Publications

UNLABELLED

Streptococcus pneumoniae is an important commensal and pathogen responsible for almost a million deaths annually in children under five. The formation of biofilms by S. pneumoniae is important in nasopharyngeal colonization, pneumonia, and otitis media. Pneumolysin (Ply) is a toxin that contributes significantly to the virulence of S. pneumoniae and is an important candidate as a serotype-independent vaccine target. Having previously demonstrated that a luxS knockout mutant was unable to form early biofilms and expressed less ply mRNA than the wild type, we conducted a study to investigate the role of Ply in biofilm formation. We found that Ply was expressed in early phases of biofilm development and localized to cellular aggregates as early as 4 h postinoculation. S. pneumoniae ply knockout mutants in D39 and TIGR4 backgrounds produced significantly less biofilm biomass than wild-type strains at early time points, both on polystyrene and on human respiratory epithelial cells, cultured under static or continuous-flow conditions. Ply's role in biofilm formation appears to be independent of its hemolytic activity, as S. pneumoniae serotype 1 strains, which produce a nonhemolytic variant of Ply, were still able to form biofilms. Transmission electron microscopy of biofilms grown on A549 lung cells using immunogold demonstrated that Ply was located both on the surfaces of pneumococcal cells and in the extracellular biofilm matrix. Altogether, our studies demonstrate a novel role for pneumolysin in the assembly of S. pneumoniae biofilms that is likely important during both carriage and disease and therefore significant for pneumolysin-targeting vaccines under development.

IMPORTANCE

The bacterium Streptococcus pneumoniae (commonly known as the pneumococcus) is commonly carried in the human nasopharynx and can spread to other body sites to cause disease. In the nasopharynx, middle ear, and lungs, the pneumococcus forms multicellular surface-associated structures called biofilms. Pneumolysin is an important toxin produced by almost all S. pneumoniae strains, extensively studied for its ability to cause damage to human tissue. In this paper, we demonstrate that pneumolysin has a previously unrecognized role in biofilm formation by showing that strains without pneumolysin are unable to form the same amount of biofilm on plastic and human cell substrates. Furthermore, we show that the role of pneumolysin in biofilm formation is separate from the hemolytic activity responsible for tissue damage during pneumococcal diseases. This novel role for pneumolysin suggests that pneumococcal vaccines directed against this protein should be investigated for their potential impact on biofilms formed during carriage and disease.

Previous studies have demonstrated that increased gastric pH from the use of acid-reducing agents, such as proton-pump inhibitors or H2-receptor antagonists, can significantly impact the absorption of weakly basic drugs that exhibit pH-dependent solubility. Clinically practical strategies to mitigate this interaction have not been developed. This pilot study evaluated the extent and time course of gastric reacidification after a solid oral dosage form of anhydrous betaine HCl in healthy volunteers with pharmacologically induced hypochlorhydria. Six healthy volunteers with baseline normochlorhydria (fasting gastric pH < 4) were enrolled in this single period study. Hypochlorhydria was induced via 20 mg oral rabeprazole twice daily for four days. On the fifth day, an additional 20 mg dose of oral rabeprazole was given and gastric pH was monitored continuously using the Heidelberg pH capsule. After gastric pH > 4 was confirmed for 15 min, 1500 mg of betaine HCl was given orally with 90 mL of water and gastric pH was continuously monitored for 2 h. Betaine HCl significantly lowered gastric pH by 4.5 (± 0.5) units from 5.2 (± 0.5) to 0.6 (± 0.2) (P < 0.001) during the 30 min interval after administration. The onset of effect of betaine HCl was rapid, with a mean time to pH < 3 of 6.3 (± 4.3) min. The reacidification period was temporary with a gastric pH < 3 and < 4 lasting 73 (± 33) and 77 (± 30) min, respectively. Betaine HCl was well tolerated by all subjects. In healthy volunteers with pharmacologically induced hypochlorhydria, betaine HCl was effective at temporarily lowering gastric pH. The rapid onset and relatively short duration of gastric pH reduction gives betaine HCl the potential to aid the absorption of orally administered weakly basic drugs that exhibit pH-dependent solubility when administered under hypochlorhydric conditions.

BACKGROUND

Cystatin C could improve chronic kidney disease (CKD) classification in HIV-infected women relative to serum creatinine.

DESIGN

Retrospective cohort analysis.

METHODS

Cystatin C and creatinine were measured from specimens taken and stored during the 1999-2000 examination among 908 HIV-infected participants in the Women's Interagency HIV study (WIHS). Mean follow-up was 10.2 years. Predictors of differential glomerular filtration rate (GFR) estimates were evaluated with multivariable linear regression. The associations of baseline categories (<60, 60-90, and >90 ml/min per 1.73 m) of creatinine estimated GFR (eGFRcr), cystatin C eGFR (eGFRcys), and combined creatinine-cystatin C eGFR (eGFRcr-cys) with all-cause mortality were evaluated using multivariable Cox regression. The net reclassification index (NRI) was calculated to evaluate the effect of cystatin C on reclassification of CKD staging.

RESULTS

CKD risk factors were associated with lower eGFRcys and eGFRcr-cys values compared with eGFRcr. Relative to eGFR more than 90, the eGFR less than 60 category by eGFRcys (Adjusted hazard ratio: 2.56; 95% confidence interval: 1.63-4.02), eGFRcr-cys (3.11; 1.94-5.00), and eGFRcr (2.34; 1.44-3.79) was associated with increased mortality risk. However, the eGFR 60-90 category was associated with increased mortality risk for eGFRcys (1.80; 1.28-2.53) and eGFRcr-cys (1.91; 1.38-2.66) but not eGFRcr (1.20; 0.85-1.67). The overall NRI for mortality was 26% when reclassifying from eGFRcr to eGFRcys (P < 0.001) and was 20% when reclassifying from eGFRcr to eGFRcr-cys (P < 0.001).

CONCLUSION

The addition of cystatin C may improve mortality risk prediction by stages of kidney function relative to creatinine. CKD risk factors are associated with an overestimate of GFR by serum creatinine relative to cystatin C.

BACKGROUND

Experts can accurately characterize the histology of diminutive polyps with narrow-band imaging (NBI). There are limited data on the performance of non-experts.

OBJECTIVE

To assess the impact of a computer-based teaching module on the accuracy of predicting polyp histology with NBI by non-experts (in academics and community practice) by using video clips.

DESIGN

Prospective, observational study.

SETTING

Academic and community practice.

PARTICIPANTS

A total of 15 gastroenterologists participated-5 experts in NBI, 5 non-experts in academic practice, and 5 non-experts in community practice.

INTERVENTION

Participants reviewed a 20-minute, computer-based teaching module outlining the different NBI features for hyperplastic and adenomatous polyps.

MAIN OUTCOME MEASUREMENTS

Performance characteristics in characterizing the histology of diminutive polyps with NBI by using short video clips before (pretest) and after (posttest) reviewing the teaching module.

RESULTS

Non-experts in academic practice showed a significant improvement in the sensitivity (54% vs 79%; P < .001), accuracy (64% vs 81%; P < .001), and proportion of high-confidence diagnoses (49% vs 69%; P < .001) in the posttest. Non-experts in community practice had significantly higher sensitivity (58% vs 75%; P = .004), specificity (76% vs 90%; P = .04), accuracy (64% vs 81%; P < .001), and proportion of high-confidence diagnoses (49% vs 72%; P < .001) in the posttest. Performance of experts in NBI was significantly better than non-experts in both academic and community practice.

LIMITATIONS

Selection bias in selecting good quality videos. Performance not assessed during live colonoscopy.

CONCLUSION

Academic and community gastroenterologists without prior experience in NBI can achieve significant improvements in characterizing diminutive polyp histology after a brief computer-based training. The durability of these results and applicability in everyday practice are uncertain.

Fibrosis is characterized by accumulation of activated fibroblasts and pathological deposition of fibrillar collagens. Activated fibroblasts overexpress matrix proteins and release factors that promote further recruitment of activated fibroblasts, leading to progressive fibrosis. The contribution of epithelial cells to this process remains unknown. Epithelium-directed injury may lead to activation of epithelial cells with phenotypes and functions similar to activated fibroblasts. Prior reports that used a reporter gene fate-mapping strategy are limited in their ability to investigate the functional significance of epithelial cell-derived mesenchymal proteins during fibrogenesis. We found that lung epithelial cell-derived collagen I activates fibroblast collagen receptor discoidin domain receptor-2, contributes significantly to fibrogenesis, and promotes resolution of lung inflammation. Alveolar epithelial cells undergoing transforming growth factor-β-mediated mesenchymal transition express several other secreted profibrotic factors and are capable of activating lung fibroblasts. These studies provide direct evidence that activated epithelial cells produce mesenchymal proteins that initiate a cycle of fibrogenic effector cell activation, leading to progressive fibrosis. Therapy targeted at epithelial cell production of type I collagen offers a novel pathway for abrogating this progressive cycle and for limiting tissue fibrosis but may lead to sustained lung injury/inflammation.