5-Ethynyl-2′-deoxyuridine as a molecular probe of cell proliferation for high-content siRNA screening assay by “click” chemistry

Labelling and identification of proliferating cells is important for the study of physiological or pathological processes in high-content screening (HCS) assays. Here we describe ethynyl deoxyuridine (EdU) as a biomarker for the assessment of cell proliferation and clearly demonstrate the feasibility of the EdU-labelling method for use in HCS assays. EdU detection is highly robust, reproducible, technically simple, and well suited for automated segmentation, which provides an excellent al- ternative for setting up multiplexed HCS assays of siRNA, miRNA and small-molecule libraries.

Akt／mTOR通路相关蛋白在卡萨巴赫-梅里特综合征中的表达及意义

目的初步探讨Akt/mTOR通路在卡萨巴赫-梅里特综合征（Kasabach-Merritt syndrome, KMS）发病机制中的作用。方法采用免疫组织化学及免疫印迹方法检测15例KMS病灶组织及15例正常皮肤组织内Akt/mTOR信号通路关键蛋白Akt、pAkt、mTOR、pmTOR、S6、pS6、4E-BP1及p4E-BP1的表达，分析Akt/mTOR通路在KMS发病机制中的作用及意义。结果Western-blot检测结果示，KMS病灶组织中Akt、pAkt蛋白表达强度为0.718±0.034和0.646±0.029，较正常皮肤组织0.382±0.028和0.426±0.027高，且差异均有统计学意义（P＝0.017及0.032）。病灶组织中mTOR、pmTOR蛋白表达强度为0.693±0.018和0.597±0.037，较正常皮肤组织0.491±0.042和0.438±0.015高，且差异均有统计学意义（P＝0.041及0.037）。病灶组织中S6、pS6蛋白表达强度为0.820±0.023和0.463±0.025，较正常皮肤组织0.537±0.018和0.238±0.019高，且差异均有统计学意义（P＝0.029及0.038）。病灶组织中4E-BP1蛋白表达强度为0.580±0.018，较正常皮肤组织0.874±0.031低；p4E-BP1的蛋白表达强度为0.733±0.020，较正常皮肤组织0.430±0.033高，且差异均有统计学意义（P＝0.028及0.035）。免疫组织化学染色结果示，Akt、pAkt、mTOR、pmTOR、S6、pS6、4E-BP1及p4E-BP1蛋白主要表达于细胞胞浆内，呈棕褐色，与正常皮肤组织相比，KMS病灶组织中Akt、pAkt、mTOR、pmTOR、S6、pS6、p4E-BP1蛋白阳性表达率增高，4E-BP1蛋白阳性表达率降低，且差异均有统计学意义（Ridit值分别为0.856、0.789、0.850、0.733、0.659、0.700、0.311和0.690，各组间95%CI值无重叠）。结论Akt、pAkt、mTOR、pmTOR、S6、pS6、p4E-BP1蛋白在KMS病灶组织中表达增高，而4E-BP1蛋白在KMS病灶组织中表达降低，提示Akt/mTOR信号通路在KMS中处于异常激活状态，Akt/mTOR信号通路可能参与KMS的发生发展。