Mitochondria Transplantation Facilitates Neuronal Survival and Neurite Outgrowth - Implication for Glaucoma Therapy

Abstract

Purpose : Mitochondrial dysfunction is associated with neurodegenerative diseases, including glaucoma, a leading cause of irreversible blindness worldwide. It is thought to be a key contributing factor to retinal ganglion cell (RGC) loss in glaucoma. Meeting the energy needs of diseased RGCs by replenishing active mitochondrial levels through mitochondrial transplantation may hold a promise for neuroprotective and regenerative medicine. In our current research work, we tested the efficacy of transplanted mitochondria in facilitating neuronal survival and axonal outgrowth in cultured neurons.

Methods : Mitochondria homeostasis in RGCs isolated from 3-month and 12-month old mice were evaluated by quantitative polymerase chain reaction (qPCR) to assess the levels of mitochondria associated genes. PC12 cells (from ATCC) were induced to differentiate using nerve growth factor (NGF); mitochondrial function was measured using Seahorse Analytics. Mitochondria were isolated from PC12 cells, stained using MitoTracker™ Red, and transplanted to cultured PC12 cells using Mitoception. Endogenous mitochondria were stained using MitoTracker™ Green FM. PC12 cell neurite outgrowth was evaluated at day 6-10 post transplantation, using Leica fluorescent microscope and EVOS M7000 Imaging System.

Results : qPCR revealed a significant decrease in levels of PINK1 (gene involved in mitochondrial quality control) in aging RGCs, implicating mitochondrial homeostatic dysfunction in aging RGCs. A marked increase in mitochondrial function was observed following NGF treatment in PC12 cells, suggesting an active role of mitochondrial dynamics in neuronal differentiation and neurite outgrowth. Effective mitochondrial uptake by PC12 cells were validated using MitoTracker™ intensity analysis in cells. Significantly increased number and average length of neurites and sustained neurite outgrowth were observed in differentiating PC12 cells following mitochondria transplantation. Transplanted mitochondria were seen to transport along the neuronal processes all the way to the growth cones.

Conclusions : Our results point to a dynamic role of mitochondria in neuronal differentiation and neurite outgrowth. Mitochondrial transplantation can pave the way to design effective therapeutics to address glaucoma induced neuronal degeneration.

This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.