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Platelet activation by Histophilus somni and its lipooligosaccharide induces endothelial cell proinflammatory responses and platelet internalization.

ABSTRACT: Histophilus somni is a gram-negative coccobacillus that causes respiratory and reproductive disease in cattle. The hallmark of systemic H. somni infection is diffuse vascular inflammation that can lead to an acute central nervous system disease known as thrombotic meningoencephalitis. Previously, we demonstrated that H. somni and its lipooligosaccharide (LOS) activate bovine platelets, leading to expression of P selectin, CD40L, and FasL. Because activated platelets have been reported to induce endothelial cell cytokine production and adhesion molecule expression, we sought to determine if bovine platelets induce proinflammatory and procoagulative changes in bovine pulmonary artery endothelial cells. Endothelial cells were incubated with platelets activated with adenosine diphosphate, H. somni, or H. somni LOS. Incubation with activated bovine platelets significantly increased expression of in adhesion molecules (intercellular adhesion molecule 1, E selectin) and tissue factor, as measured by flow cytometry, real-time polymerase chain reaction, and Western blot analysis. Activated platelets also up-regulated expression of endothelial cell IL-1beta, monocyte chemoattractant protein 1, and macrophage inflammatory protein 1alpha as determined by real-time polymerase chain reaction and an IL-1beta enzyme-linked immunosorbent assay. An interesting and surprising finding was that bovine platelets activated by H. somni or its LOS were internalized by bovine endothelial cells as visualized by transmission electron microscopy. This internalization seemed to correlate with endothelial cell activation and morphological changes indicative of cell stress. These findings suggest that activated platelets might play a role in promoting vascular inflammation during H. somni infection.

SUBMITTER: Kuckleburg CJ

PROVIDER: S-EPMC2405899 | biostudies-literature | 2008 Feb

REPOSITORIES: biostudies-literature

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Platelet activation by Histophilus somni and its lipooligosaccharide induces endothelial cell proinflammatory responses and platelet internalization.

Kuckleburg Christopher J CJ McClenahan Dave J DJ Czuprynski Charles J CJ

Shock (Augusta, Ga.) 20080201 2

Histophilus somni is a gram-negative coccobacillus that causes respiratory and reproductive disease in cattle. The hallmark of systemic H. somni infection is diffuse vascular inflammation that can lead to an acute central nervous system disease known as thrombotic meningoencephalitis. Previously, we demonstrated that H. somni and its lipooligosaccharide (LOS) activate bovine platelets, leading to expression of P selectin, CD40L, and FasL. Because activated platelets have been reported to induce ...[more]

PMID: 18386389

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Project description:Histophilus somni is a Gram-negative bacterial organism that acts as an opportunistic pathogen and is a fastidious member of the Pasteurellaceae family associated with diseases of respiratory, reproductive, cardiac, and other tissues of ruminants. We identified an intervening sequence (IVS) embedded in all five copies of the 23S rRNA gene in the closed genome sequence of the H. somni isolate USDA-ARS-USMARC-63250 that may play an important role in affecting the biology of the organism. Sequencing the RNA from this isolate shows that most of the IVS is cleaved from the transcript, resulting in independent fragments of this structural rRNA that remain functional within the bacterial ribosome. The IVS lies between positions 1170 and 1278 bp of the 3,017-bp gene and exhibits self-complementarity between its 5' and 3' ends that predicts a stem-loop structure interrupting helix-45 in the transcribed 23S rRNA. Excision removes a 94-nucleotide (nt) stem-loop structure that displays an unusual 1-nt 3' end overhang instead of the more typical 2-nt overhang commonly observed at the ends of other excised IVS stem-loops. A comparison with genomes of other H. somni isolates indicates that this IVS is highly conserved, with 31 of 32 complete genomes having similar interruptions of canonical 23S rRNA genes. The potential biological effects of either the released IVS or the fragmentation of the functional 23S rRNA are unknown, but fragmentation may enhance rRNA degradation in ways that contribute to the regulation of gene expression. IMPORTANCE The genome biology underlying H. somni virulence, pathogenicity, environmental adaptability, and broad tissue tropism is understood poorly. We identified a novel H. somni 109-nt IVS stem-loop structure, of which the central portion is excised from the 23S rRNA transcript, resulting in the fragmentation of this rRNA in the H. somni isolate USDA-ARS-USMARC-63250 and the release of a 94-nt structured RNA of unknown function. We determined that this peculiar rRNA biology is widespread among sequenced H. somni isolates, suggesting it has importance to organism biology. The fragmented 23S rRNA molecules remain functional in the ribosome, given that the isolate grows in culture. The structured excised portion of the IVS, presumably due to the action of the endoribonuclease III, has an unusual 1-nt 3' end overhang. This newly discovered H. somni 23S rRNA fragmentation may enhance rRNA degradation providing a previously unrecognized avenue for regulating H. somni biological processes.

Project description:Background and aimHistophilus somni is a Gram-negative bacterium belonging to the Pasteurellaceae family that can cause bovine histophilosis. Histophilus may act as a commensal or opportunistic bacterial cattle pathogen. Comparing genomes of the pathogenic strain 2336 with the non-pathogenic preputial 129Pt isolate revealed some putative virulence factors. The study of the complete genomes of H. somni strains circulating in Russia has never been conducted before. This study aimed to identify genetic features of the H. somni strains isolated in Russia and evaluate the possibility of using strains for vaccine development.Materials and methodsThree strains of H. somni were isolated from different sources. Strain 188-VIEV was isolated from a vaginal swab sample of cattle with endometritis. 532-VIEV and 551-VIEV were cultured from the cryopreserved bull semen samples imported from Canada. Histophilus somni strain ATCC 700025 provided by ATCC (American Type Culture Collection) was also used in the study. DNA extraction was performed using QIAamp DNA Mini Kit (QIAGEN, USA). The whole-genome sequencing of the four strains was performed using Illumina Miseq. The comparison of the resulting sequences with the complete genomes of H. somni 2336 and 129Pt, and detection of the resistance genes and virulence factors, was performed using the ResFinder and Virulence Factor Database web services.ResultsThe genome size of the samples varied from 1.9 to 2.3 Mb. The number of coding sequences varied from 1795 to 2256. The average sequence density was 90%. The total guanine-cytosine (GC) content was 36.8%-37.2%, which coincided with data previously obtained for H. somni. Three out of four studied strains encoded putative virulence factors such as filamentous hemagglutinin homologs, lipooligosaccharide biosynthesis proteins, and proteins involved in iron transport and utilization. The Ser83Ile substitution was identified in the DNA topoisomerase II (gyrA) in H. somni strains 532-VIEV and 551-VIEV cultured from bull semen which led to resistance to fluoroquinolones. The gene (AAC-6-Ia + APH-2'') encoding a bifunctional aminoglycoside modification enzyme was detected in strain 551-VIEV.ConclusionStrains with virulence genes identified could be candidates for designing vaccines and potentially represent antigen sources. The results show that antibiotic-resistant H. somni can be spread with semen used for artificial insemination.

Dataset Information

Platelet activation by Histophilus somni and its lipooligosaccharide induces endothelial cell proinflammatory responses and platelet internalization.

Publications

Platelet activation by Histophilus somni and its lipooligosaccharide induces endothelial cell proinflammatory responses and platelet internalization.

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