Gerckens Michael, Alsafadi Hani N, Wagner Darcy E, Lindner Michael, Burgstaller Gerald, Königshoff Melanie
Comprehensive Pneumology Center, Ludwig-Maximilians-Universität and Helmholtz Zentrum Munich; German Center of Lung Research (DZL); Translational Lung Research and CPC-M bioArchive, Helmholtz Zentrum München, Comprehensive Pneumology Center Munich DZL/CPC-M.
Department of Experimental Medical Science, Lung Bioengineering and Regeneration, Lund University; Wallenberg Center for Molecular Medicine, Lund University; Stem Cell Centre, Lund University.
J Vis Exp. 2019 Feb 12(144). doi: 10.3791/58437.
Translation of novel discoveries to human disease is limited by the availability of human tissue-based models of disease. Precision-cut lung slices (PCLS) used as 3D lung tissue cultures (3D-LTCs) represent an elegant and biologically highly relevant 3D cell culture model, which highly resemble in situ tissue due to their complexity, biomechanics and molecular composition. Tissue slicing is widely applied in various animal models. 3D-LTCs derived from human PCLS can be used to analyze responses to novel drugs, which might further help to better understand the mechanisms and functional effects of drugs in human tissue. The preparation of PCLS from surgically resected lung tissue samples of patients, who experienced lung lobectomy, increases the accessibility of diseased and peritumoral tissue. Here, we describe a detailed protocol for the generation of human PCLS from surgically resected soft-elastic patient lung tissue. Agarose was introduced into the bronchoalveolar space of the resectates, thus preserving lung structure and increasing the tissue's stiffness, which is crucial for subsequent slicing. 500 µm thick slices were prepared from the tissue block with a vibratome. Biopsy punches taken from PCLS ensure comparable tissue sample sizes and further increase the amount of tissue samples. The generated lung tissue cultures can be applied in a variety of studies in human lung biology, including the pathophysiology and mechanisms of different diseases, such as fibrotic processes at its best at (sub-)cellular levels. The highest benefit of the 3D-LTC ex vivo model is its close representation of the in situ human lung in respect of 3D tissue architecture, cell type diversity and lung anatomy as well as the potential for assessment of tissue from individual patients, which is relevant to further develop novel strategies for precision medicine.
新发现向人类疾病的转化受到基于人体组织的疾病模型可用性的限制。用作三维肺组织培养（3D-LTC）的精密切割肺片（PCLS）代表了一种精巧且生物学相关性极高的三维细胞培养模型，由于其复杂性、生物力学和分子组成，它与原位组织高度相似。组织切片在各种动物模型中广泛应用。源自人PCLS的3D-LTC可用于分析对新药的反应，这可能有助于更好地理解药物在人体组织中的作用机制和功能效果。从接受肺叶切除术的患者手术切除的肺组织样本中制备PCLS，增加了病变组织和瘤周组织的可及性。在此，我们描述了一种从手术切除的软弹性患者肺组织生成人PCLS的详细方案。将琼脂糖引入切除组织的支气管肺泡间隙，从而保留肺结构并增加组织硬度，这对后续切片至关重要。用振动切片机从组织块制备500μm厚的切片。从PCLS获取的活检穿孔确保了可比的组织样本大小，并进一步增加了组织样本数量。所生成的肺组织培养物可应用于人类肺生物学的各种研究，包括不同疾病的病理生理学和机制，例如在（亚）细胞水平上最典型的纤维化过程。3D-LTC体外模型的最大优势在于，就三维组织结构、细胞类型多样性和肺解剖结构而言，它与原位人肺非常相似，并且具有评估个体患者组织的潜力，这对于进一步开发精准医学的新策略具有重要意义。
