Rickettsia rickettsii infection of cultured endothelial cells induces release of large von Willebrand factor multimers from Weibel-Palade bodies

Abstract

The clinical manifestations of Rocky Mountain spotted fever (RMSF) result from Rickettsia rickettsii (R rickettsii) infection of endothelial cells and are mediated by pathologic changes localized to the vessel, including in situ thrombosis and tissue ischemia. This study uses in vitro infection of cultured human umbilical vein endothelial cells with R rickettsii to test the hypothesis that such infection induces von Willebrand factor (vWF) release from Weibel- Palade bodies, a process that could contribute to thrombotic changes. At 24 hours postinfection, there was an increase in metabolically prelabeled large multimers of vWF in the culture medium, with a concomitant decrease of these forms in the cell lysate samples. This release reaction was specific for the large multimer pool of vWF, localized to Weibel-Palade bodies, because no change in the distribution of dimeric forms between cells and culture medium was detected. Double-label immunofluorescence staining showed an inverse correlation between the number of R rickettsii and the number of Weibel- Palade bodies in infected cells. Cell lysis was minimal at 24 hours postinfection, as no detectable intracellular precursor forms (molecular weight 260,000) of vWF were released into the culture medium, there was no decrease in cell viability as measured by trypan blue exclusion, and no increase in 51Cr-release into the culture medium was observed when compared with uninfected controls. Release was likely a direct effect of the intracellular presence of the organism, rather than due to a noxious soluble factor such as endotoxin, because culture medium conditioned by infected endothelial cells was ineffective at inducing release in uninfected endothelial cell cultures. In summary, in vitro infection of endothelial cells by R rickettsii induces release of Weibel-Palade body contents, a process that may contribute to the pathogenesis of RMSF.

Streptococcus pneumoniae induces exocytosis of Weibel-Palade bodies in pulmonary endothelial cells

“…Similar clinical symptoms like fibrinous hepatization were described for pneumococcal pneumonia (van der Poll and Opal, 2009), indicating that endothelial cell activation accompanied by vWF release may also be involved in this disease process. In accordance with results obtained by double‐label immunofluorescence staining showing an inverse correlation between the number of internalized R. rickettsii and the number of WPB in infected cells (Sporn et al ., 1991), in our study the release of WPB proteins was predominantly detected for HPMEC to which high amounts of pneumococci had adhered. The same tendency has been observed for the internalization activity of pneumococci, which is higher in WPB‐lacking cells, although pneumococcal internalization remains negligibly low irrespective of the presence of WPB.…”

“…Only few data have been reported with respect to the induction of WPB exocytosis by microorganisms and viruses and these reports are restricted to the Herpes simplex virus (Etingin et al ., 1993) or to the intracellular bacteria Rickettsia rickettsii and Rickettsia conorii causing the Rocky Mountain spotted fever (Walker et al ., 1985). Rickettsia ‐induced infection leads to elevated levels of vWF and fibrinogen in plasma resulting in activation of coagulation and a widespread ‘consumption coagulopathy’ (Sporn et al ., 1991). Similar clinical symptoms like fibrinous hepatization were described for pneumococcal pneumonia (van der Poll and Opal, 2009), indicating that endothelial cell activation accompanied by vWF release may also be involved in this disease process.…”

Expression and Functional Significance of Adhesion Molecules on Cultured Endothelial Cells in Response to Ionizing Radiation

“…HUVEC was isolated using the methods of Gimbrone and colleagues (13) and Wagner and colleagues (43) as modified by Sporn and colleagues (39). HUVEC were maintained in M199 supplemented with 10% FBS, 2 mM L-glutamine, 100 g/ mL heparin, 50 g/mL endothelial growth factor, and 100 U/mL penicillin/streptomycin.…”

Interleukin-1 alpha production during Rickettsia rickettsii infection of cultured endothelial cells: potential role in autocrine cell stimulation

“…Since the vascular endothelium is a primary early target of infection of this obligate intracellular parasite, localized pathologic changes may be influenced or even initiated by proinflammatory and procoagulant responses of this cell type. Indeed, responses of cultured endothelial cells to R. rickettsii infection include expression of E-selectin with resultant increased neutrophil adherence (42), increased platelet adherence (38), increased expression of tissue factor (41) and plasminogen activator inhibitor (13), and release of von Willebrand factor from Weibel-Palade bodies (43).…”