Objective: Thrombin-induced platelet microbicidal protein (tPMP) is a cationic antimicrobial protein released from platelets by stimulation with thrombin. This peptide has potent microbicidal activities against a broad spectrum of microbial pathogens in infective endocarditis. Earlier evidence suggested that tPMP targets and disrupts the bacterial membrane. In this study, it was investigated whether membrane disruption is sufficient to kill streptococci or whether subsequent, presumably intracellular, events are also involved in killing. Methods: tPMP was prepared by stimulating rabbit platelets with thrombin. We examined the microbicidal activity of tPMP on
S. rattus BHT cells in the presence or absence of pretreatment with antibiotics that differ in their mechanisms of action. Effect of the exposure of
S. rattus BHT simultaneously to each antibiotics and tPMP was also studied. Results: Pretreatment of
S. rattus BHT cells with either penicillin or amoxicillin (inhibitors of bacterial cell wall synthesis) significantly enhanced the anti-
S. rattus BHT effects of tPMP. On the other hand, pretreament of
S. rattus BHT cells with tetracycline, doxycycline (30S ribosomal subunit inhibitors) or rifampin (an inhibitor of DNA-dependent RNA polymerase) significantly decreased the streptocidal effect of tPMP. Pretreatment with novobiocin (an inhibitor of bacterial DNA gyrase subunit B) did not affect the
S. rattus BHT killing. The exposure of
S. rattus BHT simultaneously to each antibiotics and tPMP, compared with the control (tPMP alone), resulted in the increase in the anti-streptococcal effect with penicillin or amoxicillin, and decrease in the anti-
S. rattus BHT effect with tetracycline, doxycycline or rifampin. Conclusion: These results suggest that tPMP exerts anti-
S. rattus BHT activity through mechanisms involving both the cell membrane and intracellular targets.
This study was supported by a grant of the Korea Health 21 R&D Project, Ministry of Health & Welfare, Republic of Korea. (02-PJ1-PG3-20501-0008)