BACKGROUND: Neurodevelopmental disorders (NDDs) and/or associated multiple congenital abnormalities (MCAs) represent a genetically heterogeneous group of conditions with an adverse prognosis for the quality of intellectual and social abilities and common daily functioning. The rapid development of exome sequencing (ES) techniques, together with trio-based analysis, nowadays leads to up to 50% diagnostic yield. Therefore, it is considered as the state-of-the-art approach in these diagnoses. RESULTS: In our study, we present the results of ES in a cohort of 85 families with 90 children with severe NDDs and MCAs. The interconnection of the in-house bioinformatic pipeline and a unique algorithm for variant prioritization resulted in a diagnostic yield of up to 48.9% (44/90), including rare and novel causative variants (41/90) and intragenic copy-number variations (CNVs) (3/90). Of the total number of 47 causative variants, 53.2% (25/47) were novel, highlighting the clinical benefit of ES for unexplained NDDs. Moreover, trio-based ES was verified as a reliable tool for the detection of rare CNVs, ranging from intragenic exon deletions (GRIN2A, ZC4H2 genes) to a 6-Mb duplication. The functional analysis using PANTHER Gene Ontology confirmed the involvement of genes with causative variants in a wide spectrum of developmental processes and molecular pathways, which form essential structural and functional components of the central nervous system. CONCLUSION: Taken together, we present one of the first ES studies of this scale from the central European region. Based on the high diagnostic yield for paediatric NDDs in this study, 48.9%, we confirm trio-based ES as an effective and reliable first-tier diagnostic test in the genetic evaluation of children with NDDs.

• MeSH
• dítě MeSH
• lidé MeSH
• mnohočetné abnormality * MeSH
• molekulární patologie MeSH
• neurovývojové poruchy * genetika   MeSH
• sekvenování exomu MeSH
• variabilita počtu kopií segmentů DNA MeSH
• Check Tag
• dítě MeSH
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Pathogenic variants affecting the BLM gene are responsible for the manifestation of extremely rare cancer‐predisposing Bloom syndrome. The present study reports on a case of an infant with a congenital hypotrophy, short stature and abnormal facial appearance. Initially she was examined using a routine molecular diagnostic algorithm, including the cytogenetic analysis of her karyotype, microarray analysis and methylation‐specific MLPA, however, she remained undiagnosed on a molecular level. Therefore, she and her parents were enrolled in the project of trio‐based exome sequencing (ES) using Human Core Exome kit. She was revealed as a carrier of an extremely rare combination of causative sequence variants altering the BLM gene (NM_000057.4), c.1642C>T and c.2207_2212delinsTAGATTC in the compound heterozygosity, resulting in a diagnosis of Bloom syndrome. Simultaneously, a mosaic loss of heterozygosity of chromosome 11p was detected and then confirmed as a borderline imprinting center 1 hypermethylation on chromosome 11p15. The diagnosis of Bloom syndrome and mosaic copy‐number neutral loss of heterozygosity of chromosome 11p increases a lifetime risk to develop any types of malignancy. This case demonstrates the trio‐based ES as a complex approach for the molecular diagnostics of rare pediatric diseases.

• MeSH
• Bloomův syndrom * diagnóza   genetika   patologie   MeSH
• dítě MeSH
• heterozygot MeSH
• kojenec MeSH
• lidé MeSH
• lidský chromozom Y MeSH
• mozaicismus MeSH
• sekvenování exomu MeSH
• Check Tag
• dítě MeSH
• kojenec MeSH
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH

Pathogenic sequence variant in the GNAI1 gene were recently introduced as a cause of novel syndrome with a manifestation of variable developmental delay and autistic features. In our study, we report a case of monozygotic twins with severe intellectual disability and motor delay and developmental dysphasia. Both probands and their parents were examined using multi-step molecular diagnostic algorithm including whole-exome sequencing (WES), resulting in the identification of a novel, de novo pathogenic sequence variant in the GNAI1 gene, NM_002069.6:c.815 A>G, p.(Asp272Gly) in probands. Using WES we also verified the microarray findings of a familial 8q24.23q24.3 duplication and heterozygous 5q13.2 deletion, not associated with clinical symptoms in probands. Our results confirmed the role of the GNAI1 gene in the pathogenesis of syndromic neurodevelopmental disorders. They support trio- or quatro-based WES as a suitable molecular diagnostics method for the simultaneous detection of clinically relevant sequence variants and CNVs in individuals with neurodevelopmental disorders and rare diseases.

• MeSH
• heterozygot MeSH
• lidé MeSH
• mentální retardace * diagnóza   genetika   MeSH
• neurovývojové poruchy * genetika   MeSH
• sekvenování exomu MeSH
• variabilita počtu kopií segmentů DNA MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH

Alport syndrome with intellectual disability (ATS-ID, AMME complex; OMIM #300194) is an X-linked contiguous gene deletion syndrome associated with an Xq22.3 locus mainly characterized by hematuria, renal failure, hearing loss/deafness, neurodevelopmental disorder (NDD), midface retrusion, and elliptocytosis. It is thought that ATS-ID is caused by the loss of function of COL4A5 (ATS) and FACL4 (ACSL4) genes through the interstitial (micro)deletion of chromosomal band Xq22.3. We report detailed phenotypic description and results from genome-wide screening of a Czech family with diagnosis ATS-ID (proband, maternal uncle, and two female carriers). Female carriers showed mild clinical features of microscopic hematuria only, while affected males displayed several novel clinical features associated with ATS-ID. Utilization of whole-exome sequencing discovered the presence of approximately 3 Mb of deletion in the Xq23 area, which affected 19 genes from TSC22D3 to CHRDL1. We compared the clinical phenotype with previously reported three ATS-ID families worldwide and correlated their clinical manifestations with the incidence of genes in both telomeric and centromeric regions of the deleted chromosomal area. In addition to previously described phenotypes associated with aberrations in AMMECR1 and FACL4, we identified two genes, members of tripartite motif family MID2 and subunit of the proteasome PA700/19S complex (PSMD10), respectively, as prime candidate genes responsible for additional clinical features observed in our patients with ATS-ID. Overall, our findings further improve the knowledge about the clinical impact of Xq23 deletions and bring novel information about phenotype/genotype association of this chromosomal aberration.

• Publikační typ
• kazuistiky MeSH

Neurodegenerative disorders present a broad group of neurological diseases and remain one of the greatest challenges and burdens to mankind. Maladies like amyotrophic lateral sclerosis, Alzheimer's disease, stroke or spinal cord injury commonly features astroglia involvement (astrogliosis) with signs of inflammation. Regenerative, paracrine and immunomodulatory properties of human mesenchymal stromal cells (hMSCs) could target the above components, thus opening new therapeutic possibilities for regenerative medicine. A special interest should be given to hMSCs derived from the umbilical cord (UC) tissue, due to their origin, properties and lack of ethical paradigms. The aim of this study was to establish standard operating and scale-up good manufacturing practice (GMP) protocols of UC-hMSCs isolation, characterization, expansion and comparison of cells' properties when harvested on T-flasks versus using a large-scale bioreactor system. Human UC-hMSCs, isolated by tissue explant culture technique from Wharton's jelly, were harvested after reaching 75% confluence and cultured using tissue culture flasks. Obtained UC-hMSCs prior/after the cryopreservation and after harvesting in a bioreactor, were fully characterized for "mesenchymness" immunomodulatory, tumorigenicity and genetic stability, senescence and cell-doubling properties, as well as gene expression features. Our study demonstrates an efficient and simple technique for large scale UC-hMSCs expansion. Harvesting of UC-hMSCs' using classic and large scale methods did not alter UC-hMSCs' senescence, genetic stability or in vitro tumorigenicity features. We observed comparable growth and immunomodulatory capacities of fresh, frozen and expanded UC-hMSCs. We found no difference in the ability to differentiate toward adipogenic, osteogenic and chondrogenic lineages between classic and large scale UC-hMSCs expansion methods. Both, methods enabled derivation of genetically stabile cells with typical mesenchymal features. Interestingly, we found significantly increased mRNA expression levels of neural growth factor (NGF) and downregulated insulin growth factor (IGF) in UC-hMSCs cultured in bioreactor, while IL4, IL6, IL8, TGFb and VEGF expression levels remained at the similar levels. A culturing of UC-hMSCs using a large-scale automated closed bioreactor expansion system under the GMP conditions does not alter basic "mesenchymal" features and quality of the cells. Our study has been designed to pave a road toward translation of basic research data known about human UC-MSCs for the future clinical testing in patients with neurological and immunocompromised disorders. An industrial manufacturing of UC-hMSCs next will undergo regulatory approval following advanced therapy medicinal products (ATMP) criteria prior to clinical application and approval to be used in patients.

• MeSH
• bioreaktory * MeSH
• kultivované buňky MeSH
• lidé MeSH
• mezenchymální kmenové buňky fyziologie   MeSH
• nemoci nervového systému patologie   terapie   MeSH
• proliferace buněk fyziologie   MeSH
• pupečník cytologie   fyziologie   transplantace   MeSH
• transplantace mezenchymálních kmenových buněk * trendy   MeSH
• Whartonův rosol cytologie   fyziologie   transplantace   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Pathogenic sequence variants in the IQ motif- and Sec7 domain-containing protein 2 (IQSEC2) gene have been confirmed as causative in the aetiopathogenesis of neurodevelopmental disorders (intellectual disability, autism) and epilepsy. We report on a case of a family with three sons; two of them manifest delayed psychomotor development and epilepsy. Initially proband A was examined using a multistep molecular diagnostics algorithm, including karyotype and array-comparative genomic hybridization analysis, both with negative results. Therefore, probands A and B and their unaffected parents were enrolled for an analysis using targeted "next-generation" sequencing (NGS) with a gene panel ClearSeq Inherited DiseaseXT (Agilent Technologies) and verification analysis by Sanger sequencing. A novel frameshift variant in the X-linked IQSEC2 gene NM_001111125.2:c.1813_1814del, p.(Asp605Profs*3) on protein level, was identified in both affected probands and their asymptomatic mother, having skewed X chromosome inactivation (XCI) (100:0). As the IQSEC2 gene is a known gene escaping from XCI in humans, we expect the existence of mechanisms maintaining the normal or enough level of the IQSEC2 protein in the asymptomatic mother. Further analyses may help to the characterization of the presented novel frameshift variant in the IQSEC2 gene as well as to elucidate the mechanisms leading to the rare asymptomatic phenotypes in females.

• MeSH
• algoritmy MeSH
• delece genu MeSH
• dítě MeSH
• epilepsie komplikace   genetika   MeSH
• fenotyp MeSH
• genetická variace * MeSH
• inaktivace chromozomu X MeSH
• karyotypizace MeSH
• lidé MeSH
• neurovývojové poruchy komplikace   genetika   MeSH
• posunová mutace MeSH
• předškolní dítě MeSH
• pruhování chromozomů MeSH
• sekvenční analýza hybridizací s uspořádaným souborem oligonukleotidů MeSH
• srovnávací genomová hybridizace * MeSH
• výměnné faktory guaninnukleotidů genetika   MeSH
• vysoce účinné nukleotidové sekvenování MeSH
• Check Tag
• dítě MeSH
• lidé MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH
• práce podpořená grantem MeSH

Stárnutí je nesmírně komplexním dějem, který má mnoho projevů na úrovni molekulární, buněčné i celého organismu a týká se v určité podobě prakticky všech živých bytostí. Jde o proces charakterizovaný všeobecným progresivním zhoršováním fyziologických funkcí organismu vedoucím ke zvýšené náchylnosti k nemocem. Množství genů asociovaných s dlouhověkostí je určitým způsobem zapojeno do metabolismu. Sirtuiny a AMPK figurují v katabolických drahách, geny dráhy IIS a mTOR jsou zapojené do anabolismu. Intenzivně zkoumány jsou geny zapojené do oprav poškození DNA, geny regulující délku telomer, CETP, ACE, cytokiny IL-10 a IL-6, geny systému HLA, TP53, SOD3 a mnohé další. Látky jako rapamycin, inhibující dráhu mTOR, resveratrol a další aktivátory sirtuinů (SRT1720, SRT2104) a metformin aktivující AMPK mají schopnost u modelových organismů prodloužit život a zpomalit stárnutí. U člověka lze podobného účinku docílit i obyčejnou změnou životosprávy. Jde například o omezení příjmu energie, konzumaci košťálové zeleniny (obsahující flavonoidy a polyfenoly, jako je sulforafan a genistein) nebo o pravidelnou pohybovou aktivitu.

Aging is an extremely complex phenomenon that has many manifes-tations at the molecular, cellular, and whole-body levels, and in some form involves virtually all living beings. It is a process characterized by a general progressive deterioration of the physiological functions of the organism leading to increased susceptibility to diseases. This article summarizes the basic features and molecular hallmarks of aging and describes some of the genetic mechanisms of this phenomenon. It deals with the particular genes and molecular pathways involved in the regulation of aging as well as promising possibilities of interventions affecting this process

• MeSH
• lidé MeSH
• sirtuiny fyziologie   genetika   MeSH
• stárnutí buněk fyziologie   genetika   MeSH
• stárnutí * fyziologie   genetika   MeSH
• telomery patologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• přehledy MeSH

• Publikační typ
• abstrakt z konference MeSH

BACKGROUND: Chromosomal microarray analysis has been shown to be a valuable and cost effective assay for elucidating copy number variants (CNVs) in children with intellectual disability and developmental delay (ID/DD). METHODS: In our study, we performed array-based comparative genomic hybridization (array-CGH) analysis using oligonucleotide-based platforms in 542 Czech patients with ID/DD, autism spectrum disorders and multiple congenital abnormalities. Prior to the array-CGH analysis, all the patients were first examined karyotypically using G-banding. The presence of CNVs and their putative derivation was confirmed using fluorescence in situ hybridization (FISH), multiplex ligation-dependent probe amplification (MLPA) and predominantly relative quantitative polymerase chain reaction (qPCR). RESULTS: In total, 5.9% (32/542) patients were positive for karyotypic abnormalities. Pathogenic/likely pathogenic CNVs were identified in 17.7% of them (96/542), variants of uncertain significance (VOUS) were detected in 4.8% (26/542) and likely benign CNVs in 9.2% of cases (50/542). We identified 6.6% (36/542) patients with known recurrent microdeletion (24 cases) and microduplication (12 cases) syndromes, as well as 4.8% (26/542) patients with non-recurrent rare microdeletions (21 cases) and microduplications (5 cases). In the group of patients with submicroscopic pathogenic/likely pathogenic CNVs (13.3%; 68/510) we identified 91.2% (62/68) patients with one CNV, 5.9% (4/68) patients with two likely independent CNVs and 2.9% (2/68) patients with two CNVs resulting from cryptic unbalanced translocations. Of all detected CNVs, 21% (31/147) had a de novo origin, 51% (75/147) were inherited and 28% (41/147) of unknown origin. In our cohort pathogenic/likely pathogenic microdeletions were more frequent than microduplications (69%; 51/74 vs. 31%; 23/74) ranging in size from 0.395 Mb to 10.676 Mb (microdeletions) and 0.544 Mb to 8.156 Mb (microduplications), but their sizes were not significantly different (P = 0.83). The pathogenic/likely pathogenic CNVs (median 2.663 Mb) were significantly larger than benign CNVs (median 0.394 Mb) (P < 0.00001) and likewise the pathogenic/likely pathogenic CNVs (median 2.663 Mb) were significantly larger in size than VOUS (median 0.469 Mb) (P < 0.00001). CONCLUSIONS: Our results confirm the benefit of array-CGH in the current clinical genetic diagnostics leading to identification of the genetic cause of ID/DD in affected children.

• MeSH
• dítě MeSH
• kohortové studie MeSH
• kojenec MeSH
• lidé MeSH
• mentální retardace genetika   MeSH
• mladiství MeSH
• novorozenec MeSH
• předškolní dítě MeSH
• sekvenční analýza hybridizací s uspořádaným souborem oligonukleotidů * MeSH
• srovnávací genomová hybridizace * MeSH
• variabilita počtu kopií segmentů DNA * MeSH
• vývojové poruchy u dětí genetika   MeSH
• Check Tag
• dítě MeSH
• kojenec MeSH
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• novorozenec MeSH
• předškolní dítě MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH

De novo sequence variants, including truncating and splicing variants, in the additional sex‑combs like 3 gene (ASXL3) have been described as the cause of Bainbridge‑Ropers syndrome (BRS). This pathology is characterized by delayed psychomotor development, severe intellectual disability, growth delay, hypotonia and facial dimorphism. The present study reports a case of a girl (born in 2013) with severe global developmental delay, central hypotonia, microcephaly and poor speech. The proband was examined using a multi‑step molecular diagnostics algorithm, including karyotype and array‑comparative genomic hybridization analysis, with negative results. Therefore, the proband and her unaffected parents were enrolled for a pilot study using targeted next‑generation sequencing technology (NGS) with gene panel ClearSeq Inherited DiseaseXT and subsequent validation by Sanger sequencing. A novel de novo heterozygous frameshift variant in the ASXL3 gene (c.3006delT, p.R1004Efs*21), predicted to result in a premature termination codon, was identified. In conclusion, the present study demonstrated that targeted NGS using a suitable, gene‑rich panel may provide a conclusive molecular genetics diagnosis in children with severe global developmental delays.

• MeSH
• dítě MeSH
• lidé MeSH
• mikrocefalie genetika   MeSH
• pilotní projekty MeSH
• poruchy řeči genetika   MeSH
• posunová mutace MeSH
• rodokmen MeSH
• svalová hypotonie genetika   MeSH
• transkripční faktory genetika   MeSH
• vývojové poruchy u dětí genetika   MeSH
• Check Tag
• dítě MeSH
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH