| 1. |
- Agholme, Lotta, et al.
(författare)
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Proteasome Inhibition Induces Stress Kinase Dependent Transport Deficits – Implications for Alzheimer’s Disease
- 2014
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Ingår i: Molecular and Cellular Neuroscience. - : Elsevier. - 1044-7431 .- 1095-9327. ; 58, s. 29-39
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Tidskriftsartikel (refereegranskat)abstract
- Alzheimer’s disease (AD) is characterized by accumulation of two misfolded and aggregated proteins, β-amyloid and hyperphosphorylated tau. Both cellular systems responsible for clearance of misfolded and aggregated proteins, the lysosomal and the proteasomal, have been shown to be malfunctioning in the aged brain and more so in AD patients. This malfunction could be the cause of β-amyloid and tau accumulation, eventually aggregating in plaques and tangles. We have investigated how decreased proteasome activity affects AD related pathophysiological changes of microtubule transport and stability, as well as tau phosphorylation. To do this, we used our recently developed neuronal model where human SH-SY5Y cells obtain neuronal morphology and function through differentiation. We found that exposure to low doses of the proteasome inhibitor MG-115 caused disturbed neuritic transport, together with microtubule destabilization and tau phosphorylation. Furthermore, reduced proteasome activity activated several kinases implicated in AD pathology, including JNK, c-Jun and ERK 1/2. Restoration of the microtubule transport was achieved by inhibiting ERK 1/2 activation, and simultaneous inhibition of both ERK 1/2 and c-Jun reversed the proteasome inhibition-induced tau phosphorylation. Taken together, this study suggests that a decrease in proteasome activity can, through activation of c-Jun and ERK 1/2, result in several events contributing to AD pathology. Restoring proteasome function or inhibiting ERK 1/2 and c-Jun could therefore be used as novel treatments against AD.
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| 2. |
- Agholme, Lotta, et al.
(författare)
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Low-dose γ-secretase inhibition increases secretion of Aβ peptides and intracellular oligomeric Aβ.
- 2017
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Ingår i: Molecular and cellular neurosciences. - : Elsevier BV. - 1095-9327 .- 1044-7431. ; 85, s. 211-219
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Tidskriftsartikel (refereegranskat)abstract
- γ-Secretase inhibitors have been considered promising drug candidates against Alzheimer's disease (AD) due to their ability to reduce amyloid-β (Aβ) production. However, clinical trials have been halted due to lack of clinical efficacy and/or side effects. Recent in vitro studies suggest that low doses of γ-secretase inhibitors may instead increase Aβ production. Using a stem cell-derived human model of cortical neurons and low doses of the γ-secretase inhibitor DAPT, the effects on a variety of Aβ peptides were studied using mass spectrometry. One major focus was to develop a novel method for specific detection of oligomeric Aβ (oAβ), and this was used to study the effects of low-dose γ-secretase inhibitor treatment on intracellular oAβ accumulation. Low-dose treatment (2 and 20nM) with DAPT increased the secretion of several Aβ peptides, especially Aβx-42. Furthermore, using the novel method for oAβ detection, we found that 2nM DAPT treatment of cortical neurons resulted in increased oAβ accumulation. Thus, low dose-treatment with DAPT causes both increased production of long, aggregation-prone Aβ peptides and accumulation of intracellular Aβ oligomers, both believed to contribute to AD pathology.
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| 3. |
- Agholme, Lotta, et al.
(författare)
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Getting rid of intracellular Aβ- loss of cellular degradation leads to transfer between connected neurons
- 2014
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Ingår i: Current pharmaceutical design. - : Bentham Science Publishers. - 1381-6128 .- 1873-4286. ; 20:15, s. 2458-2468
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Tidskriftsartikel (refereegranskat)abstract
- The sporadic, late onset form of Alzheimers disease (AD) shares pathological hallmarks with the familial form; however, no clear reason for increased beta-amyloid (A beta) generation has been found in the former. It has long been speculated that the late onset form of AD is caused by reduced degradation and/or clearance of A beta. Indeed, both intracellular degradation systems, the proteasomal and lysosomal systems, have been shown to be defective in AD. Reduced proteasome activity increases levels of intracellular and secreted A beta. Furthermore, accumulation of improperly degraded A beta in the lysosomes causes lysosomal disruption and cell death. We recently showed that oligomeric A beta can be transmitted from one neuron to another, which causes neurotoxicity. In both the donating and receiving cells, A beta accumulates in the endo-lysosomal compartment. It is possible that ineffective degradation of A beta causes its transfer to neighboring neurons, thereby spreading AD pathology. This review summarizes the data underlying the idea of reduced A beta clearance and subsequent A beta spread in AD, and also suggests new therapeutic methods, which are aimed at targeting the degradation systems and synaptic transfer. By enhancing degradation of intracellular accumulated A beta, it can be possible to remove it and avoid A beta-induced neurodegeneration without disturbing the endogenously important pool of secreted A beta. Additionally, drugs targeted to inhibit the spread of intracellular toxic A beta aggregates may also be useful in stopping the progression of pathology, without affecting the level of A beta that normally occurs in the brain.
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| 4. |
- Nazir, Faisal Hayat, et al.
(författare)
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Expression and secretion of synaptic proteins during stem cell differentiation to cortical neurons.
- 2018
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Ingår i: Neurochemistry international. - : Elsevier BV. - 1872-9754 .- 0197-0186. ; 121, s. 38-49
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Tidskriftsartikel (refereegranskat)abstract
- Synaptic function and neurotransmitter release are regulated by specific proteins. Cortical neuronal differentiation of human induced pluripotent stem cells (hiPSC) provides an experimental model to obtain more information about synaptic development and physiology in vitro. In this study, expression and secretion of the synaptic proteins, neurogranin (NRGN), growth-associated protein-43 (GAP-43), synaptosomal-associated protein-25 (SNAP-25) and synaptotagmin-1 (SYT-1) were analyzed during cortical neuronal differentiation. Protein levels were measured in cells, modeling fetal cortical development and in cell-conditioned media which was used as a model of cerebrospinal fluid (CSF), respectively. Human iPSC-derived cortical neurons were maintained over a period of at least 150 days, which encompasses the different stages of neuronal development. The differentiation was divided into the following stages: hiPSC, neuro-progenitors, immature and mature cortical neurons. We show that NRGN was first expressed and secreted by neuro-progenitors while the maximum was reached in mature cortical neurons. GAP-43 was expressed and secreted first by neuro-progenitors and its expression increased markedly in immature cortical neurons. SYT-1 was expressed and secreted already by hiPSC but its expression and secretion peaked in mature neurons. SNAP-25 was first detected in neuro-progenitors and the expression and secretion increased gradually during neuronal stages reaching a maximum in mature neurons. The sensitive analytical techniques used to monitor the secretion of these synaptic proteins during cortical development make these data unique, since the secretion of these synaptic proteins has not been investigated before in such experimental models. The secretory profile of synaptic proteins, together with low release of intracellular content, implies that mature neurons actively secrete these synaptic proteins that previously have been associated with neurodegenerative disorders, including Alzheimer's disease. These data support further studies of human neuronal and synaptic development in vitro, and would potentially shed light on the mechanisms underlying altered concentrations of the proteins in bio-fluids in neurodegenerative diseases.
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| 5. |
- Cicognola, Claudia, et al.
(författare)
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Tauopathy-Associated Tau Fragment Ending at Amino Acid 224 Is Generated by Calpain-2 Cleavage.
- 2020
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Ingår i: Journal of Alzheimer's disease : JAD. - 1875-8908. ; 74:4, s. 1143-1156
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Tidskriftsartikel (refereegranskat)abstract
- Tau aggregation in neurons and glial cells characterizes tauopathies as Alzheimer's disease (AD), progressive supranuclear palsy (PSP), and corticobasal degeneration (CBD). Tau proteolysis has been proposed as a trigger for tau aggregation and tau fragments have been observed in brain and cerebrospinal fluid (CSF). Our group identified a major tau cleavage at amino acid (aa) 224 in CSF; N-terminal tau fragments ending at aa 224 (N-224) were significantly increased in AD and lacked correlation to total tau (t-tau) and phosphorylated tau (p-tau) in PSP and CBD.Previous studies have shown cleavage from calpain proteases at sites adjacent to aa 224. Our aim was to investigate if calpain-1 or -2 could be responsible for cleavage at aa 224.Proteolytic activity of calpain-1, calpain-2, and brain protein extract was assessed on a custom tau peptide (aa 220-228), engineered with fluorescence resonance energy transfer (FRET) technology. Findings were confirmed with in-gel trypsination and mass spectrometry (MS) analysis of brain-derived bands with proteolytic activity on the FRET substrate. Finally, knock-down of the calpain-2 catalytic subunit gene (CAPN2) was performed in a neuroblastoma cell line (SH-SY5Y).Calpain-2 and brain protein extract, but not calpain-1, showed proteolytic activity on the FRET substrate. MS analysis of active gel bands revealed presence of calpain-2 subunits, but not calpain-1. Calpain-2 depletion and chemical inhibition suppressed proteolysis of the FRET substrate. CAPN2 knock-down caused a 76.4% reduction of N-224 tau in the cell-conditioned media.Further investigation of the calpain-2 pathway in the pathogenesis of tauopathies is encouraged.
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| 6. |
- Bergström, Petra, et al.
(författare)
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Herpes Simplex Virus 1 and 2 Infections during Differentiation of Human Cortical Neurons
- 2021
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Ingår i: Viruses-Basel. - : MDPI AG. - 1999-4915. ; 13:10
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Tidskriftsartikel (refereegranskat)abstract
- Herpes simplex virus 1 (HSV-1) and 2 (HSV-2) can infect the central nervous system (CNS) with dire consequences; in children and adults, HSV-1 may cause focal encephalitis, while HSV-2 causes meningitis. In neonates, both viruses can cause severe, disseminated CNS infections with high mortality rates. Here, we differentiated human induced pluripotent stem cells (iPSCs) towards cortical neurons for infection with clinical CNS strains of HSV-1 or HSV-2. Progenies from both viruses were produced at equal quantities in iPSCs, neuroprogenitors and cortical neurons. HSV-1 and HSV-2 decreased viability of neuroprogenitors by 36.0% and 57.6% (p < 0.0001), respectively, 48 h post-infection, while cortical neurons were resilient to infection by both viruses. However, in these functional neurons, both HSV-1 and HSV-2 decreased gene expression of two markers of synaptic activity, CAMK2B and ARC, and affected synaptic activity negatively in multielectrode array experiments. However, unaltered secretion levels of the neurodegeneration markers tau and NfL suggested intact axonal integrity. Viral replication of both viruses was found after six days, coinciding with 6-fold and 22-fold increase in gene expression of cellular RNA polymerase II by HSV-1 and HSV-2, respectively. Our results suggest a resilience of human cortical neurons relative to the replication of HSV-1 and HSV-2.
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| 7. |
- Bodda, C., et al.
(författare)
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HSV1 VP1-2 deubiquitinates STING to block type I interferon expression and promote brain infection
- 2020
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Ingår i: The Journal of experimental medicine. - : Rockefeller University Press. - 1540-9538 .- 0022-1007. ; 217:7
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Tidskriftsartikel (refereegranskat)abstract
- Herpes simplex virus (HSV) is the main cause of viral encephalitis in the Western world, and the type I interferon (IFN) system is important for antiviral control in the brain. Here, we have compared Ifnb induction in mixed murine brain cell cultures by a panel of HSV1 mutants, each devoid of one mechanism to counteract the IFN-stimulating cGAS-STING pathway. We found that a mutant lacking the deubiquitinase (DUB) activity of the VP1-2 protein induced particularly strong expression of Ifnb and IFN-stimulated genes. HSV1 ΔDUB also induced elevated IFN expression in murine and human microglia and exhibited reduced viral replication in the brain. This was associated with increased ubiquitination of STING and elevated phosphorylation of STING, TBK1, and IRF3. VP1-2 associated directly with STING, leading to its deubiquitination. Recruitment of VP1-2 to STING was dependent on K150 of STING, which was ubiquitinated by TRIM32. Thus, the DUB activity of HSV1 VP1-2 is a major viral immune-evasion mechanism in the brain. © 2020 Bodda et al.
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| 8. |
- Sofou, Kalliopi, et al.
(författare)
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Bi-allelic VPS16 variants limit HOPS/CORVET levels and cause a mucopolysaccharidosis-like disease.
- 2021
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Ingår i: EMBO molecular medicine. - : EMBO. - 1757-4684 .- 1757-4676. ; 13
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Tidskriftsartikel (refereegranskat)abstract
- Lysosomal storage diseases, including mucopolysaccharidoses, result from genetic defects that impair lysosomal catabolism. Here, we describe two patients from two independent families presenting with progressive psychomotor regression, delayed myelination, brain atrophy, neutropenia, skeletal abnormalities, and mucopolysaccharidosis-like dysmorphic features. Both patients were homozygous for the same intronic variant in VPS16, a gene encoding a subunit of the HOPS and CORVET complexes. The variant impaired normal mRNA splicing and led to an ~85% reduction in VPS16 protein levels in patient-derived fibroblasts. Levels of other HOPS/CORVET subunits, including VPS33A, were similarly reduced, but restored upon re-expression of VPS16. Patient-derived fibroblasts showed defects in the uptake and endosomal trafficking of transferrin as well as accumulation of autophagosomes and lysosomal compartments. Re-expression of VPS16 rescued the cellular phenotypes. Zebrafish with disrupted vps16 expression showed impaired development, reduced myelination, and a similar accumulation of lysosomes and autophagosomes in the brain, particularly in glia cells. This disorder resembles previously reported patients with mutations in VPS33A, thus expanding the family of mucopolysaccharidosis-like diseases that result from mutations in HOPS/CORVET subunits.
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| 9. |
- Mogren, Å, et al.
(författare)
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Orofacial function in children with Speech Sound Disorders (SSD) persisting after the age of six years
- 2020
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Ingår i: International Journal of Speech-Language Pathology. - : Informa UK Limited. - 1754-9515 .- 1754-9507. ; 22:5, s. 526-536
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Tidskriftsartikel (refereegranskat)abstract
- Purpose: The aim was to investigate, describe and analyse speech characteristics, intelligibility, orofacial function and co-existing neurodevelopmental symptoms in children with SSD of unknown origin, persisting after six years of age. Method: The study included 61 children with SSD (6–17 years) of unknown origin, referred for a speech and oral motor examination. The severity of SSD was estimated using Percentage Consonants Correct (PCC) and Percentage Vowels Correct (PVC) and assessments of resonance based on Swedish Articulation and Nasality Test (SVANTE). Orofacial function was screened using the Nordic Orofacial Test-Screening (NOT-S). Parents completed the Intelligibility in Context Scale (ICS) and a questionnaire including questions about heredity, medical and neurodevelopmental conditions, and speech development. Result: SSD varied according to PCC (8–95%) and PVC (55–100%) measurements. Percentages of co-occurring disorders included: 51% resonance deviations, 90% intelligibility issues, and 87% orofacial difficulties. The most affected orofacial domains were “Chewing and swallowing” (41%), “Masticatory muscles and jaw function” (38%) and “Sensory function” (38%). The majority (64%) had co-existing dysfunctions relating to general motor and neurodevelopmental disorders. Conclusion: Children with persistent SSD are at risk for orofacial dysfunction, general motor difficulties and other neurodevelopmental disorders and therefore should be screened for co-occurring disorders. © 2020, © 2020 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group.
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| 10. |
- Armstrong, Andrea, et al.
(författare)
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Lysosomal Network Proteins as Potential Novel CSF Biomarkers for Alzheimers Disease
- 2014
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Ingår i: Neuromolecular medicine. - : Humana Press. - 1535-1084 .- 1559-1174. ; 16:1, s. 150-160
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Tidskriftsartikel (refereegranskat)abstract
- The success of future intervention strategies for Alzheimers disease (AD) will likely rely on the development of treatments starting early in the disease course, before irreversible brain damage occurs. The pre-symptomatic stage of AD occurs at least one decade before the clinical onset, highlighting the need for validated biomarkers that reflect this early period. Reliable biomarkers for AD are also needed in research and clinics for diagnosis, patient stratification, clinical trials, monitoring of disease progression and the development of new treatments. Changes in the lysosomal network, i.e., the endosomal, lysosomal and autophagy systems, are among the first alterations observed in an AD brain. In this study, we performed a targeted search for lysosomal network proteins in human cerebrospinal fluid (CSF). Thirty-four proteins were investigated, and six of them, early endosomal antigen 1 (EEA1), lysosomal-associated membrane proteins 1 and 2 (LAMP-1, LAMP-2), microtubule-associated protein 1 light chain 3 (LC3), Rab3 and Rab7, were significantly increased in the CSF from AD patients compared with neurological controls. These results were confirmed in a validation cohort of CSF samples, and patients with no neurochemical evidence of AD, apart from increased total-tau, were found to have EEA1 levels corresponding to the increased total-tau levels. These findings indicate that increased levels of LAMP-1, LAMP-2, LC3, Rab3 and Rab7 in the CSF might be specific for AD, and increased EEA1 levels may be a sign of general neurodegeneration. These six lysosomal network proteins are potential AD biomarkers and may be used to investigate lysosomal involvement in AD pathogenesis.
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