In a recent study published in Scientific Reports, researchers investigated the mechanisms behind the anti-tumor activity of Chaga mushroom extracts in HSC-4 human oral cancer cells.
Study: Chaga mushroom extract suppresses oral cancer cell growth via inhibition of energy metabolism. Image Credit: Kyrylo Vasyliev/Shutterstock.com
Background
Oral cancer is a global health concern with limited treatment options owing to its side effects and consequences. Surgery, radiation therapy, and chemotherapy are the primary therapies, although they can harm healthy tissues, affect speech, and reduce quality of life.
Understanding and directing metabolic pathways in tumor cells provides a possible avenue for creating novel therapeutics. Chaga mushroom has anti-cancer properties across several cancer types; however, the mechanism is unclear.
About the study
In the present study, researchers examined whether Chaga mushrooms influence oral cancer development and metabolism.
Following treatment with the mushroom extract, the researchers examined cell survival, proliferative capability, glycolytic pathways, apoptosis, and mitochondrial respiratory mechanisms.
They treated HSC-4 cells with the mushroom extract at 0 µg/mL, 160 µg/mL, 200 µg/mL, 400 µg/mL, and 800.0 µg/mL doses for a day to assess its impact on oral malignancy cellular behavior, including cell cycle, proliferation, viability, mitochondrial respiration, apoptosis, and glycolysis.
The team analyzed the treated cells in terms of their cell cycle, using cell counting kit-8 (CCK-8) assays to determine cell viability.
To investigate whether the suppressive impact of Chaga mushrooms on tumor proliferation and survival in the treated cells included signal transducer and activator of transcription 3 (STAT3) signaling, they measured STAT3 activation after treatment with 200.0 µg/mL dose of the extract.
Further, they performed flow cytometry to analyze cell distribution and Western blotting to extract total cellular proteins.
The researchers used liquid chromatography-tandem mass spectrometry (LC-MS) to determine the components responsible for Chaga mushroom extract's anti-cancer properties.
They determined candidate compounds' concentrations using high-performance liquid chromatography with photodiode array detection (HPLC-DAD).
They investigated glycolysis regulation by the extracts among treated cells using extracellular acidification rate (ECAR) assays. They recorded real-time ECAR measurements in treated cells following glucose, oligomycin, and 2-deoxy-d-glucose (2-DG) injections.
The team investigated the activation of an energy sensor called adenosine monophosphate-activated protein kinase (AMPK) and the cells' oxygen consumption rate (OCR).
They also evaluated the impact of continuous energy deprivation on autophagy-associated apoptotic cell death in the treated cells.
They examined whether 200.0 µg/mL of the Chaga extract affected p38 mitogen-activated protein kinases (MAPKs)- and nuclear factor kappa B (NF-Κb)-stimulated apoptosis in the treated cells.
Results
The extract slowed HSC-4 cellular growth by inhibiting the cellular cycle and proliferation, reducing cancer cell energy consumption, and boosting autophagy-mediated cell death by apoptosis.
The extract considerably boosted oral cancer cell growth (G0/G1) phases while decreasing the synthesis (S) phase. Western blot examination revealed that the extract significantly reduced the expression of phospho-STAT3 after 15 minutes and maintained it for 120 minutes.
LC-MS identified three possible anti-cancer substances: 2-hydroxy-3,4-dimethoxybenzoic acid, syringic acid, and protocatechuic acid. The extract inhibited glycolysis, glycolytic capability, and glycolytic reserves in treated cells.
It also activated AMPK, promoting autophagy and inhibiting glycolytic pathways in the treated cells. Autophagy induction by the extract showed dose-dependent increases in mitochondrial basal respiratory rates and adenosine triphosphate (ATP) turnover.
However, there were no significant changes in maximal mitochondrial respiratory rates except at the highest concentration of the extract. Additionally, the researchers observed dose-dependent, significant reductions in mitochondrial spare respiration capacity.
The findings indicated that Chaga mushrooms reduce mitochondrial membrane potentials in the treated cells via continuous autophagy mediated by glycolysis inhibition, implying that mitochondrial malfunction causes apoptosis.
The extract-induced activation of NF-κB and p38 MAPKs increased apoptosis. The extract boosted the early apoptosis of the treated cells in a dosage-dependent fashion.
However, there was no significant difference in late apoptosis over 0-400 µg/mL extract concentration. High amounts of Chaga extract might impact other cell physiologies and decrease mitochondrial maximum respirations.
Conclusions
The researchers found that Chaga extract suppressed mitochondrial membrane potentials and glycolytic activity in the HSC-4 cell line, resulting in lower ATP levels and autophagy.
AMPK activation drove the effects, causing autophagy. STAT3 dephosphorylation inhibits the cellular cycle, stimulating apoptotic pathways via NF-κB and p38 MAPK activation.
Various cell-signaling mechanisms mediated the extract's inhibitory effects. The extract included three anti-cancer compounds: 2-hydroxy-3,4-dimethoxybenzoic acid, syringic acid, and protocatechuic acid.
While more pre-clinical research is needed to determine whether the extract suppresses tumor growth, the study findings imply that the mushroom extract has the potential to be a supplemental therapeutic option to treat oral cancer patients.
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