Abstract
Niemann-Pick type C (NPC) disease is an autosomal recessive lysosomal storage disease that is caused by a mutation of the NPC1 or NPC2 gene, in which un-esterified cholesterol and sphingolipids accumulate mainly in the liver, spleen, and brain. Abnormal lysosomal storage leads to cell damage, neurological problems, and premature death. The time of onset and severity of symptoms of NPC disease are highly variable. The molecular mechanisms that are responsible for NPC disease pathology are far from being understood. The present study generated and characterized a zebrafish mutant that lacks Npc2 protein that may be useful for studies at the organismal, cellular, and molecular levels and both small-scale and high-throughput screens. Using CRISPR/Cas9 technology, we knocked out the zebrafish homolog of NPC2. Five-day-old npc2 mutants were morphologically indistinguishable from wildtype larvae. We found that live npc2–/– larvae exhibited stronger Nile blue staining. The npc2–/– larvae exhibited low mobility and a high anxiety-related response. These behavioral changes correlated with downregulation of the mcu (mitochondrial calcium uniporter) gene, ppp3ca (calcineurin) gene, and genes that are involved in myelination (mbp and mpz). Histological analysis of adult npc2–/– zebrafish revealed that pathological changes in the nervous system, kidney, liver, and pancreas correlated with inflammatory responses (i.e., the upregulation of il1, nfκβ, and mpeg; i.e., hallmarks of NPC disease). These findings suggest that the npc2 mutant zebrafish may be a model of NPC disease.
Highlights
Cholesterol is a lipid-type organic molecule that is critical for life
The 2-month-old npc2−/− fish were smaller than their siblings, and the sex ratio of homozygotes was skewed toward males
We found that npc2−/− larvae show slightly increased expression level of becn1, which is in line with previous finding of Pacheco and colleagues who demonstrated that enhanced basal autophagy in NPC1 deficiency is mediated by increased expression of Beclin-1 (Pacheco et al, 2007)
Summary
Cholesterol is a lipid-type organic molecule that is critical for life. It builds, maintains, and modulates cell membranes, where it forms lipid rafts and is involved in endocytosis. Maintains, and modulates cell membranes, where it forms lipid rafts and is involved in endocytosis It serves as a precursor of the biosynthesis of essential biomolecules (e.g., vitamin D, sex hormones, and corticosteroids). In NPC disease, advanced hypotonia, locomotor dysfunction, ataxia, spasticity, dystonia, dysphagia, and dementia, together with developmental delay and regression, often develop in the first decade of life. These changes coincide with a lower volume of the cerebellum, hippocampus, basal ganglia, and thalamus (Geberhiwot et al, 2018). NPC1 disease has other hallmarks, including immune dysfunction (Baudry et al, 2003; Platt et al, 2016), early neuroinflammation, and microglia activation (Cologna et al, 2014; Kavetsky et al, 2019)
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