胰岛类器官研究进展

类器官是将具有多向分化潜能的干细胞或组织细胞在特定环境下培养分化成为能够模拟原生器官结构和功能的三维结构。类器官在各种疾病模型研究及药物筛选中发挥至关重要的作用。近年来,通过体外诱导胰腺组织或多能干细胞分化形成具有胰岛细胞功能的胰岛类器官研究成为热点,为胰岛相关疾病模型、药物研究以及糖尿病的治疗提供了新的手段。本文针对胰岛类器官的体外诱导方法及应用前景作一综述。     

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。     

多能干细胞诱导分化为肾脏类器官的研究进展与挑战

用于分化为多种类型细胞的多能干细胞(PSC)体外培养技术已被广泛应用于生物学领域中。由PSC分化而来的肾脏类器官可基本还原生物体内肾脏的组织结构和部分功能,在肾脏疾病模型研究和药物筛选中有重要作用,继续改善肾脏类器官的结构、功能和成熟度将会对肾脏再生治疗提供极大的帮助。研究肾脏类器官的重点在于体外准确模拟体内肾脏的发育过程。本文着重归纳了近十年来对胚胎肾发育过程研究的重点,对肾脏类器官分化技术的几个关键方案进行总结、分析和比较,并探讨肾脏类器官在分化研究和应用中将面临的挑战。     

类器官的应用研究进展

类器官是利用干细胞的自我更新和分化能力,在体外培养形成的一种微小组织器官类似物,在很大程度上具有体内相应器官的功能。迄今为止,在3D培养条件下,已经成功培养出多种类器官如肺、胃、肠、肝和肾等类器官。它们不仅可作为组织器官的替代品用于药物和临床研究,还可用于体内器官移植。本文综述了类器官在药物毒性检测、药效评价和新药筛选中的作用以及利用类器官建立疾病模型、研究组织器官发育和类器官在精准医疗、再生医学中的价值。     

类器官的研究进展及应用

随着人类生物学研究的不断深入,需建立新的模型系统为研究提供了有力的工具。虽然传统的研究模型已被广泛应用,但难以准确反映组织、器官在机体中的生理现象。类器官 (Organoid) 是来源于干细胞或器官祖细胞的三维细胞聚集体,可分化和自组织形成具有人体相应器官的部分特定功能和结构。由于类器官具有人源性,可模拟器官发育和形成,在体外长期扩增中具有基因组稳定性,并能够形成活体生物库进行高通量筛选等优势,成为近年来备受关注的体外模型。目前,利用类器官模型结合新兴的基因编辑、器官芯片、单细胞RNA测序技术等,能够突破传统模型的瓶颈,在器官水平上为疾病模型的建立、药物研发、精准医疗以及再生医学等提供有价值的信息。文中就类器官分类及特性、研究应用、与其他技术结合应用及展望这4个方面进行综述。     

人源脑类器官模型在孤独症相关神经发育缺陷研究中的应用与进展

孤独症谱系障碍(autism spectrum disorder, ASD)是一种以社交障碍及刻板行为为核心症状的神经发育疾病。其受遗传和环境等多种因素影响,病因复杂、患者异质性极高,这给疾病机制研究和治疗靶点的研究造成了阻碍。3D人脑类器官是由多能干细胞(pluripotent stem cell, PSCs)经悬浮培养及诱导分化后自组织形成的器官样三维组织,其可携带患者完整的遗传信息,可在体外模拟胚胎的早期脑发育过程并反映病理过程。脑类器官是研究孤独症谱系障碍的理想模型与平台,该文将对近年来人脑类器官在孤独症研究中的进展与成果作综述。     

生物制造领域的机遇与挑战：发展类器官构建及培养的新技术

类器官构建及培养技术是近年来新兴的前沿性科学技术，该技术已经被广泛用到组织器官发育、疾病发病机制、药物开发和再生医学等领域的研究之中。干细胞及组织器官发育分化调控的研究成果为类器官构建及培养技术的建立提供了重要的信息。体外借助细胞外基质成分及细胞因子等构建出适宜于干细胞增殖、分化的三维微环境是类器官构建及培养技术的核心。在适宜的微环境中，干细胞及其分化的多种类型细胞可通过自组织形成与体内相应组织结构和功能相似的类器官。当前，多种类型的类器官构建及培养技术虽然得到广泛应用，但其技术体系仍具有操作的复杂性、产量的不确定性及获得的类器官结构和功能与体内组织存在较大差异性等难题。生物制造领域先进技术的引入推动了类器官技术的发展。本文将综述基质成分与细胞因子构建的三维微环境的研究进展，并讨论生物制造领域的先进技术在类器官构建与培养技术中的应用，例如微孔限定的培养技术可以控制类器官的生长发育，能用于制备大小均一及生物学特性相似的类器官；图案化技术使细胞按图案特征响应性地增殖与分化，可以精准控制类器官的生成；三维生物打印技术可以精确组装各类细胞，有助于构建具有复杂结构和区域特异性的类器官。类器官构建...     

肠道类器官在精准医学中的应用

类器官3D培养是一种新兴的用来研究组织成体干细胞生长、分化、器官形成的体外研究系统.目前,肠道类器官的3D培养是将分离的肠道隐窝或干细胞植入含有多种生长因子的基质胶中,在基质3D支撑下生成具有肠道上皮样结构的微型空心球体,这些球体被称为肠道类器官.该类器官包含有所有种类的肠道功能上皮细胞,能最大程度模拟肠道组织,故也称之为"迷你肠".结直肠肿瘤细胞也可以利用该3D体系培养得到肿瘤类器官.这些肠道类器官可被广泛应用于炎症性肠病、肠道损伤再生、肠癌等多种肠道疾病的研究.本综述讨论了关于肠道干细胞的最新研究进展,正常类器官和肿瘤类器官的培养,同时还将探讨类器官在疾病建模和组织再生、基因修复、肿瘤个性化治疗等精准医学方面的应用.     

胰腺类器官技术的发展及应用

在人类生物学和疾病的研究过程中,动物模型扮演了重要的角色。但随着研究的深入,其局限性也逐渐凸显。新兴的人类类器官(organoid)技术较好地弥补了动物模型的不足。类器官主要是指由干细胞衍生出的3D多细胞微器官。它能在体外模拟组织器官自发的谱系分化与稳态维持,并具备类似于体内组织器官的生理功能。目前,类器官培养技术在很多器官(比如胃肠道、食管、肝、胆、脑和膀胱等)中都取得了较好的进展。胰腺是人体内唯一的一个既是外分泌腺又是内分泌腺的特殊脏器。因此,胰腺类器官技术的发展面临更大的挑战。目前,胰腺导管类器官和胰岛类器官技术日渐优化,但如何构建复合型胰腺类器官仍是当前研究的难点。该综述将主要回顾胰腺的发育过程、体外定向分化技术以及胰腺类器官模型构建与应用等最新研究成果,同时简要探讨胰腺类器官模型具有潜力的发展方向。     

类器官的构建与应用进展

类器官是在体外经由干细胞驱动的, 形成具有来源器官显微解剖特征的多细胞三维结构且能自我更新的微组织。类器官能分化产生器官特异性的多种细胞类型,能重现对应器官的部分功能和空间架构,它的诞生为生命医学研究和临床应用注入了新动能,在癌症基础与临床研究、再生医学等领域表现出广阔的应用前景。对近些年国内外类器官研究进展进行综述,介绍其构建过程与培养体系,并详细阐述其作为体外研究模型的优缺点,为基于类器官的科学研究与应用提供了参考。     

胸腺类器官培养及应用进展

胸腺是人体重要的免疫器官,是T细胞分化成熟的场所,受损后容易引发自身免疫性疾病甚至恶性肿瘤。多年来,研究人员主要通过T细胞体外单层培养系统探索T细胞的发育过程,揭示胸腺损伤和再生的机制。但单层培养系统既不能重现胸腺独特的三维上皮性网状结构,也无法充分提供造血干细胞定向分化为T细胞所需的细胞因子和生长因子。胸腺类器官技术利用具有干细胞潜能的细胞,在体外通过三维培养模拟胸腺的解剖结构和胸腺上皮细胞介导的信号通路,与体内胸腺微环境十分接近。在研究T细胞分化和发育、胸腺相关疾病、重建机体免疫功能以及细胞治疗等方面,胸腺类器官呈现出巨大潜力。文中系统介绍了胸腺类器官的培养方法,比较了培养所用支架的优缺点;同时探讨了胸腺类器官在疾病建模、肿瘤靶向治疗、再生医学和器官移植等领域的应用,并对其前景进行展望。     

In vitro growth and differentiation of mammalian sensory hair cell progenitors: a requirement for EGF and periotic mesenchyme

The sensory hair cells and supporting cells of the organ of Corti are generated by a precise program of coordinated cell division and differentiation. Since no regeneration occurs in the mature organ of Corti, loss of hair cells leads to deafness. To investigate the molecular basis of hair cell differentiation and their lack of regeneration, we have established a dissociated cell culture system in which sensory hair cells and supporting cells can be generated from mitotic precursors. By incorporating a Math1-GFP transgene expressed exclusively in hair cells, we have used this system to characterize the conditions required for the growth and differentiation of hair cells in culture. These conditions include a requirement for epidermal growth factor, as well as the presence of periotic mesenchymal cells. Lastly, we show that early postnatal cochlear tissue also contains cells that can divide and generate new sensory hair cells in vitro.     

Developmental biology and tissue engineering

Morphogenesis implies the controlled spatial organization of cells that gives rise to tissues and organs in early embryonic development. While morphogenesis is under strict genetic control, the formation of specialized biological structures of specific shape hinges on physical processes. Tissue engineering (TE) aims at reproducing morphogenesis in the laboratory, i.e., in vitro, to fabricate replacement organs for regenerative medicine. The classical approach to generate tissues/organs is by seeding and expanding cells in appropriately shaped biocompatible scaffolds, in the hope that the maturation process will result in the desired structure. To accomplish this goal more naturally and efficiently, we set up and implemented a novel TE method that is based on principles of developmental biology and employs bioprinting, the automated delivery of cellular composites into a three-dimensional (3D) biocompatible environment. The novel technology relies on the concept of tissue liquidity according to which multicellular aggregates composed of adhesive and motile cells behave in analogy with liquids: in particular, they fuse. We emphasize the major role played by tissue fusion in the embryo and explain how the parameters (surface tension, viscosity) that govern tissue fusion can be used both experimentally and theoretically to control and simulate the self-assembly of cellular spheroids into 3D living structures. The experimentally observed postprinting shape evolution of tube- and sheet-like constructs is presented. Computer simulations, based on a liquid model, support the idea that tissue liquidity may provide a mechanism for in vitro organ building.     

Differentiation of cartilage cells studied by quantitative [3H]thymidine autoradiography in vitro

The apical segments of the mandibular condylar cartilage of newborn ICR mice, containing the intact zones of progenitor cells along with a few rows of chondroblasts were initially prelabelled in vitro with [3H]thymidine and were subsequently chased and cultured for as long as eight days. Such explants underwent a process of tissue regeneration, as after three days in culture they reconstituted the original structure of the organ, thus resembling the in vivo appearance of neonatal mandibular condylar cartilage. Cellular proliferation with subsequent differentiation in the regenerating tissue was followed by means of quantitative autoradiography. Immediately after labelling, the autoradiography-positive grains were confined exclusively to progenitor cells. The latter revealed a substantial ability to proliferate in vitro, a fact that was manifested by a progressive increase in the labelling index along the course of the culture. The latter process was followed by cellular differentiation thereby obtaining hypertrophic chondrocytes. The increase in the rate of labelling index and in the total number of [3H]thymidine-labelled cells was significantly correlated with the overall growth of the regenerating explants.     

骨髓间充质干细胞在组织工程研究中的应用新进展

骨髓间充质干细胞又称为骨髓源性间充质干细胞,是指存在于骨髓基质细胞系统中的一类干细胞,具有高度稳定的体外扩增能力和多向分化潜能等特点。骨髓间充质干细胞因其取材方便,易于分离和培养,以及在适当条件下可诱导分化为皮肤、骨骼、内脏、血液、神经等多种组织细胞的独特优势,目前被广泛应用于药物开发、免疫调节、组织修复、器官重建等多个研究领域。近年来,骨髓间充质干细胞作为种子细胞在组织工程领域有着非常诱人的潜在应用前景。本文就骨髓间充质干细胞在组织工程学研究中应用的最新进展作一综述。     

Multiple differentiation of clonal teratocarcinoma stem cells following embryoid body formation in vitro

We have described the differentiation in vitro of clonal pluripotent teratocarcinoma stem cells derived from isolated single cells. By using solvent-resistant plastic petri dishes as a substratum for cell growth, it is possible to prepare histological sections of the cultures which can be compared with sections of teratocarcinomas formed in vivo by the same cells. Our results indicate that almost all of the cell types found in the tumors are formed in vitro, including cartilage, keratinizing epithelium, pigmented epithelium, neural tissue, and muscle. The cells are organized in a tissue structure which is remarkably similar to that found in vivo.     

Stromal cells of hemopoietic origin

Hemopoiesis is a multistep process involving stem cell renewal, commitment, differentiation, maturation and consequent positioning of the cells within the tissue. Stromal cells are a major component of the hemopoietic microenvironment. The in vitro culture of cloned stromal cells has enabled detailed analysis of their functions and has provided answers relating to the contribution of stromal cells to the control of hemopoiesis. Cultured stromal cells were found to support the renewal of stem cells through a mechanism that did not seem to involve already known cytokines. Cloned stromal cells from both marrow and thymus supported the in vitro accumulation of myeloid as well as T and B lymphoid cells. Thus, cloned stromal cells had the ability to induce multilineage hemopoiesis, irrespective of the organ from which they were derived. Invariably, stromal cells tended to select in culture for hemopoietic cells at early differentiation stages and restricted the accumulation of mature cells. These functions may be part of the mechanism that protects the stem cell pool from excess differentiation.     

Somatotropic cell differentiation in an organ culture of the chick embryo adenohypophysis

Morphogenetic potencies of the adenohypophysis tissue from 4.5 to 11-day old chicken embryos used for the differentiation of somatotropic cells were investigated by methods of organ tissue culture. STG-cells were detected in cultures by immunofluorescence using an antiserum to human STG. In vitro studies of organ cultures revealed differentiation of STG-cells when adenohypophysis tissue was cultured from the 5.5th, 7th, 9th and 11th day of development in the absence of the diencephalon. Differentiation of STG-cells occurred predominantly in embryo caudal lobe transplants after chorion-allantois culturing of Rathke's pocket fragments from 4.5-, 5.0- and 5.5-day old embryos. The data obtained suggest that at late stages of Rathke's pocket development differentiation of STG-cells is preprogrammed and that determined precursors of these cells are located in the caudal lobe of the germ.