Quantitative targeted absolute proteomics of human blood–brain barrier transporters and receptors

Uchida, Yasuo; Ohtsuki, Sumio; Katsukura, Yuki; Ikeda, Chiemi; Suzuki, Takashi; Kamiie, Junichi; Terasaki, Tetsuya

doi:10.1111/j.1471-4159.2011.07208.x

Cited by 742 publications

(908 citation statements)

References 59 publications

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“…Undetectable or low level expression of ABCC1 associated with cerebral blood vessels in humans without brain disease, 28,29 and the above referenced reports of multiple CNS cell types expressing ABCC1 in humans with brain disease, are consistent with our immunohistochemical data (Fig. 2).…”

Section: Discussionsupporting

confidence: 91%

Expression of ATP-Binding Cassette Transporters B1 and C1 after Severe Traumatic Brain Injury in Humans

Willyerd

Empey

Philbrick

et al. 2016

Journal of Neurotrauma

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Adenosine triphosphate-binding cassette (ABC) transport proteins ABCC1 and ABCB1 (also known as multidrug resistance-associated protein 1 and p-glycoprotein, respectively), are key membrane efflux transporters of drugs and endogenous substrates, including in the brain. The impact of traumatic brain injury (TBI) on ABCC1 and ABCB1 expression in humans is unknown. We hypothesized that ABCC1 and ABCB1 expression would be altered in brain tissue from patients acutely after severe TBI. Archived TBI samples (n = 10) from our Brain Trauma Research Center and control samples (n = 7) from our Alzheimer Disease Research Center were obtained under Institutional Review Board approval. Protein was extracted from fresh frozen cortical brain tissue for Western blot analysis and sections were obtained from fixed cortical tissue for immunohistochemistry. Relative abundance of ABCC1 was increased in samples from TBI versus controls (2.8 -2.5 fold; p = 0.005). ABCC1 immunohistochemistry was consistent with Western blot data, with increased immunoreactivity in cerebral blood vessel walls, as well as cells with the morphological appearance of neurons and glia in TBI versus controls. Relative abundance of ABCB1 was similar between TBI and controls ( p = 0.76), and ABCB1 immunoreactivity was primarily associated with cerebral blood vessels in both groups. These human data show that TBI increases ABCC1 expression in the brain, consistent with possible implications for both patients receiving pharmacological inhibitors and/or substrates of ABCC1 after TBI.

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Section: Discussionsupporting

confidence: 91%

Expression of ATP-Binding Cassette Transporters B1 and C1 after Severe Traumatic Brain Injury in Humans

Willyerd

Empey

Philbrick

et al. 2016

Journal of Neurotrauma

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“…In SIL methods such as the absolute quantification (AQUA) method (30), the IS is commonly spiked during or postdigestion and is the most feasible and rapid approach. Indeed, SIL peptides are among the most common IS used to evaluate the expression of drug transporters (5)(6)(7)18,19,(46)(47)(48)(49)(50)(51). A comparison of the SIL peptide with SILAM approach was recently carried out in the LC-MS/ MS method development in our laboratory (37).…”

Section: Selection Of Internal Standard (Is)mentioning

confidence: 99%

Quantitative Targeted Proteomics for Membrane Transporter Proteins: Method and Application

Qiu

Zhang

Lai

2014

AAPS J

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Abstract. Although global proteomics has shown promise for discovery of many new proteins, biomarkers, protein modifications, and polymorphisms, targeted proteomics is emerging in the proteomics research field as a complement to untargeted shotgun proteomics, particularly when a determined set of low-abundance functional proteins need to be measured. The function and expression of proteins related to drug absorption, distribution, metabolism, and excretion (ADME) such as cytochrome P450 enzymes and membrane transporters are of great interest in biopharmaceutical research. Since the variation in ADME-related protein expression is known to be a major complicating factor encountered during in vitro-in vivo and in vivo-in vivo extrapolations (IVIVE), the accurate quantification of the ADME proteins in complex biological systems becomes a fundamental element in establishing IVIVE for pharmacokinetic predictions. In this review, we provide an overview of relevant methodologies followed by a summary of recent applications encompassing mass spectrometry-based targeted quantifications of membrane transporters.KEY WORDS: drug absorption, distribution, metabolism, and excretion (ADME); drug transporters; in vitro-in vivo and in vivo-in vivo extrapolations (IVIVE); LC-MS/MS; quantitative targeted proteomics.

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“…[9][10][11] To determine how BBB functions are modulated by obesity, we performed comparative proteomic studies to determine the…”

mentioning

confidence: 99%

Diet-Induced Obesity Suppresses Expression of Many Proteins at the Blood–Brain Barrier

Ouyang

Hsuchou

Kastin

et al. 2013

J Cereb Blood Flow Metab

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The blood-brain barrier (BBB) is a regulatory interface between the central nervous system and the rest of the body. However, BBB changes in obesity and metabolic syndrome have not been fully elucidated. We hypothesized that obesity reduces energy metabolism in the cerebral microvessels composing the BBB, reflected by downregulation of protein expression and function. We performed comparative proteomic analyses in enriched microvessels from the cerebral cortex of mice 2 months after ingestion of a high-fat diet or regular rodent chow. In mice with diet-induced obesity (DIO), there was downregulation of 47 proteins in the cerebral microvessels, including cytoskeletal proteins, chaperons, enzymes, transport-related proteins, and regulators for transcriptional and translational activities. Only two proteins, involved in messenger RNA (mRNA) transport and processing, were upregulated. The changes of these proteins were further validated by quantitative polymerase chain reaction (qPCR), western blotting, and immunofluorescent staining of freshly isolated microvessels, in samples obtained from different batches of mice. The predominant downregulation suggests that DIO suppresses metabolic activity of BBB microvessels. The finding of a hypometabolic state of the BBB in mice at the chronic stage of DIO is unexpected and unprecedented; it may provide novel mechanistic insight into how obesity influences CNS function via regulatory changes of the BBB. Keywords: blood-brain barrier; cerebral microvessels; high-fat diet; mass spectrometry; obesity; proteomics INTRODUCTION Obesity has become one of the most critical problems in human health and is associated with many complications of metabolic diseases. 1,2 Proteomic methods have been used to investigate human and mouse obesity, but most reports have focused on changes in adipose tissue. [3][4][5][6] Recent results suggest that the blood-brain barrier (BBB) is activated in obesity; for example, mice with adult-onset obesity show resistance to leptin transport across the BBB, region-specific astrogliosis, and upregulation of astrocytic leptin receptors. 7,8 The BBB is a regulatory interface between the central nervous system (CNS) and the peripheral circulation. It is a complex threedimensional structure composed of specialized endothelial cells that are reinforced by pericytes, astrocytic endfeet, and extracellular matrix. The tight junctions between the endothelia and the continuous basement membrane prevent diffusion of large non-lipophilic molecules between the CNS parenchyma (brain and spinal cord) and the circulation. The function of the BBB is further reinforced by a high mitochondrial content that ensures sufficient energy, an abundance of enzymes, and metabolically active astrocytes. The functions of the BBB in obesity, however, remain unclear. We predicted that BBB homeostasis is disturbed by metabolic changes in obesity.Although normal transport and secretory functions of the BBB are essential for CNS function, proteomic analyses have shown that BBB activity is ch...

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Quantitative targeted absolute proteomics of human blood–brain barrier transporters and receptors

Cited by 742 publications

References 59 publications

Expression of ATP-Binding Cassette Transporters B1 and C1 after Severe Traumatic Brain Injury in Humans

Expression of ATP-Binding Cassette Transporters B1 and C1 after Severe Traumatic Brain Injury in Humans

Quantitative Targeted Proteomics for Membrane Transporter Proteins: Method and Application

Diet-Induced Obesity Suppresses Expression of Many Proteins at the Blood–Brain Barrier

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