胚胎干细胞技术的研究进展

干细胞可以分为成体干细胞和胚胎干细胞两类,是当前生物医学领域的一个热点。由于干细胞在医学领域有巨大应用前景,世界各国都对此给予了相当的重视;投入大量资金支持本国的科学家从事干细胞的科学研究。目前,干细胞的研究还处于试验室阶段,很多技术难题还没有解决,尤其是胚胎干细胞的不定向分化问题,以及一些伦理方面的争议。宗教界人士和社会论理学方面的人士对于干细胞的研究还有异议。集中论述了胚胎干细胞的研究进展和医学功用。     

Quantitative identification of teratoma tissues formed by human embryonic stem cells with TeratomEye

An automated vision system, TeratomEye, was developed for the identification of three representative tissue types: muscle, gut and neural epithelia which are commonly found in teratomas formed from human embryonic stem cells. Muscle tissue, a common structure was identified with an accuracy of 90.3% with high specificity and sensitivity greater than 90%. Gut epithelia were identified with an accuracy of 87.5% with specificity and sensitivity greater than 80%. Neural epithelia which were the most difficult structures to distinguish gave an accuracy of 47.6%. TeratomEye is therefore useful for the automated identification of differentiated tissues in teratoma sections.     

多能干细胞分化来源视网膜色素上皮细胞移植治疗视网膜变性研究进展

视网膜色素上皮（RPE）对视觉功能的维持起着至关重要的作用。视网膜变性是全球不可治愈性致盲疾病的重要原因,它由视网膜色素上皮功能失常所引起。因此,视网膜色素上皮移植是视网膜变性患者恢复视力的一种最有前景的手段之一。随着干细胞技术的快速发展,从多能干细胞（PSC）到有功能的视网膜色素上皮细胞的体外分化诱导技术已经成熟,其中包括胚胎干细胞（ESCs）和诱导多能干细胞（iPSCs）等。此外,从患者特异性iPSCs分化而来的RPE更能用于阐明发病机理并有针对性地个体治疗。更值得一提的是,经诱导得到RPE的移植不论在动物模型中,还是在临床试验里都已经得到了可喜的治疗效果。本文回顾PSC来源RPE干预治疗视网膜变性的最新研究进展。     

从胚胎干细胞到视网膜

视网膜退行性病变影响着全世界数百万人。然而,视网膜是人体再生能力很差的一类组织,成年机体无法自我更新那些病变中丢失的视网膜细胞,导致视网膜退行性病变的不可逆性。因此,恢复患者视觉将依赖于引入外源细胞替代丢失的视网膜神经元。胚胎干细胞（ES细胞）具有无限的自我更新能力和形成机体所有类型细胞的巨大分化潜力。这两个特性使得ES细胞成为细胞替代疗法的理想供体细胞。近年来,人们在探索将ES和诱导多能干细胞（iPS细胞）体外定向诱导分化为视网膜神经元,甚至整个视网膜方面已取得多项进展,并且体外形成的视网膜细胞可以与宿主视网膜整合。在此篇综述中,首先简要概括哺乳动物视网膜的组织结构、发育过程和调控机制,然后,重点阐述近年来科研工作者探索ES/iPS细胞体外诱导分化为视网膜细胞和组织的研究进展。     

The cardiac differentiation of bone marrow stem cells might be not important for heart repair

Lots of evidence showed that bone marrow stem cells can differentiate into cardiac myocytes so as to treat damaged hearts. However, the following studies revealed that bone marrow stem cells also produced protective effects on hearts by releasing some beneficial cytokines and suppressing inflammatory effects and so on. Therefore, we speculated that the cardiac differentiation of bone marrow stem cells did not play an important role in cardiac repair.     

多能干细胞诱导分化为肾脏类器官的分子机制及应用前景分析

干细胞具有自我更新和多向分化潜能,在再生医学领域发挥着越来越大的作用。肾脏类器官是一种由干细胞分化而来具有一定肾脏功能的组织结构,可用于肾脏疾病的细胞修复治疗,也可以模拟肾脏发育和疾病发生及用于筛选改善肾功能的药物。肾脏类器官的体外培育成为了当前研究热点,其体外培育可分为几个阶段：干细胞-原始体节中胚层-中间中胚层-输尿管芽（后肾间质）-集合管（肾单位）。本文重点介绍了目前两种较为成熟的肾脏类器官体外诱导方法,并对肾脏类器官的应用前景进行了综述。     

多能干细胞向雄性生殖细胞定向分化的研究进展

生殖健康是生命科学领域关注的核心之一,各种原因所致男性不育亟待解决,然而由于伦理限制等原因,缺少合适的具有人类遗传背景的研究模型开展研究。胚胎干细胞（ESCs）和诱导多能干细胞（iPSCs）均属于多能干细胞,具有多向分化潜能。一方面,可利用ESCs?/?iPSCs向生殖细胞分化的模型研究人类生殖细胞的发育规律,另一方面,在此基础上,可建立带有人类疾病遗传背景的iPSCs模型,体外诱导其向雄性生殖细胞分化,利用该模型研究男性不育的发病机制。由于精子在体内的形成遵循一定规律,体外环境中不同发育阶段的生殖细胞在不同诱导因子作用下才可稳定地往下一阶段定向分化,因此,诱导ESCs?/?iPSCs向雄性生殖细胞方向分化时,诱导因子的种类和加入时间的选择应根据生殖细胞的体内发育特征而定,并且在诱导的不同阶段循序加入,以此模拟精子在体内的形成过程,进而更好地研究男性不育的发病机制。本文将对多能干细胞向雄性生殖细胞定向分化的常用诱导因子及存在问题和展望进行综述,为相关研究的开展提供借鉴。     

Development of a decoy immunization strategy to identify cell-surface molecules expressed on undifferentiated human embryonic stem cells

Little is known about the cell-surface molecules that are related to the undifferentiated and pluripotent state of human embryonic stem cells (hESCs). Here, we generated a panel of murine monoclonal antibodies (MAb) against undifferentiated hESCs by a modification of a previously described decoy immunization strategy. H9 hESCs were differentiated in the presence of retinoic acid and used as a decoy immunogen. Twelve Balb/c mice were immunized in the right hind footpads with differentiated H9 cells and in the left hind footpads with undifferentiated H9 cells. After immunization, the left popliteal lymph node cells were collected and were fused with mouse myeloma cells. The fusion resulted in 79 hybridomas secreting MAbs that bound to the undifferentiated H9 cells as shown by flow cytometric analysis. Of these, 70 MAbs bound to the undifferentiated H9 cells, but only weakly or not at all to the differentiated H9 cells. We characterized 37 MAbs (32 IgGs, 5 IgMs) recognizing surface molecules that were down-regulated during embryoid body cell formation. One of the MAbs, L125-C2, was confirmed to immunoprecipitate CD9, previously known as a surface molecule on the undifferentiated hESCs. To investigate the relationship between the MAbs and hESC-specific antibodies, two representative MAbs, viz., L125-C2 and 291-D4, were selected and studied by multi-color flow cytometric analysis. This showed that more than 60% of L125-C2- and 291-D4-positive cells were also positive for the expression of hESC-specific surface molecules such as SSEA3, SSEA4, TRA-1-60, and TRA-1-81, indicating the close relationship between the two MAbs and the hESC-specific surface molecules. Our results suggest that the decoy immunization strategy is an efficient method for isolating a panel of MAbs against undifferentiated hESCs, and that the generated MAbs should be useful for studying the surface molecules on hESCs in the pluripotent and undifferentiated state.     

CD44+CD24-/low乳腺癌干细胞的流式分选及其肿瘤干细胞特性的初步鉴定

目的:通过表面标志分选法富集乳腺癌干细胞,并初步鉴定其肿瘤干细胞特性。方法:采用流式细胞分选术从人乳腺癌细胞系MCF-7中分选CD44+CD24-/low乳腺癌干细胞,并进行干细胞比例分析;用免疫荧光法检测、比较分选获得的细胞和对照细胞的干性和分化标记物Oct-4、SOX-2、CK-18和α-SMA的表达状态。结果:分选获得的CD44+CD24-/low乳腺癌干细胞阳性比例达90%以上;免疫荧光检测结果显示,CD44+CD24-/low细胞亚群比non-CD44+CD24-/low细胞亚群高表达干细胞转录因子Oct-4、SOX-2,低表达分化因子CK-18、α-SMA;体外实验表明,CD44+CD24-/low细胞亚群具有更强的成球生长能力,并具有双向分化潜能。结论:CD44+CD24-/low表面标记物分选的方法可以富集高纯度的乳腺癌干细胞,且呈现干性因子Oct-4和SOX-2高表达。     

Dendritic cells derived from pluripotent stem cells: Potential of large scale production简

Human pluripotent stem cells (hPSCs), including human embryonic stem cells and human induced pluripotent stem cells, are promising sources for hematopoietic cells due to their unlimited growth capacity and the pluripotency. Dendritic cells (DCs), the unique immune cells in the hematopoietic system, can be loaded with tumor specific antigen and used as vaccine for cancer immunotherapy. While autologous DCs from peripheral blood are limited in cell number, hPSC-derived DCs provide a novel alternative cell source which has the potential for large scale production. This review summarizes recent advances in differentiating hPSCs to DCs through the intermediate stage of hematopoietic stem cells. Step-wise growth factor induction has been used to derive DCs from hPSCs either in suspension culture of embryoid bodies (EBs) or in co-culture with stromal cells. To fulfill the clinical potential of the DCs derived from hPSCs, the bioprocess needs to be scaled up to produce a large number of cells economically under tight quality control. This requires the development of novel bioreactor systems combining guided EB-based differentiation with engineered culture environment. Hence, recent progress in using bioreactors for hPSC lineage-specific differentiation is reviewed. In particular, the potential scale up strategies for the multistage DC differentiation and the effect of shear stress on hPSC differentiation in bioreactors are discussed in detail.     

Dendritic cells derived from pluripotent stem cells:Potential of large scale production

Human pluripotent stem cells(hPSCs), including human embryonic stem cells and human induced pluripotent stem cells, are promising sources for hematopoietic cells due to their unlimited growth capacity and the pluripotency. Dendritic cells(DCs), the unique immune cells in the hematopoietic system, can be loaded with tumor specific antigen and used as vaccine for cancer immunotherapy. While autologous DCs from peripheral blood are limited in cell number, hPSC-derived DCs provide a novel alternative cell source which has the potential for large scale production. This review summarizes recent advances in differentiating hPSCs to DCs through the intermediate stage of hematopoietic stem cells. Step-wise growth factor induction has been used to derive DCs from hPSCs either in suspension cultureof embryoid bodies(EBs) or in co-culture with stromal cells. To fulfill the clinical potential of the DCs derived from hPSCs, the bioprocess needs to be scaled up to produce a large number of cells economically under tight quality control. This requires the development of novel bioreactor systems combining guided EB-based differentiation with engineered culture environment. Hence, recent progress in using bioreactors for hPSC lineage-specific differentiation is reviewed. In particular, the potential scale up strategies for the multistage DC differentiation and the effect of shear stress on hPSC differentiation in bioreactors are discussed in detail.     

Feeder-free Derivation of Neural Crest Progenitor Cells from Human Pluripotent Stem Cells

Human pluripotent stem cells (hPSCs) have great potential for studying human embryonic development, for modeling human diseases in the dish and as a source of transplantable cells for regenerative applications after disease or accidents. Neural crest (NC) cells are the precursors for a large variety of adult somatic cells, such as cells from the peripheral nervous system and glia, melanocytes and mesenchymal cells. They are a valuable source of cells to study aspects of human embryonic development, including cell fate specification and migration. Further differentiation of NC progenitor cells into terminally differentiated cell types offers the possibility to model human diseases in vitro, investigate disease mechanisms and generate cells for regenerative medicine. This article presents the adaptation of a currently available in vitro differentiation protocol for the derivation of NC cells from hPSCs. This new protocol requires 18 days of differentiation, is feeder-free, easily scalable and highly reproducible among human embryonic stem cell (hESC) lines as well as human induced pluripotent stem cell (hiPSC) lines. Both old and new protocols yield NC cells of equal identity.     

成体干细胞肺重建研究进展

成体干细胞来源广泛,无伦理争议,成为近几年的关注热点。研究表明以骨髓来源的间充质干细胞为代表的成体干细胞具有较强的多系分化潜能,可以广泛的参与包括肺在内的受损组织的修复与重建。在动物实验中已观察到,供体来源的成体干细胞可以定向分化为受损肺组织的多种功能细胞,并且有抑制纤维化等病变产生的能力。在本文中,回顾了近年来与肺损伤重建和疾病治疗相关的干细胞研究的最新进展,并探讨了成体干细胞治疗肺疾病与损伤的临床应用前景。     

诱导多能干细胞最新进展

通过导入特定基因诱导完全分化的体细胞重编程为诱导多能干细胞(iPS),这为干细胞的研究及应用带来了革命性的变化。短短3年时间,细胞重编程的机理研究、探索疾病的发生机制以及临床医学的应用等领域取得了很多突破性的进展,主要从iPS诱导机理、效率以及诱导新技术上作一综述,以期对iPS的研究提供参考。