Abstract W P359: Activation of Prostacyclin Receptor (IP) in Human Brain Microvascular Endothelial Cells Stimulates Production of SAPPa

Stroke ◽  
2014 ◽  
Vol 45 (suppl_1) ◽  
Author(s):  
Tongrong He ◽  
Tong Lu ◽  
Zvinomir Katusic
Keyword(s):  
Endothelial Cells ◽  
Protein Expression ◽  
Human Brain ◽  
Microvascular Endothelial Cells ◽  
Mrna Levels ◽  
Brain Microvascular Endothelial Cells ◽  
Camp Pathway ◽  
Processing Enzyme ◽  
Amyloidogenic Processing ◽  
Adam10 Expression

Previous studies have established that cerebral blood vessels possess high capacity to synthesize prostacyclin (PGI 2 ). Substantial evidence indicates that amyloid precursor protein (APP) and soluble APPα (sAPPα) have neurotrophic and nueroprotective properties. We hypothesized that activation of IP-cAMP pathway stimulates non-amyloidogenic processing of APP in cerebral endothelial cells. Western blotting was used to detect protein expression; real time RT-PCR was employed to measure mRNA levels of APP, and a disintegrin and metalloprotease 10 (ADAM10, an α processing enzyme); sAPPα secretion was detected in condition medium derived from culturing human brain microvascular endothelial cells (BMVECs), by ELISA. After human BMVECs were treated with PGI 2 analog iloprost (0.01, 0.05, 0.1, or 0.5 μM) for 3 days, protein levels of ADAM10 (n=7-9, P<0.05) and APP (n=6, P<0.05) were significantly increased in a concentration dependent manner, while β amyloidogenic processing enzyme BACE1, and other α processing enzymes such as ADAM9 and ADAM17 were not changed. mRNA levels of APP (n=7, P<0.05) and ADAM10 (n=4, P<0.05), as well as sAPPα production in human BMVECs (n=6, P<0.05) were also significantly augmented in response to iloprost treatment. Moreover, stimulation of human BMVECs with forskolin (40μM) significantly enhanced APP and ADAM10 protein expression. Most notably, iloprost significantly augmented protein expression of peroxisome proliferation-activated receptor-δ (PPARδ) and SIRT1. We further explored the role of PPARδ in APP processing. ADAM10 expression (n=6, P<0.05), but not APP and BACE1, was significantly increased in human BMVECs treated with PPARδ ligand GW501516 (30 or 100 nM). GW501516 (100 nM) also significantly increase sAPPα production (n=9-10, P<0.05). siRNA against PPARδ significantly suppressed basal levels of ADAM10, and blocked iloprost-induced ADAM10 expression (n=4, P<0.05). These findings suggest that activation of IP-cAMP pathway enhances non-amyloidogenic processing of APP and consequently increases production of neurotrophic molecule sAPPα in human BMVECs. Activation of PPARδ in part mediates iloprost-induced α processing of APP.

scholarly journals Neuregulin-1 Signaling in Brain Endothelial Cells

2008 ◽  
Vol 29 (1) ◽  
pp. 39-43 ◽  
Author(s):  
Josephine Lok ◽  
S Pablo Sardi ◽  
Shuzhen Guo ◽  
Elaine Besancon ◽  
Duy M Ha ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Cell Death ◽  
Protein Expression ◽  
Human Brain ◽  
Microvascular Endothelial Cells ◽  
Brain Microvascular Endothelial Cells ◽  
Brain Endothelial Cells ◽  
Neuregulin 1 ◽  
Multiple Functions ◽  
Mrna And Protein Expression

Neuregulin-1 (NRG1) signaling has multiple functions in neurons and glia. The data in this study show that NRG1 may also possess significant signaling and cytoprotective properties in human brain microvascular endothelial cells (BMECs). Neuregulin-1 mRNA and protein expression are present in these cells, and NRG1 receptors erbB2 and erbB3 are phosphorylated in response to NRG1. Neuregulin-1 triggers clear biologic responses in BMECs—elevated phospho-Akt levels, increased ring formation in a Matrigel assay, and decreased cell death after oxidative injury with H2O2. These data suggest that NRG1 signaling is functional and cytoprotective in BMECs.

scholarly journals Effects of ompA Deletion on Expression of Type 1 Fimbriae in Escherichia coli K1 Strain RS218 and on the Association of E. coli with Human Brain Microvascular Endothelial Cells

2006 ◽  
Vol 74 (10) ◽  
pp. 5609-5616 ◽  
Author(s):  
Ching-Hao Teng ◽  
Yi Xie ◽  
Sooan Shin ◽  
Francescopaolo Di Cello ◽  
Maneesh Paul-Satyaseela ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Endothelial Cells ◽  
Human Brain ◽  
Microvascular Endothelial Cells ◽  
Mrna Levels ◽  
Brain Microvascular Endothelial Cells ◽  
E Coli ◽  
Type 1 Fimbriae ◽  
Microvascular Endothelial

ABSTRACT We have previously shown that outer membrane protein A (OmpA) and type 1 fimbriae are the bacterial determinants involved in Escherichia coli K1 binding to human brain microvascular endothelial cells (HBMEC), which constitute the blood-brain barrier. In investigating the role of OmpA in E. coli K1 binding to HBMEC, we showed for the first time that ompA deletion decreased the expression of type 1 fimbriae in E. coli K1. Decreased expression of type 1 fimbriae in the ompA deletion mutant was largely the result of driving the fim promoter toward the type 1 fimbrial phase-OFF orientation. mRNA levels of fimB and fimE were found to be decreased with the OmpA mutant compared to the parent strain. Of interest, the ompA deletion further decreased the abilities of E. coli K1 to bind to and invade HBMEC under the conditions of fixing type 1 fimbria expression in the phase-ON or phase-OFF status. These findings suggest that the decreased ability of the OmpA mutant to interact with HBMEC is not entirely due to its decreased type 1 fimbrial expression and that OmpA and type 1 fimbriae facilitate the interaction of E. coli K1 with HBMEC at least in an additive manner.

scholarly journals Early gene response of human brain microvascular endothelial cells toListeria monocytogenesinfection

2011 ◽  
Vol 57 (5) ◽  
pp. 441-446 ◽  
Author(s):  
Chinling Wang ◽  
Chung-Hsi Chou ◽  
Charles Tseng ◽  
Xijin Ge ◽  
Lesya M. Pinchuk
Keyword(s):  
Endothelial Cells ◽  
Human Brain ◽  
Early Gene ◽  
Microvascular Endothelial Cells ◽  
Mrna Levels ◽  
Brain Microvascular Endothelial Cells ◽  
Reverse Transcription Pcr ◽  
Infected Cells ◽  
Microvascular Endothelial ◽  
Upregulated Genes

The gene expression of human brain microvascular endothelial cells (HBMEC) in response to 4 h of infection by Listeria monocytogenes was analyzed. Four hours after infection, the expression of 456 genes of HBMEC had changed (p < 0.05). We noted that many active genes were involved in the formyl-methionyl-leucyl-phenylalanine pathway in infected HBMEC. In the upregulated genes, mRNA levels of interleukin-8 and interleukin-15 in infected cells increased according to microarray and real-time reverse transcription – PCR analyses. Since both cytokines are regarded as potent chemotactic factors, the results suggest that HBMEC are capable of recruiting cells of innate and adaptive immune responses during early L. monocytogenes infection.

scholarly journals Cronobacter sakazakii ATCC 29544 Translocated Human Brain Microvascular Endothelial Cells via Endocytosis, Apoptosis Induction, and Disruption of Tight Junction

2021 ◽  
Vol 12 ◽  
Author(s):  
Tong Jin ◽  
Ning Guan ◽  
Yuhang Du ◽  
Xinpeng Zhang ◽  
Jiahui Li ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Tight Junction ◽  
Human Brain ◽  
Microvascular Endothelial Cells ◽  
No Production ◽  
Mrna Levels ◽  
Brain Microvascular Endothelial Cells ◽  
Cronobacter Sakazakii ◽  
Microvascular Endothelial ◽  
Related Proteins

Cronobacter sakazakii (C. sakazakii) is an emerging opportunistic foodborne pathogen that can cause neonatal necrotizing enterocolitis, meningitis, sepsis in neonates and infants with a relatively high mortality rate. Bacterial transcytosis across the human brain microvascular endothelial cells (HBMEC) is vital for C. sakazakii to induce neonatal meningitis. However, few studies focus on the mechanisms by which C. sakazakii translocates HBMEC. In this study, the translocation processes of C. sakazakii on HBMEC were explored. C. sakazakii strains could effectively adhere to, invade and intracellularly survive in HBMEC. The strain ATCC 29544 exhibited the highest translocation efficiency across HBMEC monolayer among four tested strains. Bacteria-contained intracellular endosomes were detected in C. sakazakii-infected HBMEC by a transmission electron microscope. Endocytosis-related proteins CD44, Rab5, Rab7, and LAMP2 were increased after infection, while the level of Cathepsin L did not change. C. sakazakii induced TLR4/NF-κB inflammatory signal pathway activation in HBMEC, with increased NO production and elevated mRNA levels of IL-8, IL-6, TNF-α, IL-1β, iNOS, and COX-2. C. sakazakii infection also caused LDH release, caspase-3 activation, and HBMEC apoptosis. Meanwhile, increased Dextran-FITC permeability and decreased trans epithelial electric resistance indicated that C. sakazakii disrupted tight junction of HBMEC monolayers, which was confirmed by the decreased levels of tight junction-related proteins ZO-1 and Occludin. These findings suggest that C. sakazakii induced intracellular bacterial endocytosis, stimulated inflammation and apoptosis, disrupted monolayer tight junction in HBMEC, which all together contribute to bacterial translocation.

scholarly journals Ureaplasma Species Modulate Cytokine and Chemokine Responses in Human Brain Microvascular Endothelial Cells

2019 ◽  
Vol 20 (14) ◽  
pp. 3583 ◽  
Author(s):  
Christine Silwedel ◽  
Christian P. Speer ◽  
Axel Haarmann ◽  
Markus Fehrholz ◽  
Heike Claus ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Human Brain ◽  
Ureaplasma Urealyticum ◽  
Microvascular Endothelial Cells ◽  
Mrna Levels ◽  
Brain Microvascular Endothelial Cells ◽  
Chemokine Ligand ◽  
Microvascular Endothelial ◽  
Ureaplasma Spp

Ureaplasma species are common colonizers of the adult genitourinary tract and often considered as low-virulence commensals. Intraamniotic Ureaplasma infections, however, facilitate chorioamnionitis and preterm birth, and cases of Ureaplasma-induced neonatal sepsis, pneumonia, and meningitis raise a growing awareness of their clinical relevance. In vitro studies are scarce but demonstrate distinct Ureaplasma-driven impacts on immune mechanisms. The current study addressed cytokine and chemokine responses upon exposure of native or lipopolysaccharide (LPS) co-stimulated human brain microvascular endothelial cells (HBMEC) to Ureaplasma urealyticum or U. parvum, using qRT-PCR, RNA sequencing, multi-analyte immunoassay, and flow cytometry. Ureaplasma exposure in native HBMEC reduced monocyte chemoattractant protein (MCP)-3 mRNA expression (p < 0.01, vs. broth). In co-stimulated HBMEC, Ureaplasma spp. attenuated LPS-evoked mRNA responses for C-X-C chemokine ligand 5, MCP-1, and MCP-3 (p < 0.05, vs. LPS) and mitigated LPS-driven interleukin (IL)-1α protein secretion, as well as IL-8 mRNA and protein responses (p < 0.05). Furthermore, Ureaplasma isolates increased C-X-C chemokine receptor 4 mRNA levels in native and LPS co-stimulated HBMEC (p < 0.05). The presented results may imply immunomodulatory capacities of Ureaplasma spp. which may ultimately promote chronic colonization and long-term neuroinflammation.

scholarly journals Ubiquinone Metabolism and Transcription HIF-1 Targets Pathway Are Toxicity Signature Pathways Present in Extracellular Vesicles of Paraquat-Exposed Human Brain Microvascular Endothelial Cells

2021 ◽  
Vol 22 (10) ◽  
pp. 5065
Author(s):  
Tatjana Vujić ◽  
Domitille Schvartz ◽  
Anton Iliuk ◽  
Jean-Charles Sanchez
Keyword(s):  
Endothelial Cells ◽  
Human Brain ◽  
Extracellular Vesicles ◽  
Physiological State ◽  
Microvascular Endothelial Cells ◽  
Brain Microvascular Endothelial Cells ◽  
Cell Of Origin ◽  
Functional Changes ◽  
Data Independent Acquisition ◽  
Microvascular Endothelial

Over the last decade, the knowledge in extracellular vesicles (EVs) biogenesis and modulation has increasingly grown. As their content reflects the physiological state of their donor cells, these “intercellular messengers” progressively became a potential source of biomarker reflecting the host cell state. However, little is known about EVs released from the human brain microvascular endothelial cells (HBMECs). The current study aimed to isolate and characterize EVs from HBMECs and to analyze their EVs proteome modulation after paraquat (PQ) stimulation, a widely used herbicide known for its neurotoxic effect. Size distribution, concentration and presence of well-known EV markers were assessed. Identification and quantification of PQ-exposed EV proteins was conducted by data-independent acquisition mass spectrometry (DIA-MS). Signature pathways of PQ-treated EVs were analyzed by gene ontology terms and pathway enrichment. Results highlighted that EVs exposed to PQ have modulated pathways, namely the ubiquinone metabolism and the transcription HIF-1 targets. These pathways may be potential molecular signatures of the PQ-induced toxicity carried by EVs that are reflecting their cell of origin by transporting with them irreversible functional changes.

scholarly journals Human Brain Microvascular Endothelial Cells and Umbilical Vein Endothelial Cells Differentially Facilitate Leukocyte Recruitment and Utilize Chemokines for T Cell Migration

2008 ◽  
Vol 2008 ◽  
pp. 1-8 ◽  
Author(s):  
Shumei Man ◽  
Eroboghene E. Ubogu ◽  
Katherine A. Williams ◽  
Barbara Tucky ◽  
Melissa K. Callahan ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Cell Migration ◽  
T Cell ◽  
Human Brain ◽  
Umbilical Vein ◽  
Microvascular Endothelial Cells ◽  
Brain Microvascular Endothelial Cells ◽  
T Cell Migration ◽  
Umbilical Vein Endothelial Cells

Endothelial cells that functionally express blood brain barrier (BBB) properties are useful surrogates for studying leukocyte-endothelial cell interactions at the BBB. In this study, we compared two different endothelial cellular models: transfected human brain microvascular endothelial cells (THBMECs) and human umbilical vein endothelial cells (HUVECs). With each grow under optimal conditions, confluent THBMEC cultures showed continuous occludin and ZO-1 immunoreactivity, while HUVEC cultures exhibited punctate ZO-1 expression at sites of cell-cell contact only. Confluent THBMEC cultures on 24-well collagen-coated transwell inserts had significantly higher transendothelial electrical resistance (TEER) and lower solute permeability than HUVECs. Confluent THBMECs were more restrictive for mononuclear cell migration than HUVECs. Only THBMECs utilized abluminal CCL5 to facilitate T-lymphocyte migration in vitro although both THBMECs and HUVECs employed CCL3 to facilitate T cell migration. These data establish baseline conditions for using THBMECs to develop in vitro BBB models for studying leukocyte-endothelial interactions during neuroinflammation.

Expression of Zinc-Finger Antiviral Protein in hCMEC/D3 Human Cerebral Microvascular Endothelial Cells: Effect of a Toll-Like Receptor 3 Agonist

2021 ◽  
pp. 1-10
Author(s):  
Mako Okudera ◽  
Minami Odawara ◽  
Masashi Arakawa ◽  
Shogo Kawaguchi ◽  
Kazuhiko Seya ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Protein Expression ◽  
Zinc Finger ◽  
Microvascular Endothelial Cells ◽  
Brain Microvascular Endothelial Cells ◽  
Toll Like Receptor ◽  
Antiviral Protein ◽  
Polycytidylic Acid ◽  
Microvascular Endothelial ◽  
The Brain

<b><i>Introduction:</i></b> Invasion of viruses into the brain causes viral encephalitis, which can be fatal and causes permanent brain damage. The blood-brain barrier (BBB) protects the brain by excluding harmful substances and microbes. Brain microvascular endothelial cells are important components of the BBB; however, the mechanisms of antiviral reactions in these cells have not been fully elucidated. Zinc-finger antiviral protein (ZAP) is a molecule that restricts the infection of various viruses, and there are 2 major isoforms: ZAPL and ZAPS. Toll-like receptor 3 (TLR3), a pattern-recognition receptor against viral double-stranded RNA, is implicated in antiviral innate immune reactions. The aim of this study was to investigate the expression of ZAP in cultured hCMEC/D3 human brain microvascular endothelial cells treated with an authentic TLR3 agonist polyinosinic-polycytidylic acid (poly IC). <b><i>Methods:</i></b> hCMEC/D3 cells were cultured and treated with poly IC. Expression of ZAPL and ZAPS mRNA was investigated using quantitative reverse transcription-polymerase chain reaction, and protein expression of these molecules was examined using western blotting. The role of nuclear factor-κB (NF-κB) was examined using the NF-κB inhibitor, SN50. The roles of interferon (IFN)-β, IFN regulatory factor 3 (IRF3), tripartite motif protein 25 (TRIM25), and retinoic acid-inducible gene-I (RIG-I) in poly IC-induced ZAPS expression were examined using RNA interference. Propagation of Japanese encephalitis virus (JEV) was examined using a focus-forming assay. <b><i>Results:</i></b> ZAPS mRNA and protein expression was upregulated by poly IC, whereas the change of ZAPL mRNA and protein levels was minimal. Knockdown of IRF3 or TRIM25 decreased the poly IC-induced upregulation of ZAPS, whereas knockdown of IFN-β or RIG-I did not affect ZAPS upregulation. SN50 did not affect ZAPS expression. Knockdown of ZAP enhanced JEV propagation. <b><i>Conclusion:</i></b> ZAPL and ZAPS were expressed in hCMEC/D3 cells, and ZAPS expression was upregulated by poly IC. IRF3 and TRIM25 are involved in poly IC-induced upregulation of ZAPS. ZAP may contribute to antiviral reactions in brain microvascular endothelial cells and protect the brain from invading viruses such as JEV.

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