8.The positive expression of MCM2 in rat breast epithelioglandular: The positive expression of MCM2 in rat breast epithelioglandular of group of nulliparity (DMBA) (16/20) higher than that of group of early reproduction (DMBA()3/18)and group of late reproduction (DMBA()6/17)(.p=0.00023,p=0.008 ) It shows no statistical difference between group of early reproduction (DMBA) and group of late reproduction.(p=0.264)Conclusion:1.The animal model of breast cancer in rat can be made by given twice DMBA intragastric administration, to simulate the incidence and development of human breast tumor, and used to study the mechanism in breast carcinogenesis and do many intervention studies.2.Reproduction can decline the incidence of breast cancer which induced by DMBA. It can put off the reproductive experimental group onset time.3.The sensitivity to DMBA in group of nulliparity (DMBA) is greater than group of reproduction, the incidence of breast cancer in group of nulliparity (DMBA) is higher than group of reproduction (DMBA).4.The sensitivity to DMBA in group of late reproduction (DMBA) is greater than group of early reproduction (DMBA), the incidence of breast cancer in group of late reproduction (DMBA) is higher than group of early reproduction (DMBA).5.AgNOR、C-erbB-2、PCNA、Ki-67、MCM2 show that reproduction gives SD rats better prognosis.更多还原