Effect of Streptococcus pneumoniae on human respiratory epithelium in vitro

Abstract

A total of 11 of 15 Streptococcus pneumoniae culture filtrates and all five bacterial autolysates produced by cell death in the stationary phase caused slowed ciliary beating and disruption of the surface integrity of human respiratory epithelium in organ culture. This effect was inhibited by cholesterol and was heat labile and reduced by standing at room temperature but was stable at -40°C. The activity was detected at the late stationary phase of culture and was associated with the presence of hemolytic activity. Gel filtration of a concentrated culture filtrate and autolysate both yielded a single fraction of approximately 50 kilodaltons which slowed ciliary beating and were the only fractions with hemolytic activity. Rabbit antiserum to pneumolysin, a sulfhydryl-activated hemolytic cytotoxin released by S. pneumoniae during autolysis, neutralized the effect of the culture filtrate on respiratory epithelium. Both native and recombinant pneumolysin caused ciliary slowing and epithelial disruption. Electron microscopy showed a toxic effect of pneumolysin on epithelial cells: cytoplasmic blebs, mitochondrial swelling, cellular extrusion, and cell death, but no change in ciliary ultrastructure. Recombinant pneumolysin (10 ,ug/ml) caused ciliary slowing in the absence of changes in cell ultrastructure. Release of pneumolysin in the respiratory tract during infection may perturb host defenses, allowing bacterial proliferation and spread.Streptococcus pneirnoniae is an important human pathogen. It is a constituent of the normal nasopharyngeal flora but under permissive conditions is responsible for lower respiratory tract infections, septicemia, and meningitis. The conditions required for its proliferation in and spread from the nasopharynx are unclear, but the determinant, are probably several, including both host and bacterial factors. Changes in status of the specific and nonspecific defenses of the host may be important (3,4). For example, on the one hand, pneumococcal infections are often preceded by damage to host respiratory tract epithelium by either viral infection (4, 25) or other agents, and acquisition of a pneumococcal serotype not previously encountered may permit infection owing to absence of specific antibody (4). On the other hand, the bacterium itself elaborates a number of proteins which have been implicated in its virulence (22). Meningitis and otitis media usually occur as sequelae to respiratory tract infections which are presumably accompanied by a transient bacteremia. Bacterial factors which permit invasion of the epithelium and thus entry into the bloodstream are poorly understood.In this study, we examined the effect of bacterial culture filtrates on human respiratory epithelium in vitro and characterized one moiety responsible for the changes observed. Pneumolysin, a thiol-activated membrane-damaging toxin. caused ciliary slowing, changes in epithelial cell ultrastructure, and disruption of epithelial integrity in a dose-dependent manner. MATERIALS AND METHODSPreparation of bacterial cultu...

Pneumolysin stimulates production of tumor necrosis factor alpha and interleukin-1 beta by human mononuclear phagocytes

“…However, pneumolysin has been shown to have other biological properties which, in vivo, may be important in the pathogenesis of pneumococcal infection. For example, pneumolysin alone is capable of causing the salient histological features of lobar pneumonia in rat lungs (4), is capable of causing a slowing of cilial beating in organ cultures of human respiratory epithelium (20), and is cytopathic for cultured endothelial (15) and epithelial (16) cells. Pneumolysin has been shown to activate the classical pathway of complement (12) in the absence of specific antibodies, and sublytic concentrations of the toxin cause an inhibition of antimicrobial activities of human phagocytes (9,11,19).…”

Use of Guidelines and Risk Stratification in Acute Exacerbations of Chronic Obstructive Pulmonary Disease

“…27,28 H. influenzae and other colonizing organisms such as P. aeruginosa produce substances that impair ciliary function, stimulate mucus production, destroy local immunoglobulins, impair phagocytic function, and damage the tracheobronchial epithelium. [29][30][31][32][33] H. influenzae synthesizes histamine and also releases an uncharacterized factor that impairs human neutrophil function. 34,35 A host inflammatory response is stimulated as a response to these organisms and their products.…”

Bacterial toxins which perturb ciliary function and respiratory epithelium

“…Other cilio-inhibitory factors produced by respiratory pathogens include: rhamnolipid from Ps. aeruginosa (Read et al 1992a); pneumolysin from Streptococcus pneumoniae (Steinfort et al 1989;Feldman et al 1990); a low molecular weight, heat stable factor from H. influenzae , that also increased the number of swollen, clumped and immotile cilia (Fujihara et al 1996). Abnormal ciliary beating has also been noted during infection of organ cultures by Neisseria meningitidis, Neisseria gonorrhoeae and Bordetella pertussis (Wilson 1988;Goldman 1994;Rayner et al 1995c).…”

“…Haemophilus influenzae lipopolysaccharide (Johnson et al 1983), Strep. pneumoniae pneumolysin (Steinfort et al 1989) and Bord. pertussis tracheal cytotoxin (Goldman 1994) damage epithelial cells in different experimental systems.…”