Browsing by Subject "Blood-brain barrier"

Now showing 1 - 10 of 12
  • Thumbnail Image
    Item
    4-Ethylguaiacol modulates neuroinflammation and Th1/Th17 differentiation to ameliorate disease severity in experimental autoimmune encephalomyelitis
    (BMC, 2021-05-11) Weng, Wen-Tsan; Kuo, Ping-Chang; Brown, Dennis A.; Scofield, Barbara A.; Furnas, Destin; Paraiso, Hallel C.; Wang, Pei-Yu; Yu, I-Chen; Yen, Jui-Hung; Anatomy and Cell Biology, School of Medicine
    Background: Multiple sclerosis (MS) is a progressive autoimmune disease characterized by the accumulation of pathogenic inflammatory immune cells in the central nervous system (CNS) that subsequently causes focal inflammation, demyelination, axonal injury, and neuronal damage. Experimental autoimmune encephalomyelitis (EAE) is a well-established murine model that mimics the key features of MS. Presently, the dietary consumption of foods rich in phenols has been reported to offer numerous health benefits, including anti-inflammatory activity. One such compound, 4-ethylguaiacol (4-EG), found in various foods, is known to attenuate inflammatory immune responses. However, whether 4-EG exerts anti-inflammatory effects on modulating the CNS inflammatory immune responses remains unknown. Thus, in this study, we assessed the therapeutic effect of 4-EG in EAE using both chronic and relapsing-remitting animal models and investigated the immunomodulatory effects of 4-EG on neuroinflammation and Th1/Th17 differentiation in EAE. Methods: Chronic C57BL/6 EAE and relapsing-remitting SJL/J EAE were induced followed by 4-EG treatment. The effects of 4-EG on disease progression, peripheral Th1/Th17 differentiation, CNS Th1/Th17 infiltration, microglia (MG) activation, and blood-brain barrier (BBB) disruption in EAE were evaluated. In addition, the expression of MMP9, MMP3, HO-1, and Nrf2 was assessed in the CNS of C57BL/6 EAE mice. Results: Our results showed that 4-EG not only ameliorated disease severity in C57BL/6 chronic EAE but also mitigated disease progression in SJL/J relapsing-remitting EAE. Further investigations of the cellular and molecular mechanisms revealed that 4-EG suppressed MG activation, mitigated BBB disruption, repressed MMP3/MMP9 production, and inhibited Th1 and Th17 infiltration in the CNS of EAE. Furthermore, 4-EG suppressed Th1 and Th17 differentiation in the periphery of EAE and in vitro Th1 and Th17 cultures. Finally, we found 4-EG induced HO-1 expression in the CNS of EAE in vivo as well as in MG, BV2 cells, and macrophages in vitro. Conclusions: Our work demonstrates that 4-EG confers protection against autoimmune disease EAE through modulating neuroinflammation and inhibiting Th1 and Th17 differentiation, suggesting 4-EG, a natural compound, could be potentially developed as a therapeutic agent for the treatment of MS/EAE.
  • Thumbnail Image
    Item
  • Thumbnail Image
    Item
    A high-throughput single-cell RNA expression profiling method identifies human pericyte markers
    (Wiley, 2023) Sziraki, Andras; Zhong, Yu; Neltner, Allison M.; Niedowicz, Dana; Rogers, Colin B.; Wilcock, Donna M.; Nehra, Geetika; Neltner, Janna H.; Smith, Rebecca R.; Hartz, Anika M.; Cao, Junyue; Nelson, Peter T.; Neurology, School of Medicine
    Aims: We sought to identify and optimise a universally available histological marker for pericytes in the human brain. Such a marker could be a useful tool for researchers. Further, identifying a gene expressed relatively specifically in human pericytes could provide new insights into the biological functions of this fascinating cell type. Methods: We analysed single-cell RNA expression profiles derived from different human and mouse brain regions using a high-throughput and low-cost single-cell transcriptome sequencing method called EasySci. Through this analysis, we were able to identify specific gene markers for pericytes, some of which had not been previously characterised. We then used commercially (and therefore universally) available antibodies to immunolabel the pericyte-specific gene products in formalin-fixed paraffin-embedded (FFPE) human brains and also performed immunoblots to determine whether appropriately sized proteins were recognised. Results: In the EasySci data sets, highly pericyte-enriched expression was notable for SLC6A12 and SLC19A1. Antibodies against these proteins recognised bands of approximately the correct size in immunoblots of human brain extracts. Following optimisation of the immunohistochemical technique, staining for both antibodies was strongly positive in small blood vessels and was far more effective than a PDGFRB antibody at staining pericyte-like cells in FFPE human brain sections. In an exploratory sample of other human organs (kidney, lung, liver, muscle), immunohistochemistry did not show the same pericyte-like pattern of staining. Conclusions: The SLC6A12 antibody was well suited for labelling pericytes in human FFPE brain sections, based on the combined results of single-cell RNA-seq analyses, immunoblots and immunohistochemical studies.
  • Thumbnail Image
    Item
    Dimethyl itaconate, an itaconate derivative, exhibits immunomodulatory effects on neuroinflammation in experimental autoimmune encephalomyelitis
    (BMC, 2020-04-29) Kuo, Ping-Chang; Weng, Wen-Tsan; Scofield, Barbara A.; Paraiso, Hallel C.; Brown, Dennis A.; Wang, Pei-Yu; Yu, I-Chen; Yen, Jui-Hung; Microbiology and Immunology, School of Medicine
    Background: Inflammatory stimuli induce immunoresponsive gene 1 (IRG1) expression that in turn catalyzes the production of itaconate from the tricarboxylic acid cycle. Itaconate has recently emerged as a regulator of immune cell functions, especially in macrophages. Studies show that itaconate is required for the activation of anti-inflammatory transcription factor Nrf2 by LPS in mouse and human macrophages, and LPS-activated IRG1-/- macrophages that lack endogenous itaconate production exhibit augmented inflammatory responses. Moreover, dimethyl itaconate (DMI), an itaconate derivative, inhibits IL-17-induced IκBς activation in keratinocytes and modulates IL-17-IκBς pathway-mediated skin inflammation in an animal model of psoriasis. Currently, the effect of itaconate on regulating macrophage functions and peripheral inflammatory immune responses is well established. However, its effect on microglia (MG) and CNS inflammatory immune responses remains unexplored. Thus, we investigated whether itaconate possesses an immunomodulatory effect on regulating MG activation and CNS inflammation in animal models of multiple sclerosis, experimental autoimmune encephalomyelitis (EAE). Methods: Chronic C57BL/6 EAE was induced followed by DMI treatment. The effect of DMI on disease severity, blood-brain barrier (BBB) disruption, MG activation, peripheral Th1/Th17 differentiation, and the CNS infiltration of Th1/Th17 cells in EAE was determined. Primary MG was cultured to study the effect of DMI on MG activation. Relapsing-remitting SJL/J EAE was induced to assess the therapeutic effect of DMI. Results: Our results show DMI ameliorated disease severity in the chronic C57BL/6 EAE model. Further analysis of the cellular and molecular mechanisms revealed that DMI mitigated BBB disruption, inhibited MMP3/MMP9 production, suppressed microglia activation, inhibited peripheral Th1/Th17 differentiation, and repressed the CNS infiltration of Th1 and Th17 cells. Strikingly, DMI also exhibited a therapeutic effect on alleviating severity of relapse in the relapsing-remitting SJL/J EAE model. Conclusions: We demonstrate that DMI suppresses neuroinflammation and ameliorates disease severity in EAE through multiple cellular and molecular mechanisms, suggesting that DMI can be developed as a novel therapeutic agent for the
  • Thumbnail Image
    Item
    The Effects of Propofol on a Human in vitro Blood-Brain Barrier Model
    (Frontiers Media, 2022-05-11) Hughes, Jason M.; Neese, Olivia R.; Bieber, Dylan D.; Lewis, Kirsten A.; Ahmadi, Layla M.; Parsons, Dustin W.; Canfield, Scott G.; Anatomy, Cell Biology and Physiology, School of Medicine
    Background: Recently, the safety of repeated and lengthy anesthesia administration has been called into question, a subset of these animal studies demonstrated that anesthetics induced blood-brain barrier (BBB) dysfunction. The BBB is critical in protecting the brain parenchyma from the surrounding micro-vasculature. BBB breakdown and dysfunction has been observed in several neurodegenerative diseases and may contribute to both the initiation and the progression of the disease. In this study we utilize a human induced pluripotent stem cell (iPSC) derived-BBB model, exhibiting near in vivo properties, to evaluate the effects of anesthetics on critical barrier properties. Methods: iPSC-derived brain microvascular endothelial cells (BMECs) expressed near in vivo barrier tightness assessed by trans-endothelial electrical resistance and para-cellular permeability. Efflux transporter activity was determined by substrate transport in the presence of specific inhibitors. Trans-cellular transport was measured utilizing large fluorescently tagged dextran. Tight junction localization in BMECs was evaluated with fluorescent microscopy. The anesthetic, propofol was exposed to BMECs at varying durations and concentrations and BBB properties were monitored post-exposure. Results: Following propofol exposure, BMECs displayed reduced resistance and increased permeability indicative of a leaky barrier. Reduced barrier tightness and the dysregulation of occludin, a tight junction protein, were partly the result of an elevation in matrix metalloproteinase (MMP) levels. Efflux transporter activity and trans-cellular transport were unaffected by propofol exposure. Propofol induced barrier dysfunction was partially restored following matrix metalloproteinase inhibition. Conclusion: For the first time, we have demonstrated that propofol alters BBB integrity utilizing a human in vitro BBB model that displays key in vivo characteristics. A leaky BBB enables otherwise impermeable molecules such as pathogens and toxins the ability to reach vulnerable cell types of the brain parenchyma. A robust human in vitro BBB model will allow for the evaluation of several anesthetics at fluctuating clinical scenarios and to elucidate mechanisms with the goal of ultimately improving anesthesia safety.
  • Thumbnail Image
    Item
    Fluorescein Isothiocyanate (FITC)-Dextran Extravasation as a Measure of Blood-Brain Barrier Permeability
    (Wiley, 2017-04-10) Natarajan, Reka; Northrop, Nicole; Yamamoto, Bryan; Pharmacology and Toxicology, School of Medicine
    The blood-brain barrier (BBB) is formed in part by vascular endothelial cells that constitute the capillaries and microvessels of the brain. The function of this barrier is to maintain homeostasis within the brain microenvironment and buffer the brain from changes in the periphery. A dysfunction of the BBB would permit circulating molecules and pathogens typically restricted to the periphery to enter the brain and interfere with normal brain function. As increased permeability of the BBB is associated with several neuropathologies, it is important to have a reliable and sensitive method that determines BBB permeability and the degree of BBB disruption. A detailed protocol is presented for assessing the integrity of the BBB by transcardial perfusion of a 10,000 Da FITC-labeled dextran molecule and its visualization to determine the degree of extravasation from brain microvessels.
  • Thumbnail Image
    Item
    Gliovascular transcriptional perturbations in Alzheimer's disease reveal molecular mechanisms of blood brain barrier dysfunction
    (Springer Nature, 2024-06-20) İş, Özkan; Wang, Xue; Reddy, Joseph S.; Min, Yuhao; Yilmaz, Elanur; Bhattarai, Prabesh; Patel, Tulsi; Bergman, Jeremiah; Quicksall, Zachary; Heckman, Michael G.; Tutor-New, Frederick Q.; Demirdogen, Birsen Can; White, Launia; Koga, Shunsuke; Krause, Vincent; Inoue, Yasuteru; Kanekiyo, Takahisa; Cosacak, Mehmet Ilyas; Nelson, Nastasia; Lee, Annie J.; Vardarajan, Badri; Mayeux, Richard; Kouri, Naomi; Deniz, Kaancan; Carnwath, Troy; Oatman, Stephanie R.; Lewis-Tuffin, Laura J.; Nguyen, Thuy; Alzheimer’s Disease Neuroimaging Initiative; Carrasquillo, Minerva M.; Graff-Radford, Jonathan; Petersen, Ronald C.; Jack, Clifford R., Jr.; Kantarci, Kejal; Murray, Melissa E.; Nho, Kwangsik; Saykin, Andrew J.; Dickson, Dennis W.; Kizil, Caghan; Allen, Mariet; Ertekin-Taner, Nilüfer; Radiology and Imaging Sciences, School of Medicine
    To uncover molecular changes underlying blood-brain-barrier dysfunction in Alzheimer’s disease, we performed single nucleus RNA sequencing in 24 Alzheimer’s disease and control brains and focused on vascular and astrocyte clusters as main cell types of blood-brain-barrier gliovascular-unit. The majority of the vascular transcriptional changes were in pericytes. Of the vascular molecular targets predicted to interact with astrocytic ligands, SMAD3, upregulated in Alzheimer’s disease pericytes, has the highest number of ligands including VEGFA, downregulated in Alzheimer’s disease astrocytes. We validated these findings with external datasets comprising 4,730 pericyte and 150,664 astrocyte nuclei. Blood SMAD3 levels are associated with Alzheimer’s disease-related neuroimaging outcomes. We determined inverse relationships between pericytic SMAD3 and astrocytic VEGFA in human iPSC and zebrafish models. Here, we detect vast transcriptome changes in Alzheimer’s disease at the gliovascular-unit, prioritize perturbed pericytic SMAD3-astrocytic VEGFA interactions, and validate these in cross-species models to provide a molecular mechanism of blood-brain-barrier disintegrity in Alzheimer’s disease.
  • Thumbnail Image
    Item
    Imaging of intranasal drug delivery to the brain
    (e-Century Publishing, 2020-02-25) Veronesi, Michael C.; Alhamami, Mosa; Miedema, Shelby B.; Yun, Yeonhee; Ruiz-Cardozo, Miguel; Vannier, Michael W.; Radiology and Imaging Sciences, School of Medicine
    Intranasal (IN) delivery is a rapidly developing area for therapies with great potential for the treatment of central nervous system (CNS) diseases. Moreover, in vivo imaging is becoming an important part of therapy assessment, both clinically in humans and translationally in animals. IN drug delivery is an alternative to systemic administration that uses the direct anatomic pathway between the olfactory/trigeminal neuroepithelium of the nasal mucosa and the brain. Several drugs have already been approved for IN application, while others are undergoing development and testing. To better understand which imaging modalities are being used to assess IN delivery of therapeutics, we performed a literature search with the key words “Intranasal delivery” and “Imaging” and summarized these findings in the current review. While this review does not attempt to be fully comprehensive, we intend for the examples provided to allow a well-rounded picture of the imaging tools available to assess IN delivery, with an emphasis on the nose-to-brain delivery route. Examples of in vivo imaging, for both humans and animals, include magnetic resonance imaging (MRI), positron emission tomography (PET), single-photon emission computed tomography (SPECT), gamma scintigraphy and computed tomography (CT). Additionally, some in vivo optical imaging modalities, including bioluminescence and fluorescence, have been used more in experimental testing in animals. In this review, we introduce each imaging modality, how it is being utilized and outline its strengths and weaknesses, specifically in the context of IN delivery of therapeutics to the brain.
  • Thumbnail Image
    Item
    Isolation of Mouse Cerebral Microvasculature for Molecular and Single-Cell Analysis
    (Frontiers Media, 2020) Paraiso, Hallel C.; Wang, Xueqian; Kuo, Ping-Chang; Furnas, Destin; Scofield, Barbara A.; Chang, Fen-Lei; Yen, Jui-Hung; Yu, I-Chen; Anatomy and Cell Biology, School of Medicine
    Brain microvasculature forms a specialized structure, the blood-brain barrier (BBB), to maintain homeostasis and integrity of the central nervous system (CNS). The BBB dysfunction is emerging as a critical contributor to multiple neurological disorders, including stroke, traumatic brain injury, autoimmune multiple sclerosis, and neurodegenerative diseases. The brain microvasculature exhibits highly cellular and regional heterogeneity to accommodate dynamic changes of microenvironment during homeostasis and diseases. Thus, investigating the underlying mechanisms that contribute to molecular or cellular changes of the BBB is a significant challenge. Here, we describe an optimized protocol to purify microvessels from the mouse cerebral cortex using mechanical homogenization and density-gradient centrifugation, while maintaining the structural integrity and functional activity of the BBB. We show that the isolated microvessel fragments consist of BBB cell populations, including endothelial cells, astrocyte end-feet, pericytes, as well as tight junction proteins that seal endothelial cells. Furthermore, we describe the procedures to generate single-cell suspensions from isolated microvessel fragments. We demonstrate that cells in the single-cell suspensions are highly viable and suitable for single-cell RNA-sequencing analysis. This protocol does not require transgenic mice and cell sorting equipment to isolate fluorescence-labeled endothelial cells. The optimized procedures can be applied to different disease models to generate viable cells for single-cell analysis to uncover transcriptional or epigenetic landscapes of BBB component cells.
  • Thumbnail Image
    Item
    Peripheral ammonia and blood brain barrier structure and function after methamphetamine
    (Elsevier, 2016-08) Northrop, Nicole A.; Halpin, Laura E.; Yamamoto, Bryan K.; Department of Pharmacology and Toxicology, IU School of Medicine
    An effect of the widely abuse psychostimulant, methamphetamine (Meth), is blood-brain-barrier (BBB) disruption; however, the mechanism by which Meth causes BBB disruption remains unclear. Recently it has been shown that Meth produces liver damage and consequent increases in plasma ammonia. Ammonia can mediate oxidative stress and inflammation, both of which are known to cause BBB disruption. Therefore, the current studies examined the role of peripheral ammonia in Meth-induced disruption of BBB structure and function. A neurotoxic Meth regimen (10 mg/kg, ip, q 2 h, ×4) administered to rats increased plasma ammonia and active MMP-9 in the cortex 2 h after the last Meth injection, compared to saline treated rats. At 24 h after Meth treatment, decreased immunoreactivity of BBB structural proteins, occludin and claudin-5, and increased extravasation of 10,000 Da FITC-dextran were observed, as compared to saline controls. Pretreatment with lactulose (5.3 g/kg, po, q 12 h), a drug that remains in the lumen of the intestine and promotes ammonia excretion, prevented the Meth-induced increases in plasma ammonia. These results were paralleled by the prevention of decreases in BBB structural proteins, increases in extravasation of 10,000 Da FITC-dextran and increases in active MMP-9. The results indicate that Meth-induced increases in ammonia produce BBB disruption and suggest that MMP-9 activation mediates the BBB disruption. These findings identify a novel mechanism of Meth-induced BBB disruption that is mediated by plasma ammonia and are the first to identify a peripheral contribution to Meth-induced BBB disruption.