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SNaPshot Assay in Quantitative Detection of Allelic Nondisjunction in Down Syndrome.

ABSTRACT: We wished to identify markers associated with allelic nondisjunction in nuclear families with Down syndrome (DS) offspring. Since the GRIK1 and GARS-AIRS-GART genes, mapping to chromosome 21q22.1, may be informative in this regard, we genotyped four single-nucleotide polymorphisms [30952599(A/G) rs363484; 30924733(A/G) rs363506; 34901423(A/G) rs2834235; 34877070(A/G) rs7283354] present in these genes using the SNaPshot(™) assay protocol.We have reported 30952599(A/G)-rs363484 to be monomorphic in our sample population. Genotyping revealed 35/65 families to be informative for 34877070(A/G)-rs7283354 (GARS-AIRS-GART), whereas only 25/65 and 11/65 are informative for 34901423(A/G)-rs2834235 (GARS-AIRS-GART) and 30924733(A/G)-rs363506 (GRIK1) polymorphisms, respectively. The parent- and stage-of-origin of nondisjunction could be traced in 48/65 families using at least one polymorphic marker. A single trio provided internal validation for assignment of the parent- and stage-of-origin of nondisjunction whereby the nondisjoining alleles were independently identified as G-rs363506, G-rs2834235, and G-rs7283354, respectively. An enhanced ratio of meiosis-I to meiosis-II errors during maternal or paternal meioses accounts for allelic nondisjunction.The SNaPshot assay is quantitative and permits multiplexing for detection of allelic nondisjunction. Inclusion of additional informative chromosome 21-specific markers may aid rapid aneuploidy detection, screening, and prenatal counseling of parents at risk of having babies with DS.

SUBMITTER: Ghosh D

PROVIDER: S-EPMC3468158 | biostudies-literature | 2012 Oct

REPOSITORIES: biostudies-literature

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SNaPshot Assay in Quantitative Detection of Allelic Nondisjunction in Down Syndrome.

Ghosh Debarati D Gochhait Sailesh S Banerjee Disha D Chatterjee Anindita A Sinha Swagata S Nandagopal Krishnadas K

Genetic testing and molecular biomarkers 20120829 10

<h4>Aim</h4>We wished to identify markers associated with allelic nondisjunction in nuclear families with Down syndrome (DS) offspring. Since the GRIK1 and GARS-AIRS-GART genes, mapping to chromosome 21q22.1, may be informative in this regard, we genotyped four single-nucleotide polymorphisms [30952599(A/G) rs363484; 30924733(A/G) rs363506; 34901423(A/G) rs2834235; 34877070(A/G) rs7283354] present in these genes using the SNaPshot(™) assay protocol.<h4>Results</h4>We have reported 30952599(A/G)- ...[more]

PMID: 22931243

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Project description:Altered patterns of recombination on 21q have long been associated with the nondisjunction chromosome 21 within oocytes and the increased risk of having a child with Down syndrome. Unfortunately the genetic etiology of these altered patterns of recombination have yet to be elucidated. We for the first time genotyped the gene MCM9, a candidate gene for recombination regulation and DNA repair in mothers with or without children with Down syndrome. In our approach, we identified the location of recombination on the maternal chromosome 21 using short tandem repeat markers, then stratified our population by the origin of meiotic error and age at conception. We observed that twenty-five out of forty-one single nucleotide polymorphic sites within MCM9 exhibited an association with meiosis I error (N = 700), but not with meiosis II error (N = 125). This association was maternal age-independent. Several variants exhibited aprotective association with MI error, some were neutral. Maternal age stratified characterization of cases revealed that MCM9 risk variants were associated with an increased chance of reduced recombination on 21q within oocytes. The spatial distribution of single observed recombination events revealed no significant change in the location of recombination among women harbouring MCM9 risk, protective, or neutral variant. Additionally, we identified a total of six novel polymorphic variants and two novel alleles that were either risk imparting or protective against meiosis I nondisjunction. In silico analyses using five different programs suggest the risk variants either cause a change in protein function or may alter the splicing pattern of transcripts and disrupt the proportion of different isoforms of MCM9 products within oocytes. These observations bring us a significant step closer to understanding the molecular basis of recombination errors in chromosome 21 nondisjunction within oocytes that leads to birth of child with Down syndrome.

Project description:Heparan sulphate (HS) is an important component of the extracellular matrix and the vasculature basal laminar which functions as a barrier to the extravasation of metastatic and inflammatory cells. Cleavage of HS by endoglycosidase or heparanase activity produced by invading cells may assist in the disassembly of the extracellular matrix and basal laminar, and thereby facilitate cell migration. Heparanase activity has previously been shown to be related to the metastatic potential of murine and human melanoma cell lines [Nakajima, Irimura and Nicolson (1988) J. Cell. Biochem. 36, 157-167]. To determine heparanase activity, porcine mucosal HS was partially de-N-acetylated and re-N-acetylated with [3H]acetic anhydride to yield a radiolabelled substrate. This procedure prevented the masking of, or possible formation of, new heparanase-sensitive cleavage sites as has been observed with previous methods of radiolabelling. Heparanase activity in a variety of tissues and cell homogenates including human platelets, colonic carcinoma cells, umbilical vein endothelial cells and rat mammary adenocarcinoma cells (both metastatic and non-metastatic variants) and liver homogenates all degraded the substrate in a stepwise fashion from 18.5 to approximately 13, 8 and finally to 4.5 kDa fragments, as assessed by gel-filtration analysis, confirming the substrate as suitable for the detection of heparanase activity present in a variety of cells and tissues. A rapid quantitative assay was developed with the HS substrate using a novel method for separating degradation products from the substrate by taking advantage of the decreased affinity of the heparanase-cleaved products for the HS-binding plasma protein chicken histidine-rich glycoprotein (cHRG). Incubation mixtures were applied to cHRG-Sepharose columns, with unbound material corresponding to heparanase-degradation products. Heparanase activity was determined for a variety of human, rat and murine cell and tissue homogenates. The highly metastatic rat mammary adenocarcinoma and murine lung carcinoma cell lines had four to ten times the heparanase activity of non-metastatic variants, confirming the correlation of heparanase activity with metastatic potential. Human cancer patients had twice the serum heparanase levels of normal healthy adults. The assay will be valuable for the determination of heparanase activity from a variety of tissue and cell sources, as a diagnostic tool for the determination of heparanase potential, and for the development of specific inhibitors of heparanase activity and metastasis.

Project description:ImportanceNovel plasma biomarkers, especially phosphorylated tau (p-tau), can detect brain tau aggregates in Alzheimer disease.ObjectiveTo determine which plasma biomarker combinations can accurately detect tau pathological brain changes in Down syndrome (DS).Design, setting, and participantsThe cross-sectional, multicenter Alzheimer's Biomarker Consortium-Down Syndrome study included adults with DS and a control group of siblings without DS. All participants with plasma, positron emission tomography (PET), and cognitive measures available by the time of data freeze 1.0 were included. Participants were enrolled between 2016 and 2019, and data were analyzed from August 2021 to April 2022.ExposuresPlasma p-tau217, glial fibrillary acidic protein (GFAP), amyloid β42/40 (Aβ42/Aβ40), neurofilament light (NfL), and total tau (t-tau); tau positron emission tomography (tau-PET) and Aβ-PET.Main outcomes and measuresThe primary outcome was tau-PET status. Secondary outcomes included Aβ-PET status and cognitive performance.ResultsAmong 300 participants with DS and a control group of 37 non-DS siblings, mean (SD) age was 45.0 (10.1) years, and 167 (49.6%) were men. Among participants with DS who all underwent plasma p-tau217 and GFAP analyses, 258 had other plasma biomarker data available and 119, 213, and 288 participants had tau-PET, Aβ-PET, and cognitive assessments, respectively. Plasma p-tau217 and t-tau were significantly increased in Aβ-PET-positive tau-PET-positive (A+T+) DS and A+T- DS compared with A-T- DS while GFAP was only increased in A+T+ DS. Plasma p-tau217 levels were also significantly higher in A+T+ DS than A+T- DS. In participants with DS, plasma p-tau217 and GFAP (but not other plasma biomarkers) were consistently associated with abnormal tau-PET and Aβ-PET status in models covaried for age (odds ratio range, 1.59 [95% CI, 1.05-2.40] to 2.32 [95% CI, 1.36-3.96]; P < .03). A combination of p-tau217 and age performed best when detecting tau-PET abnormality in temporal and neocortical regions (area under the curve [AUC] range, 0.96-0.99). The most parsimonious model for Aβ-PET status included p-tau217, t-tau, and age (AUC range, 0.93-0.95). In multivariable models, higher p-tau217 levels but not other biomarkers were associated with worse performance on DS Mental Status Examination (β, -0.24, 95% CI, -0.36 to -0.12; P < .001) and Cued Recall Test (β, -0.40; 95% CI, -0.53 to -0.26; P < .001).Conclusions and relevancePlasma p-tau217 is a very accurate blood-based biomarker of both tau and Aβ pathological brain changes in DS that could help guide screening and enrichment strategies for inclusion of individuals with DS in future AD clinical trials, especially when it is combined with age as a covariate.

Dataset Information

SNaPshot Assay in Quantitative Detection of Allelic Nondisjunction in Down Syndrome.

Publications

SNaPshot Assay in Quantitative Detection of Allelic Nondisjunction in Down Syndrome.

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