Metabolic influence of walnut phenolic extract on mitochondria in a colon cancer stem cell model

Abstract

PurposeWalnut phenolic extract (WPE) reduces proliferation and enhances differentiation of colon cancer stem cells (CSCs). The present study investigated the metabolic influence of WPE on the mitochondrial function of colon CSCs to determine its underlying mechanism.MethodsCD133(+)CD44(+) HCT116 colon cancer cells were selected by fluorescence-activated cell sorting and were treated with or without 40 mu g/mL WPE. RNA-sequencing (RNA-Seq) was performed to identify differentially expressed genes (DEGs), which were further validated with RT-PCR. WPE-induced alterations in mitochondrial function were investigated through a mitochondrial stress test by determining cellular oxygen consumption rate (OCR), an indicator of mitochondrial respiration, and extracellular acidification rate (ECAR), an indicator of glycolysis, which were further confirmed by glucose uptake and lactate production tests.ResultsRNA-Seq analysis identified two major functional clusters: metabolic and mitochondrial clusters. WPE treatment shifted the metabolic profile of cells towards the glycolysis pathway (ECAR=36.98 mpH/min/ptn, p=0.02) and oxidative pathway (OCR=29.18pmol/min/ptn, p=0.00001). Serial mitochondrial stimulations using respiration modulators, oligomycin, carbonyl cyanide-4 (trifluoromethoxy) phenylhydrazone, and rotenone/antimycin A, found an increased potential of mitochondrial respiration (OCR=111.5pmol/min/ptn, p=0.0006). WPE treatment also increased glucose uptake (=0.39pmol/mu L, p=0.002) and lactate production (=0.08nmol/mu L, p=0.005).ConclusionsWPE treatment shifts the mitochondrial metabolism of colon CSC towards more aerobic glycolysis, which might be associated with the alterations in the characteristics of colon CSC.