Gene Expression Changes in Long-Term In Vitro Human Blood-Brain Barrier Models and Their Dependence on a Transwell Scaffold Material.

Joel D Gaston,Lisa A Fitzgerald,Kathleen D Cusick,Bradley R Ringeisen,Lauren L Bischel,Russell K Pirlo

doi:10.1155/2017/5740975

Abstract

Disruption of the blood-brain barrier (BBB) is the hallmark of many neurovascular disorders, making it a critically important focus for therapeutic options. However, testing the effects of either drugs or pathological agents is difficult due to the potentially damaging consequences of altering the normal brain microenvironment. Recently, in vitro coculture tissue models have been developed as an alternative to animal testing. Despite low cost, these platforms use synthetic scaffolds which prevent normal barrier architecture, cellular crosstalk, and tissue remodeling. We created a biodegradable electrospun gelatin mat “biopaper” (BP) as a scaffold material for an endothelial/astrocyte coculture model allowing cell-cell contact and crosstalk. To compare the BP and traditional models, we investigated the expression of 27 genes involved in BBB permeability, cellular function, and endothelial junctions at different time points. Gene expression levels demonstrated higher expression of transcripts involved in endothelial junction formation, including TJP2 and CDH5, in the BP model. The traditional model had higher expression of genes associated with extracellular matrix-associated proteins, including SPARC and COL4A1. Overall, the results demonstrate that the BP coculture model is more representative of a healthy BBB state, though both models have advantages that may be useful in disease modeling.

Highlights

The unique characteristics of the blood-brain barrier (BBB) present a considerable challenge for studying central nervous system (CNS) therapeutics and disease modeling
In order to confirm this for both the biopaper and polyethylene terephthalate (PET) models, GJB4 and GJB6 were included on the array as negative controls
The lack of GJB6 expression is readily explained and expected by the lack of cellular signaling from neurons within the culture. These findings demonstrate that both coculture models do not display gene expression not associated with the BBB, due to either constitutive lack of expression or the requirement of additional cellular cues

Summary

Introduction

The unique characteristics of the blood-brain barrier (BBB) present a considerable challenge for studying central nervous system (CNS) therapeutics and disease modeling. The intrinsic difficulty of crossing the BBB necessitates extensive testing for new drug candidates using relevant in vitro and in vivo models. In vivo model organisms, such as rats, provide valuable information in drug screening assays, with respect to complex effects in the natural CNS environment. They are expensive and time consuming, face ethical concerns, require a considerable physical space, and often do no not translate well to human results [2, 3]. In vitro model systems consisting of cell-based arrays are a valuable tool to screen molecular BBB permeability prior to animal testing

Results

Discussion

Conclusion