诱导多能干细胞向雄性生殖细胞分化诱导物的研究进展

诱导多能干细胞(induced pluripotent stem cells,iPSCs)是利用细胞重编程技术人工获得的与胚胎干细胞(embryonic stem cells,ESCs)功能类似的细胞,能分化成包括三胚层在内的所有细胞类型,并且规避了ESCs的伦理学争议和移植后的免疫排斥问题,具有十分广阔的应用前景。对iPSCs体外诱导为生殖细胞所用的诱导物及其诱导效果进行了综述,生殖细胞发育机制的研究有望促进未来生殖和发育技术的进步。     

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。     

诱导多能干细胞向巨噬细胞分化研究进展

诱导多能干细胞 (Induced pluripotent stem cells,iPSCs) 是通过体细胞重编程得到类似胚胎干细胞特性的一种细胞类型。通过iPSCs的体外分化,可以了解巨噬细胞的进化历史和各种特性。iPSCs来源的巨噬细胞不仅是药物筛选的良好模型,也是进行免疫治疗的重要手段。本文综述了近年来iPSCs及其向巨噬细胞分化的相关研究进展、所面临的问题以及未来的发展方向。     

人多能干细胞来源的肾定向分化平台的构建

肾是一种重要的人体器官,具有多种生理功能。然而,全球范围内约有10%的人口患有肾疾病。因此,建立一种接近人体肾的结构与功能的模型进行肾疾病的研究是十分必要的。多能干细胞体外定向诱导分化技术的兴起,为再生医学和精准医学领域注入了新的动力。本研究通过在体外条件下模拟体内肾发育的过程,将人多能干细胞包括胚胎干细胞和诱导多能干细胞,通过体外定向诱导分化形成肾的祖细胞,进而建立肾的结构与功能单位：肾元。该研究通过激活WNT信号通路,同时抑制TGF-β信号通路,将人多能干细胞从多能态定向诱导至原条阶段。之后通过细胞自分化的能力使其发育至中间中胚层,再通过激活FGF信号通路,将其分化至肾祖细胞阶段。流式细胞检测结果显示,肾祖细胞占总细胞数的51.5%～61.9%。通过免疫荧光检测发现：分化得到的结构中包含肾小球足细胞、近端小管、远端小管等肾组织结构。该研究建立的肾体外分化方法,具有稳定性好、分化效率高、重复性好的特点。为研究人类肾的早期发育机制,肾疾病模型构建,以及药物筛选提供了一种新的方法。     

人诱导多能干细胞来源功能性胰岛β细胞的体外定向分化方法的建立

1型糖尿病是由胰岛β细胞功能受损、胰岛素分泌不足所致,目前,主要通过外源性胰岛素补充来治疗,但外源性胰岛素无法精准调控血糖,严重低血糖可危及生命。胰岛移植是一种替代疗法,但面临器官供体不足和异种来源胰岛β细胞存在人畜共患病交叉感染风险的问题。因此,获得足量且安全的胰岛β细胞是1型糖尿病细胞治疗面临的难题。本研究旨在通过人诱导多能干细胞(human induced pluripotent stem cells, hiPSCs)在体外向胰岛β细胞分化,提供一种潜在的1型糖尿病治疗新策略。为实现这一目标,我们采用了结合2D和3D培养系统的分化策略,模拟胰岛β细胞的体内发育环境,并使用多种生长因子调节在胰腺发育和β细胞分化中发挥重要作用的关键信号包括Notch信号通路(Notch signaling pathway)、Wnt信号通路(Wnt signaling pathway)、TGF-β/Smad信号通路(TGF-β/Smad signaling pathway)等,在体外将hiPSC定向诱导分化至胰岛β细胞。结果显示,在2D、3D结合的培养条件下,分化过程中定型内胚层细胞,胰腺祖细胞,胰腺...     

胚胎干细胞及诱导多能干细胞向心肌细胞分化和调控的研究进展

胚胎干细胞（embryonic stem cells,ESCs）具有自我更新、无限增殖和多向分化的特性,包括分化成心脏组织的多种类型细胞。经体细胞重编程产生的诱导多能干细胞（induced pluripotent stem cells,iPS）也被证明有类似胚胎干细胞的特性。但这些多能干细胞向心肌细胞自发分化的效率非常低,因此,如何有效地诱导这些多能干细胞向心肌细胞的定向分化对深入认识心肌发生发育的关键调控机制和实现其在药物发现和再生医学,如心肌梗塞、心力衰竭的细胞治疗以及心肌组织工程中的应用均具有非常重要的意义。该文重点综述了近年来胚胎干细胞及诱导多能干细胞向心肌细胞分化和调控的研究进展,并探讨了这一研究领域亟待解决的关键问题和这些多能干细胞的应用前景。     

BMP7对小鼠诱导多能干细胞骨向分化作用的研究

目的:探讨骨形态蛋白(bone morphogenetic protein,BMP)超家族成员之一BMP7在小鼠诱导多能干细胞(Induced pluripotent stem cells,iPS)骨向分化过程中的作用。方法:本试验分成三组,分别是自发分化组,骨诱导组和添加BMP7的骨诱导组。每天观察各组细胞形态学特征及生长状况的差异,在诱导第14天通过茜素红染色检测基质矿化情况,判断BMP7在体外骨诱导条件下对小鼠iPS细胞骨向分化过程所发挥的作用。结果:完成了小鼠iPS细胞的培养鉴定,并诱导形成理想状态的拟胚体(Embryoid body,EB)用于分化接种。结果发现,添加BMP7的骨诱导组细胞的矿化结节阳性率明显增加。结论:BMP7在诱导小鼠iPS细胞骨向分化过程中起促进作用,而对非骨向分化的细胞无成骨促进作用。     

多顺反子质粒重编程人脂肪干细胞为诱导多能干细胞

为建立多顺反子质粒载体转染技术获得人脂肪干细胞(adipose stem cells,ASCs)来源的诱导多能干细胞(induced pluripotency stem cells,iPSCs),应用2A元件连接Oct4/Sox2/KLF4/c-Myc四因子基因,构建为单一开放阅读框的多顺反子质粒载体．使用该质粒对ASCs进行转染及重编程为iPSC．采用形态学观察、特异性抗体免疫荧光鉴定、体外拟胚体诱导分化和体内畸胎瘤形成等方法进行鉴定．结果显示,ASCs成功重编程为iPSCs,具有与人胚胎干细胞相似的形态学及多向分化潜能;通过拟胚体和畸胎瘤实验证实iPSCs能在体内外分化成三胚层细胞;DNA印迹实验显示质粒载体序列未整合至iPSCs基因组中．因此,通过多顺反子质粒载体重编程技术成功建立的人iPSCs具有多向分化潜能,可减免发生插入突变和免疫排斥问题,为iPSCs在遗传性或退行性疾病的治疗奠定了实验基础．     

诱导多能干细胞研究进展

人类诱导多能干细胞（induced pluripotent stem cells,iPS细胞）的建立被公认为目前最重要的科技进展之一。iPS细胞在动物疾病模型上的成功治疗,病患特异性iPS细胞的研究及iPS细胞的定向分化研究将有可能使人们避开治疗性克隆的伦理和技术障碍,给人类疾病的干细胞治疗带来光明的前景。本文从iPS细胞的诱导策略和方法,来源细胞及筛选、重编程机制的研究现状、应用前景以及研究中存在的问题等方面对其作一综述和讨论。     

Methods to produce induced pluripotent stem cell-derived mesenchymal stem cells: Mesenchymal stem cells from induced pluripotent stem cells

Mesenchymal stem cells (MSCs) have received significant attention in recent years due to their large potential for cell therapy. Indeed, they secrete a wide variety of immunomodulatory factors of interest for the treatment of immune-related disorders and inflammatory diseases. MSCs can be extracted from multiple tissues of the human body. However, several factors may restrict their use for clinical applications: the requirement of invasive procedures for their isolation, their limited numbers, and their heterogeneity according to the tissue of origin or donor. In addition, MSCs often present early signs of replicative senescence limiting their expansion in vitro, and their therapeutic capacity in vivo. Due to the clinical potential of MSCs, a considerable number of methods to differentiate induced pluripotent stem cells (iPSCs) into MSCs have emerged. iPSCs represent a new reliable, unlimited source to generate MSCs (MSCs derived from iPSC, iMSCs) from homogeneous and well-characterized cell lines, which would relieve many of the above mentioned technical and biological limitations. Additionally, the use of iPSCs prevents some of the ethical concerns surrounding the use of human embryonic stem cells. In this review, we analyze the main current protocols used to differentiate human iPSCs into MSCs, which we classify into five different categories: MSC Switch, Embryoid Body Formation, Specific Differentiation, Pathway Inhibitor, and Platelet Lysate. We also evaluate common and method-specific culture components and provide a list of positive and negative markers for MSC characterization. Further guidance on material requirements to produce iMSCs with these methods and on the phenotypic features of the iMSCs obtained is added. The information may help researchers identify protocol options to design and/or refine standardized procedures for large-scale production of iMSCs fitting clinical demands.     

Cardiac differentiation of human pluripotent stem cells

Due to the extremely limited proliferative capacity of adult cardiomyocytes, human embryonic (pluripotent) stem cell derived cardiomyocytes (hESC-CMs) are currently almost the only reliable source of human heart cells which are suited to large-scale production. These cells have the potential for wide-scale application in drug discovery, heart disease research and cell-based heart repair. Embryonic atrial-, ventricular- and nodal-like cardiomyocytes can be obtained from differentiated human embryonic stem cells (hESCs). In recent years, several highly efficient cardiac differentiation protocols have been developed. Significant progress has also been made on understanding cardiac subtype specification, which is the key to reducing the heterogeneity of hESC-CMs, a major obstacle to the utilization of these cells in medical research and future cell-based replacement therapies. Herein we review recent progress in cardiac differentiation of hESCs and cardiac subtype specification, and discuss potential applications in drug screening and cell-based heart regeneration.     

Induced pluripotent stem cells for modelling human diseases

Research into the pathophysiological mechanisms of human disease and the development of targeted therapies have been hindered by a lack of predictive disease models that can be experimentally manipulated in vitro. This review describes the current state of modelling human diseases with the use of human induced pluripotent stem (iPS) cell lines. To date, a variety of neurodegenerative diseases, haematopoietic disorders, metabolic conditions and cardiovascular pathologies have been captured in a Petri dish through reprogramming of patient cells into iPS cells followed by directed differentiation of disease-relevant cells and tissues. However, realizing the true promise of iPS cells for advancing our basic understanding of disease and ultimately providing novel cell-based therapies will require more refined protocols for generating the highly specialized cells affected by disease, coupled with strategies for drug discovery and cell transplantation.     

Derivation of male germ cells from induced pluripotent stem cells by inducers: A review

Induced pluripotent stem cells (iPSCs) refer to stem cells that are artificially produced using a new technology known as cellular reprogramming, which can use gene transduction in somatic cells. There are numerous potential applications for iPSCs in the field of stem cell biology becauase they are able to give rise to several different cell features of lineages such as three-germ layers. Primordial germ cells, generated via in vitro differentiation of iPSCs, have been demonstrated to produce functional gametes. Therefore, in this review we discussed past and recent advances in the in vitro differentiation of germ cells using pluripotent stem cells with an emphasis on iPSCs. Although this domain of research is still in its infancy, exploring development mechanisms of germ cells is promising, especially in humans, to promote future reproductive and developmental engineering technologies. While few studies have evaluated the ability and efficiency of iPSCs to differentiate toward male germ cells in vitro by different inducers, the given effect was investigated in this review.