Limits...
Factors affecting antimicrobial activity of MUC7 12-mer, a human salivary mucin-derived peptide.

Wei GX, Campagna AN, Bobek LA - Ann. Clin. Microbiol. Antimicrob. (2007)

Bottom Line: To examine the effect of salts or EDTA, a checkerboard microdilution technique was used.The peptide exhibited higher inhibitory activity against C. albicans at pH 7, 8, and 9 than at pH 5 and 6, and temperature up to 60 degrees C did not affect the activity.MUC7 12-mer peptide is effective anticandidal agent at physiological concentrations of variety of ions in the oral cavity.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Oral Biology, School of Dental Medicine, University at Buffalo, SUNY, 3435 Main Street, Buffalo, USA, NY 14214. gwei@buffalo.edu.

ABSTRACT

Background: MUC7 12-mer (RKSYKCLHKRCR), a cationic antimicrobial peptide derived from the human low-molecular-weight salivary mucin MUC7, possesses potent antimicrobial activity in vitro. In order to evaluate the potential therapeutic application of the MUC7 12-mer, we examined the effects of mono- and divalent cations, EDTA, pH, and temperature on its antimicrobial activity.

Methods: Minimal Inhibitory Concentrations (MICs) were determined using a liquid growth inhibition assay in 96-well microtiter plates. MUC7 12-mer was added at concentrations of 1.56-50 microM. MICs were determined at three endpoints: MIC-0, MIC-1, and MIC-2 (the lowest drug concentration showing 10%, 25% and 50% of growth, respectively). To examine the effect of salts or EDTA, a checkerboard microdilution technique was used. Fractional inhibitory concentration index (FICi) was calculated on the basis of MIC-0. The viability of microbial cells treated with MUC7 12-mer in the presence of sodium or potassium was also determined by killing assay or flow cytometry.

Results: The MICs of MUC7 12-mer against organisms tested ranged from 6.25-50 microM. For C. albicans, antagonism (FICi 4.5) was observed for the combination of MUC7 12-mer and calcium; however, there was synergism (FICi 0.22) between MUC7 12-mer and EDTA, and the synergism was retained in the presence of calcium at its physiological concentration (1-2 mM). No antagonism but additivity or indifference (FICi 0.55-2.5) was observed for the combination of MUC7 12-mer and each K+, Na+, Mg2+, or Zn2+. MUC7 12-mer peptide (at 25 microM) also exerted killing activity in the presence of NaCl, (up to 25 mM for C. albicans and up to 150 mM for E. coli, a physiological concentration of sodium in the oral cavity and serum, respectively) and retained candidacidal activity in the presence of KCl (up to 40 mM). The peptide exhibited higher inhibitory activity against C. albicans at pH 7, 8, and 9 than at pH 5 and 6, and temperature up to 60 degrees C did not affect the activity.

Conclusion: MUC7 12-mer peptide is effective anticandidal agent at physiological concentrations of variety of ions in the oral cavity. These results suggest that, especially in combination with EDTA, it could potentially be applied as an alternative therapeutic agent for the treatment of human oral candidiasis.

Show MeSH

Related in: MedlinePlus

Antimicrobial activity of MUC7 12-mer against C. albicans in the presence of potassium, monitored by flow cytometry. Fluorescence distribution of the control (untreated) C. albicans cells (a); cells incubated for 90 min at 37°C with 25 μM MUC7 12-mer (b-d), in the absence of KCl (b), in the presence of 40 mM KCl (c), in the presence of 80 mM KCl (d). The cells were stained with propidium iodide (3 μg/mL). Susceptibility to MUC7 12-mer is shown as percentage of C. albicans with increasing fluorescence; 10,000 events were analyzed.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2211505&req=5

Figure 2: Antimicrobial activity of MUC7 12-mer against C. albicans in the presence of potassium, monitored by flow cytometry. Fluorescence distribution of the control (untreated) C. albicans cells (a); cells incubated for 90 min at 37°C with 25 μM MUC7 12-mer (b-d), in the absence of KCl (b), in the presence of 40 mM KCl (c), in the presence of 80 mM KCl (d). The cells were stained with propidium iodide (3 μg/mL). Susceptibility to MUC7 12-mer is shown as percentage of C. albicans with increasing fluorescence; 10,000 events were analyzed.

Mentions: Figures 2 and 3 show the effect of K+ on the anticandidal activity of MUC7 12-mer, which was determined with flow cytometry. Untreated cells stained with PI showed a very low intensity of fluorescence, expressed as negative (viable cells, Figure 2a) for PI. In contrast, MUC7 12-mer or ethanol (data not show) treated cells showed a high red fluorescence, expressed as positive (dead cells, Figure 2b) for PI. Candida cells treated with MUC7 12-mer in the presence of 40 mM KCl still showed PI fluorescence (Figure 2c) with 96% PI positive cells (Figure 3), although the PI positive cells deceased to 47% in the presence of 80 mM KCl (Figure 2d and Figure 3). Similarly, when treated with a low concentration of MUC7 12-mer (5 μM), more than 96% of Candida cells were killed in the presence of KCl at concentrations up to 20 mM, and 56% of cells were killed at 40 mM KCl.


Factors affecting antimicrobial activity of MUC7 12-mer, a human salivary mucin-derived peptide.

Wei GX, Campagna AN, Bobek LA - Ann. Clin. Microbiol. Antimicrob. (2007)

Antimicrobial activity of MUC7 12-mer against C. albicans in the presence of potassium, monitored by flow cytometry. Fluorescence distribution of the control (untreated) C. albicans cells (a); cells incubated for 90 min at 37°C with 25 μM MUC7 12-mer (b-d), in the absence of KCl (b), in the presence of 40 mM KCl (c), in the presence of 80 mM KCl (d). The cells were stained with propidium iodide (3 μg/mL). Susceptibility to MUC7 12-mer is shown as percentage of C. albicans with increasing fluorescence; 10,000 events were analyzed.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2211505&req=5

Figure 2: Antimicrobial activity of MUC7 12-mer against C. albicans in the presence of potassium, monitored by flow cytometry. Fluorescence distribution of the control (untreated) C. albicans cells (a); cells incubated for 90 min at 37°C with 25 μM MUC7 12-mer (b-d), in the absence of KCl (b), in the presence of 40 mM KCl (c), in the presence of 80 mM KCl (d). The cells were stained with propidium iodide (3 μg/mL). Susceptibility to MUC7 12-mer is shown as percentage of C. albicans with increasing fluorescence; 10,000 events were analyzed.
Mentions: Figures 2 and 3 show the effect of K+ on the anticandidal activity of MUC7 12-mer, which was determined with flow cytometry. Untreated cells stained with PI showed a very low intensity of fluorescence, expressed as negative (viable cells, Figure 2a) for PI. In contrast, MUC7 12-mer or ethanol (data not show) treated cells showed a high red fluorescence, expressed as positive (dead cells, Figure 2b) for PI. Candida cells treated with MUC7 12-mer in the presence of 40 mM KCl still showed PI fluorescence (Figure 2c) with 96% PI positive cells (Figure 3), although the PI positive cells deceased to 47% in the presence of 80 mM KCl (Figure 2d and Figure 3). Similarly, when treated with a low concentration of MUC7 12-mer (5 μM), more than 96% of Candida cells were killed in the presence of KCl at concentrations up to 20 mM, and 56% of cells were killed at 40 mM KCl.

Bottom Line: To examine the effect of salts or EDTA, a checkerboard microdilution technique was used.The peptide exhibited higher inhibitory activity against C. albicans at pH 7, 8, and 9 than at pH 5 and 6, and temperature up to 60 degrees C did not affect the activity.MUC7 12-mer peptide is effective anticandidal agent at physiological concentrations of variety of ions in the oral cavity.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Oral Biology, School of Dental Medicine, University at Buffalo, SUNY, 3435 Main Street, Buffalo, USA, NY 14214. gwei@buffalo.edu.

ABSTRACT

Background: MUC7 12-mer (RKSYKCLHKRCR), a cationic antimicrobial peptide derived from the human low-molecular-weight salivary mucin MUC7, possesses potent antimicrobial activity in vitro. In order to evaluate the potential therapeutic application of the MUC7 12-mer, we examined the effects of mono- and divalent cations, EDTA, pH, and temperature on its antimicrobial activity.

Methods: Minimal Inhibitory Concentrations (MICs) were determined using a liquid growth inhibition assay in 96-well microtiter plates. MUC7 12-mer was added at concentrations of 1.56-50 microM. MICs were determined at three endpoints: MIC-0, MIC-1, and MIC-2 (the lowest drug concentration showing 10%, 25% and 50% of growth, respectively). To examine the effect of salts or EDTA, a checkerboard microdilution technique was used. Fractional inhibitory concentration index (FICi) was calculated on the basis of MIC-0. The viability of microbial cells treated with MUC7 12-mer in the presence of sodium or potassium was also determined by killing assay or flow cytometry.

Results: The MICs of MUC7 12-mer against organisms tested ranged from 6.25-50 microM. For C. albicans, antagonism (FICi 4.5) was observed for the combination of MUC7 12-mer and calcium; however, there was synergism (FICi 0.22) between MUC7 12-mer and EDTA, and the synergism was retained in the presence of calcium at its physiological concentration (1-2 mM). No antagonism but additivity or indifference (FICi 0.55-2.5) was observed for the combination of MUC7 12-mer and each K+, Na+, Mg2+, or Zn2+. MUC7 12-mer peptide (at 25 microM) also exerted killing activity in the presence of NaCl, (up to 25 mM for C. albicans and up to 150 mM for E. coli, a physiological concentration of sodium in the oral cavity and serum, respectively) and retained candidacidal activity in the presence of KCl (up to 40 mM). The peptide exhibited higher inhibitory activity against C. albicans at pH 7, 8, and 9 than at pH 5 and 6, and temperature up to 60 degrees C did not affect the activity.

Conclusion: MUC7 12-mer peptide is effective anticandidal agent at physiological concentrations of variety of ions in the oral cavity. These results suggest that, especially in combination with EDTA, it could potentially be applied as an alternative therapeutic agent for the treatment of human oral candidiasis.

Show MeSH
Related in: MedlinePlus