Quantitative Ultrastructural Morphometry and Gene Expression of mTOR-Related Mitochondriogenesis within Glioblastoma Cells

Rosangela Ferese, Paola Lenzi, Federica Fulceri, Francesca Biagioni, Cinzia Fabrizi, Stefano Gambardella, Pietro Familiari, Alessandro Frati, Fiona Limanaqi, Francesco Fornai

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Abstract

In glioblastoma (GBM) cells, an impairment of mitochondrial activity along with autophagy suppression occurs. Autophagy suppression in GBM promotes stemness, invasion, and poor prognosis. The autophagy deficit seems to be due, at least in part, to an abnormal up-regulation of the mammalian target of rapamycin (mTOR), which may be counteracted by pharmacological mTORC1 inhibition. Since autophagy activation is tightly bound to increased mitochondriogenesis, a defect in the synthesis of novel mitochondria is expected to occur in GBM cells. In an effort to measure a baseline deficit in mitochondria and promote mitochondriogenesis, the present study used two different GBM cell lines, both featuring mTOR hyperactivity. mTORC1 inhibition increases the expression of genes and proteins related to autophagy, mitophagy, and mitochondriogenesis. Autophagy activation was counted by RT-PCR of autophagy genes, LC3- immune-fluorescent puncta and immune-gold, as well as specific mitophagy-dependent BNIP3 stoichiometric increase in situ, within mitochondria. The activation of autophagy-related molecules and organelles after rapamycin exposure occurs concomitantly with progression of autophagosomes towards lysosomes. Remarkably, mitochondrial biogenesis and plasticity (increased mitochondrial number, integrity, and density as well as decreased mitochondrial area) was long- lasting for weeks following rapamycin withdrawal.

Original languageEnglish
Pages (from-to)4750
JournalInternational Journal of Molecular Sciences
Volume21
Issue number13
DOIs
Publication statusPublished - Jun 27 2020

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