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. 2011 Nov 1;6(11):e27091. doi: 10.1371/journal.pone.0027091

Figure 5. Elastase deficient ΔlasB mutant is attenuated in the degradation of SP-A during lung infection.

Figure 5

(A-C) The amounts of intact mSP-A were not visibly changed at 6-hr (A) or 12- hr (B) post-infection. By 18 hr post-infection (C), intact mSP-A was reduced in the BAL fluid from PAO1- or PDO240lasB-infected SP-A+/+ mice (n = 6), suggesting that mSP-A was degraded in mouse lungs. In contrast, more abundant mSP-A was clearly visible in the BAL fluids from ΔlasB (n = 8). C  =  Purified human SP-A. M1 – M8  =  BAL of mice infected with P. aeruginosa. Western blot analyses were performed using a polyclonal antibody against SP-A. (D) Densitometry analysis of mSP-A degradation by PAO1, ΔlasB and PDO240lasB in mouse lungs. The amounts of remaining mSP-A in ΔlasB were set to the value of 100%. *p<0.05 when compared the amount of mSP-A in BAL fluids from lungs infected with PAO1 or PDO240lasB against BAL fluids from ΔlasB-infected mice. (E) Mouse BAL from ΔlasB-infected animals contains intact mSP-A that permeabilizes bacterial membranes. Pooled BAL fluids (from C) (50 µg/ml total proteins) were used for membrane permeabilization assays. hSP-A (50 µg/ml) was used as positive control. BAL fluids from PAO1 and PDO240lasB infected mice failed to permeabilize E. coli membranes. hSP-A and BAL samples from ΔlasB-infected mice were able to permeabilize bacterial membranes of E. coli DH5α at higher levels. Experiments were performed independently three times in triplicates. The mean + standard deviation from one representative experiment is shown. **p<0.05 from 60 min onward when comparing the membrane permeabilization of E. coli by pure SP-A or BAL samples from ΔlasB-infected mice against BAL samples from PAO1 or PDO240lasB-infected mice.