Microenvironmental acidification by pneumococcal sugar consumption fosters barrier disruption and immune suppression in the human alveolus. Microenvironmental acidification by pneumococcal sugar consumption fosters barrier disruption and immune suppression in the human alveolus

Streptococcus pneumoniae (S.p.) is the most common causative agent of community-acquired pneumonia worldwide. A key pathogenic mechanism that exacerbates severity of disease is the disruption of the alveolar-capillary barrier. However, the specific virulence mechanisms responsible for this in the human lung are not yet fully understood. In this study, we infected living human lung tissue with S.p. and observed a significant degradation of the central junctional proteins occludin and VE-cadherin, indicating barrier disruption. Surprisingly, neither pneumolysin, bacterial hydrogen peroxide nor pro-inflammatory activation were sufficient to cause this junctional degradation. Instead, pneumococcal infection led to a significant decrease of pH (approximately 6), resulting in acidification of the alveolar microenvironment, which was linked to junctional degradation. Stabilizing the pH at physiological levels during infection reversed this effect, even in a therapeutic-like approach. Further analysis of bacterial metabolites and RNA sequencing revealed sugar consumption and subsequent lactate production were the major factors contributing to bacterially induced alveolar acidification, which also hindered the release of critical immune factors. Our findings highlight bacterial metabolite-induced acidification as an independent virulence mechanism for barrier disruption and inflammatory dysregulation in pneumonia. Thus, our data suggest that strictly monitoring and buffering alveolar pH during infections caused by fermentative bacteria could serve as an adjunctive therapeutic strategy for sustaining barrier integrity and immune response. Overall design: Lung tissue explants were obtained from 7 patients primarily diagnosed with bronchial carcinoma, who had undergone lung resection at local thoracic surgery centres. The study was approved by the ethic committee at the Charité clinic (protocol number EA2/079/13) and the written informed consent was obtained from all patients. Tumour-free peripheral lung tissue was dissected into small pieces by scalpel (5 x 5 x 5 mm, 80 - 200 mg) and incubated overnight in RPMI 1640 medium (PAN-Biotech) at 37°C with 5 % CO2 to wash off clinically applied antibiotics as described. The following infection experiments were carried out in RPMI 1640 medium supplemented (volume-controlled) with 10 % fetal calf serum at 37°C with 5 % CO2. For infection, tumour-free normal lung tissue was inoculated with culture medium as control (mock infection) or 106 CFU/mL S.p. D39 wt in the presence and absence of 25 mmol/L HEPES (volume-controlled) or exposed to pH 6 titrated with HCl for 24 h . *************************************************************** Submitter states that missing raw files are due to patient privacy concerns. ***************************************************************