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The development of drugs targeting the central nervous system (CNS) is challenging because of the presence of the Blood-Brain barrier (BBB). Developing physiologically relevant in vitro BBB models for evaluating drug permeability and predicting the activity of drug candidates is crucial. The transwell model is one of the most widely used in vitro BBB models. However, this model has limitations in mimicking in vivo conditions, particularly in the absence of shear stress. This study aimed to overcome the limitations of the transwell model using immortalized human endothelial cells (hCMEC/D3) by developing a novel dish design for an orbital shaker, providing shear stress. During optimization, we assessed cell layer integrity using trans-endothelial electrical resistance measurements and the % diffusion of lucifer yellow. The efflux transporter activity and mRNA expression of junctional proteins (claudin-5, occludin, and VE-cadherin) in the newly optimized model were verified. Additionally, the permeability of 14 compounds was evaluated and compared with published in vivo data. The cell-layer integrity was substantially increased using the newly designed annular shaking-dish model. The results demonstrate that our model provided robust conditions for evaluating the permeability of CNS drug candidates, potentially improving the reliability of in vitro BBB models in drug development. [ABSTRACT FROM AUTHOR]

[Display omitted] • A Caco-2 monolayer model was developed using medium customization and sodium valerate. • The abbreviated model showed improved transporter activity and monolayer integrity. • The model demonstrated consistent and reliable performance over multiple days. • The model has economic and practical advantages for drug permeability assessment. The human colorectal adenocarcinoma cell line Caco-2, widely used for studying intestinal drug permeability, is typically grown on permeable filter supports and matures in 21 days with frequent media changes. The process is labor-intensive, prone to contamination, and has low throughput, contributing to the overall high utilization cost. Efforts to establish a low-cost, high-throughput, and short-duration model have encountered obstacles, such as weaker tight junctions causing monolayer leaks, incomplete differentiation resulting in low transporter expression, intricate and challenging protocols, and cytotoxicity, limiting the usability. Hence, this study aimed to develop a low-cost, efficient, and short-duration model by addressing the aforementioned concerns by customizing the media and finding a safe differentiation inducer. We generated a new rapid model using sodium valerate, which demonstrated sufficient transporter activity, improved monolayer integrity, and higher levels of differentiation markers than the 21-day model. Furthermore, this model exhibited consistent and reliable results when used to evaluate drug permeability over multiple days of repeated use. This study demonstrates the potential of a sodium valerate-assisted abbreviated model for drug permeability assessment with economic and practical advantages. [ABSTRACT FROM AUTHOR]

α-mangostin is a dietary xanthone which has been shown to have antioxidant, anti-allergic, antiviral, antibacterial, anti-inflammatory and anticancer effects in various types of human cancer cells. In the present study, we aimed to elucidate the molecular mechanisms responsible for the apoptosis-inducing effects of α-mangostin on YD-15 tongue mucoepidermoid carcinoma cells. The results from MTT assays revealed that cell proliferation significantly decreased in a dose-dependent manner in the cells treated with α-mangostin. DAPI staining illustrated that chromatin condensation in the cells treated with 15 µM α-mangostin was far greater than that in the untreated cells. Flow cytometric analysis indicated that α-mangostin suppressed YD-15 cell viability by inducing apoptosis and promoting cell cycle arrest in the sub-G1 phase. Western blot analysis of various signaling molecules revealed that α-mangostin targeted the extracellular signal‑regulated kinase 1/2 (ERK1/2) and p38 mitogen-activated protein kinase (MAPK) signaling pathways through the inhibition of ERK1/2 and p38 phosphorylation in a dose‑dependent manner. α-mangostin also increased the levels of Bax (pro-apoptotic), cleaved caspase-3, cleaved caspase-9 and cleaved-poly(ADP-ribose) polymerase (PARP), whereas the levels of the anti-apoptotic factors, Bcl-2 and c-myc, decreased in a dose-dependent manner. The anticancer effects of α-mangostin were also investigated in a tumor xenograft mouse model. The α-mangostin-treated nude mice bearing YD-15 tumor xenografts exhibited a significantly reduced tumor volume and tumor weight due to the potent promoting effects of α-mangostin on cancer cell apoptosis, as determined by TUNEL assay. Immunohistochemical analysis revealed that the level of cleaved caspase-3 increased, whereas the Ki-67, p-ERK1/2 and p-p38 levels decreased in the α-mangostin‑treated mice. Taken together, the findings of our study indicate that α-mangostin induces the apoptosis of YD-15 tongue carcinoma cells through the ERK1/2 and p38 MAPK signaling pathways.