Inheritance of one of three primary mutations at positions 11778, 3460 or 14484 of the mitochondrial genome in subunits of Complex I causes Leber's Hereditary Optic Neuropathy (LHON), a specific degeneration of the optic nerve, resulting in bilateral blindness. It has been unclear why inheritance of a systemic mitochondrial mutation would result in a specific neurodegeneration. To address the neuron-specific degenerative phenotype of the LHON genotype, we have created cybrids using a neuronal precursor cell line, Ntera 2/D1 (NT2), containing mitochondria from patient lymphoblasts bearing the most common LHON mutation (11778) and the most severe LHON mutation (3460). The undifferentiated LHON-NT2 mutant cells were not significantly different from the parental cell control in terms of mtDNA/nDNA ratio, mitochondrial membrane potential, reactive oxygen species (ROS) production or the ability to reduce Alamar Blue. Differentiation of NT2s resulted in a neuronal morphology and neuron-specific pattern of gene expression, and a 3-fold reduction in mtDNA/nDNA ratio in both mutant and control cells; however, the differentiation protocol yielded significantly less LHON cells than controls, by 30%, indicating either a decreased proliferative potential or increased cell death of the LHON-NT2 cells. Differentiation of the cells to the neuronal form also resulted in significant increases in ROS production in the LHON-NT2 neurons versus controls, which is abolished by rotenone, a specific inhibitor of Complex I. We infer that the LHON genotype requires a differentiated neuronal environment in order to induce increased mitochondrial ROS, which may be the cause of the reduced NT2 yield; and suggest that the LHON degenerative phenotype may be the result of an increase in mitochondrial superoxide which is caused by the LHON mutations, possibly mediated through neuron-specific alterations in Complex I structure.
|Original language||English (US)|
|Number of pages||8|
|Journal||Human Molecular Genetics|
|State||Published - Feb 15 2002|
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