Compound and method of treatment for fungal pathologies of the oral cavity
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example 1
[0040] The first example suggests that α-MSH (11-13) (SEQ ID NO: 1), (6-13) (SEQ ID NO: 2) and (1-13) (SEQ ID NO: 3) exhibit similar anti-candidal properties as flucanazole over an exceedingly broad range of concentrations.
[0041]C. albicans (1×106 / ml in HBSS) was incubated in the presence of absence or α-MSH (1-13) (SEQ ID NO: 3) or (11-13) (SEQ ID NO: 1) at concentrations in the range of 10−15 M to 10−6 M for 2 hours at 37° C. Cells were then washed in cold distilled waste and diluted with HBSS to a concentration of 100 organisms / ml. One-ml aliquots were dispensed on blood agar plates and incubated for 48 hours at 37° C. Organism viability was estimated from the number of colonies formed.
[0042] In subsequent experiments using familiar procedures we compared activity of α-MSH (4-10) (SEQ ID NO: 5), (6-13) (SEQ ID NO: 2), (11-13) (SEQ ID NO: 1), ACTH (1-39) (SEQ ID NO: 6), (18-39) (SEQ ID NO: 7) and fluconazole, the latter an established antifungal agent. Melanocortin peptides and ...
example 2
[0046] Example 2 demonstrates that α-MSH (1-13) (SEQ ID NO: 3), (6-13) (SEQ ID NO: 2) or (11-13) (SEQ ID NO: 1) strongly inhibits Candidal germination. C. albicans from stationary phase cultures were washed twice with distilled water and suspended in HBSS to a final concentration of 2×10−6 / ml. Hyphal growth was induced by addition of 10% inactivated horse serum (GIBCO / BRL, Great Britain) to yeast incubated for 45 minutes at 37° C. with continuous shaking. Horse serum was removed by washing cells twice with HBSS and incubation was continued for 60 minutes at 37° C. in the presence of α-MSH (1-13)_(SEQ ID NO: 3), (6-13) (SEQ ID NO: 2) or (11-13) (SEQ ID NO: 1) at a concentration of 10−6M with continuous shaking. The percentage of filamentous cells was evaluated under a light microscope with the aid of hemocytometer. Experiments were run in triplicate and at least 200 cells were scored. Photomicrographs were taken with a MC100 camera attached to an Axioskop Zeiss microscope.
[0047] FIG...
example 3
[0049] Example 3 illustrates that α-MSH and its derivatives exhibit their anti-candidal properties without compromising the ability of human neutrophils to independently combat Candida. Venous blood (20m1) from health volunteers was anticoagulated with heparin. Neutrophils were isolated using dextran sedimentation and Ficoll-Hypaque (Sigma Chemical Co., St. Louis, Mo., USA) centrifugation. Erythrocytes were lysed via hypotonic shock. Neutrophils represented at least 97% of the cell suspension. Cell viability, estimated by trypan blue exclusion, was>98%. Neutrophils were suspended to a final concentration in HBSS.
[0050]C. albicans (1×10−6) were opsonized with human AB serum in a shaking water bath for 30 minutes at 37° C. Organisms were then incubated with neutrophils in medium or in medium with α-MSH (1-13) (SEQ ID NO: 3) or (α-MSH (11-13) (SEQ ID NO: 1) in concentrations of 10×15 M to 10−4 M in a shaking water bath for 2 hours at 37° C. After incubation, the culture tubes were pla...
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