Abstract
Brain microvascular endothelial cells (BEC) constitute the blood-brain barrier (BBB) which forms a dynamic interface between the blood and the central nervous system (CNS). This highly specialized interface restricts paracellular diffusion of fluids and solutes including chemicals, toxins and drugs from entering the brain. In this study we compared the transcriptome profiles of the human immortalized brain endothelial cell line hCMEC/D3 and human primary BEC. We identified transcriptional differences in immune response genes which are directly related to the immortalization procedure of the hCMEC/D3 cells. Interestingly, astrocytic co-culturing reduced cell adhesion and migration molecules in both BECs, which possibly could be related to regulation of immune surveillance of the CNS controlled by astrocytic cells within the neurovascular unit. By matching the transcriptome data from these two cell lines with published transcriptional data from freshly isolated mouse BECs, we discovered striking differences that could explain some of the limitations of using cultured BECs to study BBB properties. Key protein classes such as tight junction proteins, transporters and cell surface receptors show differing expression profiles. For example, the claudin-5, occludin and JAM2 expression is dramatically reduced in the two human BEC lines, which likely explains their low transcellular electric resistance and paracellular leakiness. In addition, the human BEC lines express low levels of unique brain endothelial transporters such as Glut1 and Pgp. Cell surface receptors such as LRP1, RAGE and the insulin receptor that are involved in receptor-mediated transport are also expressed at very low levels. Taken together, these data illustrate that BECs lose their unique protein expression pattern outside of their native environment and display a more generic endothelial cell phenotype. A collection of key genes that seems to be highly regulated by the local surroundings of BEC within the neurovascular unit are presented and discussed.
Highlights
The specific microenvironment of the central nervous system (CNS) is vital for proper neuronal function
In this study we present evidence that those genes in Brain microvascular endothelial cells (BEC) that are highly expressed and described to be important for ensuring blood brain barrier (BBB)-like properties are affected by culturing the cells in isolation
The information was gathered by investigating how BECs respond to culture conditions without their in vivo surrounding
Summary
The specific microenvironment of the central nervous system (CNS) is vital for proper neuronal function. The anatomical constituents of the BBB are the specialized brain endothelial cells (BECs) that together with pericytes, astrocytes, neurons, and possibly other glial cells, comprise the neurovascular unit (NVU) [3,4,5,6]. All these cell types contribute to the functioning of the brain microvasculature, only the BECs are thought to control permeability directly or indirectly via stimuli from the other cells in the NVU [7,8]. There is a growing need for reliable bench models that predict important in vivo properties These models would facilitate our understanding of key biological functions of the BBB and allow study of specific transport systems potentially suitable for delivering drugs to the brain. Over the last few years, the isolation and culture of BEC have advanced significantly, resulting in a variety of BBB models
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