Population-Based Resequencing of LIPG and ZNF202 Genes in Subjects with Extreme HDL Levels

Endothelial lipase (LIPG) and zinc finger protein 202 (ZNF202) are two pivotal genes in high density lipoprotein (HDL metabolism). We sought to determine their genetic contribution to variation in HDL-cholesterol levels by comprehensive resequencing of both genes in 235 individuals with high or low HDL-C levels. The selected subjects were 141 Whites (High HDL Group: n = 68, x¯=76.90mg/dl; Low HDL Group: n = 73, x¯=32.55mg/dl) and 94 Hispanics (High HDL Group: n = 46, x¯=74.85mg/dl; Low HDL Group: n = 48, x¯=29.95mg/dl). We identified a total of 185 and 122 sequence variants in LIPG and ZNF202, respectively. We found only two missense variants in LIPG (T111I and N396S) and two in ZNF202 (A154V and K259E). In both genes, there were several variants unique to either the low or high HDL group. For LIPG, the proportion of unique variants differed between the high and low HDL groups in both Whites (p = 0.022) and Hispanics (p = 0.017), but for ZNF202 this difference was observed only in Hispanics (p = 0.021). We also identified a common haplotype in ZNF202 among Whites that was significantly associated with the high HDL group (p = 0.013). These findings provide insights into the genetics of LIPG and ZNF202, and suggest that sequence variants occurring with high frequency in non-exonic regions may play a prominent role in modulating HDL-C levels in the general population

Connecting the dots: Potential of data integration to identify regulatory snps in late-onset alzheimer's disease GWAS findings

Late-onset Alzheimer's disease (LOAD) is a multifactorial disorder with over twenty loci associated with disease risk. Given the number of genome-wide significant variants that fall outside of coding regions, it is possible that some of these variants alter some function of gene expression rather than tagging coding variants that alter protein structure and/or function. RegulomeDB is a database that annotates regulatory functions of genetic variants. In this study, we utilized RegulomeDB to investigate potential regulatory functions of lead single nucleotide polymorphisms (SNPs) identified in five genome-wide association studies (GWAS) of risk and age-at onset (AAO) of LOAD, as well as SNPs in LD (r2≥0.80) with the lead GWAS SNPs. Of a total 614 SNPs examined, 394 returned RegulomeDB scores of 1-6. Of those 394 variants, 34 showed strong evidence of regulatory function (RegulomeDB score ,3), and only 3 of them were genome-wide significant SNPs (ZCWPW1/ rs1476679, CLU/rs1532278 and ABCA7/rs3764650). This study further supports the assumption that some of the non-coding GWAS SNPs are true associations rather than tagged associations and demonstrates the application of RegulomeDB to GWAS data.©2014 Rosenthal et al

Magnetic graphene sol-gel hybrid as clean-up adsorbent for acrylamide analysis in food samples prior to GC-MS

Graphene (G) modified with magnetite (Fe3O4) and sol–gel hybrid tetraethoxysilane-methyltrimethoxysilane (TEOS-MTMOS) was used as a clean-up adsorbent in magnetic solid phase extraction (MSPE) for direct determination of acrylamide in various food samples prior to gas chromatography-mass spectrometry analysis. Good linearity (R2 = 0.9990) was achieved for all samples using matrix-matched calibration. The limit of detection (LOD = 3 × SD/m) obtained was 0.061–2.89 µg kg-1 for the studied food samples. Native acrylamide was found to be highest in fried potato with bright-fleshed (900.81 µg kg-1) and lowest in toasted bread (5.02 µg kg-1). High acrylamide relative recovery (RR = 82.7–105.2%) of acrylamide was obtained for spiked (5 and 50 µg kg-1) food samples. The Fe3O4@G-TEOS-MTMOS is reusable up to 7 times as a clean-up adsorbent with good recovery (>85%). The presence of native acrylamide was confirmed by mass analysis at m/z = 71 ([C3H5NO]+) and m/z = 55 ([C3H3O]+)

Neuro-cognitive virtual environment for children with autism (VECA).

Autism a neurological disorder which is often diagnosed during early childhood and can cause significant social, communication, and behavioral challenges over a lifetime. It is increasing day by day and people are inclining from clinical and psychological therapies to assistive technologies. We have developed an interactive virtual environment VECA that aims to enhance the cognitive skills and creativity in children with autism by playing games and interacting with the environment. The setup also incorporates the feedback of the child that whether he/she is comfortable with the environment or not. This solution is cost effective, with no side effects unlike traditional therapies, and can provide valuable insight to the behavior analysis of the autism patients

Estimation of the frequency of isoform–genotype discrepancies at the apolipoprotein E locus in heterozygotes for the isoforms

Estimates of the impact of apolipoprotein E (apo E) alleles coding for the three common isoforms on plasma lipid levels assume genetic homogeneity among the genotype classes. To test this assumption, we have determined the apo E genotype at the two common polymorphic sites (amino acids 112 and 158) by DNA amplification and hybridisation with allele‐specific oligoprobes, in 195 unrelated Caucasian participants of the Rochester Family Heart Study previously classified as heterozygotes by isoelectric focusing (IEF). Fourteen discordant samples were initially detected. Repeat typing of these samples by both methods resolved nine discrepancies and analysis of additional blood samples from the remaining five individuals eliminated a further four discrepancies. The only truly discordant allele was found in a female subject who had an E3 isoform with the common E2 (Cys 112 , Cys 158 ) genotype. Transmission of this allele from the mother was demonstrated. From these results, we estimate the frequency of discrepancies between isoforms and common genotypes to be 0.25% in this population. Allele misclassification was caused by poor amplification of the DNA in six samples and superimposition of glycosylated and nonglycosylated apo E isoforms on isoelectric focusing gels in five samples. We conclude that the assumption of genetic homogeneity among genotype classes is valid and that misclassification due to technical difficulties is more frequent than true discordancies. © 1992 Wiley‐Liss, Inc.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/101763/1/1370090403_ftp.pd

High-Level Expression of Various Apolipoprotein (a) Isoforms by "Transferrinfection". The Role of Kringle IV Sequences in the Extracellular Association with Low-Density Lipoprotein

Characterization of the assembly of lipoprotein(a) [Lp(a)] is of fundamental importance to understanding the biosynthesis and metabolism of this atherogenic lipoprotein. Since no established cell lines exist that express Lp(a) or apolipoprotein(a) [apo(a)], a "transferrinfection" system for apo(a) was developed utilizing adenovirus receptor- and transferrin receptor-mediated DNA uptake into cells. Using this method, different apo(a) cDNA constructions of variable length, due to the presence of 3, 5, 7, 9, 15, or 18 internal kringle IV sequences, were expressed in cos-7 cells or CHO cells. All constructions contained kringle IV-36, which includes the only unpaired cysteine residue (Cys-4057) in apo(a). r-Apo(a) was synthesized as a precursor and secreted as mature apolipoprotein into the medium. When medium containing r-apo(a) with 9, 15, or 18 kringle IV repeats was mixed with normal human plasma LDL, stable complexes formed that had a bouyant density typical of Lp(a). Association was substantially decreased if Cys-4057 on r-apo(a) was replaced by Arg by site-directed mutagenesis or if Cys-4057 was chemically modified. Lack of association was also observed with r-apo(a) containing only 3, 5, or 7 kringle IV repeats without "unique kringle IV sequences", although Cys-4057 was present in all of these constructions. Synthesis and secretion of r-apo(a) was not dependent on its sialic acid content. r-Apo(a) was expressed even more efficiently in sialylation-defective CHO cells than in wild-type CHO cells. In transfected CHO cells defective in the addition of N-acetylglucosamine, apo(a) secretion was found to be decreased by 50%. Extracellular association with LDL was not affected by the carbohydrate moiety of r-apo(a), indicating a protein-protein interaction between r-apo(a) and apoB. These results show that, besides kringle IV-36, other kringle IV sequences are necessary for the extracellular association of r-apo(a) with LDL. Changes in the carbohydrate moiety of apo(a), however, do not affect complex formation

Genome-wide copy-number variation study of psychosis in Alzheimer’s disease

About 40–60% of patients with late-onset Alzheimer's disease (AD) develop psychosis, which represents a distinct phenotype of more severe cognitive and functional deficits. The estimated heritability of AD+P is ~61%, which makes it a good target for genetic mapping. We performed a genome-wide copy-number variation (CNV) study on 496 AD cases with psychosis (AD+P), 639 AD subjects with intermediate psychosis (AD intermediate P) and 156 AD subjects without psychosis (AD−P) who were recruited at the University of Pittsburgh Alzheimer's Disease Research Center using over 1 million single-nucleotide polymorphisms (SNPs) and CNV markers. CNV load analysis found no significant difference in total and average CNV length and CNV number in the AD+P or AD intermediate P groups compared with the AD−P group. Our analysis revealed a marginally significant lower number of duplication events in AD+P cases compared with AD−P controls (P=0.059) using multivariable regression model. The most interesting finding was the presence of a genome-wide significant duplication in the APC2 gene on chromosome 19, which was protective against developing AD+P (odds ratio=0.42; P=7.2E−10). We also observed suggestive associations of duplications with AD+P in the SET (P=1.95E−06), JAG2 (P=5.01E−07) and ZFPM1 (P=2.13E−07) genes and marginal association of a deletion in CNTLN (P=8.87E−04). We have identified potential novel loci for psychosis in Alzheimer's disease that warrant follow-up in large-scale independent studies

Genetic risk for schizophrenia and psychosis in Alzheimer disease

Psychotic symptoms, defined as the occurrence of delusions or hallucinations, are frequent in Alzheimer disease (AD), affecting ~40 to 60% of individuals with AD (AD with psychosis (AD+P)). In comparison with AD subjects without psychosis, AD+P subjects have more rapid cognitive decline and poor outcomes. Prior studies have estimated the heritability of psychosis in AD at 61%, but the underlying genetic sources of this risk are not known. We evaluated a Discovery Cohort of 2876 AD subjects with (N=1761) or without psychosis (N=1115). All subjects were genotyped using a custom genotyping array designed to evaluate single-nucleotide polymorphisms (SNPs) with evidence of genetic association with AD+P and include SNPs affecting or putatively affecting risk for schizophrenia and AD. Results were replicated in an independent cohort of 2194 AD subjects with (N=734) or without psychosis (N=1460). We found that AD+P is associated with polygenic risk for a set of novel loci and inversely associated with polygenic risk for schizophrenia. Among the biologic pathways identified by the associations of schizophrenia SNPs with AD+P are endosomal trafficking, autophagy and calcium channel signaling. To the best of our knowledge, these findings provide the first clear demonstration that AD+P is associated with common genetic variation. In addition, they provide an unbiased link between polygenic risk for schizophrenia and a lower risk of psychosis in AD. This provides an opportunity to leverage progress made in identifying the biologic effects of schizophrenia alleles to identify novel mechanisms protecting against more rapid cognitive decline and psychosis risk in AD

Vitamin D binding protein genotype is associated with plasma 25OHD concentration in West African children

Vitamin D is well known for its role in promoting skeletal health. Vitamin D status is determined conventionally by circulating 25-dihydroxyvitamin D (25OHD) concentration. There is evidence indicating that circulating 25OHD concentration is affected by variation in Gc, the gene encoding the vitamin D binding protein (DBP). The composite genotype of two single nucleotide polymorphisms (rs7041 and rs4588) results in different DBP isotypes (Gc1f, Gc1s and Gc2). The protein configurational differences among DBP isotypes affect DBP substrate binding affinity. The aims of this study were to determine 1) Gc variant frequencies in a population from an isolated rural region of The Gambia, West Africa (n=3129) with year-round opportunity for cutaneous vitamin D synthesis and 2) the effects of Gc variants on 25OHD concentration (n=237) in a genetically representative sub-group of children (mean (SD) age: 11.9 (4.8) years). The distribution of Gc variants was Gc1f: 0.86, Gc1s: 0.11 and Gc2: 0.03. The mean (SD) concentration of 25OHD was 59.6 (12.9) nmol/L and was significantly higher in those homozygous for Gc1f compared to other Gc variants (60.7 (13.1) vs. 56.6 (12.1) nmol/L, P=0.03). Plasma 25OHD and 1,25(OH)2D concentration was significantly associated with parathyroid hormone in Gc1f-1f but not in the other Gc variants combined. This study demonstrates that different Gc variants are associated with different 25OHD concentrations in a rural Gambian population. Gc1f-1f, thought to have the highest affinity for 25OHD, had the highest 25OHD concentration compared with lower affinity Gc variants. The considerable difference in Gc1f frequency observed in Gambians compared with other non-West African populations and associated differences in plasma 25OHD concentration, may have implications for the way in which vitamin D status should be interpreted across different ancestral groups