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Development of biomimetic co-culture and tri-culture models to mimic the complex structure of the alveolar-capillary barrier

A549电池 基底膜 分泌物 细胞生物学 化学 体外 细胞培养 生物物理学 生物医学工程 病理 生物 医学 生物化学 内科学 遗传学
作者
Michela Licciardello,Viola Sgarminato,Gianluca Ciardelli,Chiara Tonda‐Turo
出处
期刊:Biomaterials advances [Elsevier BV]
卷期号:154: 213620-213620 被引量:2
标识
DOI:10.1016/j.bioadv.2023.213620
摘要

Alveoli are the functional area of respiratory system where the gaseous exchanges take place at level of the alveolar-capillary barrier. The development of safe and effective therapeutic approaches for treating lung disease is currently limited due to the lack of realistic preclinical models for their testing and validation. In this work, tissue engineering approaches were exploited to develop a biomimetic platform that provide an appropriate mimicking of the extracellular environment and the multicellular architecture of human alveoli. Here, we propose the implementation of two biomimetic in vitro models to reproduce the features of the main anatomic portions of the physiological alveolar-capillary barrier. First, a co-culture barrier model was obtained by integrating an electrospun polycaprolactone-gelatin (PCL-Gel) membrane in a modified transwell insert (PCL-Gel TW) to mimic the alveolar basement membrane (coded as thin model). Alveolar epithelial (A549) and lung microvascular endothelial (HULEC-5a) cells were cultured on the apical and basolateral side of the PCL-Gel membrane, respectively, under physiologic air-liquid interface (ALI) conditions for 7 days. The ALI condition promoted the expression of type I and type II alveolar epithelial cell markers and the secretion of mucus in A549 cells. Increased cell viability and barrier properties in co-cultures of A549 and HULEC-5a compared to mono-cultures revealed the effectiveness of the model to reproduce in vitro physiological-relevant features of the alveolar-capillary barrier. The second portion of the alveolar-capillary barrier was developed implementing a tri-culture model (coded as thick model) including a type I collagen (COLL) hydrogel formulated to host lung fibroblasts (MRC-5). The thick barrier model was implemented by seeding HULEC-5a on the basolateral side of PCL-Gel TW and then pouring sequentially MRC-5-laden COLL hydrogel and A549 cells on the apical side of the electrospun membrane. The thick model was maintained up to 7 days at ALI and immunofluorescence staining of tight and adherent junctions demonstrated the formation of a tight barrier. Lastly, the ability of models to emulate pathological inflammatory conditions was validated by exposing the apical compartment of the PCL-Gel TW to lipopolysaccharide (LPS). The damage of A549 tight junctions, the increase of barrier permeability and IL-6 pro-inflammatory cytokine release was observed after 48 h exposure to LPS.
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