Reconstruction of Ultra‐thin Alveolar‐capillary Basement Membrane Mimics

Abstract

Alveolar‐capillary basement membrane (BM) is ultra‐thin (<2 µm) extracellular matrix that maintains integral epithelial‐endothelial cell layers. In vitro reconstructions of alveolar‐capillary barrier supported on synthetic scaffolds closely resembling the fibrous and ultra‐thin natural BM are essential in mimicking the lung pathophysiology. Although BM topology and dimensions are well known to significantly influence cellular behavior, conventionally used BM mimics fail to recreate this natural niche. To overcome this, electrospun ultra‐thin 2 µm poly(caprolactone) (PCL) nanofibrous mesh is used to establish an alveolar‐capillary barrier model of lung endothelial/epithelial cells. Transepithelial electrical resistance (TEER) and permeability studies reveal integral tight junctions and improved mass transport through the highly porous PCL meshes compared to conventional dense membranes with etched pores. The chemotaxis of neutrophils is shown across the barrier in presence of inflammatory response that is naturally impeded in confined regions. Conventional requirement of 3 µm or larger pore size can lead to barrier disruption due to epithelial/endothelial cell invasion. Despite high porosity, the interconnected BM mimic prevents barrier disruption and allows neutrophil transmigration, thereby demonstrating the physiological relevance of the thin nanofibrous meshes. It is envisioned that these bipolar cultured barriers would contribute to an organ‐level in vitro model for pathological disease, environmental pollutants, and nanotoxicology.

Mimicking the Natural Basement Membrane for Advanced Tissue Engineering

“…An opposite behavior is observed on the fiber and hydrogel scaffolds where cell spreading is prominent on stiff hydrogels as compared with stiff fibers and vice versa. Similar differences in the expression of αSMA by endothelial cells on nonwoven electrospun meshes and PET membranes were observed by Jain et.al . This illustrates the influence of scaffold structure and property on cell behavior when used to mimic ECM like materials such as the BM.…”

“…by comparing ultrathin 2 μm electrospun nonwoven PCL meshes with commercial 10 μm thick PET membranes. 33 The 21 days’ stable coculture model demonstrated integral barrier formation and the absence of cell layer infiltration despite the highly porous and ultrathin nature of the PCL meshes. Interestingly, these models displayed a similar response as in vivo when induced with inflammation using IL8 where the neutrophils transmigrated across the double cell layers and the BM mimic to reach the site of cytokine addition.…”

“…Similar differences in the expression of αSMA by endothelial cells on nonwoven electrospun meshes and PET membranes were observed by Jain et.al. 33 This illustrates the influence of scaffold structure and property on cell behavior when used to mimic ECM like materials such as the BM. Furthermore, Slater et al demonstrated confluent monolayer formation of cells on the electrospun membranes as opposed to that on hydrogel systems with embedded adhesive ligands and Matrigel.…”

“…To reach the goal of a native-like scaffold, a wide variety of different biomaterials have been synthesized and manufactured, which feature typical BM properties such as intricate fibrillar architecture, the viscoelastic mechanical properties, the adhesive sequences, the dynamic enzyme-induced nature as well as the possibility of controlled storage and release of bioactive substances such as growth factors. 33 , 35 − 43 These biomaterials are mostly fabricated from natural and synthetic polymer systems and try to combine multiple BM properties that are tailored toward the needs of a desired tissue construct or a region of interest. 44 Scaffolds are used to construct in vitro models that are valuable research tools for unraveling fundamental biological processes involved in tissue homeostasis and disease development as well as serving as industrial platforms for drug screening.…”

“…To reach the goal of a native-like scaffold, a wide variety of different biomaterials have been synthesized and manufactured, which feature typical BM properties such as intricate fibrillar architecture, the viscoelastic mechanical properties, the adhesive sequences, the dynamic enzyme-induced nature as well as the possibility of controlled storage and release of bioactive substances such as growth factors. ,− These biomaterials are mostly fabricated from natural and synthetic polymer systems and try to combine multiple BM properties that are tailored toward the needs of a desired tissue construct or a region of interest . Scaffolds are used to construct in vitro models that are valuable research tools for unraveling fundamental biological processes involved in tissue homeostasis and disease development as well as serving as industrial platforms for drug screening. ,,− , Furthermore, despite the success of animal models as an invaluable source of scientific knowledge, animal models are often limited in translation to human biology and response. , Additionally, the three R’s principle when conducting animal studies (reduction, replacement, and refinement) abide by animal ethics and do promote the use of other systems such as simulation or in vitro models when possible. , Therefore, the application of in vitro models and scaffolds might not only uncover scientific knowledge and save patient lives but also reduce the necessity of animal studies.…”