Characterization of Primary Human Leptomeningeal Cells in 2D culture

Abstract

Maintaining the integrity of brain barriers is critical for a healthy central nervous system. While extensive research has focused on the blood-brain barrier (BBB) of the brain vasculature and blood-cerebrospinal fluid barrier (BCSFB) of the choroid plexus, the barriers formed by the meninges have not received as much attention. These membranes create a barrier between the brain and cerebrospinal fluid (CSF), as well as between CSF and blood. Recent studies, particularly on the arachnoid barrier, have revealed its critical role in facilitating the exchange of substances between the brain and CSF. Moreover, this barrier has been implicated in the development of neurological and immunological disorders. In order to gain a deeper comprehension of the functioning and significance of the meningeal barriers, sophisticated models of these barriers, comparable to those of the BBB and BCSFB, need to be created. The aim of this paper is to investigate the characteristics of leptomeningeal cells (LMCs) that form the meningeal barriers, in a cultured environment, including their morphology, proteomics, and barrier properties, and to determine whether passaging of these cells affects their behavior in comparison to their in vivo state. The study employed techniques such as immunofluorescence, immunoblotting, and TEER to analyze the cells. The results indicate that higher passage numbers significantly alter the morphology and protein localization and expression of the LMCs. Additionally, cultured LMCs showed an increase in Vimentin and Cytokeratin expression and a lack of junctional proteins localization on the cell membrane, which could suggest loss of epithelial properties due to culture, preventing barrier formation. This study shows that the optimal LMC passage range is between passages two and five for experimentation, as past this range showed cell properties consistent with EMT at the higher passages.