Ultrastructural studies on the interaction of haemophilus influenzae with human endothelial cells in vitro

1990 ◽  
Vol 48 (3) ◽  
pp. 636-637
Author(s):  
D.J.P. Ferguson ◽  
M. Virji ◽  
H. Kayhty ◽  
E.R. Moxon
Keyword(s):  
Endothelial Cells ◽  
Haemophilus Influenzae ◽  
Intercellular Junctions ◽  
Blood Stream ◽  
Ultrastructural Studies ◽  
Endothelial Barriers ◽  
Serotype B ◽  
Meningitis In Children ◽  
Respiratory Epithelial

Haemophilus influenzae is a human pathogen which causes meningitis in children. Systemic H. influenzae infection is largely confined to encapsulated serotype b organisms and is a major cause of meningitis in the U.K. and elsewhere. However, the pathogenesis of the disease is still poorly understood. Studies in the infant rat model, in which intranasal challenge results in bacteraemia, have shown that H. influenzae enters submucosal tissues and disseminates to the blood stream within minutes. The rapidity of these events suggests that H. influenzae penetrates both respiratory epithelial and endothelial barriers with great efficiency. It is not known whether the bacteria penetrate via the intercellular junctions, are translocated within the cells or carried across the cellular barrier in 'trojan horse' fashion within phagocytes. In the present studies, we have challenged cultured human umbilical cord_vein endothelial cells (HUVECs) with both capsulated (b+) and capsule-deficient (b-) isogenic variants of one strain of H. influenzae in order to investigate the interaction between the bacteria and HUVEC and the effect of the capsule.

Morphilogy and Ultrastructure of in Vitro Cultures of Aortic Endothelial Cells Interacting with Antibodies

1979 ◽  
Author(s):  
S. Korach ◽  
D. Ngo
Keyword(s):  
Endothelial Cells ◽  
Cell Surface ◽  
Vascular Endothelial Cells ◽  
Intercellular Junctions ◽  
Cell Types ◽  
Endothelial Damage ◽  
Antibody Concentration ◽  
High Yields ◽  
Scanning Em

Adult pig aortas, sectioned longitudinally, were incubated in 0.1% collagenase-PBS (15 mn, 37°C). Gentle scraping of the lumenal surface resulted in high yields (3-4 x 106 cell/aorta) of viable endothelial cells, essentially devoid of other cell types by morphological and immunochemical (F VIII-antigen) criteria. Confluent monolayers were incubated for various times (5 mn to 1 wk) with decomplemented rabbit antisera raised against pig endothelial cells. Changes in cell morphology appeared to depend on antibody concentration rather than on duration of contact with antiserum. High concentrations of antiserum (5 to 20%) led to cytoplasmic shredding, bulging of cells and extensive vacuolization, whereas at lower concentrations, cells appeared almost normal. Transmission EM studies by the indirect immunoperoxydase method showed antibodies reacting with unfixed cells to be distributed all over the upper cell surface, in the outer parts of intercellular junctions, and within numerous pinocytotic vesicles. Much weaker reactions could also be seen at the lower cell surface. When viewed under the Scanning EM, antiserum-treated endothelial cells also disclosed antibody concentration-dependent bulging and release of cells from their substrate. In vitro studies of gradual modifications of vascular endothelial cells acted upon by antibodies should provide a better understanding of the structural and biochemical processes underlying endothelial damage and detachment.

scholarly journals Treponema pallidum Disrupts VE-Cadherin Intercellular Junctions and Traverses Endothelial Barriers Using a Cholesterol-Dependent Mechanism

2021 ◽  
Vol 12 ◽  
Author(s):  
Karen V. Lithgow ◽  
Emily Tsao ◽  
Ethan Schovanek ◽  
Alloysius Gomez ◽  
Leigh Anne Swayne ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Lipid Raft ◽  
Vascular Endothelium ◽  
Immunofluorescence Microscopy ◽  
Treponema Pallidum ◽  
Intercellular Junctions ◽  
Endothelial Barriers ◽  
Junctional Protein ◽  
Gain Access ◽  
Dependent Mechanism

Treponema pallidum subspecies pallidum, the causative agent of syphilis, traverses the vascular endothelium to gain access to underlying tissue sites. Herein, we investigate the mechanisms associated with T. pallidum traversal of endothelial barriers. Immunofluorescence microscopy reveals that a subpopulation of T. pallidum localizes to intercellular junctions and that viable T. pallidum, as well as a T. pallidum vascular adhesin (Tp0751), disrupts the architecture of the main endothelial junctional protein VE-cadherin. Intriguingly, in this study we show that T. pallidum traverses endothelial barriers with no disruption in barrier permeability. Furthermore, barrier traversal by T. pallidum is reduced by pretreatment of endothelial cells with filipin, an inhibitor that blocks cholesterol-mediated endocytosis. Collectively, these results suggest that T. pallidum can use a cholesterol-dependent, lipid raft-mediated endocytosis mechanism to traverse endothelial barriers. Further, treponemal localization to, and disruption of, intercellular junctions suggests that a paracellular route may also be utilized, a dual traversal strategy that has also been observed to occur for leukocytes and other invasive bacteria.

Interactions of Haemophilus influenzae with Human Endothelial Cells In Vitro

1992 ◽  
Vol 165 (Supplement 1) ◽  
pp. S115-S116 ◽  
Author(s):  
M. Virji ◽  
H. Kayhty ◽  
D. J. P. Ferguson ◽  
C. Alexandrescu ◽  
E. R. Moxon
Keyword(s):  
Endothelial Cells ◽  
Haemophilus Influenzae ◽  
Human Endothelial Cells

scholarly journals Emergence of Non-Serotype b EncapsulatedHaemophilus influenzaeas a Cause of Pediatric Meningitis in Northwestern Ontario

2013 ◽  
Vol 24 (1) ◽  
pp. 13-16 ◽  
Author(s):  
Pouya Sadeghi-Aval ◽  
Raymond SW Tsang ◽  
Frances B Jamieson ◽  
Marina Ulanova
Keyword(s):  
Bacterial Meningitis ◽  
Haemophilus Influenzae ◽  
Present Article ◽  
Genetic Background ◽  
Antibiotic Sensitivity ◽  
Invasive Disease ◽  
Northwestern Ontario ◽  
Clonal Nature ◽  
Serotype B ◽  
Meningitis In Children

Before the introduction of the conjugate vaccine,Haemophilus influenzaeserotype b (Hib) was the leading cause of bacterial meningitis in children. Although successful in reducing Hib cases, the vaccine confers no protection against other serotypes ofH influenzae, such as a (Hia), or f (Hif). The emergence of invasive disease caused by non-Hib in northwestern Ontario (38 cases between 2002 and 2008) with predominance of Hia was previously reported by the authors. At that time, no cases of pediatric meningitis caused byH influenzaewere recorded in the region. Continued surveillance identified 12 new cases of invasive non-Hib between January 2009 and July 2011. Among these cases, three young children developed meningitis with severe complications caused by Hia or Hif. The present article describes these cases along with the characteristics of recentH influenzaeisolates from the region, (ie, their genetic background and antibiotic sensitivity). The findings point to the clonal nature of circulating Hia strains as well as to an increase in frequency and severity of pediatric invasiveH influenzaeinfections in northwestern Ontario.

scholarly journals ULTRASTRUCTURAL STUDIES ON THE PERMEABILITY OF THE MESOTHELIUM TO HORSERADISH PEROXIDASE

1968 ◽  
Vol 37 (1) ◽  
pp. 123-138 ◽  
Author(s):  
Ramzi S. Cotran ◽  
Morris J. Karnovsky
Keyword(s):  
Electron Microscope ◽  
Horseradish Peroxidase ◽  
Intraperitoneal Injection ◽  
Intercellular Junctions ◽  
Ultrastructural Studies ◽  
Intercellular Clefts ◽  
Peritoneal Mesothelium ◽  
Physiologic Data

Peritoneal mesothelium was exposed for 2–60 min to solutions of horseradish peroxidase by incubation in vitro, or after intraperitoneal injection in vivo. Peroxidase was localized, with the electron microscope in the intercellular clefts of the mesothelium, often along their entire lengths, in vesicles adjoining or contiguous with the clefts, and along the peritoneal and basal surfaces of the cell, and also in intracytoplasmic vacuoles. The intercellular junctions of peroxidase-treated mesothelium did not differ from those of controls: open and closed junctions were present in both groups. Intercellular localization was also obtained when the mesothelium was exposed to peroxidase during or after fixation. Although intracellular absorption of peroxidase and its incorporation into larger vacuoles were observed, there was no clearcut evidence of vesicular transport across the mesothelium in these experiments. These findings are consistent with physiologic data which postulate that mesothelial transport can be accounted for, at least in part, by passive diffusion through a system of pores, and they suggest that these pores are located in the intercellular clefts.

scholarly journals Clostridium perfringens type D epsilon toxin produces a rapid and dose-dependent cytotoxic effect on cerebral microvascular endothelial cells in vitro

2019 ◽  
Vol 32 (2) ◽  
pp. 277-281 ◽  
Author(s):  
Kimberley A. Mander ◽  
Francisco A. Uzal ◽  
Ruth Williams ◽  
John W. Finnie
Keyword(s):  
Endothelial Cells ◽  
Clostridium Perfringens ◽  
Vasogenic Edema ◽  
Ultrastructural Studies ◽  
Type D ◽  
Epsilon Toxin ◽  
Capillary Endothelial Cells ◽  
Dose Dependent ◽  
Ruminant Livestock

Clostridium perfringens type D epsilon toxin (ETX) is responsible for a severe and frequently fatal neurologic disorder in ruminant livestock. Light microscopic, immunohistochemical, and ultrastructural studies have suggested that ETX injury to the cerebral microvasculature, with subsequent severe, generalized vasogenic edema and increased intracranial pressure, is critically important in producing neurologic dysfunction. However, the effect of ETX on brain capillary endothelial cells in vitro has not been examined previously, to our knowledge. We exposed a well-characterized human blood–brain barrier cell line to increasing concentrations of ETX, and demonstrated a direct and dose-dependent endotheliotoxic effect. Our findings are concordant with the primacy of vasculocentric brain lesions in the diagnosis of acute epsilon toxin enterotoxemia in ruminant livestock.

scholarly journals TAL-1/SCL and Its Partners E47 and LMO2 Up-Regulate VE-Cadherin Expression in Endothelial Cells

2007 ◽  
Vol 27 (7) ◽  
pp. 2687-2697 ◽  
Author(s):  
Virginie Deleuze ◽  
Elias Chalhoub ◽  
Rawan El-Hajj ◽  
Christiane Dohet ◽  
Mikaël Le Clech ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Umbilical Vein ◽  
Ectopic Expression ◽  
Intercellular Junctions ◽  
Hek 293 ◽  
Helix Loop Helix ◽  
293 Cells ◽  
Functional Consequences ◽  
Bona Fide

ABSTRACT The basic helix-loop-helix TAL-1/SCL essential for hematopoietic development is also required during vascular development for embryonic angiogenesis. We reported that TAL-1 acts positively on postnatal angiogenesis by stimulating endothelial morphogenesis. Here, we investigated the functional consequences of TAL-1 silencing in human primary endothelial cells. We found that TAL-1 knockdown caused the inhibition of in vitro tubulomorphogenesis, which was associated with a dramatic reduction in vascular endothelial cadherin (VE-cadherin) at intercellular junctions. Consistently, silencing of TAL-1 as well as of its cofactors E47 and LMO2 down-regulated VE-cadherin at both the mRNA and the protein level. Endogenous VE-cadherin transcription could be activated in nonendothelial HEK-293 cells by the sole concomitant ectopic expression of TAL-1, E47, and LMO2. Transient transfections in human primary endothelial cells derived from umbilical vein (HUVECs) demonstrated that VE-cadherin promoter activity was dependent on the integrity of a specialized E-box associated with a GATA motif and was maximal with the coexpression of the different components of the TAL-1 complex. Finally, chromatin immunoprecipitation assays showed that TAL-1 and its cofactors occupied the VE-cadherin promoter in HUVECs. Together, these data identify VE-cadherin as a bona fide target gene of the TAL-1 complex in the endothelial lineage, providing a first clue to TAL-1 function in angiogenesis.

scholarly journals Structural Characterization of Haemophilus influenzae Enolase and Its Interaction with Human Plasminogen by In Silico and In Vitro Assays

Pathogens ◽  
2021 ◽  
Vol 10 (12) ◽  
pp. 1614
Author(s):  
Yesenia Osorio-Aguilar ◽  
Maria Cristina Gonzalez-Vazquez ◽  
Diana Elizabeth Hernandez-Ceron ◽  
Patricia Lozano-Zarain ◽  
Ygnacio Martinez-Laguna ◽  
...  
Keyword(s):  
Haemophilus Influenzae ◽  
Host Cells ◽  
In Vitro Assays ◽  
Binding Motif ◽  
Kringle Domains ◽  
Multifunctional Protein ◽  
Pediatric Diseases ◽  
Human Plasminogen ◽  
Serotype B

Haemophilus influenzae is the causal agent of invasive pediatric diseases, such as meningitis, epiglottitis, pneumonia, septic arthritis, pericarditis, cellulitis, and bacteremia (serotype b). Non-typeable H. influenzae (NTHi) strains are associated with localized infections, such as otitis media, conjunctivitis, sinusitis, bronchitis, and pneumonia, and can cause invasive diseases, such as as meningitis and sepsis in immunocompromised hosts. Enolase is a multifunctional protein and can act as a receptor for plasminogen, promoting its activation to plasmin, which leads to the degradation of components of the extracellular matrix, favoring host tissue invasion. In this study, using molecular docking, three important residues involved in plasminogen interaction through the plasminogen-binding motif (251EFYNKENGMYE262) were identified in non-typeable H. influenzae enolase (NTHiENO). Interaction with the human plasminogen kringle domains is conformationally stable due to the formation of four hydrogen bonds corresponding to enoTYR253-plgGLU1 (K2), enoTYR253-plgGLY310 (K3), and enoLYS255-plgARG471/enoGLU251-plgLYS468 (K5). On the other hand, in vitro assays, such as ELISA and far-western blot, showed that NTHiENO is a plasminogen-binding protein. The inhibition of this interaction using polyclonal anti-NTHiENO antibodies was significant. With these results, we can propose that NTHiENO–plasminogen interaction could be one of the mechanisms used by H. influenzae to adhere to and invade host cells.

scholarly journals DNAM-1 and PVR Regulate Monocyte Migration through Endothelial Junctions

2004 ◽  
Vol 199 (10) ◽  
pp. 1331-1341 ◽  
Author(s):  
Nicolas Reymond ◽  
Anne-Marie Imbert ◽  
Elisabeth Devilard ◽  
Stéphanie Fabre ◽  
Christian Chabannon ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Nk Cell ◽  
Vascular Endothelial Cells ◽  
Specific Binding ◽  
Intercellular Junctions ◽  
Cell Junctions ◽  
Apical Surface ◽  
Monocyte Migration ◽  
Endothelial Junctions

DNAX accessory molecule 1 (DNAM-1; CD226) is a transmembrane glycoprotein involved in T cell and natural killer (NK) cell cytotoxicity. We demonstrated recently that DNAM-1 triggers NK cell–mediated killing of tumor cells upon engagement by its two ligands, poliovirus receptor (PVR; CD155) and Nectin-2 (CD112). In the present paper, we show that PVR and Nectin-2 are expressed at cell junctions on primary vascular endothelial cells. Moreover, the specific binding of a soluble DNAM-1–Fc molecule was detected at endothelial junctions. This binding was almost completely abrogated by anti-PVR monoclonal antibodies (mAbs), but not modified by anti–Nectin-2 mAbs, which demonstrates that PVR is the major DNAM-1 ligand on endothelial cells. Because DNAM-1 is highly expressed on leukocytes, we investigated the role of the DNAM-1–PVR interaction during the monocyte transendothelial migration process. In vitro, both anti–DNAM-1 and anti-PVR mAbs strongly blocked the transmigration of monocytes through the endothelium. Moreover, after anti–DNAM-1 or anti-PVR mAb treatment, monocytes were arrested at the apical surface of the endothelium over intercellular junctions, which strongly suggests that the DNAM-1–PVR interaction occurs during the diapedesis step. Altogether, our results demonstrate that DNAM-1 regulates monocyte extravasation via its interaction with PVR expressed at endothelial junctions on normal cells.

scholarly journals Structural, Functional, and Metabolic Alterations in Human Cerebrovascular Endothelial Cells during Toxoplasma gondii Infection and Amelioration by Verapamil In Vitro

2020 ◽  
Vol 8 (9) ◽  
pp. 1386
Author(s):  
Alaa T. Al-sandaqchi ◽  
Victoria Marsh ◽  
Huw E. L. Williams ◽  
Carl W. Stevenson ◽  
Hany M. Elsheikha
Keyword(s):  
Endothelial Cells ◽  
Toxoplasma Gondii ◽  
Intercellular Junctions ◽  
Proton Nuclear Magnetic Resonance ◽  
Functional Changes ◽  
Brain Infection ◽  
Toxoplasma Gondii Infection ◽  
Cerebrovascular Endothelial Cells ◽  
Treatment Application

Toxoplasma gondii (T. gondii), the causative agent of toxoplasmosis, is a frequent cause of brain infection. Despite its known ability to invade the brain, there is still a dire need to better understand the mechanisms by which this parasite interacts with and crosses the blood–brain barrier (BBB). The present study revealed structural and functional changes associated with infection and replication of T. gondii within human brain microvascular endothelial cells (BMECs) in vitro. T. gondii proliferated within the BMECs and disrupted the integrity of the cerebrovascular barrier through diminishing the cellular viability, disruption of the intercellular junctions and increasing permeability of the BMEC monolayer, as well as altering lipid homeostasis. Proton nuclear magnetic resonance (1H NMR)-based metabolomics combined with multivariate data analysis revealed profiles that can be attributed to infection and variations in the amounts of certain metabolites (e.g., amino acids, fatty acids) in the extracts of infected compared to control cells. Notably, treatment with the Ca2+ channel blocker verapamil rescued BMEC barrier integrity and restricted intracellular replication of the tachyzoites regardless of the time of treatment application (i.e., prior to infection, early- and late-infection). This study provides new insights into the structural and functional changes that accompany T. gondii infection of the BMECs, and sheds light upon the ability of verapamil to inhibit the parasite proliferation and to ameliorate the adverse effects caused by T. gondii infection.

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