CFTR ion channel activity modulators cannot restore SHS impaired bacterial phagocytosis and limit survival.

Abstract

<p>(A) RAW264.7 cells were seeded on a 24-well plate and treated overnight with CFTR(inh)-172 (10 μM). In addition, cells were infected with <i>PA01</i>-GFP (MOI 10) for 150min followed by microscopy. Representative bright field (top) and fluorescent images (bottom) are shown (magnification 40X, n = 6, white bar = 20μm). (B) Next we quantified (from 2A) the percentage of macrophages that were infected with <i>PA01</i>-GFP and found that inhibiting CFTR ion channel activity does not impair bacterial phagocytosis. (C) RAW264.7 cells were seeded on a 24-well plate and treated overnight with the flavonoids, Rutin Hydrate (RH, 10μM) or Quercetin (Q, 10μM). Cells were also infected with <i>PA01</i>-GFP (MOI 10) and/or treated with cigarette smoke extract (CSE; 10%; SHS model) for 150 min followed by microscopy. The representative bright field (top) and fluorescent images (bottom) are shown (magnification 40X, n = 6, white bar = 20μm). (D) The quantification (from 2C) of bacterial phagocytosis shows significant (*p<0.05, **p<0.01) increase in Q or RH and CSE treated groups as compared to the CSE treated control group. But this change is not sufficient to restore SHS impaired bacterial phagocytosis to the levels seen in the control group. (E) In a separate experiment (similar to one described in 2C), media (100μl) was collected and spread on 2% LB agar plates. The plates were incubated overnight at 37°C and colony forming unit (CFU) counts were used to quantify bacterial survival. CSE treatment significantly (**p<0.01) improves bacterial survival, but RH or Q treatment of CSE group does not significantly affect bacterial counts.</p

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