Abstract
European mistletoe (Viscum album) is a medicinal plant with significant anticancer properties. In vitro callus production provides an essential alternative for biomass production and medicinal compounds propagation. In this study, we investigated (a) the biotechnological production of somaclonal proteins from mistletoe callus propagation and (b) the in vitro cytotoxic properties of these mistletoe protein extracts against cervical cancer. Mistletoe proteins (MPEs) were extracted from natural resources (plant leaves and stems) and in vitro propagated variant calluses. Potential cytotoxic effects of the extracted proteins on HeLa cells were assessed by the MTT proliferation assay, whereas oxidative damage and induction of apoptosis were measured fluorometrically by the reactive oxygen species (ROS) sensitive carboxy-H2-DCFDA method and caspase-3 activation assays, respectively. Electrophoretic results indicated differences in the protein bands between callus and donor plant tissues. Leaf and stem protein extracts (PEs) demonstrated a strong impact on HeLa cells viability by promoting oxidative damage and inducing apoptosis in contrast to callus PEs. This study shows that MPEs application in HeLa cells causes oxidative alterations and apoptosis through caspase-3 activation and increased intracellular ROS levels. These findings may be useful for uncovering novel bioactive products of pharmaceutical interest as well as for gaining an understanding of the biochemical pathways of mistletoe proteins in cancer treatment.
Key message
In spite of the low cytotoxicity observed for biotechnologically propagated somaclonal variant calluses, towards cervical cancer, the ability to establish biotechnological production of said mistletoe extracts is important because it allows for standardization of protein production.
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Abbreviations
- Ac-DEVD-CHO:
-
Acetyl-Asp-Glu-Val-Asp-al
- Ac-DEVD-pNA:
-
Acetyl-Asp-Glu-Val-Asp p-nitroanilide
- BAP:
-
Benzyladenine
- CHAPS:
-
3-[(3-Cholamidopropyl)dimethylammonio]-1-propanesulfonate hydrate
- DCF:
-
2,7-Dichlorofluorescein
- DMEM:
-
Dulbecco’s modified Eagle’s medium
- DOX:
-
Doxorubicin hydrochloride
- DMSO:
-
Dimethyl sulfoxide
- FBS:
-
Fetal bovine serum
- HEPES:
-
2-[4-(2-Hydroxyethyl)piperazin-1-yl]ethanesulfonic acid
- kDa:
-
Kilodaltons
- ML-I,-II,-III:
-
Mistletoe lectins I,-II,-III
- MPEs:
-
Mistletoe protein extracts
- MTT:
-
3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide
- NAA:
-
a-Naphthaleneacetic acid
- NAC:
-
N-acetyl l-cysteine
- PBS:
-
Phosphate-buffered saline
- ROS:
-
Reactive oxygen species
- SDS:
-
Sodium dodecyl sulfate
- SOD:
-
Superoxide dismutase
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Acknowledgements
We warmly acknowledge the support of Dr. Penelopi Bouzioti (Institute of Nuclear & Radiological Sciences & Technology, Energy & Safety, National Center for Scientific Research “Demokritos) for her critical review of our experiments and associated results.
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This work is supported by the Operational Program “Human Resources Development, Education and Lifelong Learning”, Priority Axes 6, 8, 9 co-financed by European Social Fund entitled “Support of Researchers with Emphasis on Senior Researchers” (NSRF 2014-2020).
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Methodology, SM, VT, M-AK, GM; investigation, SM, VT, M-AK, GM; data curation, SM, VT; writing—original draft preparation, SM, VT, SK, GM; writing—review and editing, SK; supervision, SK; project administration, GM.
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Communicated by Silvia Moreno.
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Mavrikou, S., Tsekouras, V., Karageorgou, MA. et al. Anticancer and biochemical effects of Viscum album L. protein extracts on HeLa cells. Plant Cell Tiss Organ Cult 140, 369–378 (2020). https://doi.org/10.1007/s11240-019-01733-0
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DOI: https://doi.org/10.1007/s11240-019-01733-0