Východiska:Chromozomové změny patří u chronické lymfocytární leukemie (CLL) mezi významné prognostické faktory. Hlavní metodou využívanou k detekci těchto chromozomových změn je fluorescenční in situ hybridizace (FISH), klasické cytogenetické vyšetření vyžadující buňky v metafázi je u CLL problematické kvůli nízké proliferační schopnosti maligních B-lymfocytů in vitro. V roce 2006 byla publikována metoda umožňující získat metafázní B-lymfocyty u CLL s využitím stimulace pomocí IL‐2 a CpG oligonukleotidem DSP30. Cílem naší studie bylo ověřit účinnost stimulace a zhodnotit využitelnost této metody v rutinní praxi. Soubor pacientů a metody: Celkem u 369 pacientů s dia­gnózou CLL byla vyšetřována periferní krev klasickou cytogenetickou metodou a současně metodou FISH pro oblasti 13q14, 11q22–23, CEP12 a 17p13. Výsledky: Pro klasické cytogenetické vyšetření se stimulací podařilo získat metafázní buňky u 307 (83 %) z 369 pacientů. Chromozomové změny byly nalezeny u 243 (79 %) ze 307 hodnocených pacientů. Kromě aberací vyšetřovaných metodou FISH byly nalezeny další specifické změny v karyotypu např. del(6q), del(14q), t(14;18)(q32;q21), t(11;14)(q13;q32) a t(18;22)(q21;q11). U 103 (42 %) pacientů byly cytogenetickým vyšetřením nalezeny komplexní změny karyotypu, které metodou FISH detekovány nebyly. Závěr: Stimulace pomocí IL‐2 a oligonukleotidu DSP30 účinně navozuje dělení maligních B‐lymfocytů a umožňuje zachytit velké množství chromozomových změn u CLL, které by při vyšetřování metodou FISH pro základní čtyři aberace zůstaly skryty. Používání této metody v rutinní praxi se osvědčilo hlavně při identifikaci pacientů s komplexními změnami karyotypu.

Background: Chromosomal aberrations play an important role as prognostic factors in chronic lymphocytic leukemia (CLL). These aberrations are mostly detected by fluorescent in situ hybridization (FISH), as chromosomal banding analysis has been scarce due to low proliferative activity of malignant B-lymphocytes in vitro. In 2006, a new method using stimulation with IL-2 and CpG oligonucleotide DSP30 for metaphase generation in CLL was published [1]. The objective of our study was to verify the efficacy of stimulation and to evaluate if the method is suitable for routine diagnostics. Patients and Methods: In total, peripheral blood samples of 369 CLL patients were analyzed in parallel by chromosomal banding analysis and by FISH probes for 13q14, 11q22–23, CEP12 and 17p13. Results: Out of 369 patients, 307 (83%) were successfully stimulated for metaphase generation. Chromosomal aberrations were detected in 243 (79%) out of 307 patients evaluated by chromosomal banding analysis. Other aberrations that are not included into standard FISH panel were detected in patients’ karyotypes, e.g. del(6q), del(14q), t(14;18)(q32;q21), t(11;14)(q13;q32) and t(18;22)(q21;q11). One hundred and three (42%) patients showed complex aberrant karyotype not detected by FISH analysis. Conclusion: Stimulation with IL-2 and oligonucleotide DSP30 is an efficient method how to induce proliferation of malignant B-lymphocytes and allows detection of a substantial number of chromosomal aberrations in addition to those detected by standard FISH panel. Using this method in routine diagnostics is helpful particularly in identification of patients with complex aberrant karyotype.

• Klíčová slova
• CpG‐ODN DSP30,
• MeSH
• chromozomální aberace * MeSH
• chromozomální delece MeSH
• chronická lymfatická leukemie * genetika   MeSH
• cytogenetické vyšetření * statistika a číselné údaje   využití   MeSH
• dospělí MeSH
• hybridizace in situ fluorescenční * MeSH
• interleukin-2 MeSH
• karyotyp MeSH
• krev MeSH
• kultivované buňky MeSH
• lidé středního věku MeSH
• lidé MeSH
• oligonukleotidy MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• translokace genetická MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• práce podpořená grantem MeSH

Syncytin-1 is a captive envelope glycoprotein encoded by one of human endogenous retroviruses W. It is expressed exclusively in the placental trophoblast where it participates in cell-to-cell fusion during differentiation of syncytiotrophobast. In other tissues, however, syncytin-1 expression must be kept in check because inadvertent cell fusion might be dangerous for tissue organization and integrity. We describe here an inverse correlation between CpG methylation of syncytin-1 5' long terminal repeat and its expression. Hypomethylation of the syncytin-1 5' long terminal repeat in the placenta and in the choriocarcinoma-derived cell line BeWo was detected. However, other analyzed primary cells and cell lines non-expressing syncytin-1 contain proviruses heavily methylated in this sequence. CpG methylation of syncytin-1 is resistant to the effect of the demethylating agent 5-azacytidine. The inhibitory role of CpG methylation is further confirmed by transient transfection of in-vitro-methylated syncytin-1 promoter-driven reporter construct. Altogether, we conclude that CpG methylation plays a principal role in the transcriptional suppression of syncytin-1 in non-placental tissues, and, in contrast, demethylation of the syncytin-1 promoter in trophoblast is a prerequisite for its expression and differentiation of multinucleated syncytiotrophoblast.

• MeSH
• buněčné linie MeSH
• CpG ostrůvky fyziologie   MeSH
• down regulace imunologie   MeSH
• financování organizované MeSH
• genové produkty env antagonisté a inhibitory   biosyntéza   genetika   MeSH
• HeLa buňky MeSH
• koncové repetice MeSH
• lidé MeSH
• metylace DNA MeSH
• placenta metabolismus   MeSH
• promotorové oblasti (genetika) MeSH
• těhotenské proteiny antagonisté a inhibitory   biosyntéza   genetika   MeSH
• transkripční faktory antagonisté a inhibitory   biosyntéza   genetika   MeSH
• trofoblasty metabolismus   MeSH
• Check Tag
• lidé MeSH

SARS-CoV-2 is an intensively investigated virus from the order Nidovirales (Coronaviridae family) that causes COVID-19 disease in humans. Through enormous scientific effort, thousands of viral strains have been sequenced to date, thereby creating a strong background for deep bioinformatics studies of the SARS-CoV-2 genome. In this study, we inspected high-frequency mutations of SARS-CoV-2 and carried out systematic analyses of their overlay with inverted repeat (IR) loci and CpG islands. The main conclusion of our study is that SARS-CoV-2 hot-spot mutations are significantly enriched within both IRs and CpG island loci. This points to their role in genomic instability and may predict further mutational drive of the SARS-CoV-2 genome. Moreover, CpG islands are strongly enriched upstream from viral ORFs and thus could play important roles in transcription and the viral life cycle. We hypothesize that hypermethylation of these loci will decrease the transcription of viral ORFs and could therefore limit the progression of the disease.

• MeSH
• COVID-19 virologie   MeSH
• CpG ostrůvky * MeSH
• genom virový MeSH
• lidé MeSH
• metylace DNA MeSH
• mutace * MeSH
• SARS-CoV-2 genetika   MeSH
• vazba proteinů MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Unmethylated oligodeoxynucleotides containing guanine-cytidine dimers (CpG ODN) have been described as potent inducers of selected antitumour immune responses and the immunotherapeutic efficacy of CpG ODN has been examined either alone or as a vaccine adjuvant. We hypothesized that CpG ODN therapy could be an effective tool for immunotherapy of not only conventional MHC class I(+) tumours but also of those tumours that have lost MHC class I expression during their progression. To address this hypothesis, we employed the animal model resembling MHC class I-proficient and -deficient human papilloma virus (HPV) 16-associated tumours. A cell line transformed with HPV16 E6 and E7 oncogenes, TC-1, as a prototype of MHC class I-positive line, and its MHC class I-deficient sublines TC-1/A9 and TC-1/P3C10 were injected into syngeneic C57BL/6 mice and the growing tumours were subjected to immunotherapy with CpG ODN 1826. The therapy started either 1 day after the challenge with the tumour cells or later, when the tumours had reached a palpable size. In both settings, CpG ODN 1826 significantly reduced the growth of MHC class I-proficient and -deficient tumours. Furthermore, we demonstrated that CpG ODN 1585, whose mechanism of action preferably involves indirect activation of the natural killer cells, induced regression of the MHC class I-deficient tumours TC1/A9 but not of the MHC class I-proficient tumours TC-1. This study infers that synthetic CpG ODN have a potential for the therapy of both MHC class I-proficient and -deficient tumours and thus could be also used against tumours that tend to down-regulate their MHC class I expression.

• MeSH
• CpG ostrůvky MeSH
• down regulace MeSH
• financování organizované MeSH
• geny MHC třídy I MeSH
• imunoterapie metody   MeSH
• infekce papilomavirem MeSH
• lidský papilomavirus 16 MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• nádory terapie   virologie   MeSH
• oligonukleotidy MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH

BACKGROUND: CpG Island Methylator Phenotype (CIMP) is an epigenetic phenotype in CRC characterized by hypermethylation of CpG islands in promoter regions of tumor suppressor genes, leading to their transcriptional silencing and loss of function. While the prevalence of CRC differs across geographical regions, no studies have compared prevalence of CIMP-High phenotype across regions. The purpose of this project was to compare the prevalence of CIMP across geographical regions after adjusting for variations in methodologies to measure CIMP in a meta-analysis. METHODS: We searched PubMed, Medline, and Embase for articles focusing on CIMP published from 2000 to 2018. Two reviewers independently identified 111 articles to be included in final meta-analysis. We classified methods used to quantify CIMP into 4 categories: a) Classical (MINT marker) Panel group b) Weisenberg-Ogino (W-O) group c) Human Methylation Arrays group and d) Miscellaneous group. We compared the prevalence of CIMP across geographical regions after correcting for methodological variations using meta-regression techniques. RESULTS: The pooled prevalence of CIMP-High across all studies was 22% (95% confidence interval:21-24%; I2 = 94.75%). Pooled prevalence of CIMP-H across Asia, Australia, Europe, North America and South America was 22, 21, 21, 27 and 25%, respectively. Meta-regression analysis identified no significant differences in the prevalence of CIMP-H across geographical regions after correction for methodological variations. In exploratory analysis, we observed variations in CIMP-H prevalence across countries. CONCLUSION: Although no differences were found for CIMP-H prevalence across countries, further studies are needed to compare the influence of demographic, lifestyle and environmental factors in relation to the prevalence of CIMP across geographical regions.

• MeSH
• CpG ostrůvky genetika   MeSH
• fenotyp * MeSH
• genetická heterogenita MeSH
• kohortové studie MeSH
• kolorektální nádory genetika   MeSH
• lidé MeSH
• metylace DNA genetika   MeSH
• pití alkoholu škodlivé účinky   genetika   MeSH
• prevalence MeSH
• promotorové oblasti (genetika) genetika   MeSH
• publikační zkreslení MeSH
• rizikové faktory MeSH
• umlčování genů MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• metaanalýza MeSH
• systematický přehled MeSH
• Geografické názvy
• Česká republika MeSH
• Indie MeSH

Unmethylated CpG islands are known to keep adjacent promoters transcriptionally active. In the CpG island adjacent to the adenosine phosphoribosyltransferase gene, the protection against transcriptional silencing can be attributed to the short CpG-rich core element containing Sp1 binding sites. We report here the insertion of this CpG island core element, IE, into the long terminal repeat of a retroviral vector derived from Rous sarcoma virus, which normally suffers from progressive transcriptional silencing in mammalian cells. IE insertion into a specific position between enhancer and promoter sequences led to efficient protection of the integrated vector from silencing and gradual CpG methylation in rodent and human cells. Individual cell clones with IE-modified reporter vectors display high levels of reporter expression for a sustained period and without substantial variegation in the cell culture. The presence of Sp1 binding sites is important for the protective effect of IE, but at least some part of the entire antisilencing capacity is maintained in IE with mutated Sp1 sites. We suggest that this strategy of antisilencing protection by the CpG island core element may prove generally useful in retroviral vectors.

• MeSH
• biologické modely MeSH
• CpG ostrůvky MeSH
• financování organizované MeSH
• genetická transkripce MeSH
• koncové repetice MeSH
• lidé MeSH
• mutace MeSH
• průtoková cytometrie MeSH
• ptačí sarkom genetika   virologie   MeSH
• reportérové geny MeSH
• transkripční faktor Sp1 metabolismus   MeSH
• umlčování genů MeSH
• vazebná místa MeSH
• virus ptačí leukózy metabolismus   MeSH
• virus Rousova sarkomu metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH

• MeSH
• CpG ostrůvky MeSH
• genetická transkripce MeSH
• genetické vektory genetika   MeSH
• regulace genové exprese MeSH
• viry ptačího sarkomu genetika   MeSH
• viry genetika   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH

Accurate assessment of neuroblastoma outcome prediction remains challenging. Therefore, this study aims at establishing novel prognostic tumor DNA methylation biomarkers. In total, 396 low- and high-risk primary tumors were analyzed, of which 87 were profiled using methyl-CpG-binding domain (MBD) sequencing for differential methylation analysis between prognostic patient groups. Subsequently, methylation-specific PCR (MSP) assays were developed for 78 top-ranking differentially methylated regions and tested on two independent cohorts of 132 and 177 samples, respectively. Further, a new statistical framework was used to identify a robust set of MSP assays of which the methylation score (i.e. the percentage of methylated assays) allows accurate outcome prediction. Survival analyses were performed on the individual target level, as well as on the combined multimarker signature. As a result of the differential DNA methylation assessment by MBD sequencing, 58 of the 78 MSP assays were designed in regions previously unexplored in neuroblastoma, and 36 are located in non-promoter or non-coding regions. In total, 5 individual MSP assays (located in CCDC177, NXPH1, lnc-MRPL3-2, lnc-TREX1-1 and one on a region from chromosome 8 with no further annotation) predict event-free survival and 4 additional assays (located in SPRED3, TNFAIP2, NPM2 and CYYR1) also predict overall survival. Furthermore, a robust 58-marker methylation signature predicting overall and event-free survival was established. In conclusion, this study encompasses the largest DNA methylation biomarker study in neuroblastoma so far. We identified and independently validated several novel prognostic biomarkers, as well as a prognostic 58-marker methylation signature.

• MeSH
• biologické markery analýza   MeSH
• CpG ostrůvky genetika   MeSH
• DNA nádorová genetika   MeSH
• kohortové studie MeSH
• kojenec MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• lidé MeSH
• metylace DNA * MeSH
• nádorové buňky kultivované MeSH
• neuroblastom diagnóza   genetika   MeSH
• prognóza MeSH
• staging nádorů MeSH
• vazebná místa MeSH
• výpočetní biologie MeSH
• Check Tag
• kojenec MeSH
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• multicentrická studie MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH

Epigenetic modifications are essential regulators of biological processes. Decreased DNA methylation of OAS2 (2'-5'-Oligoadenylate Synthetase 2), encoding an antiviral protein, has been seen in psoriasis. To provide further insight into the epigenetic regulation of OAS2, we performed pyrosequencing to detect OAS2 DNA methylation status at 11 promoter and first exon located CpG sites in psoriasis (n = 12) and two common subtypes of squamous cell carcinoma (SCC) of the head and neck: tongue (n = 12) and tonsillar (n = 11). Compared to corresponding controls, a general hypomethylation was seen in psoriasis. In tongue and tonsillar SCC, hypomethylation was found at only two CpG sites, the same two sites that were least demethylated in psoriasis. Despite differences in the specific residues targeted for methylation/demethylation, OAS2 expression was upregulated in all conditions and correlations between methylation and expression were seen in psoriasis and tongue SCC. Distinctive methylation status at four successively located CpG sites within a genomic area of 63 bp reveals a delicately integrated epigenetic program and indicates that detailed analysis of individual CpGs provides additional information into the mechanisms of epigenetic regulation in specific disease states. Methylation analyses as clinical biomarkers need to be tailored according to disease-specific sites.

• MeSH
• 2',5'-oligoadenylátsynthetasa genetika   metabolismus   MeSH
• CpG ostrůvky * MeSH
• epigeneze genetická * MeSH
• lidé MeSH
• metylace DNA MeSH
• nádory hlavy a krku genetika   metabolismus   patologie   MeSH
• nádory jazyka genetika   metabolismus   patologie   MeSH
• promotorové oblasti (genetika) MeSH
• psoriáza genetika   metabolismus   patologie   MeSH
• retrospektivní studie MeSH
• spinocelulární karcinom genetika   metabolismus   patologie   MeSH
• studie případů a kontrol MeSH
• tonzilární nádory genetika   metabolismus   patologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

CTG repeat expansions in DMPK cause myotonic dystrophy (DM1) with a continuum of severity and ages of onset. Congenital DM1 (CDM1), the most severe form, presents distinct clinical features, large expansions, and almost exclusive maternal transmission. The correlation between CDM1 and expansion size is not absolute, suggesting contributions of other factors. We determined CpG methylation flanking the CTG repeat in 79 blood samples from 20 CDM1-affected individuals; 21, 27, and 11 individuals with DM1 but not CDM1 (henceforth non-CDM1) with maternal, paternal, and unknown inheritance; and collections of maternally and paternally derived chorionic villus samples (7 CVSs) and human embryonic stem cells (4 hESCs). All but two CDM1-affected individuals showed high levels of methylation upstream and downstream of the repeat, greater than non-CDM1 individuals (p = 7.04958 × 10(-12)). Most non-CDM1 individuals were devoid of methylation, where one in six showed downstream methylation. Only two non-CDM1 individuals showed upstream methylation, and these were maternally derived childhood onset, suggesting a continuum of methylation with age of onset. Only maternally derived hESCs and CVSs showed upstream methylation. In contrast, paternally derived samples (27 blood samples, 3 CVSs, and 2 hESCs) never showed upstream methylation. CTG tract length did not strictly correlate with CDM1 or methylation. Thus, methylation patterns flanking the CTG repeat are stronger indicators of CDM1 than repeat size. Spermatogonia with upstream methylation may not survive due to methylation-induced reduced expression of the adjacent SIX5, thereby protecting DM1-affected fathers from having CDM1-affected children. Thus, DMPK methylation may account for the maternal bias for CDM1 transmission, larger maternal CTG expansions, age of onset, and clinical continuum, and may serve as a diagnostic indicator.

• MeSH
• buněčné linie MeSH
• CpG ostrůvky * MeSH
• dítě MeSH
• DM-kinasa genetika   MeSH
• dospělí MeSH
• lidé MeSH
• lidské embryonální kmenové buňky chemie   MeSH
• lineární modely MeSH
• metylace DNA * MeSH
• mladiství MeSH
• mladý dospělý MeSH
• myotonická dystrofie genetika   MeSH
• promotorové oblasti (genetika) MeSH
• rodokmen MeSH
• sekvence nukleotidů MeSH
• sekvenční analýza DNA MeSH
• těhotenství MeSH
• Check Tag
• dítě MeSH
• dospělí MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• těhotenství MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH