BACKGROUND INFORMATION: Cellular prion protein (PrPC ) is infamous for its role in prion diseases. The physiological function of PrPC remains enigmatic, but several studies point to its involvement in cell differentiation processes. To test this possibility, we monitored PrPC changes during the differentiation of prion-susceptible CAD 5 cells, and then we analysed the effect of PrPC ablation on the differentiation process. RESULTS: Neuronal CAD 5 cells differentiate within 5 days of serum withdrawal, with the majority of the cells developing long neurites. This process is accompanied by an up to sixfold increase in PrPC expression and enhanced N-terminal β-cleavage of the protein, which suggests a role for the PrPC in the differentiation process. Moreover, the majority of PrPC in differentiated cells is inside the cell, and a large proportion of the protein does not associate with membrane lipid rafts. In contrast, PrPC in proliferating cells is found mostly on the cytoplasmic membrane and is predominantly associated with lipid rafts. To determine the importance of PrPC in cell differentiation, a CAD 5 PrP-/- cell line with ablated PrPC expression was created using the CRISPR/Cas9 system. We observed no considerable difference in morphology, proliferation rate or expression of molecular markers between CAD 5 and CAD 5 PrP-/- cells during the differentiation initiated by serum withdrawal. CONCLUSIONS: PrPC characteristics, such as cell localisation, level of expression and posttranslational modifications, change during CAD 5 cell differentiation, but PrPC ablation does not change the course of the differentiation process. SIGNIFICANCE: Ablation of PrPC expression does not affect CAD 5 cell differentiation, although we observed many intriguing changes in PrPC features during the process. Our study does not support the concept that PrPC is important for neuronal cell differentiation, at least in simple in vitro conditions.

• MeSH
• buněčná diferenciace * MeSH
• buněčné linie MeSH
• membránové mikrodomény MeSH
• myši MeSH
• neurony cytologie   metabolismus   MeSH
• posttranslační úpravy proteinů MeSH
• priony metabolismus   MeSH
• PrPC proteiny metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

BACKGROUND: In this retrospective study, we analysed clinical, biochemical and molecular genetic data of 47 Czech patients with Single, Large-Scale Mitochondrial DNA Deletions (SLSMD). METHODS: The diagnosis was based on the long-range PCR (LX-PCR) screening of mtDNA isolated from muscle biopsy in 15 patients, and from the buccal swab, urinary epithelial cells and blood in 32 patients. RESULTS: A total of 57% patients manifested before the age of 16. We did not find any significant difference between paediatric and adult manifestation in either the proportion of patients that would develop extraocular symptoms, or the timespan of its progression. The survival rate in patients with Pearson Syndrome reached 60%. Altogether, five patients manifested with atypical phenotype not fulfilling the latest criteria for SLSMD. No correlation was found between the disease severity and all heteroplasmy levels, lengths of the deletion and respiratory chain activities in muscle. CONCLUSIONS: Paediatric manifestation of Progressive External Ophthalmoplegia (PEO) is not associated with a higher risk of multisystemic involvement. Contrary to PEO and Kearns-Sayre Syndrome Spectrum, Pearson Syndrome still contributes to a significant childhood mortality. SLSMD should be considered even in cases with atypical presentation. To successfully identify carriers of SLSMD, a repeated combined analysis of buccal swab and urinary epithelial cells is needed.

• Publikační typ
• časopisecké články MeSH

BACKGROUND: Maternally inherited complex I deficiencies due to mutations in MT-ND genes represent a heterogeneous group of multisystem mitochondrial disorders (MD) with a unfavourable prognosis. The aim of the study was to characterize the impact of the mutations in MT-ND genes, including the novel m.13091 T > C variant, on the course of the disease, and to analyse the activities of respiratory chain complexes, the amount of protein subunits, and the mitochondrial energy-generating system (MEGS) in available muscle biopsies and cultivated fibroblasts. METHODS: The respiratory chain complex activities were measured by spectrophotometry, MEGS were analysed using radiolabelled substrates, and protein amount by SDS-PAGE or BN-PAGE in muscle or fibroblasts. RESULTS: In our cohort of 106 unrelated families carrying different mtDNA mutations, we found heteroplasmic mutations in the genes MT-ND1, MT-ND3, and MT-ND5, including the novel variant m.13091 T > C, in 13 patients with MD from 12 families. First symptoms developed between early childhood and adolescence and progressed to multisystem disease with a phenotype of Leigh or MELAS syndromes. MRI revealed bilateral symmetrical involvement of deep grey matter typical of Leigh syndrome in 6 children, cortical/white matter stroke-like lesions suggesting MELAS syndrome in 3 patients, and a combination of cortico-subcortical lesions and grey matter involvement in 4 patients. MEGS indicated mitochondrial disturbances in all available muscle samples, as well as a significantly decreased oxidation of [1-14C] pyruvate in fibroblasts. Spectrophotometric analyses revealed a low activity of complex I and/or complex I + III in all muscle samples except one, but the activities in fibroblasts were mostly normal. No correlation was found between complex I activities and mtDNA mutation load, but higher levels of heteroplasmy were generally found in more severely affected patients. CONCLUSIONS: Maternally inherited complex I deficiencies were found in 11% of families with mitochondrial diseases in our region. Six patients manifested with Leigh, three with MELAS. The remaining four patients presented with an overlap between these two syndromes. MEGS, especially the oxidation of [1-14C] pyruvate in fibroblasts might serve as a sensitive indicator of functional impairment due to MT-ND mutations. Early onset of the disease and higher level of mtDNA heteroplasmy were associated with a worse prognosis.

• MeSH
• biopsie MeSH
• dítě MeSH
• dospělí MeSH
• fibroblasty metabolismus   MeSH
• kojenec MeSH
• kosterní svaly metabolismus   MeSH
• kultivované buňky MeSH
• Leighova nemoc genetika   MeSH
• lidé MeSH
• magnetická rezonanční tomografie MeSH
• mitochondriální DNA * MeSH
• mitochondriální nemoci genetika   MeSH
• mladiství MeSH
• mozek diagnostické zobrazování   patologie   MeSH
• mutace * MeSH
• novorozenec MeSH
• respirační komplex I nedostatek   genetika   metabolismus   MeSH
• syndrom MELAS genetika   MeSH
• věk při počátku nemoci MeSH
• Check Tag
• dítě MeSH
• dospělí MeSH
• kojenec MeSH
• lidé MeSH
• mladiství MeSH
• novorozenec MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH