Design of Case-controls Studies with Unscreened Controls

Traditionally in genetic case‐control studies controls have been screened to exclude subjects with a personal history of illness. This control group has the advantage of optimal power to detect loci involved in illness, but requires more work and may incur substantial cost in recruitment. An alternative approach to screening is to use unscreened controls sampled from the general population. Such controls are generally plentiful and inexpensive, but in general there is a risk that some may have the same disease as the cases, which will reduce power to detect associations. We have quantified the extent of this power loss, and produced mathematical formulae for the number of unscreened controls necessary to achieve the same power as a fixed sample of screened controls. The effect of using unscreened controls will also depend on the ratio of the number of screened controls to cases specified in the original study design, and this is also investigated. We have also investigated the cost‐benefits of the screened and unscreened approaches, according to variation in the relative costs of sampling screened and unscreened controls, together with genotyping costs. We have, thus, identified the range of situations in which using unscreened controls is a cost‐effective alternative to the screened control method and could be considered when designing a study. In many of the typical, real‐world situations in complex genetics, the use of unscreened controls is potentially cost‐effective and can, in general, be considered for disorders with population prevalence K_p < 0.2. With the steady reduction in genotyping costs and the availability of common sets of “population controls” this design is likely to become increasingly cost effective.     

Detection of the circulation of Crimean-Congo hemorrhagic fever virus in the piedmont steppes of the North Caucasus

The laboratory verified cases of Crimean-Congo hemorrhagic fever (CCHF) in the piedmont steppes of the North Caucasus (Malgobeksky District, Republic of Ingushetia) are first described. The source of the first infection was Ixodidae ticks; three subsequent sources were contacts with the bloody discharges from patients. CCHF virus genome was detected in the blood of the cattle from an epidemic focus and in the pools of the Ixodes ticks Haemaphysalis parva Neum., 1897 and Boophilus annulatus Say, 1821, taken from cattle. The problem of including the piedmont steppes of the North Caucasus into the CCHF nosological area is discussed.     

Inhibition of adenylyl cyclase by neuronal P2Y receptors

P2Y receptors inhibiting adenylyl cyclase have been found in blood platelets, glioma cells, and endothelial cells. In platelets and glioma cells, these receptors were identified as P2Y(12). Here, we have used PC12 cells to search for adenylyl cyclase inhibiting P2Y receptors in a neuronal cellular environment. ADP and ATP (0.1 - 100 microM) left basal cyclic AMP accumulation unaltered, but reduced cyclic AMP synthesis stimulated by activation of endogenous A(2A) or recombinant beta(2) receptors. Forskolin-dependent cyclic AMP production was reduced by ADPbetaS (71 nM)>ATP (164 nM)=ADP (244 nM). The inhibition by ADP was not antagonized by suramin, pyridoxal-phosphate-6-azophenyl-2',4'-disulphonic acid, or adenosine-3'-phosphate-5'-phosphate, but attenuated by reactive blue 2, ATP(alpha)S, and 2-methylthio-AMP. RT - PCR demonstrated the expression of P2Y(2), P2Y(4), P2Y(6), and P2Y(12), but not P2Y(1), receptors in PC12 cells. In Northern blots, only P2Y(2) and P2Y(12) were detectable. Differentiation with NGF did not alter these hybridization signals and left the nucleotide inhibition of adenylyl cyclase unchanged. We conclude that P2Y(12) receptors are expressed in neuronal cells and inhibit adenylyl cyclase activity.     

Streamlined analysis of pooled genotype data in SNP-based association studies

Several groups have developed methods for estimating allele frequencies in DNA pools as a fast and cheap way for detecting allelic association between genetic markers and disease. To obtain accurate estimates of allele frequencies, a correction factor k for the degree to which measurement of allele-specific products is biased is generally applied. Factor k is usually obtained as the ratio of the two allele-specific signals in samples from heterozygous individuals, a step that can significantly impair throughput and increase cost. We have systematically investigated the properties of k through the use of empirical and simulated data. We show that for the dye terminator primer extension genotyping method we have applied, the correction factor k is substantially influenced by the dye terminators incorporated, but also by the terminal 3' base of the extension primer. We also show that the variation in k is large enough to result in unacceptable error rates if association studies are conducted without regard to k. We show that the impact of ignoring k can be neutralized by applying a correction factor k(max) that can be easily derived, but this at the potential cost of an increase in type I error. Finally, based upon observed distributions for k we derive a method allowing the estimation of the probability pooled data reflects significant differences in the allele frequencies between the subjects comprising the pools. By controlling the error rates in the absence of knowledge of the appropriate SNP-specific correction factors, each approach enhances the performance of DNA pooling, while considerably streamlining the method by reducing time and cost.     

Allelic expression of APOE in human brain: effects of epsilon status and promoter haplotypes

The epsilon4 haplotype of APOE is the only undisputed genetic risk factor for late-onset Alzheimer's disease (LOAD). It has been proposed that at least two other polymorphisms in the promoter of the APOE gene (-219G>T and -491A>T) might also contribute to disease susceptibility, and modulate the impact of structural changes in the ApoE protein, by altering its expression. In order to assess the extent of cis-acting influences on APOE expression in human brain, highly quantitative measures of allele discrimination were applied to cortical RNA from individuals heterozygous for the epsilon alleles. A small, but significant, increase in the expression of epsilon4 allele was observed relative to that of the epsilon3 and epsilon2 alleles (P<0.0001). Similar differences were observed in brain tissue from confirmed LOAD subjects, and between cortical regions BA10 (frontopolar) and BA20 (inferior temporal). Stratification of epsilon4/epsilon3 allelic expression ratios according to heterozygosity for the -219G>T promoter polymorphism revealed significantly lower relative expression of haplotypes containing the -219T allele (P=0.02). Our data indicate that, in human brain, most of the cis-acting variance in APOE expression is accounted for by the epsilon4 haplotype, but there are additional, small, cis-acting influences associated with promoter genotype.     

Familiality of symptom dimensions in depression

BACKGROUND: Depression is a clinically heterogeneous disorder thought to result from multiple genes interacting with environmental and developmental components. A dimensional rather than a categorical approach to depressive phenotype definition may be more useful for identification of susceptibility genes. OBJECTIVES: To perform an exploratory factor analysis on a range of depressive and anxiety symptoms in a large, well-defined sample of depressed siblings, as well as a confirmatory factor analysis in a separate large group of unrelated depressed subjects, and to analyze correlations of identified symptom dimensions between depressed siblings. DESIGN: Subjects (N = 1034), including 475 sibling pairs, with a history of at least 2 depressive episodes were recruited from the Depression Network Study, a large-scale multicenter collection of families affected by recurrent unipolar depression. Subjects were interviewed using the Schedules for Clinical Assessment in Neuropsychiatry (SCAN) and diagnosed according to the DSM-IV and the International Classification of Diseases, 10th Revision, using a computerized scoring program (CATEGO5). Factor analysis was carried out on 26 depression symptom items, including 4 anxiety screening items. Confirmatory factor analysis was performed on an independent sample of 485 depressed individuals. RESULTS: Four interpretable factors were identified: (1) mood symptoms and psychomotor retardation; (2) anxiety; (3) psychomotor agitation, guilt, and suicidality; and (4) appetite gain and hypersomnia. For each symptom group, a quantitative scale was constructed, and correlations between siblings were calculated. There was a moderate degree of sibling homotypia for some depressive symptoms, and factors 1, 2, and 3 showed significant positive familial correlation (0.145 [P =.001], 0.335 [P<.001], and 0.362 [P<.001], respectively). CONCLUSIONS: This is the first study of large, well-defined samples of depressed subjects in whom symptom dimensions have been derived and then confirmed using independent material. The significant correlations between siblings for 3 of the dimensions suggest substantial familial, perhaps genetic, etiologies.     

Effectiveness of treating periodontitis in patients with thyroid dysfunction

Efficiency of some drugs in the treatment of periodontitis in combination with corrective treatment of thyroid function was evaluated in 70 patients with hypo- and hyperthyrosis with different initial level of nonspecific resistance. The therapeutic complex including drugs commonly used in the treatment of periodontitis and irrigation of the periodontium with lithium chloride and chlorohexidine solutions was highly effective in patients with thyroid dysfunction and relatively favorable status of nonspecific resistance of the organism. In patients with hypo- and hyperthyrosis with poor nonspecific resistance the best effect in the treatment of periodontitis was attained with potassium orotate as an immunomodulator and lithium chloride.