Mitochondrial abnormality in astrocytes leads to optic nerve degeneration: a mechanism mediated by CXCL14

Abstract

Purpose : Neuroinflammation is a common feature of most optic nerve (ON) disorders. To unravel the mechanisms of how astrocytes contribute to ON neuroinflammation, we have developed a rat model where two separate mutations act synergistically to produce severe ON degeneration. One mutation is in the Cryba1 gene that is crucial for lysosomal function and is expressed only in astrocytes in ON. The other mutation is in the Bckdk gene that is crucial for mitochondrial function and is expressed in all cell types. This study reveals the key role of mitophagy in astrocytes in the homeostasis of ON.

Methods : ONs were dissected from wild type, both single mutants, and double mutant (DM) rats at postnatal days 21 (P21) and P120. Primary astrocyte cultures from ONs were subjected to scRNA-seq, immunostaining, and blotting. Live cell imaging was used to measure mitochondrial membrane potential (MMP), mitochondrial reactive oxidative stress (mitoROS) and mitophagy flux. Urolithin A (UA) was used to induce mitophagy in DM astrocytes.

Results : Immunoblotting and staining revealed severe ON degeneration associated with inflammation in DM ONs compared to other genotypes at P120, but not at P21. RNA-seq data showed that CXCL14 was the most up-regulated gene in the ON of P120 DM rats. scRNA-seq analysis on mixed primary cultures of glial cells showed that astrocytes are the main source of CXCL14 production in DM ONs. Overexpression of CXCL14 in cultured DM astrocytes is associated with elevation of mitoROS and reduction of MMP. Moreover, the number of fragmented mitochondria (required for mitophagy) was significantly increased, but mitophagy flux was inhibited in DM astrocytes relative to other genotypes. Stimulation of mitophagy flux by UA in DM astrocytes led to significant reductions in AKT/mTORC1 activation, the number of fragmented mitochondria and the level of mitoROS generation. Interestingly, mitophagy induction by UA led to reduction of CXCL14 expression in DM astrocytes, indicating a role of dysfunctional mitochondria in CXCL14 expression.

Conclusions : Astrocytes are the only cells in DM ON where the function of both mitochondria and lysosomes is impaired. Defective mitophagy in DM astrocytes leads to accumulation of dysfunctional mitochondria, which in turn, promotes recruitment of immune cells into ON through upregulated CXCL14 expression.

This is a 2020 ARVO Annual Meeting abstract.