图1 荧光显微镜下的类器官（图片源自网络）
类器官（Organoids）是一种在体外环境下培育而成的具备三维结构的微器官（图1），拥有类似真实器官的复杂结构，并能部分模拟来源组织或器官的生理功能。借助类器官，研究人员可深入观察人体组织的变化，更好地理解发育过程，并可用于再生医学以及药物的疗效筛选。因此，类器官研究具有广阔的发展前景。
图2 “Organoid Modeling of the Tumor Immune Microenvironment”文章封面
生物中心针对2018年12月美国斯坦福大学Calvin J. Kuo教授团队发表在国际顶尖杂志Cell的文章“Organoid Modeling of the Tumor Immune Microenvironment”（图2），特邀诺贝尔医学和生理学奖获得者Thomas C. Südhof教授、著名环境毒理学及神经类器官专家Ellen Fritsche教授就类器官研究的现状、瓶颈、应用价值以及未来的发展方向展开评述。下面将为读者呈现访谈内容。
1. 问：什么是类器官？类器官的主要类型和他们各自的关键特征是什么？
Question：What is organoid? What are the main organoid types and their key characteristics?
Thomas C. Südhof答：目前有很多该领域的综述和专著可供参考，以下是我推荐的一些综述。(编者按：综述列表见英文回答)
Thomas C. Südhof’s answer：There are innumerable review articles and textbooks on organoids that I would suggest you consult.
Here are some reviews:
[1] Little MH, Hale LJ, Howden SE, Kumar SV. Generating Kidney from Stem Cells. Annu Rev Physiol. 2019 Feb 10;81:335-357
[2] Rowe RG, Daley GQ. Induced pluripotent stem cells in disease modeling and drug discovery. Nat Rev Genet. 2019 Feb 8.
[3] Sontheimer-Phelps A, Hassell BA, Ingber DE. Modeling cancer in microfluidic human organs-on-chips. Nat Rev Cancer. 2019 Feb;19(2):65-81.
[4] Mittal R, Woo FW, Castro CS, Cohen MA, Karanxha J, Mittal J, Chhibber T, Jhaveri VM. Organ-on-chip models: Implications in drug discovery and clinical applications. J Cell Physiol. 2019 Jun;234(6):8352-8380.
[5] Amin ND, Paşca SP. Building Models of Brain Disorders with Three-Dimensional Organoids. Neuron. 2018 Oct 24;100(2):389-405.
Ellen Fritsche答：类器官是一种在体外培育而成的具有来源器官显微解剖特征的多细胞三维结构。迄今为止，不同组织、疾病模型及模拟发育的类器官已问世。类器官的工具细胞主要为组织特异性多能干细胞。类器官的主要特征包括基于细胞类别的自我组织及空间限制的定向分化，与体内发育过程相似。他们（类器官）含有多种器官特异性细胞，这些细胞的空间组织、排列与来源器官类似。另外，他们（类器官）具有一些来源器官特有的功能。迄今，来源于多种器官的类器官业已面世，包括脑、肠道、胃、舌、甲状腺、胸腺、睾丸、肝脏、胰腺、皮肤、肺、肾、心脏及视网膜。除了来源于健康组织的类器官，大量疾病模型（包括肿瘤模型）的类器官也不断涌现。最后，类器官为科研人员进行发育生物学研究提供了绝佳模型。
Ellen Fritsche’s answer：An organoid is a three-dimensional (3D), multicellular structure with microanatomical features of the organ of origin produced in vitro. So far, organoids of different tissues, disease models, as well as organoids resembling development have been created. Cellular basis for organoids are mainly pluripotent or tissue-specific stem cells. Key features of organoids include their self-organization through cell sorting and spatially restricted lineage commitment in a manner similar to in vivo. They contain multiple, organ-specific cell types which are spatially organized in a manner similar to the organ. In addition, they recapitulate some specific organ functions. Organoids from multiple organs have so far been created. These include brain, intestine, stomach, tongue, thyroid, thymus, testis, liver, pancreas, skin, lung, kidney, heart and retina. In addition to the healthy organoids, a plethora of disease models including tumor models, have been developed. Last, organoids offer researchers an exceptional model to study developmental biology.
2. 问：可否谈谈类器官在生物医学领域的主要应用？
Question：What are the main applications of the organoids in the field of biomedicine?
Thomas C. Südhof答：类器官的价值在于其具有在体外培养环境下构建人类器官疾病模型的潜力。这非常适用于像心脏这样的组织，在人类早期脑发育的研究上也逐渐变得可行。但类器官在再生医学上的应用依然前路漫漫。
Thomas C. Südhof’s answer：The attraction of organoids is that they potentially allow disease modeling of human organs in a dish. This works best for tissues such as heart, and is becoming feasible for early human brain development. The use of organoids in regenerative medicine is still far in the future.
Ellen Fritsche答：作为一项重大的技术突破，类器官目前已被公认为生物研究的重要工具,并具有重要的临床应用价值。类器官允许在一个模拟内源性细胞组织和器官结构的环境中进行组织生物学、发育、再生、疾病建模 (包括癌症研究)、器官移植技术改良、药物发现/疗效评估以及毒理学的研究。
Ellen Fritsche’s answer：Starting as a major technological breakthrough, organoids are now well-established as an essential tool in biological research and also have important implications for clinical use. Organoids allow research on tissue biology, development, regeneration, disease modeling (including cancer research), improvements in organ transplantation, drug discovery/response as well as toxicological studies in an environment that mimics endogenous cell organization and organ structures.
3. 问：在肿瘤生物学及新药开发领域，类器官相对于细胞系、动物模型的主要优势在哪里？
Question：What are the main advantages of using organoids instead of cell lines, or animal models in the field of tumor biology and new drug development?
Thomas C. Südhof答：相对于细胞系而言，类器官构建了一个具备三维结构的器官样组织，尽管并不完全（模拟人类器官）。相较于动物模型，类器官的优势体现在其实现了应用人源性组织进行实验研究。
Thomas C. Südhof’s answer：The advantage over cell lines is that organoids model a three dimensional organ, although not completely. The advantage over animal models is that organoids enable studies of human material.
Ellen Fritsche答：传统的二维 (2D) 肿瘤细胞系培养和动物人源性肿瘤异种移植物 (PDTXs) 长期以来一直被用作肿瘤模型, 并为癌症研究做出了巨大贡献。然而, 各种缺点阻碍了这些模型的临床应用，这主要是由于与肿瘤治疗相关的药物开发是成功率最低的。二维细胞培养体系不具备免疫细胞、微环境、间质成分和器官特异性的功能。其他限制包括肿瘤细胞系经多次传代后缺乏来源肿瘤的遗传异质性, 原因是细胞在培养皿二维生长的环境下会发生优势克隆选择，但这并不符合生理。此外, PDTX 模型还经历了小鼠特异性的肿瘤演化。在资源方面, 这些模型也是极度的费钱费时。类器官可以克服其中的一些限制。类器官的基因修饰可实现在接近生理环境的情况下进行疾病建模。比如, 将肿瘤性突变导入健康干细胞可以产生遗传控制的肿瘤类器官。此外, 类器官可以从患者来源的健康组织和肿瘤组织中迅速培育，从而使患者特异性药物检测和个性化治疗方案的开发成为可能。在这种患者特异性的肿瘤类器官中，可观察到组织稳态(histostasis)，如3D培养保留了与来源患者肿瘤相一致的组织病理学特征，为未来个性化肿瘤治疗的发展提供了希望。与 PDTX 不同，类器官维护便利，具有整合免疫细胞的可能性，易进行基因改造 (遗传性肿瘤建模)，支持匹配对照的研究，并可用于高通量药物筛选和生物库的建设。
除了肿瘤学, 类器官也为新药开发提供了绝佳模型。新药开发的失败率很高，这在一定程度上是由于动物药代动力学和药效学的差异或动物疾病模型并不能完全模拟人体病理过程。具有人体特异生理和病理特征的类器官有助于克服这些问题。基于特定疾病，甚至特定个体，以高通量方式培育的类器官预计将发展成为精确治疗的强大工具。未来可借助生物库进行筛选，不仅是为了鉴定新药，还可揭示哪些患者可以从某些 (现有) 药物的治疗中受益。此外，对潜在药物的重点检测可为制药业提供新的指引。另外，类器官未来可能用于毒理学检测, 以作为动物试验的有力补充(如果不是部分取代的话)。
Ellen Fritsche’s answer：Traditional two-dimensional (2D) tumor cell line cultures and patient-derived tumor xenografts (PDTXs) in animals have long been employed as tumor models and have made tremendous contribution to cancer research. However, a variety of drawbacks hamper these models for clinical use as success rates for tumor therapeutics are lowest in the field of drug development. 2D cell line cultures do not contain immune cells, microenvironment, stromal compartments, and organ-specific functions. Other limitations include the lack of genetic heterogeneity of original tumors after many passages for cancer cell lines because clonal selection in the dish happens for superiority in 2D growth, which is not physiologic. Moreover, PDTX models experience mouse-specific tumor evolution. On the resource side, such models are highly money- and time-consuming. Organoids can overcome some of these constraints. Genetic modification of organoids allows disease modeling in a setting that approaches the physiological environment. Here, insertions of tumor mutations into healthy stem cells allow generation of genetically-controlled tumoroids. Additionally, organoids can be grown with high efficiency from patient-derived healthy and tumour tissues, potentially enabling patient-specific drug testing and the development of individualized treatment regimens. In such patient-specific tumoroids, histostasis is observed, i.e. conservation of histopathological traits between 3D cultures and the matched patient tumor, promising advances in personalized tumor therapies in the future. In contrast to PDTX, organoids are of easier maintenance, bear the possibility to integrate immune cells, are amenable to genetic modification (genetic cancer modeling), allow study of matched controls, can be used for high throughput drug screening and biobanking.
Besides oncology, organoids are promising models for drug development. Attrition rates in new drug development are high. This is partly reasoned indifferences between animal pharmacokinetics and –dynamics or in animal disease models that do not correctly resemble human pathology. Organoids with human-relevant physiology and pathology are thought to help overcoming these issues. Organoid cultures based on a specific disease and even on a specific individual used in a high-throughput manner are expected to develop into powerful tools for precision therapy. Future screens may be performed using biobanks with the aim of not only identifying new drugs but also revealing which patients may benefit from treatment with certain (existing) drugs. In addition, focused tests of potential drugs should identify new leads for the pharmaceutical industry. Furthermore, organoids may be used in the future for toxicology testing to complement, if not in part replace, animal testing.
4. 问：当前类器官的局限是什么？为了满足肿瘤生物学、干细胞生物学、移植、新药开发领域的研究需要，类器官需要在哪些方面进一步改进？
Question：What are the limitations of organoids and what aspects of organoids can be further improved to meet the demand for research in tumor biology, stem cell biology, transplantation and drug development?
Thomas C. Südhof答: 类器官领域的研究仍在起步阶段。即使对于如心脏和肝脏这样的组织，类器官也很不成熟，仅能部分模拟人体器官。对于脑组织则更甚。许多基本的（脑组织）生理功能，如细胞生理、生化功能仍有待突破。这将花费数年的时间。
Thomas C. Südhof’s answer：The field of organoid research is still in the beginning. Even for tissues like heart and liver, organoids are very immature and only partly model the human organ. This is worse for brain. Much fundamental biology, such as cell biology and biochemistry, is needed to advance the field. This will take many years.
Ellen Fritsche答：类器官是融合了各种器官特异性细胞类型、组织形态和功能的组织模型。但类器官仅为有限度的模拟，困扰这项技术应用的一个重要限制是它的体积。当类器官体积增加时，缺氧和缺乏可溶性因子所致的组织坏死是亟需解决的问题。解决这个问题的一个可能方案是激活血管生成途径, 从而使类器官血管化。这已经在hiPSC衍生的肝脏类器官上成功实现。类器官领域的另一个挑战在于一个完整的有机体中所自然存在的器官“对话”。类器官研究可满足生物工程的要求, 通过培育包含不同类型hiPSC衍生类器官（呈现多个器官系统的结构和功能）的器官芯片设备，用以在更类似于体内的环境中筛选药物。另外，通过在类器官中添加免疫细胞, 还可模拟具有免疫系统的组织间“对话”。另外，在药理和毒理学研究中，物质的肝脏代谢至关重要，这可通过以器官芯片的形式包含肝脏代谢来实现。
Ellen Fritsche’s answer：Organoids are organ models recapitulating an assortment of organ-specific cell types, tissue morphogenesis and functions. Yet, there are limitations in their mimicry. One important limitation plaguing the application of this technology is their size. When the organoids’ volume increases, the issue of tissue necrosis caused by the lack of diffusion of oxygen and soluble factors needs to be addressed. One solution for this problem might be the activation of angiogenic pathways that will lead to vascularised organoids. This was already succeeded with hiPSC-derived liver organoids. One more challenge of the organoid field lies in organ crosstalk, which is naturally present in an intact organism. Here, organoid research meets bioengineering by producing organ-on-a-chip devices containing different types of hiPSC-derived organoids representing the structure and function of multiple organ systems for screening the effects of drugs in more in vivo-like settings. The crosstalk of tissues with the immune system can be modelled by adding immune cells to the organoids. For pharmacological and toxicological applications, liver metabolism of compounds is crucial. Including such metabolism via an organ-on-a-chip approach can solve this issue.
5. 问：当前类器官研究的发展方向如何？
Question：What are the current trends for organoids research?
Thomas C. Südhof答:（当前的现状是）所有人都在盲目追求应用，却忽略了一个坚实的科学基础。我认为未来会有数以百计的公司在那里贩卖希望，但他们大多数将以失败告终。因为相关生理学研究成果并不足以支撑这些应用项目。类器官最有前景的应用领域应是用于肝脏、心脏和肿瘤的药物筛选。
Thomas C. Südhof’s answer：Everybody rushes towards applications, without a solid scientific basis. I think hundreds of companies will be founded that will sell hope, but will mostly fail because the biology isn't there to support applications. Most promising are drug screens in tissue organoids such as liver or heart and in cancer.
Ellen Fritsche答：目前类器官研究的趋势包括建立用于高通量筛选的类器官库和平台, 建立其他疾病模型, 以及建立用于整个生物体建模的器官芯片和微流体芯片。在此特别要强调的是培养基的限制亟待解决。对微流体芯片来讲，需要一种芯片上所有类器官均适用的通用培养基。此外，根据器官系统的不同, 需要开发与生理过程相关的来自类器官的高通量数据输出装置。在临床方面，为了开发最佳个体化治疗方案，使用源自患者特异性hiPSC类器官的个体化医疗研究亟需开展。在毒理学领域，类器官目前已被用来替代动物进行毒性测试。
Ellen Fritsche’s answer：Current trends for organoid research include generation of organoid banks and platforms for high-throughput screening approaches, generation of additional disease models, and set up of organ-on-a-chip and microfluidics devices for whole organism modeling. Here, especially medium constrictions have to be solved. For microfluidics a common medium for all organoids on the chip is needed. Moreover, depending on the organ system, physiologically relevant high-throughput readouts from organoids need to be developed. On the clinical side, research on personalized medicine using organoids derived from patient-specific hiPSC is warranted for optimal individual treatment regimes. In the toxicology field, organoids as substrates for toxicity testing replacing animals is currently exploited.
6. 问：可否预测一下接下来5年内类器官研究领域的发展？
Question：How the organoids research field will be look like in 5 years?
Thomas C. Südhof答: 我的预测是在接下来的5年内好的实验室将学会如何促进类器官的成熟，并明确该方法的局限。我认为，尽管类器官为干细胞研究提供了巨大的机会，如促进新发现和疗法的出现；但这将耗费10年或更长的时间来发展。到那时候，也只有到那时候，走向应用才能真正成为可能。在那之前，大量初创公司将会烧掉数以亿计的资金，他们中的少部分将会走向成功，并找到增加营收的新途径。
Thomas C. Südhof’s answer：My prediction is that in 5 years, good science labs will have learned how to mature organoids and the limitations of the approach will have been defined. I think organoids are a tremendous opportunity in stem cell approaches that will enable novel discoveries and therapies, but that this will take at least 10 years to develop. Then and only then will it be possible to rationally move towards applications. Until then, lots of start-ups will have spent hundreds of millions of dollars, and a few of them will have been successful in generating some future avenue of revenue.
Ellen Fritsche答：在5年内，类器官的遗传操作与类器官库相结合将给生物医学研究带来翻天覆地的变化。购买来源于具有不同遗传背景患者的疾病特异性类器官将成为可能。器官芯片平台将具有特定标准，由合同研究组织（CRO）以与目前动物试验类似的方式提供。类器官将极大地促进药物疗效试验和安全性测试的开展，因此也将进入药物开发和化学安全性评估研究的监管领域。
Ellen Fritsche’s answer：In 5 years, genetic manipulation of organoids in combination with organoid banking will have revolutionized biomedical research. It will be possible to purchase disease-specific organoids from broad ranges of patients with distinct genetic backgrounds. Organ-on-a-chip platforms will be standard and offered by CROs in a similar manner than currently animal testing. Organoids will have tremendously facilitated drug efficacy and safety testing and thus will have entered also into the regulatory areas of research for drug development as well as chemical safety assessment.
附：
1. Thomas C. Südhof教授简介
斯坦福大学医学院教授、霍华德-休斯医学研究所 (HHMI) 研究员、美国科学院院士、美国医学科学院院士、英国皇家学会外籍院士、2013年诺贝尔医学和生理学奖获得者。1955年生于德国哥廷根，1982年获得哥廷根大学医学博士学位。Südhof教授的研究主要聚焦于突触前神经递质释放的分子机制，为该领域的顶级科学家。他发现了囊泡内神经递质释放过程中的多种关键蛋白，并阐明了神经递质释放的具体分子机制。鉴于在囊泡转运领域的开创性工作，他先后荣获拉斯克基础医学奖及诺贝尔生理学和医学奖等重要医学奖项。
2. Ellen Fritsche教授简介
德国IUF-莱布尼茨环境医学研究所（IUF-Leibniz Research Institute for Environmental Medicine）环境毒理学教授，球模型和风险评估专家组组长。1998年获雷根斯堡大学和杜塞尔多夫大学医学博士学位，曾先后在美国国立环境卫生研究所（NIEHS)和IUF-莱布尼茨环境医学研究所完成博士后研究工作，2009-2012年任亚琛工业大学皮肤毒理学教授。目前为Neurotoxicology杂志副主编、欧洲化学理事会（cefic）顾问、欧盟地平线2020计划专家组成员、欧洲替代动物试验研究中心（CERST-NRW）项目牵头人、替代法信托大会（ACT）成员和OECD发育神经毒性专家组委员。历任欧洲替代动物试验协会（EUSAAT）副主席、主席。
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