Medvedeva Snezhanna, Achasova Kseniya, Boldyreva Lidiya, Ogienko Anna, Kozhevnikova Elena
Institute of Molecular and Cellular Biology SB RAS, Novosibirsk, Russia.
Scientific Research Institute of Neurosciences and Medicine, Novosibirsk, Russia.
Tissue Barriers. 2024 Nov 5:2423137. doi: 10.1080/21688370.2024.2423137.
models are of great importance in advancing our understanding of human diseases, especially complex disorders with unknown etiologies like inflammatory bowel diseases (IBD). One of the key IBD features is the increased intestinal permeability. The disruption of the intestinal barrier can occur due to a destructive inflammatory response involving intestinal cell death. Alternatively, proteins that form tight junctions (TJ) fail to form function complexes and promote epithelial barrier disruption. The mechanisms behind this process are not fully understood. Thus, models that facilitate studying the intestinal barrier and its molecular components are of particular importance in the context of IBD. There are and models that can be used to recapitulate some aspects of IBD. Among these are intestinal explants, crypts, and epithelial 3D-organoids. Here we describe some practical limitations of isolated crypts, gut tissue explants, and intestinal organoids as models in epithelial barrier biology, and TJ in particular. Our findings demonstrate that only 3D intestinal organoids formed from single cells are suitable to study barrier permeability , as primary crypt-derived organoids do not retain epithelial integrity due to cell death. Importantly, 3D organoids raised in culture conditions may fail to recapitulate inflammatory and barrier phenotypes of the source mouse model. To study the features of the inflamed epithelium, intestinal explants and crypts were employed. We show here that isolated crypts do not preserve native TJ structure in a long-term experimental setting and tend to disintegrate in the unsupported culture environment. However, intestinal explants were stable in culture conditions for about 24 hours and demonstrated their applicability for short-term living tissue imaging and fluorescence recovery after photobleaching (FRAP). Thus, a combination of 3D organoids and intestinal explants provides a more accurate experimental platform to understand the intestinal epithelial barrier.
模型对于增进我们对人类疾病的理解非常重要，尤其是对于病因不明的复杂疾病，如炎症性肠病（IBD）。IBD的一个关键特征是肠道通透性增加。肠道屏障的破坏可能是由于涉及肠道细胞死亡的破坏性炎症反应所致。或者，形成紧密连接（TJ）的蛋白质无法形成功能复合物并导致上皮屏障破坏。这一过程背后的机制尚未完全了解。因此，有助于研究肠道屏障及其分子成分的模型在IBD背景下尤为重要。有一些模型可用于概括IBD的某些方面。其中包括肠道外植体、隐窝和上皮3D类器官。在这里，我们描述了分离的隐窝、肠道组织外植体和肠道类器官作为上皮屏障生物学模型，特别是TJ模型的一些实际局限性。我们的研究结果表明，只有由单细胞形成的3D肠道类器官适合研究屏障通透性，因为源自原始隐窝的类器官由于细胞死亡而无法保持上皮完整性。重要的是，在培养条件下培养的3D类器官可能无法概括源小鼠模型的炎症和屏障表型。为了研究炎症上皮的特征，我们采用了肠道外植体和隐窝。我们在此表明，在长期实验环境中，分离的隐窝无法保留天然TJ结构，并且在无支撑的培养环境中容易解体。然而，肠道外植体在培养条件下可稳定约24小时，并证明其适用于短期活组织成像和光漂白后荧光恢复（FRAP）。因此，3D类器官和肠道外植体的组合提供了一个更准确的实验平台来理解肠道上皮屏障。
