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1.
J Med Genet ; 2019 Aug 22.
Artigo em Inglês | MEDLINE | ID: mdl-31439720

RESUMO

BACKGROUND: Autistic spectrum disorders (ASDs) with developmental delay and seizures are a genetically heterogeneous group of diseases caused by at least 700 different genes. Still, a number of cases remain genetically undiagnosed. OBJECTIVE: The objective of this study was to identify and characterise pathogenic variants in two individuals from unrelated families, both of whom presented a similar clinical phenotype that included an ASD, intellectual disability (ID) and seizures. METHODS: Whole-exome sequencing was used to identify pathogenic variants in the two individuals. Functional studies performed in the Drosophila melanogaster model was used to assess the protein function in vivo. RESULTS: Probands shared a heterozygous de novo secretory carrier membrane protein (SCAMP5) variant (NM_001178111.1:c.538G>T) resulting in a p.Gly180Trp missense variant. SCAMP5 belongs to a family of tetraspanin membrane proteins found in secretory and endocytic compartments of neuronal synapses. In the fly SCAMP orthologue, the p.Gly302Trp genotype corresponds to human p.Gly180Trp. Western blot analysis of proteins overexpressed in the Drosophila fat body showed strongly reduced levels of the SCAMP p.Gly302Trp protein compared with the wild-type protein, indicating that the mutant either reduced expression or increased turnover of the protein. The expression of the fly homologue of the human SCAMP5 p.Gly180Trp mutation caused similar eye and neuronal phenotypes as the expression of SCAMP RNAi, suggesting a dominant-negative effect. CONCLUSION: Our study identifies SCAMP5 deficiency as a cause for ASD and ID and underscores the importance of synaptic vesicular trafficking in neurodevelopmental disorders.

2.
Genet Med ; 2019 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-31363182

RESUMO

PURPOSE: Kabuki syndrome (KS) (OMIM 147920 and 300867) is a rare genetic disorder characterized by specific facial features, intellectual disability, and various malformations. Immunopathological manifestations seem prevalent and increase the morbimortality. To assess the frequency and severity of the manifestations, we measured the prevalence of immunopathological manifestations as well as genotype-phenotype correlations in KS individuals from a registry. METHODS: Data were for 177 KS individuals with KDM6A or KMT2D pathogenic variants. Questionnaires to clinicians were used to assess the presence of immunodeficiency and autoimmune diseases both on a clinical and biological basis. RESULTS: Overall, 44.1% (78/177) and 58.2% (46/79) of KS individuals exhibited infection susceptibility and hypogammaglobulinemia, respectively; 13.6% (24/177) had autoimmune disease (AID; 25.6% [11/43] in adults), 5.6% (10/177) with ≥2 AID manifestations. The most frequent AID manifestations were immune thrombocytopenic purpura (7.3% [13/177]) and autoimmune hemolytic anemia (4.0% [7/177]). Among nonhematological manifestations, vitiligo was frequent. Immune thrombocytopenic purpura was frequent with missense versus other types of variants (p = 0.027). CONCLUSION: The high prevalence of immunopathological manifestations in KS demonstrates the importance of systematic screening and efficient preventive management of these treatable and sometimes life-threatening conditions.

3.
Neurochem Res ; 44(8): 1986-1998, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31309393

RESUMO

Parkinson's disease (PD) is a progressive neurodegenerative disorder associated with oxidative stress. Therefore, finding new antioxidant sources might be beneficial for its treatment. Avocado Persea americana is a fruit widely cultivated in tropical and subtropical climates worldwide. Although avocado by-products in the form of peel, seed coat and seeds are currently of no commercial use, they constitute a natural source of bioactive compounds. Methanolic (80%) extract obtained from lyophilized ground peels, seed coats, and seeds of the avocado Hass, Fuerte, Reed and Colinred varieties were analyzed for their total phenolic content (TPC) and their correlations with antioxidant capacity (AC) were assessed by ABTS, FRAP, and ORAC assays. For all varieties, the var. Colinred peel shows the highest TPC and AC. Further analysis showed that the var. Colinred peel presented major phenolic compounds B-type procyanidins and epicatechin according to HPLC-MS. The antioxidant effect of peel extract was evaluated upon in vivo oxidative stress (OS) model. We show for the first time that the peel extract can protect and/or prevent transgenic parkinDrosophila melanogaster fly against paraquat-induced OS, movement impairment and lipid peroxidation, as model of PD. Our findings offer an exceptional opportunity to test natural disease-modifying substances from avocado's by-products.

4.
Am J Med Genet A ; 179(7): 1126-1138, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31058441

RESUMO

CHOPS syndrome is a multisystem disorder caused by missense mutations in AFF4. Previously, we reported three individuals whose primary phenotype included cognitive impairment and coarse facies, heart defects, obesity, pulmonary involvement, and short stature. This syndrome overlaps phenotypically with Cornelia de Lange syndrome, but presents distinct differences including facial features, pulmonary involvement, and obesity. Here, we provide clinical descriptions of an additional eight individuals with CHOPS syndrome, as well as neurocognitive analysis of three individuals. All 11 individuals presented with features reminiscent of Cornelia de Lange syndrome such as synophrys, upturned nasal tip, arched eyebrows, and long eyelashes. All 11 individuals had short stature and obesity. Congenital heart disease and pulmonary involvement were common, and those were seen in about 70% of individuals with CHOPS syndrome. Skeletal abnormalities are also common, and those include abnormal shape of vertebral bodies, hypoplastic long bones, and low bone mineral density. Our observation indicates that obesity, pulmonary involvement, skeletal findings are the most notable features distinguishing CHOPS syndrome from Cornelia de Lange syndrome. In fact, two out of eight of our newly identified patients were found to have AFF4 mutations by targeted AFF4 mutational analysis rather than exome sequencing. These phenotypic findings establish CHOPS syndrome as a distinct, clinically recognizable disorder. Additionally, we report three novel missense mutations causative for CHOPS syndrome that lie within the highly conserved, 14 amino acid sequence of the ALF homology domain of the AFF4 gene, emphasizing the critical functional role of this region in human development.

5.
Int J Biol Macromol ; 134: 1052-1062, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31129208

RESUMO

The effect of Micrurus mipartitus snake venom as a therapeutic alternative for T-acute lymphoblastic leukemia (ALL) is still unknown. This study was aimed to evaluate the cytotoxic effect of M. mipartitus snake venom and a new L-amino acid oxidase (LAAO), named MipLAAO, on human peripheral blood lymphocytes (PBL) and on T-ALL cells (Jurkat), and its mechanism of action. PBL and Jurkat cells were treated with venom and MipLAAO, and morphological changes in the cell nucleus/DNA, mitochondrial membrane potential, levels of intracellular reactive oxygen species and cellular apoptosis markers were determined by fluorescence microscopy, flow cytometry and pharmacological inhibition. Venom and MipLAAO induced apoptotic cell death in Jurkat cells, but not in PBL, in a dose-response manner. Additionally, venom and MipLAAO increased dichlorofluorescein fluorescence intensity, indicative of H2O2 production, increased DJ-1 Cys106-sulfonate, as a marker of intracellular stress and induced the up-regulation of PUMA, p53 and phosphorylation of c-JUN. Additionally, it increased the expression of apoptotic CASPASE-3. In conclusion, M. mipartitus venom and MipLAAO selectively induces apoptosis in Jurkat cells through a H2O2-mediated signaling pathway dependent mostly on CASPASE-3 pathway. Our findings support the potential use of M. mipartitus snake venom compounds as a potential treatment for T-ALL.

7.
Neurosci Lett ; 703: 111-118, 2019 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-30904577

RESUMO

Alzheimer's disease (AD) is a progressive, degenerative disorder that mainly results in memory loss and a cognitive disorder. Although the cause of AD is still unknown, a minor percentage of AD cases are produced by genetic mutations in the presenilin-1 (PSEN1) gene. Differentiated neuronal cells derived from induced pluripotent stem cells (iPSCs) of patients can recapitulate key pathological features of AD in vitro; however, iPSCs studies focused on the p.E280 A mutation, which afflicts the largest family in the world with familial AD, have not been carried out yet. Although a link between the loss of the Y (LOY) chromosome in peripheral blood cells and risk for AD has been reported, LOY-associated phenotype has not been previously studied in PSEN1 E280 A carriers. Here, we report the reprogramming of fibroblast cells into iPSCs from a familial AD patient with the PSEN1 E280 A mutation, followed by neuronal differentiation into neural precursor cells (NPCs), and the differentiation of NPCs into differentiated neurons that lacked a Y chromosome. Although the PSEN1 E280 A iPSCs and NPCs were successfully obtained, after 8 days of differentiation, PSEN1 E280 A differentiated neurons massively died reflected by release and/ or activation of death markers, and failed to reach complete neural differentiation compared to PSEN 1 wild type cells.

8.
Parkinsonism Relat Disord ; 63: 204-208, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30765263

RESUMO

BACKGROUND: Mutations in the glucocerebrosidase (GBA) gene are an important risk factor for Parkinson's disease (PD). However, most GBA genetic studies in PD have been performed in patients of European origin and very few data are available in other populations. METHODS: We sequenced the entire GBA coding region in 602 PD patients and 319 controls from Colombia and Peru enrolled as part of the Latin American Research Consortium on the Genetics of Parkinson's disease (LARGE-PD). RESULTS: We observed a significantly higher proportion of GBA mutation carriers in patients compared to healthy controls (5.5% vs 1.6%; OR = 4.3, p = 0.004). Interestingly, the frequency of mutations in Colombian patients (9.9%) was more than two-fold greater than in Peruvian patients (4.2%) and other European-derived populations reported in the literature (4-5%). This was primarily due to the presence of a population-specific mutation (p.K198E) found only in the Colombian cohort. We also observed that the age at onset was significantly earlier in GBA carriers when compared to non-carriers (47.1 ±â€¯14.2 y vs. 55.9 ±â€¯14.2 y; p = 0.0004). CONCLUSION: These findings suggest that GBA mutations are strongly associated with PD risk and earlier age at onset in Peru and Colombia. The high frequency of GBA carriers among Colombian PD patients (∼10%) makes this population especially well-suited for novel therapeutic approaches that target GBA-related PD.

10.
Am J Hum Genet ; 104(2): 319-330, 2019 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-30639322

RESUMO

ZMIZ1 is a coactivator of several transcription factors, including p53, the androgen receptor, and NOTCH1. Here, we report 19 subjects with intellectual disability and developmental delay carrying variants in ZMIZ1. The associated features include growth failure, feeding difficulties, microcephaly, facial dysmorphism, and various other congenital malformations. Of these 19, 14 unrelated subjects carried de novo heterozygous single-nucleotide variants (SNVs) or single-base insertions/deletions, 3 siblings harbored a heterozygous single-base insertion, and 2 subjects had a balanced translocation disrupting ZMIZ1 or involving a regulatory region of ZMIZ1. In total, we identified 13 point mutations that affect key protein regions, including a SUMO acceptor site, a central disordered alanine-rich motif, a proline-rich domain, and a transactivation domain. All identified variants were absent from all available exome and genome databases. In vitro, ZMIZ1 showed impaired coactivation of the androgen receptor. In vivo, overexpression of ZMIZ1 mutant alleles in developing mouse brains using in utero electroporation resulted in abnormal pyramidal neuron morphology, polarization, and positioning, underscoring the importance of ZMIZ1 in neural development and supporting mutations in ZMIZ1 as the cause of a rare neurodevelopmental syndrome.

11.
Am J Hum Genet ; 104(1): 139-156, 2019 Jan 03.
Artigo em Inglês | MEDLINE | ID: mdl-30595372

RESUMO

Type 2A protein phosphatases (PP2As) are highly expressed in the brain and regulate neuronal signaling by catalyzing phospho-Ser/Thr dephosphorylations in diverse substrates. PP2A holoenzymes comprise catalytic C-, scaffolding A-, and regulatory B-type subunits, which determine substrate specificity and physiological function. Interestingly, de novo mutations in genes encoding A- and B-type subunits have recently been implicated in intellectual disability (ID) and developmental delay (DD). We now report 16 individuals with mild to profound ID and DD and a de novo mutation in PPP2CA, encoding the catalytic Cα subunit. Other frequently observed features were severe language delay (71%), hypotonia (69%), epilepsy (63%), and brain abnormalities such as ventriculomegaly and a small corpus callosum (67%). Behavioral problems, including autism spectrum disorders, were reported in 47% of individuals, and three individuals had a congenital heart defect. PPP2CA de novo mutations included a partial gene deletion, a frameshift, three nonsense mutations, a single amino acid duplication, a recurrent mutation, and eight non-recurrent missense mutations. Functional studies showed complete PP2A dysfunction in four individuals with seemingly milder ID, hinting at haploinsufficiency. Ten other individuals showed mutation-specific biochemical distortions, including poor expression, altered binding to the A subunit and specific B-type subunits, and impaired phosphatase activity and C-terminal methylation. Four were suspected to have a dominant-negative mechanism, which correlated with severe ID. Two missense variants affecting the same residue largely behaved as wild-type in our functional assays. Overall, we found that pathogenic PPP2CA variants impair PP2A-B56(δ) functionality, suggesting that PP2A-related neurodevelopmental disorders constitute functionally converging ID syndromes.

12.
Hum Mol Genet ; 28(9): 1445-1462, 2019 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-30566640

RESUMO

Mitochondria contain a dedicated translation system, which is responsible for the intramitochondrial synthesis of 13 mitochondrial DNA (mtDNA)-encoded polypeptides essential for the biogenesis of oxidative phosphorylation (OXPHOS) complexes I and III-V. Mutations in nuclear genes encoding factors involved in mitochondrial translation result in isolated or multiple OXPHOS deficiencies and mitochondrial disease. Here, we report the identification of disease-causing variants in the MRPS28 gene, encoding the small mitoribosomal subunit (mtSSU) protein bS1m in a patient with intrauterine growth retardation, craniofacial dysmorphism and developmental delay. Whole exome sequencing helped identify a seemingly homozygous missense variant NM_014018.2:c.356A>G, p.(Lys119Arg) which affected a highly conserved lysine residue. The variant was present in the mother in a heterozygous state, but not in the father who likely carried a large deletion spanning exon 2 and parts of introns 1 and 2 that could account for the apparent homozygosity of the patient. Polymerase chain reaction (PCR) amplification and Sanger sequencing of MRPS28 cDNA from patient fibroblasts revealed the presence of a truncated MRPS28 transcript, which lacked exon 2. Molecular and biochemical characterization of patient fibroblasts revealed a decrease in the abundance of the bS1m protein, decreased abundance of assembled mtSSU and inhibited mitochondrial translation. Consequently, OXPHOS biogenesis and cellular respiration were compromised in these cells. Expression of wild-type MRPS28 restored mitoribosomal assembly, mitochondrial translation and OXPHOS biogenesis, thereby demonstrating the deleterious nature of the identified MRPS28 variants. Thus, MRPS28 joins the increasing number of nuclear genes encoding mitoribosomal structural proteins linked to mitochondrial disease.

13.
Neuroimage Clin ; 2018 Nov 19.
Artigo em Inglês | MEDLINE | ID: mdl-30497982

RESUMO

Kabuki syndrome (KS) is a rare congenital disorder (1/32000 births) characterized by distinctive facial features, intellectual disability, short stature, and dermatoglyphic and skeletal abnormalities. In the last decade, mutations in KMT2D and KDM6A were identified as a major cause of kabuki syndrome. Although genetic abnormalities have been highlighted in KS, brain abnormalities have been little explored. Here, we have investigated brain abnormalities in 6 patients with KS (4 males; Mage = 10.96 years, SD = 2.97 years) with KMT2D mutation in comparison with 26 healthy controls (17 males; Mage = 10.31 years, SD = 2.96 years). We have used MRI to explore anatomical and functional brain abnormalities in patients with KS. Anatomical abnormalities in grey matter volume were assessed by cortical and subcortical analyses. Functional abnormalities were assessed by comparing rest cerebral blood flow measured with arterial spin labeling-MRI. When compared to healthy controls, KS patients had anatomical alterations characterized by grey matter decrease localized in the bilateral precentral gyrus and middle frontal gyrus. In addition, KS patients also presented functional alterations characterized by cerebral blood flow decrease in the left precentral gyrus and middle frontal gyrus. Moreover, subcortical analyses revealed significantly decreased grey matter volume in the bilateral hippocampus and dentate gyrus in patients with KS. Our results strongly indicate anatomical and functional brain abnormalities in KS. They suggest a possible neural basis of the cognitive symptoms observed in KS, such as fine motor impairment, and indicate the need to further explore the consequences of such brain abnormalities in this disorder. Finally, our results encourage further imaging-genetics studies investigating the link between genetics, anatomical and functional brain alterations in KS.

14.
PLoS Genet ; 14(11): e1007671, 2018 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-30500825

RESUMO

Mutations that alter signaling of RAS/MAPK-family proteins give rise to a group of Mendelian diseases known as RASopathies. However, among RASopathies, the matrix of genotype-phenotype relationships is still incomplete, in part because there are many RAS-related proteins and in part because the phenotypic consequences may be variable and/or pleiotropic. Here, we describe a cohort of ten cases, drawn from six clinical sites and over 16,000 sequenced probands, with de novo protein-altering variation in RALA, a RAS-like small GTPase. All probands present with speech and motor delays, and most have intellectual disability, low weight, short stature, and facial dysmorphism. The observed rate of de novo RALA variants in affected probands is significantly higher (p = 4.93 x 10-11) than expected from the estimated random mutation rate. Further, all de novo variants described here affect residues within the GTP/GDP-binding region of RALA; in fact, six alleles arose at only two codons, Val25 and Lys128. The affected residues are highly conserved across both RAL- and RAS-family genes, are devoid of variation in large human population datasets, and several are homologous to positions at which disease-associated variants have been observed in other GTPase genes. We directly assayed GTP hydrolysis and RALA effector-protein binding of the observed variants, and found that all but one tested variant significantly reduced both activities compared to wild-type. The one exception, S157A, reduced GTP hydrolysis but significantly increased RALA-effector binding, an observation similar to that seen for oncogenic RAS variants. These results show the power of data sharing for the interpretation and analysis of rare variation, expand the spectrum of molecular causes of developmental disability to include RALA, and provide additional insight into the pathogenesis of human disease caused by mutations in small GTPases.

15.
J Neurosci Methods ; 312: 126-138, 2018 Nov 22.
Artigo em Inglês | MEDLINE | ID: mdl-30472070

RESUMO

Barckground Alzheimer's disease (AD) is mainly caused by cellular loss and dysfunction of the basal forebrain cholinergic neurons and cholinergic axons in the cortex leading to slowly progressive decline in learning and memory performance. Unfortunately, no definitive treatment to halt neural cell loss exists to date. Therefore, it is necessary to obtain an unlimited source of cholinergic neurons for future pharmacological applications in AD. Human mesenchymal stromal cells (hMSCs) represent a unique source of cholinergic-like neurons (ChLNs). New method hWJ-MSCs were incubated with Cholinergic-N-Run medium for 4 and 7 days. Results hWJ-MSCs cultured with Cholinergic-N-Run medium differentiated into ChLNs in 4 days as evidenced by high levels of protein expression of the neuronal markers ChAT, VAChT, AChE, MAP2, ß-Tubulin III, NeuN, TUC-4, NF-L and no expression of the immature marker SOX2, the dopaminergic marker TH, GABAergic marker GAD67 and glial marker GFAP. Comparison with existing method(s) The hWJ-MSCs form ChLNs (e.g., ∼26% IF+) within 20 days by using complex conditioned mediums that are expensive and time-consuming. We report for the first time, to our best knowledge, a direct method of hWJ-MSCs transdifferentiation into ChLNs (∼76% ChAT /VAChT assessed by immunofluorescence microscopy and flow cytometry) in an economic, efficient and timely fashion. Conclusions The fastest method to obtain ChLNs from hWJ-MSCs takes only four days using the one-step incubation medium Cholinergic-N-Run.

17.
Cell ; 175(2): 488-501.e22, 2018 Oct 04.
Artigo em Inglês | MEDLINE | ID: mdl-30270045

RESUMO

Detection of viruses by innate immune sensors induces protective antiviral immunity. The viral DNA sensor cyclic GMP-AMP synthase (cGAS) is necessary for detection of HIV by human dendritic cells and macrophages. However, synthesis of HIV DNA during infection is not sufficient for immune activation. The capsid protein, which associates with viral DNA, has a pivotal role in enabling cGAS-mediated immune activation. We now find that NONO is an essential sensor of the HIV capsid in the nucleus. NONO protein directly binds capsid with higher affinity for weakly pathogenic HIV-2 than highly pathogenic HIV-1. Upon infection, NONO is essential for cGAS activation by HIV and cGAS association with HIV DNA in the nucleus. NONO recognizes a conserved region in HIV capsid with limited tolerance for escape mutations. Detection of nuclear viral capsid by NONO to promote DNA sensing by cGAS reveals an innate strategy to achieve distinction of viruses from self in the nucleus.

18.
Hum Mutat ; 39(12): 2047-2059, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30252186

RESUMO

Aminoacyl-tRNA synthetases are ubiquitous enzymes, which universally charge tRNAs with their cognate amino acids for use in cytosolic or organellar translation. In humans, mutations in mitochondrial tRNA synthetases have been linked to different tissue-specific pathologies. Mutations in the KARS gene, which encodes both the cytosolic and mitochondrial isoform of lysyl-tRNA synthetase, cause predominantly neurological diseases that often involve deafness, but have also been linked to cardiomyopathy, developmental delay, and lactic acidosis. Using whole exome sequencing, we identified two compound heterozygous mutations, NM_001130089.1:c.683C>T p.(Pro228Leu) and NM_001130089.1:c.1438del p.(Leu480TrpfsX3), in a patient presenting with sensorineural deafness, developmental delay, hypotonia, and lactic acidosis. Nonsense-mediated mRNA decay eliminated the truncated mRNA transcript, rendering the patient hemizygous for the missense mutation. The c.683C>T mutation was previously described, but its pathogenicity remained unexamined. Molecular characterization of patient fibroblasts revealed a multiple oxidative phosphorylation deficiency due to impaired mitochondrial translation, but no evidence of inhibition of cytosolic translation. Reintroduction of wild-type mitochondrial KARS, but not the cytosolic isoform, rescued this phenotype confirming the disease-causing nature of p.(Pro228Leu) exchange and demonstrating the mitochondrial etiology of the disease. We propose that mitochondrial translation deficiency is the probable disease culprit in this and possibly other patients with mutations in KARS.

19.
Chem Res Toxicol ; 31(9): 945-953, 2018 Sep 17.
Artigo em Inglês | MEDLINE | ID: mdl-30092128

RESUMO

Neuroblastoma (NB) is the most common neoplasm during infancy. Unfortunately, NB is still a lethal cancer. Therefore, innovative curative therapies are immediately required. In this study, we showed the prodeath activity of TPGS in human NB SK-N-SH cancer cells. NB cells were exposed to TPGS (10-80 µM). We report for the first time that TPGS induces cell death by apoptosis in NB cells via a pro-oxidant-mediated signaling pathway. Certainly, H2O2 directly oxidizes DJ-1 cysteine106-thiolate into DJ-1 cysteine106-sulfonate, indirectly activates the transcription factors NF-kappaB, p53, and c-JUN, induces the upregulation of mitochondria regulator proteins BAX/PUMA, and provokes the loss of mitochondrial membrane potential (ΔΨm) and the activation of caspase-3/AIF, leading to nuclear disintegration, demonstrated by cellular and biochemical techniques such as fluorescence microscopy, flow cytometry, and Western blot analysis. Since TPGS is a U.S. Food and Drug Administration (FDA)-approved pharmaceutical excipient, this molecule might be used in clinical trials for NB treatment.

20.
Neurotox Res ; 34(3): 417, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-29859003

RESUMO

The original version of this article contained mistakes, and the authors would like to publish this erratum. The "Acknowledgement" section was not included in the aforementioned manuscript.

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