The human genetic mutant cell repository : list of genetic variants, chromosomal aberrations and normal cell cultures submitted to the repository / Institute for Medical Research, Camden, New Jersey.

4th ed.

Genetics for clinicians: from candidate genes to whole genome scans (technological advances).

Human genetics has progressed at an unprecedented pace during the past 10 years. DNA microarrays currently allow screening of the entire human genome with high level of coverage and we are now entering the era of high-throughput sequencing. These remarkable technical advances are influencing the way medical research is conducted and have boosted our understanding of the structure of the human genome as well as of disease biology. In this context, it is crucial for clinicians to understand the main concepts and limitations of modern genetics. This review will describe key concepts in genetics, including the different types of genetic markers in the human genome, review current methods to detect DNA variation, describe major online public databases in genetics, explain key concepts in statistical genetics and finally present commonly used study designs in clinical and epidemiological research. This review will therefore concentrate on human genetic variation analysis.

Functional and transcriptional induction of aquaporin-1 gene by hypoxia; analysis of promoter and role of Hif-1α.

Aquaporin-1 (AQP1) is a water channel that is highly expressed in tissues with rapid O(2) transport. It has been reported that this protein contributes to gas permeation (CO(2), NO and O(2)) through the plasma membrane. We show that hypoxia increases Aqp1 mRNA and protein levels in tissues, namely mouse brain and lung, and in cultured cells, the 9L glioma cell line. Stopped-flow light-scattering experiments confirmed an increase in the water permeability of 9L cells exposed to hypoxia, supporting the view that hypoxic Aqp1 up-regulation has a functional role. To investigate the molecular mechanisms underlying this regulatory process, transcriptional regulation was studied by transient transfections of mouse endothelial cells with a 1297 bp 5' proximal Aqp1 promoter-luciferase construct. Incubation in hypoxia produced a dose- and time-dependent induction of luciferase activity that was also obtained after treatments with hypoxia mimetics (DMOG and CoCl(2)) and by overexpressing stabilized mutated forms of HIF-1α. Single mutations or full deletions of the three putative HIF binding domains present in the Aqp1 promoter partially reduced its responsiveness to hypoxia, and transfection with Hif-1α siRNA decreased the in vitro hypoxia induction of Aqp1 mRNA and protein levels. Our results indicate that HIF-1α participates in the hypoxic induction of AQP1. However, we also demonstrate that the activation of Aqp1 promoter by hypoxia is complex and multifactorial and suggest that besides HIF-1α other transcription factors might contribute to this regulatory process. These data provide a conceptual framework to support future research on the involvement of AQP1 in a range of pathophysiological conditions, including edema, tumor growth, and respiratory diseases.

How accurate is the current picture of human genetic variation?

Our understanding of the distribution of worldwide human genomic diversity has greatly increased over recent years thanks to the availability of large data sets derived from short tandem repeats (STRs), insertion deletion polymorphisms (indels) and single nucleotide polymorphisms (SNPs). A concern, however, is that the current picture of worldwide human genomic diversity may be inaccurate because of biases in the selection process of genetic markers (so-called 'ascertainment bias'). To evaluate this problem, we first compared the distribution of genomic diversity between these three types of genetic markers in the populations from the HGDP-CEPH panel for evidence of bias or incongruities. In a second step, using a very relaxed set of criteria to prevent the intrusion of bias, we developed a new set of unbiased STR markers and compared the results against those from available panels. Contrarily to recent claims, our results show that the STR markers suffer from no discernible bias, and can thus be used as a baseline reference for human genetic diversity and population differentiation. The bias on SNPs is moderate compared to that on the set of indels analysed, which we recommend should be avoided for work describing the distribution of human genetic diversity or making inference on human settlement history.

La biologie moléculaire est-elle utile au praticien [Is molecular biology useful to the practitioner?]

The relative importance of molecular biology in clinical practice is often underestimated. However, numerous procedures in clinical diagnosis and new therapeutic drugs have resulted from basic molecular research. Furthermore, understanding of the physiological and physiopathological mechanisms underlying several human diseases has been improved by the results of basic molecular research. For example, cloning of the gene encoding leptin has provided spectacular insights into the understanding of the mechanisms involved in the control of food intake and body weight maintenance in man. In cystic fibrosis, the cloning and identification of several mutations in the gene encoding the chloride channel transmembrane regulator (CFTR) have resolved several important issues in clinical practice: cystic fibrosis constitutes a molecular defect of a single gene. There is a strong correlation between the clinical manifestations or the severity of the disease (phenotype) with the type of mutations present in the CFTR gene (genotype). More recently, identification of mutations in the gene encoding a subunit of the renal sodium channel in the Liddle syndrome has provided important insight into the physiopathological understanding of mechanisms involved in this form of hereditary hypertension. Salt retention and secondary high blood pressure are the result of constitutive activation of the renal sodium channel by mutations in the gene encoding the renal sodium channel. It is speculated that less severe mutations in this channel could result in a less severe form of hypertension which may correspond to patients suffering from high blood pressure with low plasma renin activity. Several tools, most notably PCR, are derived from molecular research and are used in everyday practice, i.e. in prenatal diagnosis and in the diagnosis of several infectious diseases including tuberculosis and hepatitis. Finally, the production of recombinant proteins at lower cost and with fewer side effects is used in everyday clinical practice. Gene therapy remains an extraordinary challenge in correcting severe hereditary or acquired diseases. The use of genetically modified animal cell lines producing growth factors, insulin or erythropoetin, which are subsequently encapsulated and transferred to man, represents an attractive approach for gene therapy.

Association of MMP-2 polymorphisms with severe and very severe COPD: a case control study of MMPs-1, 9 and 12 in a European population.

BACKGROUND: Genetic factors play a role in chronic obstructive pulmonary disease (COPD) but are poorly understood. A number of candidate genes have been proposed on the basis of the pathogenesis of COPD. These include the matrix metalloproteinase (MMP) genes which play a role in tissue remodelling and fit in with the protease--antiprotease imbalance theory for the cause of COPD. Previous genetic studies of MMPs in COPD have had inadequate coverage of the genes, and have reported conflicting associations of both single nucleotide polymorphisms (SNPs) and SNP haplotypes, plausibly due to under-powered studies. METHODS: To address these issues we genotyped 26 SNPs, providing comprehensive coverage of reported SNP variation, in MMPs- 1, 9 and 12 from 977 COPD patients and 876 non-diseased smokers of European descent and evaluated their association with disease singly and in haplotype combinations. We used logistic regression to adjust for age, gender, centre and smoking history. RESULTS: Haplotypes of two SNPs in MMP-12 (rs652438 and rs2276109), showed an association with severe/very severe disease, corresponding to GOLD Stages III and IV. CONCLUSIONS: Those with the common A-A haplotype for these two SNPs were at greater risk of developing severe/very severe disease (p = 0.0039) while possession of the minor G variants at either SNP locus had a protective effect (adjusted odds ratio of 0.76; 95% CI 0.61 - 0.94). The A-A haplotype was also associated with significantly lower predicted FEV1 (42.62% versus 44.79%; p = 0.0129). This implicates haplotypes of MMP-12 as modifiers of disease severity.

Cap a una ètica per a la genètica: fonamentació

Voldria iniciar aquest article, plantejant dues qüestions bàsiques: perquè cal preguntar quina ètica per a la genètica, i perquè la genètica requereix d'una tal ètica. Pel que fa a la primera qüestió, quina ètica, cal aclarir que ha de ser una....

SINDROME DE DUBOWITZ

A presentation of two cases of Dubowitz syndrome in monozigous twin girls. The syndrome is a rare congenital disorder, the main clinical aspects of which include retarded intrauterine and post-natal growth, microcephaly, peculiar face and an eczemic rash, resulting from photosensitivity of the regions exposed to sunlight.

Canadian Cardiovascular Society 2009 Consensus Conference on the management of adults with congenital heart disease: executive summary

With advances in pediatric cardiology and cardiac surgery, the population of adults with congenital heart disease (CHD) has increased. In the current era, there are more adults with CHD than children. This population has many unique issues and needs. They have distinctive forms of heart failure, and their cardiac disease can be associated with pulmonary hypertension, thromboemboli, complex arrhythmias and sudden death.Medical aspects that need to be considered relate to the long-term and multisystemic effects of single-ventricle physiology, cyanosis, systemic right ventricles, complex intracardiac baffles and failing subpulmonary right ventricles. Since the 2001 Canadian Cardiovascular Society Consensus Conference report on the management of adults with CHD, there have been significant advances in the understanding of the late outcomes, genetics, medical therapy and interventional approaches in the field of adult CHD. Therefore, new clinical guidelines have been written by Canadian adult CHD physicians in collaboration with an international panel of experts in the field. The present executive summary is a brief overview of the new guidelines and includes the recommendations for interventions. The complete document consists of four manuscripts that are published online in the present issue of The Canadian Journal of Cardiology, including sections on genetics, clinical outcomes, recommended diagnostic workup, surgical and interventional options, treatment of arrhythmias, assessment of pregnancy and contraception risks, and follow-up requirements. The complete document and references can also be found at www.ccs.ca or www.cachnet.org.

Xenobiotic Metabolism Genes and Clubfoot

Idiopathic or isolated clubfoot is a common orthopedic birth defect that affects approximately 135,000 children worldwide. It is characterized by equinus, varus and adductus deformities of the ankle and foot. Correction of clubfoot involves months of serial manipulations, castings and bracing, with surgical correction needed in forty percent of cases. Multifactorial etiology has been suggested in numerous studies with both environmental and genetic factors playing an etiologic role. Maternal smoking during pregnancy is the only common environmental factor that has consistently been shown to increase the risk for clubfoot. Moreover, a positive family history of clubfoot and maternal smoking increases the risk of clubfoot twenty fold. These findings suggest that genetic variation in smoking metabolism genes may increase susceptibility to clubfoot. Based on this reasoning, we interrogated eight candidate genes, chosen based on their involvement in phase 1 and 2 cigarette smoke metabolism. Twenty-two SNPs and two null alleles in eight genes (CYP1A1, CYP1A2, CYP1B1, CYP2A6, EPHX1, NAT2, GSTM1 and GSTT1) were genotyped in a dataset composed of nonHispanic white and Hispanic multiplex and simplex families. Only one SNP in CYP1A1, rs1048943, had significantly altered transmission in the aggregate and multiplex NHW datasets (p=0.003 and p=0.009). Perturbation of CYP1A1 by rs1048943 polymorphism causes an increase in the amount of harmful, adduct forming metabolic intermediates. A significant gene interaction between EPHX1 and NAT2 was also found (p=0.007). This interaction may affect the metabolism of harmful metabolic intermediates. Additionally, marginal interactions were found for other xenobiotic genes and these interactions may play a contributory role in clubfoot. Importantly, for CYP1A2, significant maternal (p=0.03; RR=1.24; 95% CI: 1.04-1.44) and fetal (p=0.01; RR=1.33; 95% CI: 1.13-1.54) genotypic effects were identified suggesting that both maternal and fetal genotypes impact normal limb development. No association was found for maternal smoking status and tobacco metabolism genes. Together, these results suggest that xenobiotic metabolism genes may play a contributory role in the etiology of clubfoot regardless of maternal smoking status and may impact foot development through perturbation of tobacco metabolic pathways.

Prevalence of Premature Ovarian Failure in Women with Tuberous Sclerosis

Tuberous Sclerosis Complex (TSC) is an autosomal dominant tumor suppressor disorder characterized by hamartomas, or benign growths, in various organ systems. Inactivating mutations in either the TSC1 or the TSC2 gene cause most cases of TSC. Recently, the use of ovarian specific conditional knock-out mouse models has demonstrated a crucial role of the TSC genes in ovarian function. Mice with complete deletion of Tsc1 or Tsc2 showed accelerated ovarian follicle activation and subsequent premature follicular depletion, consistent with the human condition premature ovarian failure (POF). POF is defined in women as the cessation of menses before the age of 40 and elevated levels of follicle stimulating hormone (FSH). The prevalence of POF is estimated to be 1%, affecting a substantial number of women in the general population. Nonetheless, the etiology of most cases of POF remains unknown. Based on the mouse model results, we hypothesized that the human TSC1 and TSC2 genes are likely to be crucial for ovarian development and function. Moreover, since women with TSC already have one inactivated TSC gene, we further hypothesized that they may show a higher prevalence of POF. To test this hypothesis, we surveyed 1000 women with TSC belonging to the Tuberous Sclerosis Alliance, a national support organization. 182 questionnaires were analyzed for information on menstrual and reproductive function, as well as TSC. This self-reported data revealed 8 women (4.4%) with possible POF, as determined by menstrual history report and additional supportive data. This prevalence is much higher than 1% in the general population. Data from all women suggested other reproductive pathology associated with TSC such as a high rate of miscarriage (41.2%) and menstrual irregularity of any kind (31.2%). These results establish a previously unappreciated effect of TSC on women’s reproductive health. Moreover, these data suggest that perturbations in the cellular pathways regulated by the TSC genes may play an important role in reproductive function.

Gene by BMI Interactions Influencing C-Reactive Protein Levels in European-Americans

C-Reactive Protein (CRP) is a biomarker indicating tissue damage, inflammation, and infection. High-sensitivity CRP (hsCRP) is an emerging biomarker often used to estimate an individual’s risk for future coronary heart disease (CHD). hsCRP levels falling below 1.00 mg/l indicate a low risk for developing CHD, levels ranging between 1.00 mg/l and 3.00 mg/l indicate an elevated risk, and levels exceeding 3.00 mg/l indicate high risk. Multiple Genome-Wide Association Studies (GWAS) have identified a number of genetic polymorphisms which influence CRP levels. SNPs implicated in such studies have been found in or near genes of interest including: CRP, APOE, APOC, IL-6, HNF1A, LEPR, and GCKR. A strong positive correlation has also been found to exist between CRP levels and BMI, a known risk factor for CHD and a state of chronic inflammation. We conducted a series of analyses designed to identify loci which interact with BMI to influence CRP levels in a subsample of European-Americans in the ARIC cohort. In a stratified GWA analysis, 15 genetic regions were identified as having significantly (p-value < 2.00*10-3) distinct effects on hsCRP levels between the two obesity strata: lean (18.50 kg/m2 < BMI < 24.99 kg/m2) and obese (BMI ≥ 30.00 kg/m2). A GWA analysis performed on all individuals combined (i.e. not a priori stratified for obesity status) with the inclusion of an additional parameter for BMI by gene interaction, identified 11 regions which interact with BMI to influence hsCRP levels. Two regions containing the genes GJA5 and GJA8 (on chromosome 1) and FBXO11 (on chromosome 2) were identified in both methods of analysis suggesting that these genes possibly interact with BMI to influence hsCRP levels. We speculate that atrial fibrillation (AF), age-related cataracts and the TGF-β pathway may be the biological processes influenced by the interaction of GJA5, GJA8 and FBXO11, respectively, with BMI to cause changes in hsCRP levels. Future studies should focus on the influence of gene x bmi interaction on AF, age-related cataracts and TGF-β.

Predictors of Contralateral Breast Cancer in BRCA Negative Women

Breast cancer is the most common cancer diagnosis and second leading cause of death in women. Risk factors associated with breast cancer include: increased age, alcohol consumption, cigarette smoking, white race, physical inactivity, benign breast conditions, reproductive and hormonal factors, dietary factors, and family history. Hereditary breast and ovarian cancer syndrome (HBOC) is caused by mutations in the BRCA1 and BRCA2 genes. Women carrying a mutation in these genes are at an increased risk to develop a second breast cancer. Contralateral breast cancer is the most common second primary cancer in patients treated for a first breast cancer. Other risk factors for developing contralateral breast cancer include a strong family history of breast cancer, age of onset of first primary breast cancer, and if the first primary was a lobular carcinoma, which has an increased risk of being bilateral. A retrospective chart review was performed on a select cohort of women in an IRB approved database at MD Anderson Cancer Center. The final cohort contained 572 women who tested negative for a BRCA1 or BRCA2 mutation, had their primary invasive breast cancer diagnosed under the age of 50, and had a BRCAPro risk assessment number over 10%. Of the 572 women, 97 women developed contralateral breast cancer. A number of predictors of contralateral breast cancer were looked at between the two groups. Using univariable Cox Proportional Hazard model, thirteen statistically interesting risk factors were found, defined as having a p-value under 0.2. Multivariable stepwise Cox Proportional Hazard model found four statistically significant variables out of the thirteen found in the univariable analysis. In our study population, the incidence of contralateral breast cancer was 17%. Four statistically significant variables were identified. Undergoing a prophylactic mastectomy was found to reduce the risk of developing contralateral breast cancer, while not having a prophylactic mastecomy, a young age at primary diagnosis, having a positive estrogen receptor status of the primary tumor, and having a family history of breast cancer increased a woman’s risk to develop contralateral breast cancer.

Post-transcriptional Regulation of Mammalian Gene Expression in Non-coding Region of Target RNA

Tumor Suppressor Candidate 2 (TUSC2) is a novel tumor suppressor gene located in the human chromosome 3p21.3 region. TUSC2 mRNA transcripts could be detected on Northern blots in both normal lung and some lung cancer cell lines, but no endogenous TUSC2 protein could be detected in a majority of lung cancer cell lines. Mechanisms regulating TUSC2 protein expression and its inactivation in primary lung cancer cells are largely unknown. We investigated the role of the 5’- and 3’-untranslated regions (UTRs) of the TUSC2 gene in the regulation of TUSC2 protein expression. We found that two small upstream open-reading frames (uORFs) in the 5’UTR of TUSC2 could markedly inhibit the translational initiation of TUSC2 protein by interfering with the “scanning” of the ribosome initiation complexes. Site-specific stem-loop array reverse transcription-polymerase chain reaction (SLA-RT-PCR) verified several micoRNAs (miRNAs) targeted at 3’UTR and directed TUSC2 cleavage and degradation. In addition, we used the established let-7-targeted high mobility group A2 (Hmga2) mRNA as a model system to study the mechanism of regulation of target mRNA by miRNAs in mammalian cells under physiological conditions. There have been no evidence of direct link between mRNA downregulation and mRNA cleavages mediated by miRNAs. Here we showed that the endonucleolytic cleavages on mRNAs were initiated by mammalian miRNA in seed pairing style. Let-7 directed cleavage activities among the eight predicted potential target sites have varied efficiency, which are influenced by the positional and the structural contexts in the UTR. The 5’ cleaved RNA fragments were mostly oligouridylated at their 3’-termini and accumulated for delayed 5’–3’ degradation. RNA fragment oligouridylation played important roles in marking RNA fragments for delayed bulk degradation and in converting RNA degradation mode from 3’–5’ to 5’–3’ with cooperative efforts from both endonucleolytic and non-catalytic miRNA-induced silencing complex (miRISC). Our findings point to a mammalian miRNA-mediated mechanism for the regulation of mRNA that miRNA can decrease target mRNA through target mRNA cleavage and uridine addition