scholarly journals An Atlas of the Quantitative Protein Expression of Anti-Epileptic-Drug Transporters, Metabolizing Enzymes and Tight Junctions at the Blood–Brain Barrier in Epileptic Patients

Pharmaceutics ◽  
2021 ◽  
Vol 13 (12) ◽  
pp. 2122
Author(s):  
Risa Sato ◽  
Kotaro Ohmori ◽  
Mina Umetsu ◽  
Masaki Takao ◽  
Mitsutoshi Tano ◽  
...  
Keyword(s):  
Protein Expression ◽  
Blood Brain Barrier ◽  
Brain Barrier ◽  
Brain Capillaries ◽  
Metabolizing Enzymes ◽  
Expression Levels ◽  
Blood Brain ◽  
Anti Epileptic Drug ◽  
Epileptic Patients ◽  
The Brain

The purpose of the present study was to quantitatively elucidate the levels of protein expression of anti-epileptic-drug (AED) transporters, metabolizing enzymes and tight junction molecules at the blood–brain barrier (BBB) in the focal site of epilepsy patients using accurate SWATH (sequential window acquisition of all theoretical fragment ion spectra) proteomics. Brain capillaries were isolated from focal sites in six epilepsy patients and five normal brains; tryptic digests were produced and subjected to SWATH analysis. MDR1 and BCRP were significantly downregulated in the epilepsy group compared to the normal group. Out of 16 AED-metabolizing enzymes detected, the protein expression levels of GSTP1, GSTO1, CYP2E1, ALDH1A1, ALDH6A1, ALDH7A1, ALDH9A1 and ADH5 were significantly 2.13-, 6.23-, 2.16-, 2.80-, 1.73-, 1.67-, 2.47- and 2.23-fold greater in the brain capillaries of epileptic patients than those of normal brains, respectively. The protein expression levels of Claudin-5, ZO-1, Catenin alpha-1, beta-1 and delta-1 were significantly lower, 1.97-, 2.51-, 2.44-, 1.90- and 1.63-fold, in the brain capillaries of epileptic patients compared to those of normal brains, respectively. Consistent with these observations, leakage of blood proteins was also observed. These results provide for a better understanding of the therapeutic effect of AEDs and molecular mechanisms of AED resistance in epileptic patients.

scholarly journals EXTH-21. DIFFERENTIAL PROTEIN EXPRESSION LEVELS OF ABC AND SOLUTE CARRIER TRANSPORTERS AT THE BLOOD-BRAIN BARRIER OF HUMAN BRAIN AND GLIOBLASTOMA

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi86-vi86
Author(s):  
Xun Bao ◽  
Jianmei Wu ◽  
Youming Xie ◽  
Seongho Kim ◽  
Sharon Michelhaugh ◽  
...  
Keyword(s):  
Protein Expression ◽  
Human Brain ◽  
Blood Brain Barrier ◽  
Mouse Brain ◽  
Brain Barrier ◽  
Normal Brain ◽  
Drug Penetration ◽  
Expression Levels ◽  
Transporter Protein ◽  
Blood Brain

Abstract BACKGROUND Mechanistic understanding and quantitative prediction of drug penetration across the human blood-brain barrier (BBB) is critical to rational drug development and treatment for brain cancer especially glioblastoma. However, prediction of drug brain/tumor penetration has been significantly hindered mainly due to the lack of quantitation data on transporter protein expression levels at the human BBB. This study was to determine protein expression levels of major transporters and markers at the BBB of human brain and glioblastoma. METHOD The absolute protein expression levels of major transporters and markers were determined in isolated microvessels of human brain (N=30), glioblastoma (N=47), rat (N=10) and mouse brain (N=10), using liquid chromatography with tandem mass spectrometry (LC-MS/MS) based targeted proteomics. RESULTS In isolated microvessels of 30 human brain specimens, the median protein abundances for ABCB1, ABCG2, GLUT1, GLUT3, LAT1, MCT1, Na/K ATPase, and Claudin-5 were 3.38, 6.21, 54.51, 7.17, 3.42, 5.71, 32.14, and 1.15 fmol/µg protein, respectively. In glioblastoma microvessels, ABCB1, ABCG2, MCT1, GLUT1, Na/K ATPase, and Claudin-5 protein levels were significantly reduced, while LAT1 was increased and GLU1 remained the same. ABCC4, OATP1A2, OATP2B1, and OAT3 were undetectable in isolated microvessels of both human brain and glioblastoma. There was species difference in transporter protein expression levels in isolated microvessels of human, rat and mouse brain. Specifically, rodent BBB expressed significantly higher ABCB1 but similar ABCG2, as compared to human BBB. CONCLUSION The physical and biochemical barriers of the BBB in glioblastomas are largely disrupted, as indicated by the loss or significant reduction in protein expression of the tight junction marker (claudin-5), brain endothelial cell marker (GLUT1), and major efflux transporters (ABCB1 and ABCG2) as compared to normal human BBB. Differential BBB transporter protein expression levels provides mechanistic and quantitative basis for the prediction of heterogeneous drug penetration into human normal brain and glioblastoma.

Changes in the glucose transporter of brain capillaries

1992 ◽  
Vol 70 (S1) ◽  
pp. S113-S117 ◽  
Author(s):  
Sami I. Harik
Keyword(s):  
Alzheimer Disease ◽  
Blood Brain Barrier ◽  
Glucose Transporter ◽  
Glucose Transporters ◽  
Brain Barrier ◽  
Capillary Endothelium ◽  
Brain Capillaries ◽  
Glut 1 ◽  
Blood Brain ◽  
The Brain

Brain capillary endothelium has a high density of the GLUT-1 facilitative glucose transporter protein. This is reasonable in view of the brain's high metabolic rate for glucose and its isolation behind unique capillaries with blood – brain barrier properties. Thus, the brain endothelium, which constitutes less than 0.1% of the brain weight, has to transport glucose for the much larger mass of surrounding neurons and glia. I describe here the changes that occur in the density of glucose transporters in brain capillaries of subjects with Alzheimer disease, where there is a decreased cerebral metabolic rate for glucose, and in a novel clinical entity characterized by defective glucose transport at the blood – brain barrier. In subjects with Alzheimer disease, cerebral microvessels showed a marked decrease in the density of the glucose transporter when compared with age-matched controls, but there was no change in the density of glucose transporters in erythrocyte membranes. Thus, I believe that the decreased density of glucose transporters in the brains of subjects with Alzheimer disease is the result rather than the cause of the disease. In contradistinction, the primary defect in glucose transport at the blood – brain barrier in subjects with the recently described entity is associated with decreased density of GLUT-1 in erythrocyte membranes.Key words: brain microvessels, capillary endothelium, blood – brain barrier, glucose transporter, Alzheimer disease, hypoglycorrhachia.

scholarly journals Protecting P-glycoprotein at the blood–brain barrier from degradation in an Alzheimer’s disease mouse model

2021 ◽  
Vol 18 (1) ◽  
Author(s):  
Yujie Ding ◽  
Yu Zhong ◽  
Andrea Baldeshwiler ◽  
Erin L. Abner ◽  
Björn Bauer ◽  
...  
Keyword(s):  
Mouse Model ◽  
Protein Expression ◽  
Blood Brain Barrier ◽  
Brain Barrier ◽  
Activity Levels ◽  
Transport Activity ◽  
Brain Capillaries ◽  
P Glycoprotein ◽  
Blood Brain

AbstractBackgroundFailure to clear Aβ from the brain is partly responsible for Aβ brain accumulation in Alzheimer’s disease (AD). A critical protein for clearing Aβ across the blood-brain barrier is the efflux transporter P-glycoprotein (P-gp). In AD, P-gp levels are reduced, which contributes to impaired Aβ brain clearance. However, the mechanism responsible for decreased P-gp levels is poorly understood and there are no strategies available to protect P-gp. We previously demonstrated in isolated brain capillariesex vivothat human Aβ40 (hAβ40) triggers P-gp degradation by activating the ubiquitin-proteasome pathway. In this pathway, hAβ40 initiates P-gp ubiquitination, leading to internalization and proteasomal degradation of P-gp, which then results in decreased P-gp protein expression and transport activity levels. Here, we extend this line of research and present results from anin vivostudy using a transgenic mouse model of AD (human amyloid precursor protein (hAPP)-overexpressing mice; Tg2576).MethodsIn our study, hAPP mice were treated with vehicle, nocodazole (NCZ, microtubule inhibitor to block P-gp internalization), or a combination of NCZ and the P-gp inhibitor cyclosporin A (CSA). We determined P-gp protein expression and transport activity levels in isolated mouse brain capillaries and Aβ levels in plasma and brain tissue.ResultsTreating hAPP mice with 5 mg/kg NCZ for 14 days increased P-gp levels to levels found in WT mice. Consistent with this, P-gp-mediated hAβ42 transport in brain capillaries was increased in NCZ-treated hAPP mice compared to untreated hAPP mice. Importantly, NCZ treatment significantly lowered hAβ40 and hAβ42 brain levels in hAPP mice, whereas hAβ40 and hAβ42 levels in plasma remained unchanged.ConclusionsThese findings provide in vivo evidence that microtubule inhibition maintains P-gp protein expression and transport activity levels, which in turn helps to lower hAβ brain levels in hAPP mice. Thus, protecting P-gp at the blood-brain barrier may provide a novel therapeutic strategy for AD and other Aβ-based pathologies.

scholarly journals Activation of PKC Isoform βI at the Blood–Brain Barrier Rapidly Decreases P-Glycoprotein Activity and Enhances Drug Delivery to the Brain

2010 ◽  
Vol 30 (7) ◽  
pp. 1373-1383 ◽  
Author(s):  
Robert R Rigor ◽  
Brian T Hawkins ◽  
David S Miller
Keyword(s):  
Rat Brain ◽  
Blood Brain Barrier ◽  
Brain Barrier ◽  
Brain Capillaries ◽  
P Glycoprotein ◽  
Blood Brain ◽  
Tnf Α ◽  
The Brain

P-glycoprotein is an ATP (adenosine triphosphate)-driven drug efflux transporter that is highly expressed at the blood–brain barrier (BBB) and is a major obstacle to the pharmacotherapy of central nervous system diseases, including brain tumors, neuro-AIDS, and epilepsy. Previous studies have shown that P-glycoprotein transport activity in rat brain capillaries is rapidly reduced by the proinflammatory cytokine, tumor necrosis factor-α (TNF-α) acting through protein kinase C (PKC)-dependent signaling. In this study, we used isolated rat brain capillaries to show that the TNF-α-induced reduction of P-glycoprotein activity was prevented by a PKCβI/II inhibitor, LY333531, and mimicked by a PKCβI/II activator, 12-deoxyphorbol-13-phenylacetate-20-acetate (dPPA). Western blotting of brain capillary extracts with phospho-specific antibodies showed that dPPA activated PKCβI, but not PKCβII. Moreover, in intact rats, intracarotid infusion of dPPA potently increased brain accumulation of the P-glycoprotein substrate, [3H]-verapamil without compromising tight junction integrity. Thus, PKCβI activation selectively reduced P-glycoprotein activity both in vitro and in vivo. Targeting PKCβI at the BBB may prove to be an effective strategy for enhancing the delivery of small molecule therapeutics to the brain.

scholarly journals Alterations in blood-brain barrier ICAM-1 expression and brain microglial activation after λ-carrageenan-induced inflammatory pain

2006 ◽  
Vol 290 (2) ◽  
pp. H732-H740 ◽  
Author(s):  
J. D. Huber ◽  
C. R. Campos ◽  
K. S. Mark ◽  
T. P. Davis
Keyword(s):  
Protein Expression ◽  
Blood Brain Barrier ◽  
Proinflammatory Cytokines ◽  
Inflammatory Pain ◽  
Microglial Activation ◽  
Brain Barrier ◽  
Cell Counts ◽  
Blood Brain ◽  
Junction Protein ◽  
The Brain

Previous studies showed that peripheral inflammatory pain increased blood-brain barrier (BBB) permeability and altered tight junction protein expression and the delivery of opioid analgesics to the brain. What remains unknown is which pathways and mediators during peripheral inflammation affect BBB function and structure. The current study investigated effects of λ-carrageenan-induced inflammatory pain (CIP) on BBB expression of ICAM-1. We also examined the systemic contribution of a number of proinflammatory cytokines and microglial activation in the brain to elucidate pathways involved in BBB disruption during CIP. We investigated ICAM-1 RNA and protein expression levels in isolated rat brain microvessels after CIP using RT-PCR and Western blot analyses, screened inflammatory cytokines during the time course of inflammation, assessed white blood cell counts, and probed for BBB and central nervous system stimulation and leukocyte transmigration using immunohistochemistry and flow cytometry. Results showed an early increase in ICAM-1 RNA and protein expression after CIP with no change in circulating levels of several proinflammatory cytokines. Changes in ICAM-1 protein expression were noted at 48 h. Immunohistochemistry showed that the induction of ICAM-1 was region specific with increased expression noted in the thalamus and frontal and parietal cortices, which directly correlated with increased expression of activated microglia. The findings of the present study were that CIP induces increased ICAM-1 mRNA and protein expression at the BBB and that systemic proinflammatory mediators play no apparent role in the early response (1–6 h); however, brain region-specific increases in microglial activation suggest a potential for a central-mediated response.

scholarly journals Protecting P-glycoprotein at the Blood-Brain Barrier from Degradation in an Alzheimer’s Disease Mouse Model

2020 ◽  
Author(s):  
Yujie Ding ◽  
Yu Zhong ◽  
Andrea Baldeshwiler ◽  
Erin L. Abner ◽  
Björn Bauer ◽  
...  
Keyword(s):  
Mouse Model ◽  
Protein Expression ◽  
Blood Brain Barrier ◽  
Brain Barrier ◽  
Activity Levels ◽  
Transport Activity ◽  
Brain Capillaries ◽  
P Glycoprotein ◽  
Blood Brain

Abstract Background. Failure to clear Aβ from the brain is partly responsible for Aβ brain accumulation in Alzheimer’s disease (AD). A critical protein for clearing Aβ across the blood-brain barrier is the efflux transporter P-glycoprotein (P-gp). In AD, P-gp levels are reduced, which contributes to impaired Aβ brain clearance. However, the mechanism responsible for decreased P-gp levels is poorly understood and there are no strategies available to protect P-gp. We previously demonstrated in isolated brain capillaries ex vivo that human Aβ40 (hAβ40) triggers P-gp degradation by activating the ubiquitin-proteasome pathway. In this pathway, hAβ40 initiates P-gp ubiquitination, leading to internalization and proteasomal degradation of P-gp, which then results in decreased P-gp protein expression and transport activity levels. Here, we extend this line of research and present results from an in vivo study using a transgenic mouse model of AD (human amyloid precursor protein (hAPP)-overexpressing mice; Tg2576). Methods. In our study, hAPP mice were treated with vehicle, nocodazole (NCZ, microtubule inhibitor to block P-gp internalization), or a combination of NCZ and the P-gp inhibitor cyclosporin A (CSA). We determined P-gp protein expression and transport activity levels in isolated mouse brain capillaries and Aβ levels in plasma and brain tissue. Results. Treating hAPP mice with 5 mg/kg NCZ for 14 days protected P-gp from degradation. Consistent with this, P-gp-mediated hAβ42 transport in brain capillaries was increased in NCZ-treated hAPP mice compared to untreated hAPP mice. Importantly, NCZ treatment significantly lowered hAβ40 and hAβ42 brain levels in hAPP mice, whereas hAβ40 and hAβ42 levels in plasma remained unchanged. Conclusions. These findings provide in vivo proof that blocking P-gp internalization protects P-gp from degradation and maintains P-gp protein expression and transport activity levels, which in turn lowers hAβ brain levels. Thus, protecting P-gp at the blood-brain barrier may provide a novel therapeutic target for AD and other Aβ-based pathologies.

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