Rapid, Reversible Modulation of Blood-Brain Barrier P-Glycoprotein Transport Activity by Vascular Endothelial Growth Factor

Laboratory of Pharmacology, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709, USA.
The Journal of Neuroscience : The Official Journal of the Society for Neuroscience (Impact Factor: 6.34). 01/2010; 30(4):1417-25. DOI: 10.1523/JNEUROSCI.5103-09.2010
Source: PubMed


Increased brain expression of vascular endothelial growth factor (VEGF) is associated with neurological disease, brain injury, and blood-brain barrier (BBB) dysfunction. However, the specific effect of VEGF on the efflux transporter P-glycoprotein, a critical component of the BBB, is not known. Using isolated rat brain capillaries and in situ rat brain perfusion, we determined the effect of VEGF exposure on P-glycoprotein activity in vitro and in vivo. In isolated capillaries, VEGF acutely and reversibly decreased P-glycoprotein transport activity without decreasing transporter protein expression or opening tight junctions. This effect was blocked by inhibitors of the VEGF receptor flk-1 and Src kinase, but not by inhibitors of phosphatidylinositol-3-kinase or protein kinase C. VEGF also increased Tyr-14 phosphorylation of caveolin-1, and this was blocked by the Src inhibitor PP2. Pharmacological activation of Src kinase activity mimicked the effects of VEGF on P-glycoprotein activity and Tyr-14 phosphorylation of caveolin-1. In vivo, intracerebroventricular injection of VEGF increased brain distribution of P-glycoprotein substrates morphine and verapamil, but not the tight junction marker, sucrose; this effect was blocked by PP2. These findings indicate that VEGF decreases P-glycoprotein activity via activation of flk-1 and Src, and suggest Src-mediated phosphorylation of caveolin-1 may play a role in downregulation of P-glycoprotein activity. These findings also imply that P-glycoprotein activity is acutely diminished in pathological conditions associated with increased brain VEGF expression and that BBB VEGF/Src signaling could be targeted to acutely modulate P-glycoprotein activity and thus improve brain drug delivery.

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    • "VEGF was increased several fold in the CSF in experimental chronic hydrocephalus and the increase was significantly correlated with ventricular volume (78). Excess VEGF in the CSF is known to decrease p-gp activity, thus contributing to hydrocephalus (79). "
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    ABSTRACT: Hydrocephalus is a common brain disorder that is treated only with surgery. The basis for surgical treatment rests on the circulation theory. However, clinical and experimental data to substantiate circulation theory have remained inconclusive. In brain tissue and in the ventricles, we see that osmotic gradients drive water diffusion in water-permeable tissue. As the osmolarity of ventricular CSF increases within the cerebral ventricles, water movement into the ventricles increases and causes hydrocephalus. Macromolecular clearance from the ventricles is a mechanism to establish the normal CSF osmolarity, and therefore ventricular volume. Efflux transporters, (p-glycoprotein), are located along the blood brain barrier and play an important role in the clearance of macromolecules (endobiotics and xenobiotics) from the brain to the blood. There is clinical and experimental data to show that macromolecules are cleared out of the brain in normal and hydrocephalic brains. This article summarizes the existing evidence to support the role of efflux transporters in the pathogenesis of hydrocephalus. The location of p-gp along the pathways of macromolecular clearance and the broad substrate specificity of this abundant transporter to a variety of different macromolecules are reviewed. Involvement of p-gp in the transport of amyloid beta in Alzheimer disease and its relation to normal pressure hydrocephalus is reviewed. Finally, individual variability of p-gp expression might explain the variability in the development of hydrocephalus following intraventricular hemorrhage.
    Croatian Medical Journal 08/2014; 55(4):366-376. DOI:10.3325/cmj.2014.55.366 · 1.31 Impact Factor
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    • "A possible explanation of this discrepancy is that the ratio of the intrinsic transport activity per P-gp molecule to the passive diffusion rate at the BBB in EL mice may be lower than that in the in vitro P-gp–transfected LLC-PK1 cells used for the reconstruction of the K p brain values in the present study. The intrinsic P-gp transport activity in brain capillaries has been suggested to decrease under inflammatory conditions (e.g., upon TNF-a stimulation) and also under pathologic conditions associated with increased expression of vascular endothelial growth factor in brain (Hawkins et al., 2010; Miller, 2010). In the rodent model of epilepsy, protein levels of vascular endothelial growth factor and inflammatory mediators, including TNF-a, in brain are increased after the induction of seizures (Croll et al., 2004; Vezzani and Granata, 2005; Rigau et al., 2007; Vezzani et al., 2011). "
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    ABSTRACT: The purpose of this study was to demonstrate experimentally that alterations of in vivo transporter function at the blood-brain barrier (BBB) in disease and during pharmacotherapy can be reconstructed from in vitro data based on our established pharmacoproteomic concept of reconstructing in vivo function by integrating intrinsic transport activity per transporter molecule and absolute protein expression level at the BBB. Pentylenetetrazole-kindled (PTZ) and EL mice were used as models of chemically induced and spontaneous epilepsy, respectively. A mouse model of anti-epileptic drug treatment was prepared by consecutive 5-week administration of phenytoin (PHT). Quantitative targeted absolute proteomic analysis of 31 membrane proteins showed that P-glycoprotein (P-gp/mdr1a) protein expression levels were significantly increased in brain capillaries of PTZ (129%), EL (143%), and PHT mice (192%) compared to controls. The brain-to-plasma concentration ratios (Kp brain) of P-gp/mdr1a substrate verapamil were 0.563, 0.394, 0.432, and 0.234 in control, PTZ, EL, and PHT mice, respectively. In vivo P-gp/mdr1a function at the BBB was reconstructed from the measured P-gp/mdr1a protein expression levels and intrinsic transport activity for verapamil per P-gp/mdr1a previously reported by our group. Then, the reconstructed P-gp/mdr1a functional activities were integrated with unbound fractions of verapamil in plasma and brain to reconstruct Kp brain of verapamil. In all mice, reconstructed Kp brain values agreed well with the observed values within a 1.21-fold range. These results demonstrate that altered P-gp functions at the BBB in epilepsy and during pharmacotherapy can be reconstructed from in vitro data by means of our pharmacoproteomic approach.
    Drug metabolism and disposition: the biological fate of chemicals 07/2014; 42(10). DOI:10.1124/dmd.114.059055 · 3.25 Impact Factor
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    • "Finally, the entry of toxins and pathogens, such as bacteria and viruses circulating in the blood, can lead to neuron cell death and hence must also be prevented (Begley and Brightman, 2003; Hawkins and Davis, 2005; Abbott et al., 2010). The tight junctions formed by brain microvascular endothelial cells (BMECs) regulate paracellular transport whereas transcellular transport is regulated by specialized transporters, pumps, and receptors (Figure 1) (Chishty et al., 2001; Demeule et al., 2002; Hawkins et al., 2002; Ohtsuki and Terasaki, 2007; Ueno, 2009; Abbott et al., 2010; Hartz and Bauer, 2011). "
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    ABSTRACT: It has been more than 100 years since Paul Ehrlich reported that various water-soluble dyes injected into the circulation did not enter the brain. Since Ehrlich's first experiments, only a small number of molecules, such as alcohol and caffeine have been found to cross the blood-brain barrier, and this selective permeability remains the major roadblock to treatment of many central nervous system diseases. At the same time, many central nervous system diseases are associated with disruption of the blood-brain barrier that can lead to changes in permeability, modulation of immune cell transport, and trafficking of pathogens into the brain. Therefore, advances in our understanding of the structure and function of the blood-brain barrier are key to developing effective treatments for a wide range of central nervous system diseases. Over the past 10 years it has become recognized that the blood-brain barrier is a complex, dynamic system that involves biomechanical and biochemical signaling between the vascular system and the brain. Here we reconstruct the structure, function, and transport properties of the blood-brain barrier from an engineering perspective. New insight into the physics of the blood-brain barrier could ultimately lead to clinical advances in the treatment of central nervous system diseases.
    Frontiers in Neuroengineering 08/2013; 6:7. DOI:10.3389/fneng.2013.00007
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