Related Experiment Videos
Staphylocidal action of thrombin-induced platelet microbicidal protein is influenced by microenvironment and target
S P Koo1, M R Yeaman, A S Bayer
1Department of Medicine, St. John's Cardiovascular Research Center, LAC-Harbor-UCLA Medical Center, Torrance 90509, USA. KOO@HUMC.EDU
Abstract:
Thrombin-induced platelet microbicidal protein (tPMP) is a small, cationic peptide released from rabbit platelets following exposure to thrombin in vitro. This peptide exerts potent in vitro microbicidal activity against a broad spectrum of bloodstream pathogens, including Staphylococcus aureus. It is known that the microbicidal actions of other cationic antimicrobial peptides (e.g., neutrophil defensins) are influenced by environmental factors and target cell growth phase. However, whether these parameters affect tPMP microbicidal activity has not been studied. Thus, we assessed the in vitro bactericidal activity of tPMP against two tPMP-susceptible strains, Bacillus subtilis ATCC 6633 and S. aureus 502A, in various target cell growth phases or under various microenvironmental conditions. The conditions studied included differing bacterial growth phase (logarithmic versus stationary), temperature (range, 4 to 42 degrees C), pH (range, 4.5 to 8.5), cationicity (range, 0.1 mM to 2 M), anionicity (range, 0.08 to 5 microM), and neutral carbohydrates ranging in molecular weight (MW) from 180 to 37,700 (range, 50 to 500 mM) as well as rabbit platelet-free plasma and serum. tPMP staphylocidal activity was greater against logarithmic- than stationary-phase cells. tPMP bactericidal activity against both B. subtilis and S. aureus was directly correlated with temperature and pH, with microbicidal activity exhibited near the physiological range (37 to 42 degrees C and pH 7.2 to 8.5, respectively). The presence of cations (Na+, K+, Ca2+, and Mg2+) decreased tPMP bactericidal activity in a time- and concentration-dependent manner, with complete inhibition at monovalent or divalent cation concentrations of > or = 250 or > or = 10 mM, respectively. Staphylocidal activity of tPMP was also inhibited by the polyanions polyanetholsulfonic acid and polyaspartic acid, at 0.1 and 0.4 microM, respectively. Coincident exposure with low-MW carbohydrates (glucose, sucrose, and melezitose) did not affect tPMP staphylocidal activity. However, higher-MW carbohydrates (raffinose and dextrans) decreased tPMP activity in a manner directly proportional to their concentration and MW. Solute-mediated inhibition of tPMP bactericidal activity was independent of solute osmolality but directly related to the duration of tPMP-solute coexposure. tPMP enhanced the staphylocidal activities of platelet-free plasma and heat-inactivated serum, while the activity of normal serum was not affected. These collective observations suggest that tPMP retains antimicrobial activities under physiological conditions which are likely to be relevant to host defense in vivo.
Insights
Thrombin-induced platelet microbicidal protein (tPMP) shows potent antimicrobial activity against bacteria, especially in logarithmic growth phases. Its efficacy is influenced by environmental factors like pH and temperature, but it retains activity under physiological conditions.
Area of Science:
- Antimicrobial Peptides
- Bacteriology
- Immunology
Background:
- Thrombin-induced platelet microbicidal protein (tPMP) is a cationic peptide with known antimicrobial activity.
- Environmental factors and bacterial growth phase can influence antimicrobial peptide efficacy.
- The impact of these factors on tPMP activity was previously unstudied.
Purpose of the Study:
- To investigate the effect of bacterial growth phase and microenvironmental conditions on tPMP's in vitro bactericidal activity.
- To determine if tPMP retains antimicrobial efficacy under physiological conditions relevant to host defense.
Main Methods:
- Assessed tPMP activity against Bacillus subtilis and Staphylococcus aureus in different growth phases (logarithmic vs. stationary).
- Tested tPMP efficacy across a range of temperatures, pH levels, cation and anion concentrations, and neutral carbohydrates.
- Evaluated tPMP activity in rabbit platelet-free plasma and serum.
Main Results:
- tPMP exhibited greater activity against logarithmic-phase bacteria compared to stationary-phase cells.
- Bactericidal activity correlated positively with temperature and pH, optimal within physiological ranges (37-42°C, pH 7.2-8.5).
- Cations and polyanions inhibited tPMP activity; higher-MW carbohydrates reduced efficacy, while low-MW carbohydrates had no effect.
- tPMP enhanced antimicrobial activity in platelet-free plasma and heat-inactivated serum.
Conclusions:
- tPMP's antimicrobial activity is modulated by bacterial growth phase and microenvironmental conditions.
- tPMP demonstrates significant antimicrobial efficacy under physiological conditions, suggesting in vivo relevance.
- These findings support tPMP's potential role in host defense mechanisms against bacterial pathogens.