STREPTOLYSIN-O AND ADHERENCE SYNERGISTICALLY MODULATE PROINFLAMMATORYRESPONSES OF KERATINOCYTES TO GROUP-A STREPTOCOCCI

In contrast to a mutant adhesin-deficient Streptococcus pyogenes (grou p A streptococcus), its isogenic parental strain binds to human kerati nocytes and promotes a vigorous proinflammatory response, characterize d by enhanced expression of several cytokines, a more rapid release of prostaglandin E-2 (PGE(2)) and damage to keratinocyte membranes. Howe ver, adherence alone is not sufficient to induce these responses. In t his study, we have begun to examine the contribution of other streptoc occal products in interactions with keratinocytes by the construction and evaluation of mutants deficient in expression of the secreted pore -forming haemolysin, streptolysin O (SLO). Inactivation of SLO did not prevent the streptococci from adhering to cultured HaCaT keratinocyte s or from expressing an unrelated second streptococcal haemolysin, str eptolysin S, during infection of keratinocytes. As measured by a quant itative reverse transcriptase polymerase chain reaction (PCR) assay, i nactivation of SLO also did not have a marked effect on the expression of interleukin 1 alpha (IL-1 alpha) during infection. However, the la ck of the ability to produce SLO was associated with a considerable re duction in expression of IL-1 beta, IL-6 and IL-8 by infected keratino cytes. Measurement of the release of PGE(2) by an enzyme-linked immuno sorbent assay demonstrated that the SLO-deficient mutants were also no t capable of promoting the rapid high level of PGE(2) release characte ristic of the adherent SLO-producing parental strain. Finally, analyse s using the fluorescent probe ethidium homodimer-l and measurements of release of keratinocyte lactate dehydrogenase indicated that the fail ure of the SLO-deficient mutants to induce responses was associated wi th the failure of these mutants to damage the integrity of the keratin ocyte membrane. These data implicate SLO as a factor that acts synergi stically with an adhesin to modulate the signalling responses of kerat inocytes during infection.