BBB Model Research Articles

Toward reconstruction of a BBB model by 3D co-culture of endothelial cells and neural stem cells

Toward reconstruction of a BBB model by 3D co-culture of endothelial cells and neural stem cells

Carboxymethyl cellulose coated magnetic nanoparticles transport across a human lung microvascular endothelial cell model of the blood-brain barrier.

Sustained and safe delivery of therapeutic agents across the blood–brain barrier (BBB) is one of the major challenges for the treatment of neurological disorders as this barrier limits the ability of most drug molecules to reach the brain. Targeted delivery of the drugs used to treat these disorders could potentially offer a considerable reduction of the common side effects of their treatment. The preparation and characterization of carboxymethyl cellulose (CMC) coated magnetic nanoparticles (Fe3O4@CMC) is reported as an alternative that meets the need for novel therapies capable of crossing the BBB. In vitro assays were used to evaluate the ability of these polysaccharide coated biocompatible, water-soluble, magnetic nanoparticles to deliver drug therapy across a model of the BBB. As a drug model, dopamine hydrochloride loading and release profiles in physiological solution were determined using UV-Vis spectroscopy. Cell viability tests in Human Lung Microvascular Endothelial (HLMVE) cell cultures showed no significant cell death, morphological changes or alterations in mitochondrial function after 24 and 48 h of exposure to the nanoparticles. Evidence of nanoparticle interactions and nanoparticle uptake by the cell membrane was obtained by electron microscopy (SEM and TEM) analyses. Permeability through a BBB model (the transwell assay) was evaluated to assess the ability of Fe3O4@CMC nanoparticles to be transported across a densely packed HLMVE cell barrier. The results suggest that these nanoparticles can be useful drug transport and release systems for the design of novel pharmaceutical agents for brain therapy.

Peptide-functionalized and high drug loaded novel nanoparticles as dual-targeting drug delivery system for modulated and controlled release of paclitaxel to brain glioma

A dual-targeting drug delivery system for paclitaxel (PTX) was developed by functionalizing novel polyester-based nanoparticles (NPs) with peptides possessing special affinity for low-density lipoprotein receptor (LDLR), overcoming the limitations of the current chemotherapeutics, to transport drug from blood to brain, and then target glioma cells. Employing novel biodegradable block co-polymers (P and 2P), PTX loaded and peptide-functionalized nanoparticles were prepared by a modified nano-co-precipitation method, carried out in one step only without emulsifier, allowing to obtain spherical nanometric (<200 nm), monodisperse (PDI ∼ 0.1), Poly (Ethylene Glycol) (PEG)-coated and high PTX loaded NPs with a slow and controlled release rate for a prolonged period of time. Peptide functionalization, confirmed by fluorimetric assay and HPLC amino acids analysis, enhanced the cellular uptake of functionalized-PTX-NPs by human primary glioblastoma cell line (U-87 MG) and Bovine Brain Endothelial Cells (BBMVECs), compared with non-functionalized-PTX-NPs. To confirm dual-targeting effect, transendothelial transport experiments in an in vitro BBB model and in vitro anti-tumoral activity against U-87 MG revealed that peptide-functionalized-PTX-NPs significantly increased the transport ratio of PTX across the BBB along with an improved anti-proliferative efficiency. Pharmacokinetics and biodistribution studies in rats, carried out by in vivo experiments with 125I radiolabelled dual-targeting PTX-NPs, confirmed the stealthy behavior of NPs and indicated slightly lower levels of penetration into brain tissue in comparison with peptides known to be able to cross the BBB. These promising results suggested that the dual-targeting drug delivery system might have great potential for glioma therapy in clinical applications.

The inhibitory effect of LPS on the expression of GPR81 lactate receptor in blood-brain barrier model in vitro

BackgroundLipopolysaccharide (LPS) is one of the main constituents of the cell wall of gram-negative bacteria. As an endotoxin, LPS induces neuroinflammation, which is associated with the blood-brain barrier impairment. Lactate is a metabolite with some significant physiological functions within the neurovascular unit/blood-brain barrier (BBB). Accumulation of extracellular and cerebrospinal fluid lactate is a specific feature of bacterial meningitis. However, the role of lactate production, transport, and sensing by lactate receptors GPR81 in the pathogenesis of bacterial neuroinflammation is still unknown.MethodsIn this study, we analyzed effects of LPS on the expression of GPR81 and MCT-1 and proliferation of cerebral endothelial cells in the BBB model in vitro. We used molecular profiling methods to measure the expression of GPR81, MCT-1, IL-1β, and Ki67 in the cerebral endothelium after treatment with different concentrations of LPS followed by measuring the level of extracellular lactate, transendothelial electric resistance, and permeability of the endothelial cell layer.ResultsOur findings showed that exposure to LPS results in neuroinflammatory changes associated with decreased expression of GPR81 and MCT-1 in endothelial cells, as well as overproduction of IL-1β and elevation of lactate concentrations in the extracellular space in a dose-dependent manner. LPS treatment reduced JAM tight junction protein expression in cerebral endothelial cells and altered BBB structural integrity in vitro.ConclusionThe impairment of lactate reception and transport might contribute to the alterations of BBB structural and functional integrity caused by LPS-mediated neuroinflammation.

Functionalization and Characterization of an MRI-Capable, Targeted Nanoparticle Platform for Delivery to the Brain

ABSTRACTNovel methods are needed to traverse the blood-brain barrier and deliver drugs to specific targets in the brain. To this end, MS2 bacteriophage was explored as a multifunctional transport and targeting vector. The MS2 capsid exterior was modified with two different targeting moieties for delivery across the BBB and targeting specific regions of interest in the brain. Successful modification of MS2 capsids with a brain targeting peptide and NMADAR2D-targeting antibody was confirmed by immunoblotting and fluorescence detection. To measure transport efficiency of MS2 particles across an in vitro BBB model, a highly sensitive RT-qPCR protocol was developed and implemented. Finally, in order to demonstrate the potential of MS2 as a drug delivery vehicle, nucleotide-mediated loading of capsids was investigated with the MRI contrast agent Gd-DOTA modified with psoralen.

Enhanced permeability of blood-brain barrier and targeting function of brain via borneol-modified chemically solid lipid nanoparticle.

IntroductionThe incidence of central nervous system disease has increased in recent years. However, the transportation of drug is restricted by the blood–brain barrier, contributing to the poor therapeutic effect in the brain. Therefore, the development of a new brain-targeting drug delivery system has become the hotspot of pharmacy.Materials and methodsBorneol, a simple bicyclic monoterpene extracted from Dryobalanops aromatica, can direct drugs to the upper body parts according to the theory of traditional Chinese medicine. Dioleoyl phosphoethanolamine (DOPE) was chemically modified by borneol as one of the lipid materials of solid lipid nanoparticle (SLN) in the present study.ResultsThe borneol-modified chemically solid lipid nanoparticle (BO-SLN/CM), borneol-modified physically solid lipid nanoparticle (BO-SLN/PM), and SLN have similar diameter (of about 87 nm) and morphological characteristics. However, BO-SLN/CM has a lower cytotoxicity, higher cell uptake, and better blood–brain barrier permeability compared with BO-SLN/PM and SLN. BO-SLN/CM has a remarkable targeting function to the brain, while BO-SLN/ PM and SLNs are concentrated at the lung.ConclusionThe present study provides an excellent drug delivery carrier, BO-SLN/CM, having the application potential of targeting to the brain and permeating to the blood–brain barrier.

A novel human induced pluripotent stem cell blood-brain barrier model: Applicability to study antibody-triggered receptor-mediated transcytosis

We have developed a renewable, scalable and transgene free human blood-brain barrier model, composed of brain endothelial cells (BECs), generated from human amniotic fluid derived induced pluripotent stem cells (AF-iPSC), which can also give rise to syngeneic neural cells of the neurovascular unit. These AF-iPSC-derived BECs (i-BEC) exhibited high transendothelial electrical resistance (up to 1500 Ω cm2) inducible by astrocyte-derived molecular cues and retinoic acid treatment, polarized expression of functional efflux transporters and receptor mediated transcytosis triggered by antibodies against specific receptors. In vitro human BBB models enable pre-clinical screening of central nervous system (CNS)-targeting drugs and are of particular importance for assessing species-specific/selective transport mechanisms. This i-BEC human BBB model discriminates species-selective antibody- mediated transcytosis mechanisms, is predictive of in vivo CNS exposure of rodent cross-reactive antibodies and can be implemented into pre-clinical CNS drug discovery and development processes.

The enhancement of siPLK1 penetration across BBB and its anti glioblastoma activity in vivo by magnet and transferrin co-modified nanoparticle

In order to enhance the penetration of small interference RNA against the polo-like kinase I (siPLK1) across BBB to treat glioblastoma (GBM), transferrin (Tf) modified magnetic nanoparticle (Tf-PEG-PLL/MNP@siPLK1) was prepared. The in vitro experiments indicated that Tf-PEG-PLL/MNP@siPLK1 enhanced the cellular uptake of siPLK1, which resulted in an increase of gene silencing effect and cytotoxicity of Tf-PEG-PLL/MNP@siPLK1 on U87 cells. Besides, Tf-PEG-PLL/MNP@siPLK1 significantly inhibited the growth of U87 glioblastoma spheroids and markedly increased the BBB penetration efficiency of siPLK1 with the application of external magnetic field in in-vitro BBB model. The in vivo experiments indicated that siPLK1 selectively accumulated in the brain tissue, and markedly reduced tumor volume and prolonged the survival time of GBM-bearing mice after Tf-PEG-PLL/MNP@siPLK1 was injected to GBM-bearing mice via tail vein. The above data indicated that magnet and transferrin co-modified nanoparticle enhanced siPLK1 penetration across BBB and increased its anti GBM activity in vivo.

The influence and mechanism of ligustilide, senkyunolide I, and senkyunolide A on echinacoside transport through MDCK-MDR1 cells as blood-brain barrier in vitro model.

Efficient transcytosis across the blood-brain-barrier is an important strategy for accessing drug targets within the central nervous system. Ligusticum chuanxiong Hort. was used as a messenger drug to increase the distribution of drugs in brain tissue in Traditional Chinese Medicine. The present study investigates the transport of echinacoside (ECH) through MDCK-MDR1 cell and the effects of ligustilide (LIG), senkyunolide A (SENA) and senkyunolide I (SENI) in chuanxiong on its transport. The results indicated that the absorption of ECH was relatively poor in MDCK-MDR1cells, and was concentration dependent and not saturable. The P-glycoprotein inhibitor verapamil could significantly increase the transport of ECH. It indicated that the transport mechanism might be passive diffusion as the dominating process with the active transportation mediated mechanism involved. The increased apparent permeability of ECH in A→B direction by ethylenediaminetetraacetic acid-Na2 suggested that ECH was absorbed via the paracellular route. The transport of ECH in A→B direction significantly increased when co-administrated with increasing concentrations of LIG, SENI and SENA. Western blot analysis and a decrease in transepithelial electrical resistance during the permeation experiment indicated that LIG, SENI and SENA had enhanced the transport of ECH in the BBB models attribute to down-regulate the expressions of claudin-5 and zonula occludens-1 expression.

Abstract 5785: Characterization of a novel microfluidic in vitro model of the blood-tumor and blood-brain barrier

Abstract Background: There is a need for a cost-effective and reliable in vitro model of the blood-tumor barrier (BBB) and blood-brain barrier (BTB) for primary and metastatic lesions in brain. Current models do not accurately recapitulate physiological conditions and are unreliable with up to a 100-fold variance in predicting drug permeability into brain tumors. Methods: Human umbilical vein endothelial cells (HUVECs) were cultured with either CTX-TRD2 rat astrocytes (BBB) or Met-1 murine breast cancer cells (BTB) in a microfluidic chip, which contained chambers separated by a porous wall. Shear stress was induced on HUVECs by perfusion of media through the apical chamber, and permeability was characterized by the diffusion of fluorescent tracers, one of which is subject to P-gp efflux. Data was compared to in-vivo work. Results: Permeability of free Texas Red was significantly higher (p&amp;lt;0.05) in BTB chips (kin= 13.1 ± 1.3 X 10-3) than in the BBB chips (Kin=2.5 ± 0.3 X 10-3). There was a similar trend observed in larger markers, but these were not significantly different. The movement of Rhodamine 123 was also restricted, indicating functional P-gp efflux in vitro, similar to data from published in vivo studies. In the BBB model, diffusion of Rho123 increased 14-fold in the presence of verapamil (kin=14.7 ± 7.5 X 10-3) and eight fold with the addition of Cyclosporine A (kin=8.8 ± 1.8 X 10-3). In the BTB model, this increase was also observed, but to a lesser extent, as only 3 and 2-fold increases in permeability were observed, respectively. The magntidue of permeability changes were not significantly different from in-vivo models in both passive diffusion and active efflux experiments. Conclusion: The novel microfluidic chip is a cost-effective in vitro model which recapitulates the BTB and BBB physiology and can predict permeability more reliably than current in-vitro models. Citation Format: Jessica Griffith, Tori Terrell-Hall, Amanda Ammer, Paul Lockman. Characterization of a novel microfluidic in vitro model of the blood-tumor and blood-brain barrier [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5785. doi:10.1158/1538-7445.AM2017-5785

Regulation of Human Brain Microvascular Endothelial Cell Adhesion and Barrier Functions by Memantine.

Vascular risk factors have been linked to cognitive decline and dementia in the elderly. Microvascular inflammation, especially of the endothelium, may contribute to the progression of neurodegenerative events in Alzheimer's disease (AD). Memantine, an uncompetitive N-methyl-D-aspartate (NMDA) receptor antagonist, is a licensed drug used for the treatment of moderate to severe AD. However, little information is available regarding its anti-inflammatory effects on the endothelium. In this study, we investigated the effects of memantine on human brain microvascular endothelial dysfunction induced by the pro-inflammatory cytokine tumor necrosis factor-α (TNF-α). Our results show that memantine prevents the attachment of monocyte THP-1 cells to human brain microvascular endothelial cells (HBMVEs). An in vitro BBB model experiment displayed that memantine could rescue TNF-α-induced disruption of the in vitro BBB model. In addition, memantine also interferes with monocyte transmigration across the BBB model. Our results indicate that TNF-α significantly increased the expression of cell adhesion molecules, such as ICAM-1, VCAM-1, and E-selectin, which was prevented by pretreatment with memantine. Mechanistically, memantine reversed activation of the transcription factor NF-κB by preventing the phosphorylation and degradation of its inhibitor IκBα. Our data is the first to describe a novel anti-inflammatory mechanism driven by the endothelial cell-mediated neuroprotective effects of memantine.

Dual-functional nanoparticles for precise drug delivery to Alzheimer’s disease lesions: Targeting mechanisms, pharmacodynamics and safety

Alzheimer’s disease (AD) is the most common form of dementia and is characterized by the cerebral accumulation of extracellular amyloid plaques. In a previous study, this histopathological hallmark was used as a target on a dual-functional nanoparticle (TQNP) to deliver biotechnological drugs, such as the H102 peptide, a β-sheet breaker, to AD lesions precisely. This delivery system could reduce the amyloid-β (Aβ) burden in the brains of AD model mice, as well as ameliorated the memory impairment of the mice. Regretfully, the mechanism how nanoparticles penetrated the BBB and subsequently targeted to the plaques is still unclear. In this study, the internalization, subcellular fate and transportation of the nanoparticles on bEnd.3 cells and an in vitro BBB model, demonstrated that TQNP could be taken up through various routes, including caveolae-mediated endocytosis, suggesting that some of TQNP were able to cross the BBB intact. Then, the TQNP were visualized to specifically bind to the Aβ plaques. TQNP targeting to amyloid plaques might lead to enhanced therapeutic efficacy, which was further evaluated in APP/PS1 transgenic mice. The TQNP/H102 obtained better ability in decreasing amyloid plaques, increasing Aβ-degrading enzymes, reducing tau protein phosphorylation, protecting synapses and improving the spatial learning and memory of transgenic mice than nanoparticles modified with a single ligand. And good biocompatibility of TQNP was indicated with subacute toxicity assays. In conclusion, TQNP was a valuable nanodevice for the precise delivery for biotechnological drugs to treat AD.

Effect of Long-term In Vitro Lithium Exposure on mRNA Levels of Claudin-3, CYP1A1, ABCG2 and GSTM3 Genes in the hCMEC/D3 Human Brain Endothelial Cell Line.

Lithium chloride (LiCl) has been shown to improve the tightness of brain endothelial cell monolayers in vitro by inhibition of the GSK-3β enzyme, activation of the Wnt/beta-catenin pathway and regulation of tight junction (TJ) protein expression. However, the effect of LiCl on the drug transporters and drug-metabolizing enzymes has not been addressed so far. The hCMEC/D3 cell line is a validated in vitro BBB model expressing transporters and drug-metabolizing enzymes (phase 1 and 2). The present study was conducted to compare the mRNAs levels corresponding to several BBB endothelial markers in hCMEC/D3 cells with and without incubation in LiCl. We used quantitative real-time polymerase chain reaction (qRT-PCR) to quantify the mRNA expression of 5 tight junction (TJ) proteins, 4 adhesion proteins, 5 solute carriers, 7 ATP-binding cassette (ABC) transporters, 8 cytochrome P450 (CYP) and 17 phase 2 conjugation enzymes in hCMEC/D3 cells with and without incubation in LiCl. Our study showed that LiCl treatment for 6days at a concentration of 10mM induced the TJ protein claudin-3, the ABC transporter BCRP/ABCG2, the cytochrome P-450 CYP1A1 and the glutathione-S-transferase GSTM3, while the other selected markers were not significantly affected. Our findings provide new insights into the effects of lithium on some drug transporters and drug-metabolizing enzymes in the BBB that may have consequences in pharmacotherapy.

Permeability across a novel microfluidic blood-tumor barrier model

BackgroundThe lack of translatable in vitro blood-tumor barrier (BTB) models creates challenges in the development of drugs to treat tumors of the CNS and our understanding of how the vascular changes at the BBB in the presence of a tumor.MethodsIn this study, we characterize a novel microfluidic model of the BTB (and BBB model as a reference) that incorporates flow and induces shear stress on endothelial cells. Cell lines utilized include human umbilical vein endothelial cells co-cultured with CTX-TNA2 rat astrocytes (BBB) or Met-1 metastatic murine breast cancer cells (BTB). Cells were capable of communicating across microfluidic compartments via a porous interface. We characterized the device by comparing permeability of three passive permeability markers and one marker subject to efflux.ResultsThe permeability of Sulforhodamine 101 was significantly (p < 0.05) higher in the BTB model (13.1 ± 1.3 × 10−3, n = 4) than the BBB model (2.5 ± 0.3 × 10−3, n = 6). Similar permeability increases were observed in the BTB model for molecules ranging from 600 Da to 60 kDa. The function of P-gp was intact in both models and consistent with recent published in vivo data. Specifically, the rate of permeability of Rhodamine 123 across the BBB model (0.6 ± 0.1 × 10−3, n = 4), increased 14-fold in the presence of the P-gp inhibitor verapamil (14.7 ± 7.5 × 10−3, n = 3) and eightfold with the addition of Cyclosporine A (8.8 ± 1.8 × 10−3, n = 3). Similar values were noted in the BTB model.ConclusionsThe dynamic microfluidic in vitro BTB model is a novel commercially available model that incorporates shear stress, and has permeability and efflux properties that are similar to in vivo data.

Substance P-mediated chemokine production promotes monocyte migration.

The neuropeptide SP has physiologic and pathophysiologic roles in CNS and peripheral tissues and is involved in crosstalk between nervous and immune systems in various conditions, including HIV and SIV infection. Increased SP levels were demonstrated in plasma of HIV+ individuals as well as in the CNS of SIV-infected, nonhuman primates. SP increases HIV infection in macrophages through interaction with its receptor, NK1R. The SP effect on immune system is both pro- and anti-inflammatory and includes up-regulation of a number of cytokines and cell receptors. The main goal of this study was to determine whether there is interplay between monocyte exposure to SP and recruitment into sites of inflammation. We now demonstrate that exposure of either human macrophages or PBMCs to SP leads to increased production of chemokines, including MCP-1, for which expression is limited to cells of the myeloid lineage. This effect is inhibited by the NK1R antagonist, aprepitant. Exposure to conditioned medium derived from SP-treated PBMCs resulted in increased monocyte migration through semipermeable membranes and an in vitro human BBB model. Monocyte migration was blocked by anti-MCP-1 antibodies. Our results suggest that increased SP levels associated with HIV and other inflammatory conditions may contribute to increased monocyte migration into the CNS and other tissues through a MCP-1-dependent mechanism.

Double-targeted polymersomes and liposomes for multiple barrier crossing

In order to treat metastasis in the brain, drug delivery systems must overcome multiple physical barriers between the point of administration and the target, such as the Blood-brain barrier, that hinder their free access across them. Multiple targeting approaches arise as a promising alternative to this barrier and target certain tissues inside the brain at a time. Herein, two surface modification methods are presented to obtain dual-targeted vesicle-like carriers functionalized with an MCF-7-specific phage protein and a BBB-specific peptide, providing the system the ability to cross a BBB model, target breast cancer cells and deliver its payload. The aim of this study was to compare new designed polymersomes with liposomes, a well-established delivery vehicle, in terms of drug loading, targeting, release and tumor cell killing. The bilayer structure of both systems allowed the conjugation with different ligands both by insertion and covalent binding. Different behaviour was observed in release, uptake and tumor cell killing corresponding to differences in membrane permeability of both vehicles and type of targeting and ligands’ combination. Preliminary results showed that both formulations were able to cross the BBB monolayer without harming it, showing cytotoxic activity in the abluminal compartment.

The Role of Nanoparticle in Brain Permeability: An in-vitro BBB Model.

Membrane permeability and P-glycoprotein (P-gp) efflux system are regulating factors in the drug brain penetration. Recently, some drug delivery systems have been developed to overcome these limitations. In this study, Metoclopramid has been encapsulated in PLGA nanoparticles using the emulsification/solvent evaporation technique for in-viro evaluation of the effect of PLGA nanoparticles on BBB permeability. Subsequently, prepared nanoparticles were characterized using PCS, TEM, FT-IR, DSC and XRD techniques and in-viro cell permeability of optimum formulation was evaluated using MDCK cell line as BBB model. Data investigation showed that prepared nanoparticles have the entrapment efficiency of 50 %. PCS investigation showed that prepared nanoparticles have an average size of approximately 150 ± 14 nm and a relatively monodisperse distribution. TEM micrographs of the samples showed spherical shape and smooth surface with a particle size of nanometric range. Through DSC thermograms and XRD diffractograms analysis, it was demonstrated that there was no crystalline form of the drug in the loaded formulation. Moreover, our results showed that the greater crossing of metoclopramide in the form of nanoparticle in comparison with the free form. The widely used rhodamine-123 transport assay performed in the MDCK cells demonstrated the presence of P-glycoprotein in this model.

Bifunctional Peptide-Based Opioid Agonist-Nociceptin Antagonist Ligands for Dual Treatment of Acute and Neuropathic Pain.

Herein, the opioid pharmacophore H-Dmt-d-Arg-Aba-β-Ala-NH2 (7) was linked to peptide ligands for the nociceptin receptor. Combination of 7 and NOP ligands (e.g., H-Arg-Tyr-Tyr-Arg-Ile-Lys-NH2) led to binding affinities in the low nanomolar domain. In vitro, the hybrids behaved as agonists at the opioid receptors and antagonists at the nociceptin receptor. Intravenous administration of hybrid 13a (H-Dmt-d-Arg-Aba-β-Ala-Arg-Tyr-Tyr-Arg-Ile-Lys-NH2) to mice resulted in potent and long lasting antinociception in the tail-flick test, indicating that 13a was able to permeate the BBB. This was further supported by a cell-based BBB model. All hybrids alleviated allodynia and hyperalgesia in neuropathic pain models. Especially with respect to hyperalgesia, they showed to be more effective than the parent compounds. Hybrid 13a did not result in significant respiratory depression, in contrast to an equipotent analgesic dose of morphine. These hybrids hence represent a promising avenue toward analgesics for the dual treatment of acute and neuropathic pain.

Elucidation of Transport Mechanism of Paeoniflorin and the Influence of Ligustilide, Senkyunolide I and Senkyunolide A on Paeoniflorin Transport through Mdck-Mdr1 Cells as Blood-Brain Barrier in Vitro Model.

The objectives of the present investigation were to: (1) elucidate the transport mechanism of paeoniflorin (PF) across MDCK-MDR1 monolayers; and (2) evaluate the effect of ligustilide (LIG), senkyunolide I (SENI) and senkyunolide A (SENA) on the transport of PF through blood–brain barrier so as to explore the enhancement mechanism. Transport studies of PF were performed in both directions, from apical to basolateral side (A→B) and from basolateral to apical sides (B→A). Drug concentrations were analyzed by LC-MS/MS. PF showed relatively poor absorption in MDCK-MDR1 cells, apparent permeability coefficients (Papp) ranging from 0.587 × 10−6 to 0.705 × 10−6 cm/s. In vitro experiments showed that the transport of PF in both directions was concentration dependent and not saturable. The B→A/A→B permeability ER of PF was more than 2 in the MDCK-MDR1 cells, which indicated that the transport mechanism of PF might be passive diffusion as the dominating process with the active transportation mediated mechanism involved. The increased Papp of PF in A→B direction by EDTA-Na2 suggested that PF was absorbed via the paracellular route. The P-gp inhibitor verapamil could significantly increase the transport of PF in A→B direction, and ER decreased from 2.210 to 0.690, which indicated that PF was P-gp substance. The transport of PF in A→B direction significantly increased when co-administrated with increasing concentrations of LIG, SENI and SENA. An increased cellular accumulation of Rho 123 and Western blot analysis indicated that LIG, SENI and SENA had increased the transport of PF in the BBB models attribute to down-regulate P-gp expression. A decrease in transepithelial electrical resistance (TEER) during the permeation experiment can be explained by the modulation and opening of the tight junctions caused by the permeation enhancer LIG, SENI and SENA.

Synthesis and in Vitro Studies of a Series of Carborane-Containing Boron Dipyrromethenes (BODIPYs).

A series of seven BODIPYs functionalized with ortho-carborane groups at the 8(meso) or 3/5(α) position were synthesized and characterized by NMR, HRMS, HPLC, and in the cases of 2b and 5b, by X-ray analysis. The BODIPYs exhibited low dark toxicity and phototoxicity toward human glioma T98G cells, and their cellular uptake varied significantly, with 5b accumulating the most and 7 the least. All BODIPYs localized mainly within the cell ER. The BODIPYs showed higher permeabilities than lucifer yellow across human hCMEC/D3 brain endothelial cell monolayers as the BBB model. Among this series, 1b showed the highest BBB permeability (Pe = 16.4 × 10(-5) cm/s), probably as a result of its lower MW (366 Da) and favorable hydrophobicity (logP = 1.5). The combination of low cytotoxicity, amphiphilicity, high boron content, high cellular uptake, and moderate BBB permeability renders these compounds promising boron delivery agents for the BNCT of brain tumors.