Porous Optically Transparent Cellulose Acetate Scaffolds for Biomimetic Blood-Brain Barrierin vitro Models

Abstract

In vitro blood-brain barrier (BBB) models represent an efficient platform to conduct high-throughput quantitative investigations on BBB crossing ability of different drugs. Such models provide a closed system where different fundamental variables can be efficaciously tuned and monitored, and issues related to scarce accessibility of animal brains and ethics can be addressed. In this work, we propose the fabrication of cellulose acetate (CA) porous bio-scaffolds by exploiting both vapor-induced phase separation (VIPS) and electrospinning methods. Parameters of fabrication have been tuned in order to obtain porous and transparent scaffolds suitable for optical/confocal microscopy, where endothelial cell monolayers are allowed to growth thus obtaining biomimetic BBB in vitro models. Concerning VIPS-based approach, CA membranes fabricated using 25% H2O + 75% EtOH as non-solvent showed submicrometer-scale porosity and an optical transmittance comparable to that one of commercially available poly(ethylene terephthalate) membranes. CA membranes fabricated via VIPS have been exploited for obtaining multicellular BBB models through the double seeding of endothelial cells and astrocytes on the two surfaces of the membrane. Electrospun CA substrates, instead, were characterized by micrometer-sized pores, and were unsuitable for double seeding approach and long term studies. However, the potential exploitation of the electrospun CA substrates for modeling blood-brain-tumor barrier and studying cell invasiveness has been speculated. The features of the obtained models have been critically compared and discussed for future applications.

Electrospun Scaffolds as Cell Culture Substrates for the Cultivation of an In Vitro Blood–Brain Barrier Model Using Human Induced Pluripotent Stem Cells

“…Few individual studies on in vitro BBB models incorporating electrospun fibers as already exist. However, these models exhibited TEER values in the range of 10-250 Ω × cm 2 , displaying weak barrier properties [51,[61][62][63]. In this study, TEER values were measured over a cultivation time of 72 h. Generally, the TEER values of hiPS-BCECs on all electrospun scaffolds increased over 72 h. Nevertheless, the characteristics of each individual scaffold strongly influenced the final barrier properties of the cell layer, as shown in Figure 9A.…”

Liposomes loaded with polyphenol-rich grape pomace extracts protect from neurodegeneration in a rotenone-basedin vitromodel of Parkinson's disease

“…The in vitro multicellular BBB model was obtained following the double seeding approach presented in a previous work. 54 Human astrocytes were seeded on the bottom surface (i.e., abluminal side) of 3 μm pore transwell inserts (Corning Incorporated) at a density of 10 000 cells per cm 2 and left for 15 h to adhere to the porous membranes in the humid chamber of the incubator. Subsequently, inserts were placed into a 24 multi-well plate and hCMEC/D3 cells were seeded at a high density (80 000 cells per cm 2 ) on the upper surface (luminal side) of the inserts.…”

“…S7 †) similarly as previously described. 54,93 In Fig. S7a, † the schematic representation of the in vitro BBB system with luminal (in blue) and abluminal (in red) compartments separated by astrocytes and by an endothelial cell layer is shown.…”

Vascularizing the brain in vitro

“…The electrospinning technology is a precise and versatile technology that allows the fabrication of a nanofiber-based platform. Several synthetic and natural polymers have been used to achieve electrospun membranes ( Chen et al., 2021 ; Marino et al., 2021 ; Ozyurt et al., 2020 ; Patient et al., 2019 ). The fibrous structure produced using this method offers essential advantages that can ameliorate the functions of BBB models.…”