In vitro porcine blood–brain barrier model for permeability studies: pCEL-X software pKaFLUX method for aqueous boundary layer correction and detailed data analysis

Abstract

In vitro blood-brain barrier (BBB) models from primary brain endothelial cells can closely resemble the in vivo BBB, offering valuable models to assay BBB functions and to screen potential central nervous system drugs. We have recently developed an in vitro BBB model using primary porcine brain endothelial cells. The model shows expression of tight junction proteins and high transendothelial electrical resistance, evidence for a restrictive paracellular pathway. Validation studies using small drug-like compounds demonstrated functional uptake and efflux transporters, showing the suitability of the model to assay drug permeability. However, one limitation of in vitro model permeability measurement is the presence of the aqueous boundary layer (ABL) resulting from inefficient stirring during the permeability assay. The ABL can be a rate-limiting step in permeation, particularly for lipophilic compounds, causing underestimation of the permeability. If the ABL effect is ignored, the permeability measured in vitro will not reflect the permeability in vivo. To address the issue, we explored the combination of in vitro permeability measurement using our porcine model with the pKa(FLUX) method in pCEL-X software to correct for the ABL effect and allow a detailed analysis of in vitro (transendothelial) permeability data, Papp. Published Papp using porcine models generated by our group and other groups are also analyzed. From the Papp, intrinsic transcellular permeability (P0) is derived by simultaneous refinement using a weighted nonlinear regression, taking into account permeability through the ABL, paracellular permeability and filter restrictions on permeation. The in vitro P0 derived for 22 compounds (35 measurements) showed good correlation with P0 derived from in situ brain perfusion data (r(2)=0.61). The analysis also gave evidence for carrier-mediated uptake of naloxone, propranolol and vinblastine. The combination of the in vitro porcine model and the software analysis provides a useful tool to better predict BBB permeability in vivo and gain better mechanistic information about BBB permeation.

Establishment of a Human Blood-Brain Barrier Co-culture Model Mimicking the Neurovascular Unit Using Induced Pluri- and Multipotent Stem Cells

“…In in vitro tests, caffeine, a more hydrophilic compound, was shown to permeate almost as fast as diazepam, a lipophilic compound often used as a fast-migrating transcellular marker that does not interact with efflux transporters (Zhao and Pollack, 2009, Mealey et al., 2010). On the contrary, in vivo data revealed that caffeine was significantly slower than diazepam, probably due to interactions with active transporters (Nakazono et al., 1992, Yusof et al., 2014). These in vivo observations were also reflected in our models.…”

Hydrothermal synthesis of nitrogen-doped carbon dots with real-time live-cell imaging and blood–brain barrier penetration capabilities

“…12,13 For instance, Wang et al used CDs as a drug delivery system for doxorubicin (DOX), which produced a pH-controlled DOX release. 14 Recently, an in vitro study demonstrated that CDs can potently inhibit insulin fibrillation, which is associated with neurological diseases.…”

Современные Возможности Идентификации Новых Молекул-Мишеней Для Фармакологической Коррекции Проницаемости Гематоэнцефалического Барьера

“…Åùå îäíèì èç ñïîñîáîâ èçó÷åíèÿ ïà-òîãåíåçà çàáîëåâàíèé ÖÍÑ è ïàòîëîãèè ÃÝÁ ÿâëÿåòñÿ èñïîëüçîâàíèå êëåòî÷íûõ ìîäåëåé ÃÝÁ in vitro [84]. Ïåðâîå ïîêîëåíèå aeèçíåñïîñîáíûõ êëåòî÷íûõ ìîäå-ëåé in vitro ïîÿâèëîñü îêîëî 30 ëåò íàçàä [61].…”