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Figure 1. Rip2 mice develop a severe and chronic lung inflammation and manifest impaired early cytokine production in response to infection.

WT and Rip2 −/− mice were infected with C. pneumoniae (1×10 6 IFU/mouse). Lungs of infected mice were isolated on days 3, 5, 14, and 35 after infection. Lungs were fixed in 10% buffered formalin, sectioned, and stained with HE to determine the extent of inflammation (A). Histological analysis of inflammation in C. pneumoniae –infected lungs of WT and Rip2 −/− mice on days 3, 5, 14, and 35 post-infection (B). Statistical significance was determined by Student's t test (* p <0.05, ** p <0.01, *** p <0.001). At 3, 5, and 14 days post-infection, the levels of IL-6, IL-12p40, and IFN-γ in BALF and lung tissue homogenates were determined (C). Statistical significance was determined by Student's t test (* p <0.05, n = 8–14).

https://doi.org/10.1371/journal.ppat.1000379.g001

Rip2 deficiency impairs the production of inflammatory cytokines during infection by day 3

Innate immune responses, and particularly IFN-γ plays an important role in host defense against acute infection and in establishment of persistence of Sale 2018 FOOTWEAR Laceup shoes Farrutx Cheap Price Original ShxgsW0
. We, therefore, determined the production of cytokines such as IL-6, IL-12 p40 and IFN-γ levels in BALF and lung homogenates from infected Rip2 and WT mice on days 3, 5 and 14. Concentrations of IL-6, IL-12p40, and IFN-γ were significantly reduced in BALF of Rip2 mice at day 3 compared to WT mice ( Online Cheapest Exclusive to mytheresacom Greta Mid suede pumps Gianvito Rossi Latest Collections Sale Outlet Clearance Best Place c2QLTd3
). However, by day 5 and day 14, IL-6, IL-12p40 and IFN-γ concentrations in the BALF and lung homogenates from Rip2 mice were significantly increased and exceeded levels in WT mice ( Lanvin Woman Metallic Patent Leathertrimmed Leather Sandals Black Size 37 Lanvin 2018 Unisex For Sale Cheap Best ptyYEh
). Thus, in addition to increased histopathological inflammation seen in Rip2 mice on days 5 and 14 and during the later stages, we observed an initial impaired and delayed kinetics in cytokine production in –infected Rip2 mice on day 3, which was also followed by a significant increased in cytokine production in the lungs on days 5 and 14 ( Womens DaisyAppliquéd Suede Caged Sandals Fabrizio Viti Websites uj61sGuzi
). We next measured IL-6 and IFN-γ levels in the supernatant of infected bone marrow–derived macrophages (BMDM) and whole lung cells . infection-induced cytokine production (IL-6, and IFN-γ release) were significantly impaired in both Rip2 macrophages and whole lung cells compared to WT macrophages and whole lung cells ( Top Quality Hyper Online FOOTWEAR Sandals MotClè Cheap Websites cdneJ7Z
).

Our data show impaired cytokine production in Rip2 −/− mice infected with C. pneumoniae early on day 3 following infection but a significant reversal and increase in cytokines and more severe and persistent lung inflammation by day 5 and 14 compared to WT mice ( Figure 1 ). We hypothesized that this more severe and persistent lung inflammation was due to an inability of Rip2 −/− mice to clear bacteria, which would then be expected to continue to provoke inflammation and cause the delayed increase in cytokine production. To test this hypothesis, we performed quantitative bacterial cultures in the lungs of mice at days 3, 5, and 14 post-infection. As anticipated, we observed significantly higher numbers of C. pneumoniae IFU in the lungs of Rip2 −/− mice on days 5 and 14 compared to WT mice ( Stretchjersey And Lace Plunge Tshirt Bra Midnight blue Stella McCartney With Paypal Sale Online AnNMEGH
). This could not be explained by higher baseline load of bacteria in Rip2 −/− mice, since on day 3, bacterial numbers in lungs were similar between WT and Rip2-deficient mice ( Cheap In China Genuine Cheap Price Tory Burch Woman Marguerite Floralappliquéd Lasercut Leather Sandals Coral Size 10 Tory Burch Professional For Sale Cheap For Nice IO5qCep5h
). Consistent with the bacterial clearance data, virtually all WT mice survived the infectious challenge, while Rip2 −/− mice had significantly increased mortality, and less than half the Rip2 −/− mice survived until the end of the experiment at day 35 ( Popular Hat for Women On Sale Black Silk 2017 Universal Size Valentino Outlet Low Price Classic Shop Sale Online Sale Footaction FTKu09MAio
). Furthermore, the lungs from the Rip2 −/− mice that succumbed to infection harbored an abundance of C. pneumoniae (data not shown), while those who survived cleared the bacteria but still manifested chronic lung inflammation at day 35 ( Figure 1B ). Collectively then, these data indicate that: 1) Rip2 importantly contributes to clearance of C. pneumoniae from the lungs; and 2) in the absence of Rip2, severe lung inflammation occurs and persists, but fails to effectively combat the infection.

Current recommendations for AOM are valid with the exception again of areas where BLNAR H. influenzae strains are found, where MIC 90 s of amoxicillin, cefaclor, cefpodoxime, and cefdinir against high-level BLNAR strains are above PK/PD breakpoints. The MIC 90 of cefditoren against high-level BLNAR strains is 0.25 μg/ml; however, the PK/PD breakpoint for this agent has not been established but is likely to be lower than the MIC 90 ( DUNGAREES Jumpsuits Momoni Free Shipping 2018 New Cost Designer L3baoy
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). Cefixime and cefpodoxime may have clinically useful activity against high-BLNAR strains, but additional information is needed and use of quinolones should be considered ( 104 , Sneakers for Women On Sale in Outlet Black Suede leather 2017 45 Hogan Buy Cheap Websites Buy Cheap Largest Supplier Extremely Online Outlet Best Wholesale Official Site Online UoeQZQ3uc5
). Similarly, current recommendations for sinusitis are valid, with the exception of areas where BLNAR H. influenzae strains are found.

Recommendations for AECB for patients with baseline risk factors, amoxicillin-clavulanate, and respiratory quinolones are valid, as H. influenzae is the predominant pathogen and these agents are active based on PK/PD breakpoints. However, of the agents recommended for patients without baseline risk factors, azithromycin, clarithromycin, telithromycin, doxycycline, cefuroxime axetil, cefpodoxime, and cefdinir, only cefpodoxime is active against H. influenzae based on PK/PD breakpoints. The rationale for these latter recommendations of agents that will not be effective for the patient group with a high probability of spontaneous resolution is unclear.

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Haemophilus influenzae is a major pathogen causing significant morbidity and mortality in all parts of the world. Meningitis and bacteremia occur in areas where the Hib vaccine is not used, whereas community-acquired respiratory tract infections caused by nontypeable strains occur worldwide. Although it has previously been assumed that little development of drug resistance has occurred in this species, we have reviewed the evidence showing that this is not the case. Apart from β-lactamase production, BLNAR strains occur, especially in Japan and France, and BLPACR strains have also been described. The clinical utility of antimicrobial agents recommended for treating diseases associated with H. influenzae are reviewed, and the limitations of the macrolide, azalide, and ketolide group of agents, tetracyclines, and selected β-lactams against infections caused by this species are noted. Quinolone resistance, though currently rare, may increase in the future. Also, widespread use of the pediatric pneumococcal conjugated vaccine has led to a relative increase in nontypeable H. influenzae compared to pneumococci in community-acquired respiratory tract infections. For all of these reasons, continued epidemiological monitoring of the susceptibility status of these strains is required. New agents are also needed, especially for penicillin-allergic patients in both pediatric patients with otitis media and older patients with AECB.

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