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A Tetrameric Peptide Derived from Bovine Lactoferricin Exhibits Specific Cytotoxic Effects against Oral Squamous-Cell Carcinoma Cell Lines.

Solarte VA, Rosas JE, Rivera ZJ, Arango-Rodríguez ML, García JE, Vernot JP - Biomed Res Int (2015)

Bottom Line: Additionally, this tetrameric peptide showed the highest specificity towards tumorigenic cells among the tested peptides.Interestingly, this effect was very fast, with cell shrinkage, severe damage to cell membrane permeability, and lysis within one hour of treatment.Our results are consistent with a necrotic effect rather than an apoptotic one and suggest that this tetrameric peptide could be considered as a new candidate for the therapeutic treatment of OSCC.

View Article: PubMed Central - PubMed

Affiliation: Cellular and Molecular Physiology Group, Biomedical Research Institute, Faculty of Medicine, Universidad Nacional de Colombia, Bogotá 111321, Colombia.

ABSTRACT
Several short linear peptides derived from cyclic bovine lactoferricin were synthesized and tested for their cytotoxic effect against the oral cavity squamous-cell carcinoma (OSCC) cell lines CAL27 and SCC15. As a control, an immortalized and nontumorigenic cell line, Het-1A, was used. Linear peptides based on the RRWQWR core sequence showed a moderate cytotoxic effect and specificity towards tumorigenic cells. A tetrameric peptide, LfcinB(20-25)4, containing the RRWQWR motif, exhibited greater cytotoxic activity (>90%) in both OSCC cell lines compared to the linear lactoferricin peptide or the lactoferrin protein. Additionally, this tetrameric peptide showed the highest specificity towards tumorigenic cells among the tested peptides. Interestingly, this effect was very fast, with cell shrinkage, severe damage to cell membrane permeability, and lysis within one hour of treatment. Our results are consistent with a necrotic effect rather than an apoptotic one and suggest that this tetrameric peptide could be considered as a new candidate for the therapeutic treatment of OSCC.

No MeSH data available.


Related in: MedlinePlus

Assessment of necrosis/apoptosis in nontumorigenic cell line Het-1A (a) and the tumorigenic cell line CAL27 (b). Cells were detached and incubated with different peptides indicated for 1 h, after which they were labeled with both Annexin V-FITC and PI, and analyzed by flow cytometry. 10 μM (4.66 μg/mL) STA and 0.2% T-X100 were used as controls. The maximum concentration of the peptides used was 100 μg/mL equivalent to LfcinB25, 32 μM; LfcinB(20–25), 101.5 μM; LfcinB-Pal, 67.3 μM; and LfcinB(20–25)4, 22.25 μM. Photomicrographs were taken with a phase-contrast microscope. Barr = 100 μm.
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fig7: Assessment of necrosis/apoptosis in nontumorigenic cell line Het-1A (a) and the tumorigenic cell line CAL27 (b). Cells were detached and incubated with different peptides indicated for 1 h, after which they were labeled with both Annexin V-FITC and PI, and analyzed by flow cytometry. 10 μM (4.66 μg/mL) STA and 0.2% T-X100 were used as controls. The maximum concentration of the peptides used was 100 μg/mL equivalent to LfcinB25, 32 μM; LfcinB(20–25), 101.5 μM; LfcinB-Pal, 67.3 μM; and LfcinB(20–25)4, 22.25 μM. Photomicrographs were taken with a phase-contrast microscope. Barr = 100 μm.

Mentions: In order to determine whether the rapid disruptive effect of the LfcinB(20–25)4 peptide is triggered by an apoptotic or necrotic process, the CAL27 tumorigenic and the Het-1A nontumorigenic cells were treated with the tetrameric peptide for 1 h, labeled with both Annexin V-FITC and PI, and analyzed by flow cytometry. The LfcinB(20–25)4-treated cells were significantly permeabilized (cellPI+/cellPI+/Annexin+) and an apoptotic cell population was not detected (cellPI−/Annexin+). A similar effect was found when the cells were treated with T-X100 (Figures 7(a) and  7(b)), indicating that the mechanism associated with the cytotoxic effect of the LfcinB(20–25)4 peptide is due to a necrotic event. Additionally, cells were treated with the caspase inhibitor Z-VAD-FMK and no differences were found (Supplementary Figure 3). These results showed that both LfcinB25 and LfcinB(20–25)4 have a cytotoxic effect possible due to a necrotic event (Figure 7). The necrotic damage was equivalent to the cytotoxicity found in the viability cell assays (Figure 2).


A Tetrameric Peptide Derived from Bovine Lactoferricin Exhibits Specific Cytotoxic Effects against Oral Squamous-Cell Carcinoma Cell Lines.

Solarte VA, Rosas JE, Rivera ZJ, Arango-Rodríguez ML, García JE, Vernot JP - Biomed Res Int (2015)

Assessment of necrosis/apoptosis in nontumorigenic cell line Het-1A (a) and the tumorigenic cell line CAL27 (b). Cells were detached and incubated with different peptides indicated for 1 h, after which they were labeled with both Annexin V-FITC and PI, and analyzed by flow cytometry. 10 μM (4.66 μg/mL) STA and 0.2% T-X100 were used as controls. The maximum concentration of the peptides used was 100 μg/mL equivalent to LfcinB25, 32 μM; LfcinB(20–25), 101.5 μM; LfcinB-Pal, 67.3 μM; and LfcinB(20–25)4, 22.25 μM. Photomicrographs were taken with a phase-contrast microscope. Barr = 100 μm.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4644816&req=5

fig7: Assessment of necrosis/apoptosis in nontumorigenic cell line Het-1A (a) and the tumorigenic cell line CAL27 (b). Cells were detached and incubated with different peptides indicated for 1 h, after which they were labeled with both Annexin V-FITC and PI, and analyzed by flow cytometry. 10 μM (4.66 μg/mL) STA and 0.2% T-X100 were used as controls. The maximum concentration of the peptides used was 100 μg/mL equivalent to LfcinB25, 32 μM; LfcinB(20–25), 101.5 μM; LfcinB-Pal, 67.3 μM; and LfcinB(20–25)4, 22.25 μM. Photomicrographs were taken with a phase-contrast microscope. Barr = 100 μm.
Mentions: In order to determine whether the rapid disruptive effect of the LfcinB(20–25)4 peptide is triggered by an apoptotic or necrotic process, the CAL27 tumorigenic and the Het-1A nontumorigenic cells were treated with the tetrameric peptide for 1 h, labeled with both Annexin V-FITC and PI, and analyzed by flow cytometry. The LfcinB(20–25)4-treated cells were significantly permeabilized (cellPI+/cellPI+/Annexin+) and an apoptotic cell population was not detected (cellPI−/Annexin+). A similar effect was found when the cells were treated with T-X100 (Figures 7(a) and  7(b)), indicating that the mechanism associated with the cytotoxic effect of the LfcinB(20–25)4 peptide is due to a necrotic event. Additionally, cells were treated with the caspase inhibitor Z-VAD-FMK and no differences were found (Supplementary Figure 3). These results showed that both LfcinB25 and LfcinB(20–25)4 have a cytotoxic effect possible due to a necrotic event (Figure 7). The necrotic damage was equivalent to the cytotoxicity found in the viability cell assays (Figure 2).

Bottom Line: Additionally, this tetrameric peptide showed the highest specificity towards tumorigenic cells among the tested peptides.Interestingly, this effect was very fast, with cell shrinkage, severe damage to cell membrane permeability, and lysis within one hour of treatment.Our results are consistent with a necrotic effect rather than an apoptotic one and suggest that this tetrameric peptide could be considered as a new candidate for the therapeutic treatment of OSCC.

View Article: PubMed Central - PubMed

Affiliation: Cellular and Molecular Physiology Group, Biomedical Research Institute, Faculty of Medicine, Universidad Nacional de Colombia, Bogotá 111321, Colombia.

ABSTRACT
Several short linear peptides derived from cyclic bovine lactoferricin were synthesized and tested for their cytotoxic effect against the oral cavity squamous-cell carcinoma (OSCC) cell lines CAL27 and SCC15. As a control, an immortalized and nontumorigenic cell line, Het-1A, was used. Linear peptides based on the RRWQWR core sequence showed a moderate cytotoxic effect and specificity towards tumorigenic cells. A tetrameric peptide, LfcinB(20-25)4, containing the RRWQWR motif, exhibited greater cytotoxic activity (>90%) in both OSCC cell lines compared to the linear lactoferricin peptide or the lactoferrin protein. Additionally, this tetrameric peptide showed the highest specificity towards tumorigenic cells among the tested peptides. Interestingly, this effect was very fast, with cell shrinkage, severe damage to cell membrane permeability, and lysis within one hour of treatment. Our results are consistent with a necrotic effect rather than an apoptotic one and suggest that this tetrameric peptide could be considered as a new candidate for the therapeutic treatment of OSCC.

No MeSH data available.


Related in: MedlinePlus