Energy metabolism of two kinds of stromal fibroblasts differently fuel the development of oral squamous cell carcinoma

Abstract

Background: Cancer is primarily a metabolic disease involving disturbances in cellular energy metabolism. “Reverse Warburg Effect” in adjacent stromal fibroblasts can be induced to feed cancer cells. Objectives: Develop a novel co-culture system to compare energy metabolism between normal oral fibroblasts (NOFs) and cancer-associated fibroblasts (CAFs), study their pathophysiological significance in microenvironment of oral squamous cell carcinoma (OSCC). Methods: CAFs and NOFs were both monolayer-cultured and co-cultured, respectively with normal oral keratinocytes (NOKs) and OSCC. Stained and analysed them by luminescent reader, flow cytometry, confocal microscope and Western blot for adenosine triphosphate (ATP) production, mitochondrial volume, mitochondrial membrane potential (MMP), reactive oxygen species (ROS), lactate, caveolin-1 and monocarboxylate transporter 4 (MCT-4). Mean and standard deviation were plotted for 3 replicates from each condition. Findings: After co-cultured with OSCC, CAFs showed decreased ATP production, ROS, caveolin-1 and lower MMP, but mitochondrial volume and MCT-4 expression were increased. Interestingly, NOFs showed increased ATP production, ROS, lactate, MCT-4, but decreased MMP, mitochondrial volume and caveolin-1 after co-cultured with OSCC. Confocal work showed that mitochondrial transfer more easily happened from NOFs to OSCC. Conclusions: OSCC can induce “Reverse Warburg Effect” in both NOFs and CAFs, which present mitochondrial dysfunction and oxidative stress in different ways that fuel ATP production in OSCC. Interestingly, metabolic coupling and mitochondrial transfer occur between stromal fibroblasts and OSCC. As CAFs’ precursor, NOFs play a more significant role in invasive tumour front than CAFs, which dramatically increase aggressive behaviour of OSCC and associate with early tumour recurrence and metastasis.