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HLA-G and HLA-E polymorphisms in an indigenous African population. The ZVITAMBO Study Group.

Studies of HLA-G and HLA-E polymorphisms in different populations from many industrialized countries have uniformly reported little sequence variation within these genes. To date, the polymorphism of these genes has not been characterized in populations from developing countries where more sequence variation would be anticipated due to greater exposure to microbial pathogens. In order to address this issue, we have recently investigated the polymorphism of these MHC genes in an indigenous African population (Shona ethnic group). HLA-G and HLA-E alleles were determined by PCR-restriction fragment length polymorphism (PCR-RFLP) and amplification refractory mutation system (ARMS) analysis in 108 unrelated women recruited from maternity clinics and hospitals in Harare (Zimbabwe). The genotyping analysis identified six different HLA-G alleles and three HLA-E alleles in this population. Of interest, the null-allele HLA-G*0105N was found with 11.1% of frequency, which is higher than in other populations tested so far. We observed the presence of 15 distinct HLA-G genotypes and 6 HLA-E genotypes. These data indicate that this African population contains low levels of allelic polymorphism similar to ethnic groups from industrialized countries. This is the first report describing HLA-G and HLA-E polymorphisms in an indigenous African population.

Africa↗

Molecular characterisation of type 1 Gaucher disease families and patients: intrafamilial heterogeneity at the clinical level.

Type 1 Gaucher disease families were studied in an attempt to establish a phenotype/genotype correlation in affected persons and also to identify carriers accurately. In the Portuguese type 1 Gaucher patients, screening for mutations N370S, L444P, R463C, and 1066 + 1 G-->A allowed the identification of 85% of the alleles among unrelated patients. A subclinical case with genotype N370S/1066 + 1 G-->A was identified in one family in which there were three other symptomatic sibs. To our knowledge this is the first subclinical case with a genotype other than N370S/N370S. No genotype-phenotype correlation could be established and considerable clinical heterogeneity was found even among sibs with the same genotype. The data collected on the origins of the Gaucher families indicated two areas in northern Portugal where a higher frequency of the disease may be expected to exist.

Adult↗

Estimation of haplotype frequencies, linkage-disequilibrium measures, and combination of haplotype copies in each pool by use of pooled DNA data.

Inference of haplotypes is important for many genetic approaches, including the process of assigning a phenotype to a genetic region. Usually, the population frequencies of haplotypes, as well as the diplotype configuration of each subject, are estimated from a set of genotypes of the subjects in a sample from the population. We have developed an algorithm to infer haplotype frequencies and the combination of haplotype copies in each pool by using pooled DNA data. The input data are the genotypes in pooled DNA samples, each of which contains the quantitative genotype data from one to six subjects. The algorithm infers by the maximum-likelihood method both frequencies of the haplotypes in the population and the combination of haplotype copies in each pool by an expectation-maximization algorithm. The algorithm was implemented in the computer program LDPooled. We also used the bootstrap method to calculate the standard errors of the estimated haplotype frequencies. Using this program, we analyzed the published genotype data for the SAA (n=156), MTHFR (n=80), and NAT2 (n=116) genes, as well as the smoothelin gene (n=102). Our study has shown that the frequencies of major (frequency >0.1 in a population) haplotypes can be inferred rather accurately from the pooled DNA data by the maximum-likelihood method, although with some limitations. The estimated D and D' values had large variations except when the /D/ values were >0.1. The estimated linkage-disequilibrium measure rho2 for 36 linked loci of the smoothelin gene when one- and two-subject pool protocols were used suggested that the gross pattern of the distribution of the measure can be reproduced using the two-subject pool data.

Algorithms↗

PPARgamma and colon and rectal cancer: associations with specific tumor mutations, aspirin, ibuprofen and insulin-related genes (United States).

We hypothesize that the peroxisome proliferator-activated receptor-gamma (PPARgamma) is associated with colorectal cancer given its association with insulin, diabetes, obesity, and inflammation. In this study, we evaluated the association between colorectal cancer and specific tumor mutations and the Pro12Ala (P12A) PPARgamma polymorphism. We also evaluated interactions between the PPARgamma gene and other insulin-related genes and use of aspirin and non-steroidal anti-inflammatory drug use. Data were available from 1,577 cases of colon cancer that were matched to 1,971 population-based controls and 794 cases of rectal cancer that were matched to 1,001 population-based controls. Colon tumors from the case subjects were evaluated for p53 and Ki-ras mutations and microsatellite instability (MSI). Insulin-related genes evaluated were the Bsm1, polyA, and Fok1 polymorphisms of the VDR gene; the G972R IRS1 polymorphism; the G1057D IRS2 polymorphism; the 19CA repeat polymorphism of the IGF1 gene; and the -200A>C IGFBP3 polymorphism. The odds ratio (OR) between the PA/AA genotypes and proximal tumors was 0.83 (95% CI: 0.69-1.01); for distal tumors was 1.00 (95% CI: 0.83-1.21); and for rectal tumors was 1.04 (95% CI: 0.86-1.25). Evaluation of specific types of tumor mutations showed that colon cancer cases with the PA or AA genotypes were less likely to have p53 tumor mutations (OR 0.78; 95% CI: 0.62-0.99), specifically transition mutations (OR 0.74; 95% CI: 0.56-0.97). Colon cancer cases also were less likely to have a tumor with MSI if they had the PA or AA PPARgamma genotype (OR 0.68; 95% CI: 0.47-0.98); differences in Ki-ras mutations were not seen in colon tumors by PPARgamma genotype. Those who did not take ibuprofen-type drugs and had the PA or AA genotypes were at a significantly greater risk of rectal cancer (OR 2.11; 95% CI: 1.52-2.92; p interaction 0.03) than people with the PP genotype regardless of ibuprofen-type drug use. There was a significant interaction between the -200A>C IGFBP3 polymorphism and the Pro12Ala PPARgamma polymorphism and risk of colon cancer (p for interaction = 0.02) with individuals being at significantly lower risk if they had both the CC IGFBP3 genotype and the PA/AA PPARgamma genotype. For rectal cancer there was a significant interaction between the Bsm1/polyA polymorphisms (p = 0.001) of the VDR gene and the PA/AA Pro12Ala PPARgamma polymorphism with the highest risk group being those with both the PA/AA Pro12Ala PPARgamma and the BB/SS VDR genotypes. These data suggest that PPARgamma may be associated with many aspects of colorectal cancer including insulin- and inflammation-related mechanisms.

Adult↗

Case/pseudocontrol analysis in genetic association studies: A unified framework for detection of genotype and haplotype associations, gene-gene and gene-environment interactions, and parent-of-origin effects.

Estimation and testing of genetic effects (genotype relative risks) are often performed conditionally on parental genotypes, using data from case-parent trios. This strategy avoids having to estimate nuisance parameters such as parental mating type frequencies, and also avoids generating spurious results due to confounding causes of association such as population stratification. For effects at a single locus, the resulting analysis is equivalent to matched case/control analysis via conditional logistic regression, using the case and three "pseudocontrols" derived from the untransmitted parental alleles. We previously showed that a similar approach can be used for analyzing genotype and haplotype effects at a set of closely linked loci, but with a required adjustment to the conditioning argument that results in varying numbers of pseudocontrols, depending on the disease model that is to be fitted. Here we extend this method to include the analysis of epistatic effects (gene-gene interactions) at unlinked loci, to include parent-of-origin effects at one or more loci, and to allow additional incorporation of gene-environment interactions. The conditional logistic approach provides a natural and flexible framework for incorporating these additional effects. By relaxing the conditioning on parental genotypes to allow exchangeability of parental genotypes, we show how the power of this approach can be increased when studying parent-of-origin effects. Simulations suggest that there is limited power to distinguish between parent-of-origin effects and effects due to interaction between genotypes of mother and child.

Algorithms↗

Genotypic and phenotypic characterization of acyclovir-resistant herpes simplex viruses isolated from haematopoietic stem cell transplant recipients.

Thirty-one herpes simplex virus type one (HSV-1) isolates from 12 haematopoietic stem cell transplant recipients with persistent HSV infections despite acyclovir (ACV) prophylaxis or treatment, were genotypically and phenotypically characterized. The relationship between drug susceptibility of the isolates and mutations in thymidine kinase (TK) and DNA polymerase (DNA pol) genes was examined. In all 12 patients, HSV infections were due to ACV-resistant, foscarnet-sensitive viruses. Out of 31 isolates examined, 23 were resistant and eight were sensitive to ACV; eight patients carried viruses with frameshift mutations in the TK gene (due to addition or deletion of single nucleotides in homopolymeric repeats). These mutations were found at codon 61 (G deletion, one patient), 146 (G insertion, five patients) and 153 or 185 (C deletion, one patient each). In four patients, viruses were selected during ACV therapy that contained novel amino acid substitutions in the TK gene (H58R, G129D, A189V, R216H, R220C). Their possible role in ACV resistance was further confirmed phenotypically and by the absence of any resistance-associated mutations in the DNA pol gene. These substitutions were located in ATP- or nucleoside-binding sites or in conserved regions of the TK gene. In addition, a single mutation, Q570R, in the delta-region C of the DNA pol gene, was identified in an isolate from a single patient with resistance to ACV. Our study confirms and expands previous data on genotypic changes associated with ACV resistance of HSV-1 clinical isolates.

Acyclovir↗

Dietary folate is associated with p16(INK4A) methylation in head and neck squamous cell carcinoma.

Inactivation of the p16(INK4A) (CDKN2A) gene in the Rb pathway is among the most common somatic alterations observed in tobacco-related solid tumors, including head and neck squamous cell carcinoma (HNSCC). In addition, a low folate diet is an important risk factor for HNSCC. Decreased dietary folate in an animal model of hepatocellular carcinoma has been associated with the induction of epigenetic silencing of the p16(INK4A) gene. In an ongoing population-based study of HNSCC, we sought to extend this observation to human disease testing the hypothesis that p16(INK4A) methylation is associated with decreased dietary folate. We also investigated the association of methylation silencing with functional polymorphisms in the folate metabolism enzyme methylene tetrahydrofolate reductase (MTHFR). In 169 HNSCCs, the odds ratio for p16(INK4A) methylation among those with low dietary folate intake was 2.3 (95% CI = 1.1-4.8) when compared with those with high folate intake. Furthermore, this increased risk for epigenetic silencing at p16(INK4A) was modified by the MTHFR alleles previously associated with diminished serum folate levels. Hence, in HNSCC low dietary intake of folate is associated with p16(INK4A) methylation, and this relationship is modified by the MTHFR genotype. Our data provides important evidence for a mechanism of action of folate deficiency in cancer.

Adult↗

[Preimplantation embryonic mortality--method of studying interstrain differences in the reparative activity of mouse ova].

Cytological analysis of preimplantation embryonic death in 101/HY, C57BL/6JY and CBA/lacY females crossed with hybird males F1 (BALB/cYxDBA/2Y) was carried out. Embryonic death was induced by thiophosphamide at a dose of 2 mg/kg, i. p. The maximum preimplantation death was recorded in 101/HY females (38.8%), the minimum in CBA/LacY females (21.9%). In C57BL/6JY females, the maximum preimplantation death accounted for 31.3%. Thus the same chromosome damage induced by thiophosphamide in late spermatids of F1CD2 males caused quantitative differences in embryonic mortality in females of different genotypes. The data obtained evidence that fertilized eggs are capable of repairing part of damage induced by paternal chromosomes. It was demonstrated that the preimplantation embryonic death can be used for studying strain differences from the reparative activity of mouse eggs.

Animals↗

Renal hemodynamic changes induced by captopril and angiotensin-converting enzyme gene polymorphism.

We studied the relationship between renal hemodynamic changes induced by a single acute administration of captopril (50 mg p.o.) and angiotensin-converting enzyme (ACE) gene insertion/deletion (I/D) polymorphism in 27 healthy human volunteers, 7 with DD genotype, 10 with ID, and 10 with II genotype. The increase in effective renal plasma flow (p < 0.02) and the fall in renal vascular resistance (p < 0.01) in response to captopril were significantly less in subjects with the DD genotype than in subjects with the other genotypes. These data suggest that intrarenal ACE inhibition by captopril differs according to ACE gene ID polymorphism in healthy humans.

Adult↗

The role of mdr2 in manganese-bilirubin induced cholestasis in mice.

The pathogenesis of manganese-bilirubin (Mn-BR) induced cholestasis has only been studied in rats and is associated with alteration in the hepatic homeostasis of cholesterol and phospholipids. Multidrug resistance-2 (mdr2) transporter, which mediates excretion of these lipids, is suggested to be involved in this phenomenon. The present study was undertaken to examine if Mn-BR induced cholestasis is reproducible in mice, then to clarify the role of mdr2 in its pathogenesis, using mice with disrupted mdr2 gene (mdr2 (-/-)). Results showed that Mn-BR combination decreased bile flow in mice. This reduction in bile flow was similar in mdr2 (-/-) and the wild type mdr2 (+/+). Furthermore, the change in biliary lipid excretion was comparable in both genotypes. These data indicate that Mn-BR induced cholestasis is reproducible in mice and provide evidence that mdr2 alteration is not a primary event in this form of cholestasis.

ATP Binding Cassette Transporter, Subfamily B↗

The relationship between a polymorphism in CYP17 with plasma hormone levels and breast cancer.

The A2 allele of CYP17 has been associated with polycystic ovarian syndrome, elevated levels of certain steroid hormones in premenopausal women, and increased breast cancer risk. We prospectively assessed the association between the A2 allele of CYP17 and breast cancer risk in a case-control study nested within the Nurses' Health Study cohort. We also evaluated associations between this CYP17 genotype and plasma steroid hormone levels among postmenopausal controls not using hormone replacement to assess the biological significance of this genetic variant. Women with the A2 allele were not at an increased risk of incident breast cancer [OR (odds ratio), 0.85; 95% CI (confidence interval), 0.65-1.12] or advanced breast cancer (OR, 0.84; 95% CI, 0.54-1.32). We did observe evidence that the inverse association of late age at menarche with breast cancer may be modified by the CYP17 A2 allele. The protective effect of later age at menarche was only observed among women without the A2 allele (A1/A1 genotype: for age at menarche > or =13 versus <13; OR, 0.57; 95% CI, 0.36-0.90; A1/A2 and A2/A2 genotypes: OR, 1.05; 95% CI, 0.76-1.45; P for interaction = 0.07). Among controls, we found women with the A2/A2 genotype to have elevated levels of estrone (+14.3%, P = 0.01), estradiol (+13.8%, P = 0.08), testosterone (+8.6%, P = 0.34), androstenedione (+17.1%, P = 0.06), dehydroepiandrosterone (+14.4%, P = 0.02), and dehydroepiandrosterone sulfate (+7.2%, P = 0.26) compared with women with the A1/A1 genotype. These data suggest that the A2 allele of CYP17 modifies endogenous hormone levels, but is not a strong independent risk factor for breast cancer.

Adult↗

Gene-environment interaction in the determination of levels of haemostatic variables involved in thrombosis and fibrinolysis.

Although a specific genotype may be associated in healthy subjects with modest differences in levels of a risk factor for thrombosis, this effect may be larger or smaller in subgroups of subjects. Documenting such gene-environment interactions is important if genotype information is ever to be used in a clinical or diagnostic setting, and understanding the molecular mechanisms of such interactions is vital to be able to use the information to develop novel therapeutic approaches for reducing the risk of myocardial infarction. This review focuses on gene-environment interactions detected to date for the G-455A beta fibrinogen gene promoter polymorphism. Carriers of the A allele, representing roughly 20% of the population, consistently have on average 7-10% higher fibrinogen levels than those with the genotype GG. Data will be presented to demonstrate interaction between gender, smoking habit, age, presence of ischaemic disease and level of physical exercise, in the determination of the magnitude of this A-raising effect and thus on an individual's plasma fibrinogen levels.

Adult↗

Dopamine D4 receptor 48-bp repeat polymorphism: no association with response to antipsychotic treatment, but association with catatonic schizophrenia.

The dopamine D4 receptor (DRD4) may play a role in the pathogenesis of neuropsychiatric disease and in the action of dopaminergic drugs. The 48-bp repeat polymorphism (48-bp VNTR) coding for a 16-amino acid segment in the third cytoplasmatic loop of the DRD4 was studied as a predictor of the therapeutic response to antipsychotics and as susceptibility factor for schizophrenia. We included 638 in-patients with acute schizophrenic, schizoaffective (mainly schizophrenic), and other nonaffective psychoses, as well as two reference groups: one with 278 in-patients with non-psychiatric diseases, and one with 474 healthy volunteers. Catatonic patients (DSM-IV 295.2) more frequently carried the DRD4 D4.2 and D4.3 allele than did all other schizophrenic cases (P < 0.001; OR: 2.7; CI: 1.5-4.9) and controls (P < 0.004; OR: 2.3; CI: 1.3-4.2). We found no significant difference in the DRD4 allele or in genotype frequencies in our comparison of all schizophrenic patients and controls. The subgroups with affected family members, and the subgroups with early or late onset of disease, also did not differ from the controls in DRD4 allele frequencies. The 48-bp VNTR was not a predictor for therapeutic outcome measured by the positive and negative symptoms scale. A total of 1390 subjects showed between 1 and 10 repeats (D4. 1 and D4.10), with 25 different genotypes. These data exclude a major role of DRD4 48-bp VNTR in response to antipsychotic therapy and as susceptibility factor for schizophrenia, but catatonic schizophrenia may be associated with the D4.2 and D4.3 alleles.

Adolescent↗

Differences in the incorporation of thymidine into DNA of normal and cystic fibrosis fibroblasts in vitro.

Although similar fractions of cells were in the S phase of the cell cycle, normal human skin fibroblasts were shown to incorporate more than twice the 3HTdR into their DNA in vitro than did cells obtained from individuals with cystic fibrosis (CF). Obligate heterozygotes incorporated an intermediate amount of the DNA precursor. Studied were initiated to determine the basis of the differential incorporation of 3HTdR among the genotypes. An analog of thymidine, BUdR, produced varied effects on the growth kinetics of the three genotypes. The growth of cells in BUdR resulted in a 50% increase in the population doubling times of all three genotypes, and caused the cell morphology to change from a spindle shape to one in which the cells became broadened and flat, with numerous cytoplasmic projections extending for distances of several cell diameters. The activities of thymidine kinase and the participation of the exogenous and de novo pathways in the synthesis of TMP were found to be approximately the same in all three genotypes. The data suggest that an alteration in the transport of thymidine into the cells may account for the differences in TdR incorporation into DNA, and this may be associated with other changes in cystic fibrosis that are apparently membrane associated.

Bromodeoxyuridine↗

HLA association with dermatitis herpetiformis is accounted for by a cis or transassociated DQ heterodimer.

HLA-DR, DQ, and DP restriction fragment genotyping was undertaken in 23 dermatitis herpetiformis patients and 53 healthy control subjects. HLA-DQw2 was present in 100% of patients with dermatitis herpetiformis (23 of 23) versus 40% of control subjects (21 of 53). Significant secondary associations occurred with HLA-DR3 (91% of patients versus 28% of control subjects) and DPw1 (39% of patients versus 11% of control subjects). Dermatitis herpetiformis and coeliac disease thus share an identical HLA class II association. It is likely that HLA class II genes directly influence the immune responses leading to mucosal damage in both diseases. The strongest candidate for disease susceptibility to dermatitis herpetiformis is DQw2. The HLA molecule most likely to be involved in coeliac disease is a specific DQ alpha/DQ beta heterodimer, encoded in cis arrangement in DR3 haplotypes or in trans arrangement in a DR5, 7 genotype. Our data on dermatitis herpetiformis patients fits this model perfectly. All these patients are capable of expressing this molecule, which may be responsible for the gluten sensitive enteropathy seen in a subgroup of patients with dermatitis herpetiformis and coeliac disease.

Celiac Disease↗

A common polymorphism in the 5'-untranslated region of the VEGF gene is associated with diabetic retinopathy in type 2 diabetes.

Vascular endothelial growth factor (VEGF), a major mediator of vascular permeability and angiogenesis, may play a pivotal role in mediating the development and progression of diabetic retinopathy. In the present study, we examined the genetic variations of the VEGF gene to assess its possible relation to diabetic retinopathy in type 2 diabetic patients. Among seven common polymorphisms in the promoter region, 5'-untranslated region (UTR) and 3'UTR of the VEGF gene, genotype distribution of the C(-634)G polymorphism differed significantly (P = 0.011) between patients with (n = 150) and without (n = 118) retinopathy, and the C allele was significantly increased in patients with retinopathy compared with those without retinopathy (P = 0.0037). The odds ratio (OR) for the CC genotype of C(-634)G to the GG genotype was 3.20 (95% CI 1.45-7.05, P = 0.0046). The -634C allele was significantly increased in patients with nonproliferative diabetic retinopathy (non-PDR) (P = 0.0026) and was insignificantly increased in patients with proliferative diabetic retinopathy (PDR) (P = 0.081) compared with patients without retinopathy, although frequencies of the allele did not differ significantly between the non-PDR and PDR groups. Logistic regression analysis revealed that the C(-634)G polymorphism was strongly associated with an increased risk of retinopathy (P = 0.0018). Furthermore, VEGF serum levels were significantly higher in healthy subjects with the CC genotype of the C(-634)G polymorphism than in those with the other genotypes. These data suggest that the C(-634)G polymorphism in the 5'UTR of the VEGF gene is a novel genetic risk factor for diabetic retinopathy.

5' Untranslated Regions↗

Replication and recombination in adenovirus-infected cells are temporally and functionally related.

We have studied the temporal and functional relationships between DNA replication and recombination in adenovirus-infected cells by using Southern blot hybridization to detect recombinant products among intracellular viral genomes. The data show that recombination can be detected soon after DNA replication has commenced and that the proportion of recombinant products increases thereafter. To determine the functional relationship between DNA replication and recombination, replication was blocked with the protein synthesis inhibitor anisomycin, the replication inhibitor cytosine arabinoside, and conditionally lethal mutations in either the virus-specified DNA-binding protein or the DNA polymerase. All treatments that directly or indirectly blocked DNA replication caused a delay in the appearance of recombinant products and a marked decline in their abundance relative to products of parental genotype. These data strongly suggest that DNA replication and recombination are interrelated, either because both processes share functions or because DNA structures produced by replication are suitable substrates for recombination. In addition, we have shown that some recombination function(s) is intrinsically thermolabile at 40.9 degrees C, even in wild-type crosses, since the appearance of recombinant products is delayed and their extent is reduced compared with that from crosses performed at 39.9 degrees C.

Adenoviruses, Human↗

Association between severity of alcoholism and the A1 allele of the dopamine D2 receptor gene TaqI A RFLP in Japanese.

The allelic association of TaqI A restriction fragment length polymorphism (RFLP) of the dopamine D2 receptor gene with alcoholism was examined in 78 Japanese alcoholics and compared with Japanese controls. A significantly higher frequency of the A1 allele (0.42) was found in 100 Japanese unscreened controls compared with those reported in white populations. Among 70 alcoholics whose severities were determined, the A1 allele was present in 77% of 43 more severe alcoholics and in 59% of 27 less severe alcoholics. The A1 allele was present significantly less frequently in the alcoholics at the age of 60 or older (42%), compared with those under the age of 60 (74%). In the subjects under the age of 60, the A1 allele was present in 83% of the 35 more severe alcoholics, being significantly more frequent than in 60% of the 35 nonalcoholic controls. All of the 7 alcoholics homozygous for the A1 allele were classified as severe. The average severity of alcoholism increased in the order A2/A2, A1/A2, and A1/A1 genotypes. These data suggest that the A1 allele is associated with severe alcoholism in the Japanese population and that the effect is related to or has a linkage disequilibrium with a genetic factor that has a small but not negligible additive effect on alcoholism.

Age Factors↗