Neisseria meningitidis Infection of Induced Pluripotent Stem-Cell Derived Brain Endothelial Cells

The blood-brain barrier (BBB), and the meningeal blood-CSF barrier (mBCSFB) are extremely tight cellular barriers that separate the circulation from the central nervous system (CNS) and are primarily comprised of highly specialized brain endothelial cells (BECs)^1,2. Together, BECs maintain proper brain homeostasis by regulating nutrients and waste products in and out of the brain, while excluding many toxins, drugs, and pathogens^1,2. Bacterial meningitis occurs when blood-borne bacteria are able to interact with, and penetrate the barrier formed by BECs and cause inflammation. Neisseria meningitidis (Nm, meningococcus) is a Gram-negative bacterium that colonizes the nasopharaynx of 10‒40 % of healthy individuals, but in some cases can cause serious systemic disease³. In affected individuals, Nm can gain access to the blood stream where it can cause purpura fulminans as well as penetrate BECs gaining access to the CNS causing meningitis³. Nm is a leading cause of bacterial meningitis world-wide, and despite vaccination efforts, is still a primary cause of meningitis⁴. Modern medical intervention, such as antibiotic treatment, have made these conditions survivable, however those affected with meningitis often are left with permanent neurological damage^5,6.

Previous studies have identified bacterial factors and host signaling that contribute to Nm-BEC interactions^7,8,9,10,11. The identified adhesins and invasins such as the opacity protein Opc, and type-IV pili, as well as receptors such as CD147, have been conducted on various BEC models in vitro, however these models lack many defining BBB properties^7,9,11,12. Complete understanding of Nm-BEC interactions remain elusive due partially to the inability to utilize in vivo models, incomplete vaccination protection, and lack of robust human BEC models in vitro.

Modeling hBECs in vitro has been challenging due to the unique properties of BECs. Compared with peripheral endothelial cells, BECs have a number of phenotypes that enhance their barrier properties such as high trans-endothelial electrical resistance (TEER) due to complex tight junctions¹². Once removed from the brain microenvironment, BECs rapidly lose their barrier properties limiting the usefulness of primary or immortalized in vitro models that only form a weak barrier^12,13. The combination of the human specificity of Nm infections, lack of robust in vivo models, and challenges modeling human BECs in vitro creates a need for better models to understand the complex host-pathogen interaction between Nm and BECs. Recently using model human induced pluripotent stem cell (iPSC) technologies BEC-like cells have been derived from iPSCs that better mimic BECs in vivo^12,13,14,15. iPSC-BECs are of human origin, easily scalable, and possess expected BEC phenotypes compared to their primary or immortalized counterparts^12,13,14,15. Additionally we and others have demonstrated that iPSC-BECs are useful for modeling various diseases of the CNS such as host-pathogen interaction, Huntington’s disease, and MCT8 deficiency that causes Allan-Hurndon-Dudley syndrome^{16,17,18,19,20,21}. Here, we demonstrate how to derive iPSC-BECs from renewable iPSC sources and the infection of iPSC-BECs with Nm leading to activation of the innate immune response. We believe that this model is useful to interrogate host-pathogen interaction that is unable to be recapitulated in other in vitro models and is especially useful when examining interactions with human specific pathogens such as Nm.