Immune features that afford protection from clinical disease versus sterilizing immunity to Bordetella pertussis infection in a nonhuman primate model of whooping cough

The respiratory bacterial infection caused by Bordetella pertussis (whooping cough) is the only vaccine-preventable disease whose incidence has been increasing over the last 3 decades. To better understand the resurgence of this infection, a baboon animal model of pertussis infection has been developed. Naïve baboons that recover from experimental pertussis infection are resistant both to clinical disease and to airway colonization when re-challenged. In contrast, animals vaccinated with acellular pertussis vaccine and experimentally challenged do not develop disease, but airways remain colonized for 4-6 weeks. We explored the possibility that the IgG antibody response to pertussis infection is qualitatively different from antibodies induced by acellular pertussis vaccination. IgG was purified from pertussis-convalescent baboons shown to be resistant to pertussis disease and airway colonization. Purified IgG contained high titers to pertussis toxin, pertactin, and filamentous hemagglutinin. This pertussis-immune IgG or control IgG was passively transferred to naïve, juvenile baboons before experimental airway pertussis inoculation. The control animal that received normal IgG developed a typical symptomatic infection including leukocytosis, cough and airway colonization for 4 weeks. In contrast, baboons that received convalescent IgG maintained normal WBC counts and were asymptomatic. However, despite remaining asymptomatic, their airways were colonized for 4-6 weeks with B. pertussis. All animals developed IgG and IgA anti-pertussis antibody responses. Interestingly, the clearance of B. pertussis from airways coincided with the emergence of a serum anti-pertussis IgA response. These studies demonstrate that passive administration of pertussis-specific IgG from previously infected animals can prevent clinical disease but does not affect prolonged airway colonization with B. pertussis. This outcome is similar to that observed following acellular pertussis vaccination. Understanding immune mechanisms—other than IgG—that are capable of preventing airway colonization with B. pertussis will be critical for developing more effective vaccines to prevent whooping cough

Comparison of Three Whole-Cell Pertussis Vaccines in the Baboon Model of Pertussis

Clear glass windows replace Stone's signature "Venetian loggia" arches near top of curved facade; The museum was founded in 1956 by the American Craft Council as the Museum of Contemporary Crafts; it was renamed in 2002 and in 2008, moved to 2 Columbus Circle. The new tenant radically altered the building's original design (1964) by Edward Durell Stone, which touched off a preservation battle. The museum's redesign was developed by Brad Cloepfil and his Portland, Oregon-based firm Allied Works Architecture and replaced the original white Vermont marble with a glazed terra-cotta (nacreous ceramic) and glass facade. Stone's signature elements like the "lollypop" shaped piers are still there, but boxed in with fritted glass. The interior was opened up and new slots in the building allow more light. Source: Wikipedia; http://en.wikipedia.org/wiki/Main_Page (accessed 8/5/2013

Anthrax Lethal Toxin Downregulates Claudin-5 Expression in Human Endothelial Tight Junctions

<div><p></p><p>Vascular leakage pathologies such as pleural effusion and hemorrhage are hallmarks of anthrax pathogenesis. We previously reported that anthrax lethal toxin (LT), the major virulence factor of anthrax, reduces barrier function in cultured primary human microvascular endothelial cells. Here, we show that LT-induced barrier dysfunction is accompanied by the reduced expression of the endothelial tight junction (TJ) protein claudin-5 but no change in the expression of other TJ components occludin, ZO-1, ZO-2, or the adherens junction (AJ) protein VE-cadherin. The downregulation of claudin-5 correlated temporally and dose-dependently with the reduction of transendothelial electrical resistance. LT-induced loss of claudin-5 was independent of cell death and preceded the appearance of actin stress fibers and altered AJ morphology. Pharmacological inhibition of MEK-1/2, two kinases that are proteolytically inactivated by LT, showed a similar reduction in claudin-5 expression. We found that LT reduced claudin-5 mRNA levels but did not accelerate the rate of claudin-5 degradation. Mice challenged with LT also showed significant reduction in claudin-5 expression. Together, these findings support a possible role for LT disruption of endothelial TJs in the vascular leakage pathologies of anthrax.</p></div

Claudin-5 expression and inhibitors of proteosome, lysosome, and matrix metalloproteinases (MMPs).

<p>(A) Cells were treated with medium alone or the combination of 100 ng/ml +500 ng/ml PA for 18 hours prior to the addition of the proteosome inhibitor MG132 (1 µM) for an additional 12 hours. Whole cell lysates were analyzed for claudin-5, MEK-1, and the accumulation of ubiquitinated proteins by Western blot. Representative immunoblots of three separate experiments are shown. Claudin-5 expression was normalized to tubulin and presented relative to control. Means ± SE for a minimum of three separate experiments are shown. (B) Cells were treated as indicated above for 18 hours prior to the addition of lysosome inhibitors, chloroquine (CQ, 20 µM) or E-64 (10 µg/ml) plus pepstatin A (10 µg/ml) (EP), or the broad spectrum MMP inhibitor marimastat (MST, 100 µM) for an additional 24 hours. Whole cell lysates were analyzed for claudin-5 and MEK-1 by Western blot. Claudin-5 expression was normalized to tubulin and presented relative to control. Means ± SE for a minimum of three separate experiments are shown. *, <i>p</i><0.05 versus control, #, <i>p</i><0.05 versus LT alone.</p

Effect of MEK and MAPK inhibitors on claudin-5 expression.

<p>Cells were treated with medium alone, 10 µM U0126, 10 µM SP600125, or 20 µM SB208580 for 48 hours. (A) Whole cell lysates were analyzed for claudin-5, VE-cadherin, and the phosphorylated and total forms of ERK 1/2, HSP27, and c-Jun by Western blot. Tubulin served as the loading control. Representative immunoblots of three separate experiments are shown. (B) Claudin-5 expression was normalized to tubulin and presented relative to control. Means ± SE for a minimum of three separate experiments are shown. *, <i>p</i><0.05 versus control.</p

Time-dependent reduction of claudin-5.

<p>(A) Cells were treated with medium alone or the combination of 100 ng/ml LF +500 ng/ml PA. Claudin-5 expression was analyzed by Western blot in whole cell lysates collected after the indicated treatment times. Claudin-5 expression was normalized to tubulin and presented relative to control. Means ± SE for a minimum of three separate experiments are shown. *, <i>p</i><0.05 versus control. (B) Immunofluorescence analysis of claudin-5. Cells were treated with medium alone, inactive mutant LT, or LT as described above for 48 hours. Monolayers were stained for claudin-5 (red) as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0062576#s2" target="_blank">Materials and Methods</a>. Nuclei were counterstained with Hoechst 33342 (blue). Images are representative of three separate experiments (400x magnification).</p