DNase activity after 6 hours incubation with tPA and antimicrobials or tPA and bacteria.

<p>The activity of DNase in the presence of different antimicrobial agents (Panels A & B) or bacteria (Panels C & D) was measured through digestion of 1 μg of pcDNA3 DNA. V+G = Vancomycin and Gentamicin, A = Amoxicillin, C = Cefazolin and F = Fluconazole. The effects of bacteria on DNase activity was assessed at baseline (T = 0) and following 6 hours incubation (T = 6) at 37°C. Samples were incubated with 5 μg/mL tPA and/or 2.5 μg/mL DNase, as applicable.</p

tPA and DNase do not directly affect the viability of <i>E</i>. <i>coli</i> and <i>S</i>. <i>aureus</i>.

<p>tPA and DNase do not directly affect the viability of <i>E</i>. <i>coli</i> and <i>S</i>. <i>aureus</i>.</p

DNase activity in the presence of vancomycin and gentamicin.

<p>The effect of increasing concentrations of gentamicin on DNase activity was measured in the presence of a constant concentration of Vancomycin. Inhibition of DNase activity, visible as smearing (incomplete digestion) of DNA (Lanes 4–8), was evident at Gentamicin concentrations >35 μg/mL. Samples contained 5 μg/mL tPA and 2.5 μg/mL DNase, as appropriate.</p

H&E sections of kidney, liver, peritoneum and spleen.

<p>Microscopic analysis of abdominal organs and peritoneum was performed following intraperitoneal administration of dialysate (A-D) or dialysate with tPA and DNase (E-H) in the presence or absence of LPS (data with LPS not shown). Samples contained 5 μg/mL tPA and 2.5 μg/mL DNase, as appropriate.</p

tPA and DNase do not influence antimicrobial activity.

<p>The effect of tPA and DNase on antimicrobial activity was assessed by measuring the survival of bacteria (with known sensitivity) to the antimicrobial agents over time in the presence or absence of tPA and DNase. All assessments were performed in triplicate and bacterial counts were calculated through spot counting of serial dilutions of bacteria. Data show is the mean number of cfu/mL at 6 and 24 hours as a % of baseline (error bars are the standard deviation). A = Amoxicillin, C = Cefazolin, V&G = Vancomycin and Gentamicin.</p

Baseline patient characteristics.

<p>Data are presented as median (quartile range) or n (%).</p><p>+Significantly higher than the respective values in other groups by <i>post-hoc</i> test.</p>*<p>Significantly lower than the respective values in other groups by <i>post-hoc</i> test.</p

Mesothelial cells express Hsp72 and Hsp73 on the cell surface.

<p>Non-permeabilised mesothelial cells were assessed for expression of cell surface-associated HSP70 proteins by immunocytochemistry. The constitutive Hsp73 and stress-induced Hsp72 forms were examined separately. The figures are representative of three independent experiments. Bar  = 20 μm.</p

Pleural mesothelial cells release Hsp72 in response to infection with <i>Streptococcus pneumoniae</i>.

<p>MeT-5A cells were treated with live (A and C) or heat-killed (B and D) <i>Streptococcus pneumoniae</i> 262 strain (A and B) and <i>S. pneumoniae</i> TIGR4 strain (C and D) and Hsp72 levels measured in culture supernatants at various time points up to 24 hr by ELISA. Significant release of Hsp72 was observed at all time points examined (p<0.05) and was still evident following treatment with heat-killed <i>S. pneumoniae.</i> * Denotes significantly higher than vehicle control cells (p<0.05). The data are presented as the mean ± SEM of three independent experiments performed in triplicate.</p

Measures of diagnostic accuracy of pleural fluid Hsp72.

<p>Values in parentheses are 95% confidence intervals.</p><p>LR, likelihood ratio; AUC, area under ROC curve.</p>*<p>This figure represents three times the upper normal limit for serum LDH.</p

Hsp72 levels in peritoneal lavage are elevated following intraperitoneal injection of mice with <i>Streptococcus pneumoniae</i>.

<p>BALB/c mice were given a single intraperitoneal injection of live <i>Streptococcus pneumoniae</i> D39 strain (∼1×107 CFU in 0.1 ml of saline; n = 11) or saline as a control (n = 10). The peritoneal cavity was lavaged 7 (n = 6) and 17 hr (n = 5) post-injection with 1 ml PBS for quantification of Hsp72 protein. The results are presented as the fold-increase in Hsp72 levels over the mean Hsp72 level in mice injected with saline alone. An approximately 2- and 2.5-fold increase in Hsp72 was shown peritoneal lavage following infection with <i>S. pneumoniae</i> for 7 and 17 hr, respectively (p<0.01 for both).</p