Hemocyanins from Helix and Rapana Snails Exhibit in Vitro Antitumor Effects in Human Colorectal Adenocarcinoma
Abstract
:1. Introduction
2. Materials and Methods
2.1. Materials
2.2. Isolation of the Hemocyanin and Isoforms from Snail Rapana Venosa
2.3. Isolation of the Native Hemocyanin and Isoforms from Snail H. Lucorum
2.4. Isolation of the Native Hemocyanin and Mucus from Snail H. Aspersa
2.5. Separation of H. Aspersa Hemocyanin Isoforms
2.6. Cell culture and Cell Viability
2.7. Fluorescent Microscopy
2.7.1. Acridine Orange/Ethidium Bromide Double Staining
2.7.2. DAPI Staining
2.8. Transmission Electron Microscopy (TEM)
2.9. Statistical Analysis
3. Results
3.1. Isolation of Bioactive Compounds from the Three Mollusk Species
3.2. Effects of the Isolated Bioactive Compounds on the Viability and Proliferative Activity of HT-29 Tumor Cells
3.3. Apotogenic Effects of the Isolated Bioactive Compounds
3.3.1. Vital Double Staining of HT-29 Tumor Cells with Acridine Orange/Ethidium Bromide Acridin
3.3.2. DAPI Staining of HT-29 Tumor Cells
3.4. Transmission Electron Microscopy
4. Discussion
5. Conclusions
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
Abbreviations
AO | Acridine orange |
ATCC | 4′,6-Diamidine-2′-phenylindole dihydrochloride |
DMEM | Dulbecco’s modified Eagle’s medium |
LD | Linear dichroism |
DMSO | Dimethyl sulfoxide |
EB | Ethidium bromide |
HaH | Helix aspersa hemocyanin |
HlH | Helix lucorum hemocyanin |
KLH | Keyhole limpet hemocyanin |
MTT | Methyl thiazol tetrazolium bromide |
PBS | Phosphate-buffered saline |
RvH | Rapana venosa hemocyanin |
TEM | Transmission electron microscopy |
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IC50 Values (µg/mL) | HT-29 | Balb/c 3T3 |
---|---|---|
HaH-total | ›1000 | 934.1 |
Subunit βc-HaH | 733.8 | ›1000 |
Subunits α-HaH | 235.3 | 514.6 |
HlH-total | ›1000 | ›1000 |
Subunit βc-HlH | ›1000 | ›1000 |
Subunits α-HlH | ›1000 | ›1000 |
Subunit RvH I | ›1000 | ›1000 |
Subunit RvH II | ›1000 | ›1000 |
Ha-mucus | 415.7 | 825.1 |
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Georgieva, A.; Todorova, K.; Iliev, I.; Dilcheva, V.; Vladov, I.; Petkova, S.; Toshkova, R.; Velkova, L.; Dolashki, A.; Dolashka, P. Hemocyanins from Helix and Rapana Snails Exhibit in Vitro Antitumor Effects in Human Colorectal Adenocarcinoma. Biomedicines 2020, 8, 194. https://doi.org/10.3390/biomedicines8070194
Georgieva A, Todorova K, Iliev I, Dilcheva V, Vladov I, Petkova S, Toshkova R, Velkova L, Dolashki A, Dolashka P. Hemocyanins from Helix and Rapana Snails Exhibit in Vitro Antitumor Effects in Human Colorectal Adenocarcinoma. Biomedicines. 2020; 8(7):194. https://doi.org/10.3390/biomedicines8070194
Chicago/Turabian StyleGeorgieva, Ani, Katerina Todorova, Ivan Iliev, Valeriya Dilcheva, Ivelin Vladov, Svetlozara Petkova, Reneta Toshkova, Lyudmila Velkova, Aleksandar Dolashki, and Pavlina Dolashka. 2020. "Hemocyanins from Helix and Rapana Snails Exhibit in Vitro Antitumor Effects in Human Colorectal Adenocarcinoma" Biomedicines 8, no. 7: 194. https://doi.org/10.3390/biomedicines8070194