乳腺癌发生模型MCF10各阶段细胞系中TIMP3基因启动子区甲基化分析

目的探讨抑癌基因TIMP3失活与乳腺癌发生和进展的关系。方法用甲基化特异性PCR技术和亚硫酸盐测序技术检测乳腺癌发生模型MCF10的增生细胞系MCF10A、癌前细胞系MCF10AT、导管内癌细胞系MCF10DCIS.com、浸润癌细胞系MCF10CA1a和转移癌细胞系MCF10CA1d、MCF10CA1h中TIMP3启动子区甲基化状态。结果甲基化特异性PCR分析显示,在上述各细胞系中,TIMP3启动子区均呈高度甲基化状态。亚硫酸盐测序显示,在上述各细胞系中,测序区内的68个CG位点几乎全部发生了甲基化,且甲基化累及了绝大部分等位基因。结论TIMP3基因启动子区甲基化在乳腺癌的发生和进展中起重要作用,可能成为早期诊断乳腺癌和判断乳腺癌预后的分子生物学标记。     

乳腺癌发生模型MCF10各细胞系中APC基因启动子区甲基化及mRNA表达检测

目的探讨乳腺癌发生模型MCF10中抑癌基因APC启动子区甲基化状态及其对mRNA表达的影响。方法应用甲基化特异性聚合酶链反应（MSP）及双亚硫酸钠基因测序技术检测MCF10模型的乳腺增生细胞系MCF10A、癌前细胞系MCF10AT、导管内癌细胞系MCF10DCIS.com、浸润癌细胞系MCF10CA1a、MCF10CA1d、MCF10CA1h及经典乳腺癌细胞系MCF-7和正常乳腺组织中APC基因启动子1A甲基化状态,然后用逆转录聚合酶链反应（RT-PCR）和实时PCR技术检测上述样品的mRNA表达水平。结果在MCF10模型的增生细胞系、癌前细胞系、导管内癌细胞系、浸润癌细胞系中,APC基因启动子1A处于低甲基化状态;与正常乳腺组织相比,各细胞系APC基因mRNA表达无明显减少,MCF10AT、MCF10CA1d、MCF10CA1h、MCF10DCIS．com的mRNA表达分别减少0．27、0．96、1．78、2．70、2．03倍,MCF10A和MCF-7分别增加0．02和0．33倍）。结论MCF10模型中乳腺癌的发生发展过程与APC基因启动子区异常甲基化无关。     

乳腺癌发生模型MCF10 抑癌基因NOEY2启动子区甲基化及mRNA表达

DNA甲基化是常见的表观遗传学变化。越来越多的证据表明,抑癌基因启动子区CpG岛甲基化可使其转录沉默,从而解除其抑癌作用,导致肿瘤的发生和发展。NOEY2是近年来发现的抑癌基因,表达于正常乳腺导管上皮细胞和卵巢生发上皮细胞,但在乳腺癌和卵巢癌细胞中表达显著减少或不表达。在本实验中检测乳腺癌发生模型MCF10中不同阶段的细胞系NOEY2基因启动子区CpG岛Ⅰ甲基化状态,并与mRNA表达水平进行比较,以研究乳腺癌的发生机制和早期诊断。     

Epigenetic modifications of human placenta associated with preterm birth: a systematic review

Objective: Preterm birth (PTB) is one of the leading causes of neonatal mortality and morbidity around the world. Epigenetic alterations of the human placenta may be involved in the causal chain of adverse pregnancy outcomes specifically PTB. In this systematic review, we investigated whether epigenetic dysregulation of the human placenta is associated with PTB.

Methods: We searched MEDLINE and EMBASE and systematically reviewed all relevant studies on epigenetic placental modifications in PTB. Two independent reviewers selected controlled human studies published in any language, evaluated their quality, and graded them using the Newcastle–Ottawa Quality Assessment Scale. We resolved disagreements by consensus with a third reviewer.

Results: Eleven observational studies of low to moderate quality met the eligibility criteria out of 60 unique studies. Most studies reported an association between placental epigenetic changes (methylation, mRNA and miRNA) and PTB, although research methods were highly heterogeneous.

Conclusions: Studies reported various associations between specific epigenetic findings and PTB, although methodological concerns limited results’ validity. Additional high quality studies are needed to assess the repeatability of these findings. The STROBE guidelines can be used to improve the quality of reporting.     

MiR‐1253 exerts tumor‐suppressive effects in medulloblastoma via inhibition of CDK6 and CD276 (B7‐H3)

Of the four primary subgroups of medulloblastoma, the most frequent cytogenetic abnormality, i17q, distinguishes Groups 3 and 4 which carry the highest mortality; haploinsufficiency of 17p13.3 is a marker for particularly poor prognosis. At the terminal end of this locus lies miR‐1253, a brain‐enriched microRNA that regulates bone morphogenic proteins during cerebellar development. We hypothesized miR‐1253 confers novel tumor‐suppressive properties in medulloblastoma. Using two different cohorts of medulloblastoma samples, we first studied the expression and methylation profiles of miR‐1253. We then explored the anti‐tumorigenic properties of miR‐1253, in parallel with a biochemical analysis of apoptosis and proliferation, and isolated oncogenic targets using high‐throughput screening. Deregulation of miR‐1253 expression was noted, both in medulloblastoma clinical samples and cell lines, by epigenetic silencing via hypermethylation; specific de‐methylation of miR‐1253 not only resulted in rapid recovery of expression but also a sharp decline in tumor cell proliferation and target gene expression. Expression restoration also led to a reduction in tumor cell virulence, concomitant with activation of apoptotic pathways, cell cycle arrest and reduction of markers of proliferation. We identified two oncogenic targets of miR‐1253, CDK6 and CD276, whose silencing replicated the negative trophic effects of miR‐1253. These data reveal novel tumor‐suppressive properties for miR‐1253, i.e., (i) loss of expression via epigenetic silencing; (ii) negative trophic effects on tumor aggressiveness; and (iii) downregulation of oncogenic targets.     

乳腺癌发生过程中NOEY2基因启动子区甲基化及mRNA表达

目的 探讨乳腺癌发生过程中抑癌基因NOEY2启动子区甲基化状态及其对mRNA表达的影响。方法 应用甲基化特异性PCR及双亚硫酸钠基因测序技术检测MCF10模型中乳腺增生细胞系MCF10A、癌前细胞系MCF10AT、导管内癌细胞系MCFIODCIS．com、浸润癌细胞系MCF10CA1a、MCF10CA1d、MCF10CA1h及正常乳腺组织中NOEY2基因启动子区CpG岛Ⅰ甲基化状态,然后用RT-PCR和实时PCR技术检测上述样品的mRNA表达水平。结果 MCF10模型的增生细胞系、癌前细胞系、导管内癌细胞系、浸润癌细胞系均发生该基因启动子区CpG岛Ⅰ高度甲基化;与正常乳腺组织相比,上述细胞系mRNA表达显著减少。结论 NOEY2基因启动子区高度甲基化及相应的mRNA表达减少是乳腺癌发生过程中的早期事件,与乳腺癌发生有关,可能成为早期诊断乳腺癌的潜在分子生物学标记。     

乳腺癌发生模型MCF10各细胞系中WT1基因启动子区甲基化及mRNA表达研究

目的通过观察乳腺癌发生模型MCF10中WT1基因启动子区甲基化状态和mRNA表达水平,探讨该基因在乳腺癌发生中的作用。方法应用甲基化特异性PCR及双亚硫酸钠基因测序技术检测MCF10模型的乳腺增生细胞系MCF10A、癌前细胞系MCF10AT、导管内癌细胞系MCF10DCIS．com、浸润癌细胞系（MCF10CAla、MCF10CA1d、MCF10CA1h）、经典乳腺癌细胞系MCF7及正常乳腺组织中WT1基因启动子区甲基化状态,然后用逆转录．聚合酶链反应（RT—PCR）和即时定量PCR技术检测上述样品的mRNA表达水平。结果在MCF10模型的增生细胞系MCF10A、癌前细胞系MCF10AT、导管内癌细胞系MCF10DCIS．com、浸润癌细胞系（MCF10CAla、MCF10CA1d、MCF10CA1h）、经典乳腺癌细胞系MCF7中,WT1基因启动子均处于高度甲基化状态。与正常乳腺组织相比,WT1基因mRNA在MCFl0模型的增生细胞系、癌前细胞系、导管内癌细胞系、浸润癌细胞系和经典乳腺癌细胞系MCF7中的表达均有不同程度的增加（MCF10A、MCF10AT、MCF10CAla、MCF10CA1d、MCF10CA1h、MCF10DCIS、MCF7的WT1基因mRNA表达量分别是正常乳腺组织3．23、1．94、4．20、1．53、4．20、4．35、28．69倍）。结论乳腺癌发生过程中WnmRNA的表达不被启动子甲基化所抑制;可WT1mRNA过表达出现于乳腺癌发生的早期阶段,提示该基因在乳腺癌发生中起作用。