An induced pluripotent stem cell line (TRNDi001-D) from a Niemann-Pick disease type C1 (NPC1) patient carrying a homozygous p. I1061T (c. 3182T>C) mutation in the NPC1 gene.
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SUBMITTER: Li R
PROVIDER: S-EPMC7218921 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:BACKGROUND: Niemann-Pick type C disease (NPC) is a rare autosomal recessive lipid storage disease characterized by progressive neurodegeneration. As only a few studies have been conducted on the impact of NPC on sensory systems, we used a mutant mouse model (NPC1(-/-)) to examine the effects of this disorder to morphologically distinct regions of the olfactory system, namely the olfactory epithelium (OE) and olfactory bulb (OB). METHODOLOGY/PRINCIPAL FINDINGS: For structural and functional analysis immunohistochemistry, electron microscopy, western blotting, and electrophysiology have been applied. For histochemistry and western blotting, we used antibodies against a series of neuronal and glia marker proteins, as well as macrophage markers. NPC1(-/-) animals present myelin-like lysosomal deposits in virtually all types of cells of the peripheral and central olfactory system. Especially supporting cells of the OE and central glia cells are affected, resulting in pronounced astrocytosis and microgliosis in the OB and other olfactory cortices. Up-regulation of Galectin-3, Cathepsin D and GFAP in the cortical layers of the OB underlines the critical role and location of the OB as a possible entrance gate for noxious substances. Unmyelinated olfactory afferents of the lamina propria seem less affected than ensheathing cells. Supporting the structural findings, electro-olfactometry of the olfactory mucosa suggests that NPC1(-/-) animals exhibit olfactory and trigeminal deficits. CONCLUSIONS/SIGNIFICANCE: Our data demonstrate a pronounced neurodegeneration and glia activation in the olfactory system of NPC1(-/-), which is accompanied by sensory deficits.
| S-EPMC3877006 | biostudies-literature
Project description:BackgroundNiemann-Pick type C1 disease (NPC1) is a rare progressive neurodegenerative disorder caused by mutations in the NPC1 gene. In this lysosomal storage disorder the intracellular transport and sequestration of several lipids like cholesterol is severely impaired, resulting in an accumulation of lipids in late endosomes and lysosomes. The neurological manifestation of the disease is caused by dysfunction and cell death in the central nervous system. Several animal models were used to analyze the impaired pathways. However, the underlying pathogenic mechanisms are still not completely understood and the genetic variability in humans cannot be reflected in these models. Therefore, a human model using patient-specific induced pluripotent stem cells provides a promising approach.MethodsWe reprogrammed human fibroblasts from a NPC1 patient and a healthy control by retroviral transduction with Oct4, Klf4, Sox2 and c-Myc. The obtained human induced pluripotent stem cells (hiPSCs) were characterized by immunocytochemical analyses. Neural progenitor cells were generated and patch clamp recordings were performed for a functional analysis of derived neuronal cells. Filipin stainings and the Amplex Red assay were used to demonstrate and quantify cholesterol accumulation.ResultsThe hiPSCs expressed different stem cell markers, e.g. Nanog, Tra-1-81 and SSEA4. Using the embryoid body assay, the cells were differentiated in cells of all three germ layers and induced teratoma in immunodeficient mice, demonstrating their pluripotency. In addition, neural progenitor cells were derived and differentiated into functional neuronal cells. Patch clamp recordings revealed voltage dependent channels, spontaneous action potentials and postsynaptic currents. The accumulation of cholesterol in different tissues is the main hallmark of NPC1. In this study we found an accumulation of cholesterol in fibroblasts of a NPC1 patient, derived hiPSCs, and neural progenitor cells, but not in cells derived from fibroblasts of a healthy individual. These findings were quantified by the Amplex Red assay, demonstrating a significantly elevated cholesterol level in cells derived from fibroblasts of a NPC1 patient.ConclusionsWe generated a neuronal model based on induced pluripotent stem cells derived from patient fibroblasts, providing a human in vitro model to study the pathogenic mechanisms of NPC1 disease.
| S-EPMC3848807 | biostudies-literature
Project description:Niemann-Pick disease type C (NPC) is associated with mutations in NPC1 and NPC2, whose gene products are key players in the endosomal/lysosomal egress of low-density lipoprotein-derived cholesterol. NPC1 is also the intracellular receptor for Ebola virus (EBOV). Here, we present a 4.4 Å structure of full-length human NPC1 and a low-resolution reconstruction of NPC1 in complex with the cleaved glycoprotein (GPcl) of EBOV, both determined by single-particle electron cryomicroscopy. NPC1 contains 13 transmembrane segments (TMs) and three distinct lumenal domains A (also designated NTD), C, and I. TMs 2-13 exhibit a typical resistance-nodulation-cell division fold, among which TMs 3-7 constitute the sterol-sensing domain conserved in several proteins involved in cholesterol metabolism and signaling. A trimeric EBOV-GPcl binds to one NPC1 monomer through the domain C. Our structural and biochemical characterizations provide an important framework for mechanistic understanding of NPC1-mediated intracellular cholesterol trafficking and Ebola virus infection.
| S-EPMC7111323 | biostudies-literature
Project description:Niemann-Pick type C (NPC) disease is a fatal neurodegenerative disorder characterized by the accumulation of unesterified cholesterol in the late endosomal/lysosomal compartments. Mutations in the NPC1 protein are implicated in 95% of patients with NPC disease. The most prevalent mutation is the missense mutation I1061T that occurs in ∼ 15-20% of the disease alleles. In our study, an isobaric labeling-based quantitative analysis of proteome of NPC1(I1061T) primary fibroblasts when compared with wild-type cells identified 281 differentially expressed proteins based on stringent data analysis criteria. Gene ontology enrichment analysis revealed that these proteins play important roles in diverse cellular processes such as protein maturation, energy metabolism, metabolism of reactive oxygen species, antioxidant activity, steroid metabolism, lipid localization, and apoptosis. The relative expression level of a subset of differentially expressed proteins (TOR4A, DHCR24, CLGN, SOD2, CHORDC1, HSPB7, and GAA) was independently and successfully substantiated by Western blotting. We observed that treating NPC1(I1061T) cells with four classes of seven different compounds that are potential NPC drugs increased the expression level of SOD2 and DHCR24. We have also shown an abnormal accumulation of glycogen in NPC1(I1061T) fibroblasts possibly triggered by defective processing of lysosomal alpha-glucosidase. Our study provides a starting point for future more focused investigations to better understand the mechanisms by which the reported dysregulated proteins triggers the pathological cascade in NPC, and furthermore, their effect upon therapeutic interventions.
| S-EPMC4587331 | biostudies-literature
Project description:People homozygous for mutations in the Niemann-Pick type C1 (NPC1) gene have physiological defects, including excess accumulation of intracellular cholesterol and other lipids, that lead to drastic neural and liver degeneration. The NPC1 multipass transmembrane protein is resident in late endosomes and lysosomes, but its functions are unknown. We find that organelles containing functional NPC1-fluorescent protein fusions undergo dramatic movements, some in association with extending strands of endoplasmic reticulum. In NPC1 mutant cells the NPC1-bearing organelles that normally move at high speed between perinuclear regions and the periphery of the cell are largely absent. Pulse-chase experiments with dialkylindocarbocyanine low-density lipoprotein showed that NPC1 organelles function late in the endocytic pathway; NPC1 protein may aid the partitioning of endocytic and lysosomal compartments. The close connection between NPC1 and the drug U18666A, which causes NPC1-like organelle defects, was established by rescuing drug-treated cells with overproduced NPC1. U18666A inhibits outward movements of NPC1 organelles, trapping membranes and cholesterol in perinuclear organelles similar to those in NPC1 mutant cells, even when cells are grown in lipoprotein-depleted serum. We conclude that NPC1 protein promotes the creation and/or movement of particular late endosomes, which rapidly transport materials to and from the cell periphery.
| S-EPMC30967 | biostudies-literature
Project description:The Niemann-Pick type C1 (NPC1) protein regulates the transport of cholesterol and fatty acids from late endosomes/lysosomes and has a central role in maintaining lipid homeostasis. NPC1 loss-of-function mutations in humans cause NPC1 disease, a rare autosomal-recessive lipid-storage disorder characterized by progressive and lethal neurodegeneration, as well as liver and lung failure, due to cholesterol infiltration. In humans, genome-wide association studies and post-genome-wide association studies highlight the implication of common variants in NPC1 in adult-onset obesity, body fat mass, and type 2 diabetes. Heterozygous human carriers of rare loss-of-function coding variants in NPC1 display an increased risk of morbid adult obesity. These associations have been confirmed in mice models, showing an important interaction with high-fat diet. In this review, we describe the current state of knowledge for NPC1 variants in relationship to pleiotropic effects on metabolism. We provide evidence that NPC1 gene variations may predispose to common metabolic diseases by modulating steroid hormone synthesis and/or lipid homeostasis. We also propose several important directions of research to further define the complex roles of NPC1 in metabolism. This review emphasizes the contribution of NPC1 to obesity and its metabolic complications.
| S-EPMC5888214 | biostudies-literature
Project description:Niemann-Pick C (NPC) disease is due to loss of NPC1 or NPC2 protein function that is required for unesterified cholesterol transport from the endosomal/lysosomal compartment. Though lung involvement is a recognized characteristic of Niemann-Pick type C disease, the pathological features are not well understood. We investigated components of the surfactant system in both NPC1 mutant mice and felines and in NPC2 mutant mice near the end of their expected life span. Histological analysis of the NPC mutant mice demonstrated thickened septae and foamy macrophages/leukocytes. At the level of electron microscopy, NPC1-mutant type II cells had uncharacteristically larger lamellar bodies (LB, mean area 2-fold larger), while NPC2-mutant cells had predominantly smaller lamellar bodies (mean area 50% of normal) than wild type. Bronchoalveolar lavage from NPC1 and NPC2 mutant mice had an approx. 4-fold and 2.5-fold enrichment in phospholipid, respectively, and an approx. 9-fold and 35-fold enrichment in cholesterol, consistent with alveolar lipidosis. Phospholipid and cholesterol also were elevated in type II cell LBs and lung tissue while phospholipid degradation was reduced. Enrichment of surfactant protein-A in the lung and surfactant of the mutant mice was found. Immunocytochemical results showed that cholesterol accumulated in the LBs of the type II cells isolated from the affected mice. Alveolar macrophages from the NPC1 and NPC2 mutant mice were enlarged compared to those from wild type mice and were enriched in phospholipid and cholesterol. Pulmonary features of NPC1 mutant felines reflected the disease described in NPC1 mutant mice. Thus, with the exception of lamellar body size, the lung phenotype seen in the NPC1 and NPC2 mutant mice were similar. The lack of NPC1 and NPC2 proteins resulted in a disruption of the type II cell surfactant system contributing to pulmonary abnormalities.
| S-EPMC3699545 | biostudies-literature
Project description:Niemann-Pick C1 (NPC1) disease is a rare, neurodegenerative lysosomal cholesterol storage disorder, typified by progressive cognitive and motor function impairment. Affected individuals usually succumb to the disease in adolescence. 2-Hydroxypropyl-β-cyclodextrin (HP-β-CD) has emerged as a promising intervention that reduces lipid storage and prolongs survival in NPC1 disease animal models. A barrier to the development of HP-β-CD and other treatments for NPC disease has been the lack of validated biochemical measures to evaluate efficacy. Here we explored whether cholesterol homeostatic responses resulting from HP-β-CD-mediated redistribution of sequestered lysosomal cholesterol could provide biomarkers to monitor treatment. Upon direct CNS delivery of HP-β-CD, we found increases in plasma 24(S)-HC in two independent NPC1 disease animal models, findings that were confirmed in human NPC1 subjects receiving HP-β-CD. Since circulating 24(S)-HC is almost exclusively CNS-derived, the increase in plasma 24(S)-HC provides a peripheral, non-invasive measure of the CNS effect of HP-β-CD. Our findings suggest that plasma 24(S)-HC, along with the other cholesterol-derived markers examined in this study, can serve as biomarkers that will accelerate development of therapeutics for NPC1 disease.
| S-EPMC4204776 | biostudies-literature
Project description:Niemann-Pick disease type B (NPB) is a rare autosomal recessive lysosomal storage disease caused by mutations in the SMPD1 gene, which encodes for acid sphingomyelinase. A human induced pluripotent stem cell (iPSC) line was generated from dermal fibroblasts of a 1-year old male patient with NPB that has a heterozygous mutation of a p.L43_A44delLA of SMPD1 using non-integrating Sendai virus technique. This iPSC line offers a useful resource to study the disease pathophysiology and as a cell-based model for drug development to treat NPB.
| S-EPMC6643268 | biostudies-literature
Project description:Data presented in this article demonstrate the generation and characterization of two novel Niemann-Pick disease Type C1 (NPC1) patient-specific induced pluripotent stem cell (iPSC) lines, related to the research article Trilck et al. (Diversity of Glycosphingolipid GM2 and Cholesterol Accumulation in NPC1 Patient-Specific iPSC-Derived Neurons; Brain Res.; 2017; 1657:52-61. doi: 10.1016/j.brainres.2016.11.031). For reprogramming fibroblasts, carrying the novel homozygous mutation c.1180T>C and the prevalent homozygous mutation c.3182T>C, were used. Reprogramming into patient-specific iPSCs was induced by retroviral transduction of the transcription factors Sox2, Klf4, Oct4 and c-Myc, and confirmed according to their pluripotency. The iPSCs were subsequently differentiated into neural progenitor cells, which were terminally differentiated into functional neurons and glial cells. The generation of these cell lines provides further valuable tools to investigate pathogenic mechanism of NPC1 in human neuronal cells carrying different NPC1 mutations.
| S-EPMC5384887 | biostudies-literature