Haemophilus somnusInduces Apoptosis in Bovine Endothelial Cells In Vitro

Abstract

Haemophilus somnus causes pneumonia, reproductive failure, infectious myocarditis, thrombotic meningoencephalitis, and other diseases in cattle. Although vasculitis is commonly seen as a result of systemic H. somnus infections, the pathogenesis of vascular damage is poorly characterized. In this study, we demonstrated that H. somnus (pathogenic isolates 649, 2336, and 8025 and asymptomatic carrier isolates 127P and 129Pt) induce apoptosis of bovine endothelial cells in a time-and dose-dependent manner, as determined by Hoechst 33342 staining, terminal deoxynucleotidyl transferase-mediated dUTP-FITC nick end labeling, DNA fragmentation, and transmission electron microscopy. H. somnus induced endothelial cell apoptosis in as little as 1 h of incubation and did not require extracellular growth of the bacteria. Viable H. somnus organisms induced greater endothelial cell apoptosis than heat-killed organisms. Since viable H. somnus cells release membrane fibrils and blebs, which contain lipooligosaccharide (LOS) and immunoglobulin binding proteins, we examined culture filtrates for their ability to induce endothelial cell apoptosis. Culture filtrates induced similar levels of endothelial cell apoptosis, as did viable H. somnus organisms. Heat inactivation of H. somnus culture filtrates partially reduced the apoptotic effect on endothelial cells, which suggested the presence of both heat-labile and heat-stable factors. We found that H. somnus LOS, which is heat stable, induced endothelial cell apoptosis in a time-and dose-dependent manner and was inhibited by the addition of polymyxin B. These data demonstrate that H. somnus and its LOS induce endothelial cell apoptosis, which may play a role in producing vasculitis in vivo.Haemophilus somnus (24, 26) is a member of the family Pasteurellaceae, which causes bovine pneumonia (3,16,17,23,32), abortion (9, 58), thrombotic meningoencephalitis (TME) (24, 26, 52), myocarditis (24), infertility (43, 53), and arthritis (26). Vasculitis is a common finding during H. somnus infections (3,17,24,26). H. somnus also can be isolated from the urogenital tracts of male and female cattle in the absence of clinical disease (27,35,36). Most of these asymptomatic carrier isolates are sensitive to in vitro killing by bovine serum (8). Conversely, most pathogenic isolates of H. somnus are resistant to in vitro killing by bovine serum (8). Administration of pathogenic strains of H. somnus to cattle reproduces clinical respiratory disease and pulmonary vasculitis (17). In light of the propensity of H. somnus infections to cause septicemia and vasculitis, it is unfortunate that few investigators have examined the in vitro interactions of H. somnus with endothelial cells. Thompson and Little (57) first noted that pathogenic isolates of H. somnus adhered to endothelial cells from carotid arterial explants to a greater extent than Escherichia coli and Salmonella enterica serovar Typhimurium, as viewed with scanning electron microscopy. These authors observed that adherence of H. somnus cells cause...

Interactions ofHaemophilus parasuisand its LOS with porcine brain microvascular endothelial cells

“…In this study, we confirmed that H. parasuis as well as its purified LOS are not toxic to PBMEC. These results differ from those with H. influenzae and H. somnus [16,35,38]. Although not toxic, we demonstrated that both live and heat-killed H. parasuis are able to induce PBMEC apoptosis in caspase-3 dependent fashion as previously demonstrated for H. somnus [35,36].…”

“…Haemophilus influenzae LOS has been shown to cause marked cytotoxicity against bovine BMEC [38]. Moreover, Histophilus (Haemophilus) somnus LOS is known to induce apoptosis of bovine endothelial cells [35]. However, little is known about the actual biological activity of LOS from H. parasuis.…”

“…Bacteriainduced overexpression of immunomodulatory mediators may be deleterious to the host and have been linked to an increase in BBB permeability in meningitis [38]. Alterations of endothelial cells could also be the result of bacteria-induced cytotoxic effects or cell destruction by apoptosis [35,38]. Research thus far has suggested that H. parasuis is not toxic to endothelial cells [41].…”

IL‐4 induces protection of vascular endothelial cells against killing by complement and melittin through lipid biosynthesis

“…We processed and embedded the EC in Polybed 812 (Electron Microscopy Sciences, Washington, PA, USA) through a graded resin series (25-100%) [46]. We selected areas of interest for dissection, thin sectioning, and double-staining with saturated, aqueous uranyl acetate and modified Reynold's lead citrate.…”

Observation of Apoptosis in Calves Experimentally Infected With Pasteurella multocida Serotype B:2

“…Although there have been no reports characterizing the interaction of P. multocida and endothelial cells, these findings suggest that apoptosis might be linked to P. multocida endotoxin. Sylte et al (2001) has demonstrated that H. somus induced endothelial cell apoptosis in vitro. These authors also reported that exposure of endothelial cells to H. somus lipooligosaccharide (LOS) may be a key virulence factor capable of mediating endothelial cell apoptosis and ultimately vasculitis.…”