Infection of primary human bronchial epithelial cells by Haemophilus influenzae: Macropinocytosis as a mechanism of airway epithelial cell entry

Nontypeable Haemophilus influenzae is an exclusive human pathogen which inf ects the respiratory epithelium. We have initiated studies to explore the i nteraction of the nontypeable H. influenzae strain 2019 with primary human airway epithelial cells by electron and confocal microscopy, Primary human airway cell cultures were established as monolayers on glass collagen-coate d coverslips or on semipermeable membranes at an air-fluid interface, Scann ing electron microscopy indicated that bacteria adhered to nonciliated cell s in the population. The surface of infected cells showed evidence of cytos keletal rearrangements manifested by microvilli and lamellipodia extending toward and engaging bacteria. Confocal microscopic analysis demonstrated th at infection induced actin polymerization with an increase in cortical acti n as well as evidence of actin strands around the bacteria. Transmission el ectron microscopic analysis showed lamellipodia and microvilli surrounding organisms, as well as organisms adherent to the cell surface. These studies also demonstrated the presence of bacteria within vacuoles inside of airwa y cells. Confocal microscopic studies with Texas red-labeled dextran (molec ular weight, 70,000) indicated that H. influenzae cells were entering cells by the process of macropinocytosis. These studies indicate that nontypeabl e H. influenzae can initiate cytoskeletal rearrangement within human airway epithelium, resulting in internalization of the bacteria within nonciliate d human airway epithelial cells by the process of macropinocytosis.