Characterization and application of a disease-cell model for a neurodegenerative lysosomal disease

Ribbens, Jameson; Moser, Ann B.; Hubbard, Walter C.; Bongarzone, Ernesto R.; Maegawa, Gustavo

doi:10.1016/j.ymgme.2013.09.011

Cited by 28 publications

(42 citation statements)

References 51 publications

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“…Lipid extraction was performed by dispersing cell pellets in 500 μL of HPLC grade methanol containing 9.9 nM 1-acetyl psychosine (purity >98%; Matreya LLC), the internal standard as previously described (Ribbens et al, 2013). Following nitrogen evaporation, lipid film weight was measured using ultra-precision scale.…”

Section: Methodsmentioning

confidence: 99%

“…The flow rate was 0.75 mL/min. The samples were analyzed using a gradient elution, which is a modification to the previously reported method (Ribbens et al, 2013). Prior to injection of the sample, the column was equilibrated for 5 minutes with a mixture of 90% mobile phase A (MPA) as water:acetonitrile:formic acid (54.5:45:0.5, v/v/v), and 10% mobile phase B (MPB), comprised of acetonitrile:chloroform:formic acid (90:10:0.5, v/v/v).…”

Section: Methodsmentioning

confidence: 99%

“…After, acetonitrile:formic acid (99.5:0.1, v/v/v) and water:acetonitrile (50:50, v/v) served as the strong wash and weak wash, respectively. Multiple reactions monitoring (MRM) was used to identify the analytes including psychosine (MRM 462/264) and 1-acetyl psychosine (MRM 504/264) as previously described (Ribbens et al, 2013). The integrated areas of the analyte (psychosine) were analyzed using the Analyst® Software, version 1.5.1, (AB SCIEX) and extrapolated in a standard curve of psychosine obtained at concentrations of standard psychosine (Matreya LLC) from 0.025 – 5 ng/mL under exact same conditions.…”

Section: Methodsmentioning

confidence: 99%

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Early axonal loss accompanied by impaired endocytosis, abnormal axonal transport, and decreased microtubule stability occur in the model of Krabbe's disease

Teixeira¹,

Miranda

Sousa

et al. 2014

Neurobiology of Disease

109

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In Krabbe’s Disease (KD), a leukodystrophy caused by β-galactosylceramidase deficiency, demyelination and a myelin-independent axonopathy contribute to the severe neuropathology. Beyond axonopathy, we show that in Twitcher mice, a model of KD, a decreased number of axons both in the PNS and CNS, and of neurons in dorsal root ganglia (DRG), occurred before the onset of demyelination. Despite the early axonal loss, and although in vitro Twitcher neurites degenerated over time, Twitcher DRG neurons displayed an initial neurite overgrowth and, following sciatic nerve injury, Twitcher axons were regeneration-competent, at a timepoint where axonopathy was already ongoing. Psychosine, the toxic substrate that accumulates in KD, induced lipid raft clustering. At the mechanistic level, TrkA recruitment to lipid rafts was dysregulated in Twitcher neurons, and defective activation of the ERK1/2 and AKT pathways was identified. Besides defective recruitment of signaling molecules to lipid rafts, the early steps of endocytosis and the transport of endocytic and synaptic vesicles were impaired in Twitcher DRG neurons. Defects in axonal transport, specifically in the retrograde component, correlated with decreased levels of dynein, abnormal levels of post-translational tubulin modifications and decreased microtubule stability. The identification of the axonal defects that precede demyelination in KD, together with the finding that Twitcher axons are regeneration-competent when axonopathy is already installed, open new windows of action to effectively correct the neuropathology that characterizes this disorder.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Early axonal loss accompanied by impaired endocytosis, abnormal axonal transport, and decreased microtubule stability occur in the model of Krabbe's disease

Teixeira¹,

Miranda

Sousa

et al. 2014

Neurobiology of Disease

109

View full text Add to dashboard Cite

show abstract

“…This suggests that an inhibitor closer to the last step of galactosylceramide biosynthesis might have a more favorable outcome, i.e., less toxicity (Biswas et al, ; Hawkins‐Salsbury et al, ). Recently, an assay to measure psychosine levels in immortalized neuroglial cells from twitcher mice was developed (Ribbens et al, ). Such an assay could be helpful in a preliminary search for novel SRT compounds.…”

Section: Srt In Kd: L‐cycloserinementioning

confidence: 99%

Substrate reduction therapy for Krabbe's disease

Sands

Levine

2016

J of Neuroscience Research

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Krabbe's disease (KD) is a lysosomal storage disorder in which galactosylceramide, a major glycosphingolipid of myelin, and psychosine (galactose‐sphingosine) cannot be adequately metabolized because of a deficiency in galactosylceramidase. Substrate reduction therapy (SRT) has been tested in preclinical studies. The premise of SRT is to reduce the synthesis of substrates that are not adequately digested so that the substrate burden is lowered, resulting in less accumulation of unmetabolized material. SRT is used for Gaucher's disease, in which inhibitors of the terminal biosynthetic step are used. Unfortunately, an inhibitor for the final step of galactosylceramide biosynthesis, i.e., UDP glycosyltransferase 8 (a.k.a. UDP‐galactose ceramide galactosyltransferase), has not been found. Approaches that inhibit an earlier biosynthetic step or that lessen the substrate burden by other means, such as genetic manipulations, have been tested in the twitcher mouse model of KD. Either as a stand‐alone therapy or in combination with other approaches, SRT slowed the disease course, indicating that this approach has potential therapeutic value. For instance, in individuals with adult‐onset disease, SRT theoretically could lessen the production of substrates so that residual enzymatic activity could adequately manage the lower substrate burden. In more severe forms of disease, SRT theoretically could be part of a combination therapy. However, SRT has the potential to impair normal function by reducing the synthesis of galactosylceramide to levels that impede myelin function, or SRT could have other deleterious effects. Thus, multiple issues need to be resolved before this approach is ready for testing in humans. © 2016 Wiley Periodicals, Inc.

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“…They obtained brain samples from twitcher mice, the natural mouse model with GALC deficiency, and immortalized the primary neuroglial cultured cells with SV40 large T antigen, generating the 145M‐Twi and the 145C‐Wt cell lines from twitcher and control mice, respectively. Both cell lines expressed specific oligodendrocyte markers, including A2B5 and GalC (Ribbens et al, ). These established 145M‐Twi cells can be a powerful research tool for investigating the neurologically relevant pathways in KD.…”

Section: Kd and Inflammationmentioning

confidence: 99%

Krabbe's leukodystrophy: Approaches and models in vitro

Avola

Graziano

Pannuzzo

et al. 2016

J of Neuroscience Research

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This Review describes some in vitro approaches used to investigate the mechanisms involved in Krabbe's disease, with particular regard to the cellular systems employed to study processes of inflammation, apoptosis, and angiogenesis. The aim was to update the knowledge on the results obtained from in vitro models of this neurodegenerative disorder and provide stimuli for future research. For a long time, the nonavailability of established neural cells has limited the understanding of neuropathogenic mechanisms in Krabbe's leukodystrophy. More recently, the development of new Krabbe's disease cell models has allowed the identification of neurologically relevant pathogenic cascades, including the major role of elevated psychosine levels. Thus, direct and/or indirect roles of psychosine in the release of cytokines, reactive oxygen species, and nitric oxide and in the activation of kinases, caspases, and angiogenic factors results should be clearer. In parallel, it is now understood that the presence of globoid cells precedes oligodendrocyte apoptosis and demyelination. The information described here will help to continue the research on Krabbe's leukodystrophy and on potential new therapeutic approaches for this disease that even today, despite numerous attempts, is without cure. © 2016 Wiley Periodicals, Inc.

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Characterization and application of a disease-cell model for a neurodegenerative lysosomal disease

Cited by 28 publications

References 51 publications

Early axonal loss accompanied by impaired endocytosis, abnormal axonal transport, and decreased microtubule stability occur in the model of Krabbe's disease

Early axonal loss accompanied by impaired endocytosis, abnormal axonal transport, and decreased microtubule stability occur in the model of Krabbe's disease

Substrate reduction therapy for Krabbe's disease

Krabbe's leukodystrophy: Approaches and models in vitro

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