TLR4-dependent early recruitment of phagocytes to airways is critical for Bordetella pertussis clearance

GRISELDA MORENO; ERREA AGUSTINA; ROY ROBERTS; AUGUSTO GRAIEB; LAURYE VAN MAELE; JEAN CLAUDE SIRARD; ARNDT BENECKE; MARTIN RUMBO; DANIELA HOZBOR

Congreso; 1st. French-Argentinean Congress of Immunology, November 2-5, 2010. Buenos Aires, Argentina.; 2010

<!-- /* Font Definitions */ @font-face {font-family:"Arial Narrow"; panose-1:2 11 5 6 2 2 2 3 2 4; mso-font-charset:0; mso-generic-font-family:swiss; mso-font-pitch:variable; mso-font-signature:647 0 0 0 159 0;} /* Style Definitions */ p.MsoNormal, li.MsoNormal, div.MsoNormal {mso-style-parent:""; margin:0cm; margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:12.0pt; font-family:"Times New Roman"; mso-fareast-font-family:"Times New Roman";} @page Section1 {size:612.0pt 792.0pt; margin:70.85pt 3.0cm 70.85pt 3.0cm; mso-header-margin:36.0pt; mso-footer-margin:36.0pt; mso-paper-source:0;} div.Section1 {page:Section1;} --> TLR4 dependent processes shape adaptative anti-pertussis immunity, however there is scarce knowledge on its contribution to the antimicrobial scenario during the early stage of B. pertussis infection. The aim of this study was to characterize the recruitment and antipicrobial capacity of cell populations from broncoalveolar lavage (BALF) at 2, 6, and 24 h after B. pertussis mice intranasal infection (1.10E8CFU/40μl). By flow cytometry we observed that the influx of neutrophils (PMN, CD11c-, CD11b+, Ly6G+) into the airways was significant in TLR4 competent mice, rising from less than 1.10E3/BALF to 7.10E5 ± 1.10E5 cells/BALF at 6 h. For TLR4 deficient mice strain (C3H/HeJ mice strain), recruitment of PMN was evident only at 24h, being in all time points analyzed lower than in TLR4 competent strain. The expression of proinflammatory chemokines such as CXCL1, CXCL2, CXCL5 was higher in TLR4 competent mice than in C3H/HeJ mice. Depletion of PMN by specific monoclonal antibodies, increased bacterial loads from 7.7±0.12 to 8.4±0.53 logCFU/lung at 24 hs (p=0.097) and 7.15 ± 0.21 to 8.36 ± 0.026 log CFU/lung at 48 hs (p=0.0023). Using CFSE labeled bacteria we observed that during the first 24 h of infection bacteria were mainly associated to PMN (27,66±0,66 %vs 11,41± 1,99% at 6h and 16,54±3,02% vs 7,29±6,33% at 24h. This result seems to be independent of TLR4 since there was no difference between TLR4 competent and deficient mice strains. As an effector mechanisms we observed that PMN contained higher level of reactive oxygen species (ROS) in comparision with AM cell population  PMN ROS+: 33.72±1.50% vs AM ROS+: 9.10±3.89% at 6hs; 57.70±10.18%  vs 18.50±5.10% at 24hs), indicated that PMN were the main producer of ROS in the first 24hs of infection. Alltogether these results indicate that TLR4 activation at early time points of B. pertussis infection is critical for eliciting anti-pertussis response and that PMNs recruitment participates actively in the bacterial clearance.