Modifications of the mitochondrial functionality in colonospheres. Response to oxaliplatin.

Abstract

[eng] Colorrectal cancer (CRC) develops following a hierarchical model of heterogeneous cell populations with subpopulations of poorly-differentiated, self-renewables cancer stem cells (CSCs). Understanding this subset of cells may be crucial in CRC research, thus CSCs have been suggested as a source of cancer recurrence, drug resistance and metastasis. Culturing cells as spheroids (3D culture) may lead to an enrichment of CSCs, preserving more faithfully the features of original tumors, including gene expression and tumor biology. Given that one of the hallmarks of human cancer is the adaptive metabolic reprogramming, mitochondrial biogenesis, dynamics and mitophagy are crucial pathways in cancer development. In this regard, the aim of this study was to obtain SW620-derived spheroids (colonospheres) and to analyze their stemness-related, mitochondrial biogenesis, functionality and dynamic gene expression, compared to cells cultured in adherent conditions. Following this objective, cells were seeded in adherent plates with standard seeding medium (DMEM) and tumorsphere medium (3DTM), to obtain adherent cell groups and also in ultra-low attachment plates with 3DTM, to obtain primary colonospheres (CS1). RNA of the three groups was extracted and retrotranscribed to perform Real-time PCR. Colonosphere forming efficiency was also evaluated under Oxaliplatin (OXA) treatment, a commonly used drug in CRC. The obtained mRNA levels showed that CS1 increased the expression of the main modulators of mitochondrial biogenesis and mitophagy and also altered genes related to fission and fusion, in comparison with their adherent counterparts. These results suggest that in the colonosphere forming process, mitochondria suffer several damages and consequently there is an enhanced demand of mitochondrial mitophagy and biogenesis. Treatment with OXA at the seeding time, reduced the colonosphere forming efficiency at highest levels. On the contrary, when treatment was applied at 48h post-seeding, colonospheres grew more rounded and defined, thus suggesting that OXA may help in the spheroid forming process.