Disruption and therapeutic rescue of autophagy in a human neuronal model of Niemann Pick type C1

Abstract

An unresolved issue about many neurodegenerative diseases is why neurons are particularly sensitive to defects in ubiquitous cellular processes. One example is Niemann Pick type C1, caused by defects in cholesterol trafficking in all cells, but where neurons are preferentially damaged. Understanding this selective failure is limited by the difficulty in obtaining live human neurons from affected patients. To solve this problem, we generated neurons with decreased function of NPC1 from human embryonic stem cells and used them to test the hypothesis that defective cholesterol handling leads to enhanced pathological phenotypes in neurons. We found that human NPC1 neurons have strong spontaneous activation of autophagy, and, contrary to previous reports in patient fibroblasts, a block of autophagic progression leading to defective mitochondrial clearance. Mitochondrial fragmentation is an exceptionally severe phenotype in NPC1 neurons compared with fibroblasts, causing abnormal accumulation of mitochondrial proteins. Contrary to expectation, these abnormal phenotypes were rescued by treatment with the autophagy inhibitor 3-methyladenine and by treatment with the potential therapeutic cyclodextrin, which mobilizes cholesterol from the lysosomal compartment. Our findings suggest that neurons are especially sensitive to lysosomal cholesterol accumulation because of autophagy disruption and accumulation of fragmented mitochondria, thus defining a new route to effective drug development for NPC1 disease.

Emerging role of autophagy in pediatric neurodegenerative and neurometabolic diseases

“…In contrast to this notion, however, autophagy activation in NP-C was recently shown to impair both mitochondrial turnover 26 and lysosomal protease function. 27 Using human embryonic stem cell-derived neurons, it was shown that both induction of basal autophagy and impaired autophagic flux concomitantly occur with NPC1 knockdown and that this significantly compromises mitochondrial clearance.…”

“…27 Using human embryonic stem cell-derived neurons, it was shown that both induction of basal autophagy and impaired autophagic flux concomitantly occur with NPC1 knockdown and that this significantly compromises mitochondrial clearance. 26 Remarkably, these changes were partially reversible through the application of the autophagy inhibitor 3-methyladenine, which reduces autophagosome formation and, by treatment with cyclodextrin, a compound capable of mobilizing cholesterol from the lysosomal compartment. 26 Concurrent with the notion that NPC deficiency interferes with the completion of the autophagic cascade, recent findings demonstrate that the autophagic response increases delivery of cholesterol to the lysosome and thereby causes dysfunction of lysosomal proteases, which in turn could impair turnover of autolysosomes, therefore creating a pathological feedback loop that promotes induction of dysfunctional autophagy 27 (Figure 3).…”

“…26 Remarkably, these changes were partially reversible through the application of the autophagy inhibitor 3-methyladenine, which reduces autophagosome formation and, by treatment with cyclodextrin, a compound capable of mobilizing cholesterol from the lysosomal compartment. 26 Concurrent with the notion that NPC deficiency interferes with the completion of the autophagic cascade, recent findings demonstrate that the autophagic response increases delivery of cholesterol to the lysosome and thereby causes dysfunction of lysosomal proteases, which in turn could impair turnover of autolysosomes, therefore creating a pathological feedback loop that promotes induction of dysfunctional autophagy 27 (Figure 3). The cascade of deleterious events was again ameliorated with pharmacological inhibition of autophagy initiation, and vice versa, unesterified cholesterol significantly increased in NP-C fibroblasts following treatment with the mTOR-dependent autophagy inducer rapamycin.…”

Niemann-Pick C disease and mobilization of lysosomal cholesterol by cyclodextrin

“…Indeed, the cholesterol content of mitochondria isolated from brains is increased, not decreased, by NPC1 deficiency ( 40,41 ). Several properties of mitochondria such as ATP production, oxidative stress, and possibly mitophagy are altered by NPC1 defi ciency (41)(42)(43), probably because of the increase in mitochondrial cholesterol. In contrast to defi ciency of NPC1, however, the movement of endosomal cholesterol to mitochondria is disrupted by NPC2 defi ciency ( 44 ).…”

Cholesterol-Lowering Effect of Octaarginine-Appended β-Cyclodextrin in <i>Npc1</i>-Trap-CHO Cells

“…In addition, a basal autophagy plays a pivotal role in the constitutive turnover of cytoplasmic components for maintaining cellular function. [26][27][28][29] Indeed, the impairment of basal autophagy has been reported in various neurodegenerative diseases and lysosomal storage disorders including NPC. 30,31) Therefore, an amelioration of impaired autophagy is still a challenging issue for the treatment of NPC.…”