Pre-diagnostic high-sensitive C-reactive protein and breast cancer risk, recurrence, and survival.

Inflammation may initiate and promote breast cancer development, and be associated with elevated circulating levels of inflammation markers. A total of eight 130 initially healthy women, participated in the population-based Tromsø study (1994-2008). Pre-diagnostic high-sensitivity C-reactive protein (hs-CRP) was assessed. During 14.6 years of follow-up, a total of 192 women developed invasive breast cancer. These cases were followed for additional 7.2 years. Detailed medical records were obtained. We observed an overall positive dose-response relationship between pre-diagnostic hs-CRP and breast cancer risk (hazard ratio (HR) = 1.06, 95 % CI 1.01-1.11). Postmenopausal women with above median levels of hs-CRP (>1.2 mg/l) had a 1.42 (95 % CI 1.01-2.00) higher breast cancer risk compared to postmenopausal women with hs-CRP below median. Postmenopausal women, who were hormone replacement therapy non-users, and were in the middle tertile (0.8-1.9 mg/l), or highest tertile of hs-CRP (>1.9 mg/l), had a 2.31 (95 % CI 1.31-4.03) and 2.08 (95 % CI 1.16-3.76) higher breast cancer risk, respectively, compared with women in the lowest tertile. For each unit increase in pre-diagnostic hs-CRP levels (mg/l), we observed an 18 % increase in disease-free interval (95 % CI 0.70-0.97), and a 22 % reduction in overall mortality (95 % CI 0.62-0.98). Our study supports a positive association between pre-diagnostic hs-CRP and breast cancer risk. In contrast, increased pre-diagnostic hs-CRP was associated with improved overall mortality, but our findings are based on a small sample size, and should be interpreted with caution.

Elevated odds of metabolic syndrome in psoriasis: a population-based study of age and sex differences.

BACKGROUND: Questions remain concerning to what extent age and sex may modify the suggested association between psoriasis and the metabolic syndrome in the general population. OBJECTIVES: To investigate the association between psoriasis and the metabolic syndrome within a large population-based cohort by age and sex. METHODS: A cross-sectional study including 10 521 participants aged 30-79 years from the Tromsø Study cohort was performed; 1137 participants reported lifetime psoriasis of a mainly mild character. The new harmonized definition of metabolic syndrome was used in the multivariable logistic regression analysis. RESULTS: There was a uniformly higher prevalence of metabolic syndrome in men and women with psoriasis compared with those without across all age groups. In women, psoriasis was associated with a 3·8-times higher odds of metabolic syndrome at age 30 years (95% confidence interval 1·5-9·7), with a decreasing odds ratio with increasing age. In men, psoriasis was associated with a stable 1·35-times higher odds of metabolic syndrome (95% confidence interval 1·1-1·6) at all ages. Abdominal obesity was the most frequent metabolic syndrome component in women in this study, and there was indication of a dose-response relationship between psoriasis severity, indicated through treatment, and having a high waistline in women. CONCLUSIONS: This study suggests age and sex variations in the risk of metabolic syndrome among individuals with psoriasis. Given the high prevalence of psoriasis and the significantly elevated burden of metabolic syndrome in this patient group, there may be a benefit from targeted screening of metabolic syndrome among individuals with psoriasis regardless of age and disease severity.

Association studies in QTL regions linked to bovine trypanotolerance in a West African crossbred population.

African animal trypanosomosis is a parasitic blood disease transmitted by tsetse flies and is widespread in sub-Saharan Africa. West African taurine breeds have the ability, known as trypanotolerance, to limit parasitaemia and anaemia and remain productive in enzootic areas. Several quantitative trait loci (QTL) underlying traits related to trypanotolerance have been identified in an experimentally infected F(2) population resulting from a cross between taurine and zebu cattle. Although this information is highly valuable, the QTL remain to be confirmed in populations subjected to natural conditions of infection, and the corresponding regions need to be refined. In our study, 360 West African cattle were phenotyped for the packed cell volume control under natural conditions of infection in south-western Burkina Faso. Phenotypes were assessed by analysing data from previous cattle monitored over 2 years in an area enzootic for trypanosomosis. We further genotyped for 64 microsatellite markers mapping within four previously reported QTL on BTA02, BTA04, BTA07 and BTA13. These data enabled us to estimate the heritability of the phenotype using the kinship matrix between individuals computed from genotyping data. Thus, depending on the estimators considered and the method used, the heritability of anaemia control ranged from 0.09 to 0.22. Finally, an analysis of association identified an allele of the MNB42 marker on BTA04 as being strongly associated with anaemia control, and a candidate gene, INHBA, as being close to that marker.

Genomic characterization and chromosomal mapping of 5 river buffalo skeletal muscle differentiation master genes.

River buffalo (Bubalus bubalis, 2n = 50, BBU) is a species of economic relevance in a number of countries. This species shows a very peculiar biology and a great capacity for environmental adaptation. There has been an increasing economic interest as well as a growing demand for a more detailed knowledge of molecular features in this species. From this perspective we report a genomic, transcriptional and cytogenetic analysis of 5 master genes involved in skeletal muscle development. Of these 5 genes, MYOD1, MYF5, MYF6 and MYOG belong to the basic helix-loop helix protein family while MSTN belongs to the TNF-B protein family. In mammals, these genes are involved in the early stages of skeletal muscle differentiation, development and regeneration. These pivotal biological functions are finely regulated in a tissue- and temporal-specific manner. We used a comparative genomic approach to obtain the buffalo specific sequences of MYOD1 and MYF6. The nucleotide sequence similarity and the protein domain conservation of the newly obtained sequences are analysed with respect to bovine and other mammalian species showing sequence similarity. The presence of a polymorphism in MYOD1 coding sequence is described and its possible effect discussed. Using a quantitative PCR approach, we compared the level of the 5 transcripts in adult and fetal muscle. These genes were physically localised on river buffalo R-banded chromosomes by FISH using bovine genomic BAC-clones. Here, we present a genomic and cytogenetic analysis which could offer a background to better characterise the buffalo genes involved in muscle function and which may be responsible for buffalo-specific meat features.

Comparative mapping and genomic annotation of the bovine oncosuppressor gene WWOX.

WWOX (WW domain-containing oxidoreductase) is the gene mapping at FRA16D HSA16q23.1, the second most active common fragile site in the human genome. In this study we characterized at a detailed molecular level WWOX in the bovine genome. First, we sequenced cDNA from various tissues and obtained evidence in support of a 9-exon structure for the gene, similar to the human gene. Then, we recovered BACs using exon tags and annotated the gene to a >1-Mb genomic region of BTA18 using the Btau 4.0 genome assembly as a reference, thus resolving an issue related to exon 9, which is not included in the genomic annotation of the gene in the Entrez database. Finally, BACs spanning WWOX were used as FISH probes to obtain comparative mapping of the gene in Bos taurus, Bubalus bubalis, Ovis aries and Capra hircus to BTA18q12.1, BBU18q13, OAR14q12.1 and CHI18q12.1, respectively. Our data show that the chromosomal location of WWOX is conserved between man and 4 major domesticated species. Moreover, the annotation of the bovine gene also suggests a highly conserved genomic arrangement, including number and size of introns.

Detection of selection signatures within candidate regions underlying trypanotolerance in outbred cattle populations.

Breeding indigenous African taurine cattle tolerant to trypanosomosis is a straightforward approach to control costs generated by this disease. A recent study identified quantitative trait loci (QTL) underlying trypanotolerance traits in experimental crosses between tolerant N'Dama and susceptible Boran zebu cattle. As trypanotolerance is thought to result from local adaptation of indigenous cattle breeds, we propose an alternative and complementary approach to study the genetic architecture of this trait, based on the identification of selection signatures within QTL or candidate genes. A panel of 92 microsatellite markers was genotyped on 509 cattle belonging to four West African trypanotolerant taurine breeds and 10 trypanosusceptible European or African cattle breeds. Some of these markers were located within previously identified QTL regions or candidate genes, while others were chosen in regions assumed to be neutral. A detailed analysis of the genetic structure of these different breeds was carried out to confirm a priori grouping of populations based on previous data. Tests based on the comparison of the observed heterozygosities and variances in microsatellite allelic size among trypanotolerant and trypanosusceptible breeds led to the identification of two significantly less variable microsatellite markers. BM4440, one of these two outlier loci, is located within the confidence interval of a previously described QTL underlying a trypanotolerance-related trait. Detection of selection signatures appears to be a straightforward approach for unravelling the molecular determinism of trypanosomosis pathogenesis. We expect that a whole genome approach will help confirm these results and achieve a higher resolving power.

Genomic location of the bovine growth hormone secretagogue receptor (GHSR) gene and investigation of genetic polymorphism.

The growth hormone secretagogue receptor (GHSR) is involved in the regulation of energetic homeostasis and GH secretion. In this study, the bovine GHSR gene was mapped to BTA1 between BL26 and BMS4004. Two different bovine GHSR CDS (GHSR1a and GHSR1b) were sequenced. Six polymorphisms (five SNPs and one 3-bp indel) were also identified, three of them leading to amino acid variations L24V, D194N, and Del R242. These variations are located in the extracellular N-terminal end, the exoloop 2, and the cytoloop 3 of the receptor, respectively.

Estimation of genetic parameters and genome scan for 15 semen characteristics traits of Holstein bulls.

A QTL detection experiment was performed in French dairy cattle to search for QTL related to male fertility. Ten families, involving a total of 515 bulls, were phenotyped for ejaculated volume and sperm concentration, number of spermatozoa, motility, velocity, percentage of motile spermatozoa after thawing and abnormal spermatozoa. A set of 148 microsatellite markers were used to realize a genome scan. First, genetic parameters were estimated for all traits. Semen production traits were found to have moderate heritabilities (from 0.15 to 0.30) while some of the semen quality traits such as motility had high heritabilities (close to 0.60). Genetic correlations among traits showed negative relationships between volume and concentration and between volume and most quality traits such as motility or abnormal sperm while correlations between concentration and these traits were rather favourable. Percentages of abnormal sperm were negatively related to quality traits, especially with motility and velocity of spermatozoa. Three QTL related to abnormal sperm frequencies were significant at p < 0.01. In total, 11 QTL (p < 0.05) were detected. However, the number of QTL detected was within the range of expected false positives. Because of the lack of power to find QTL in this design further analyses are required to confirm these QTL.

A Generalized Caprine-like Hypoplasia Syndrome is localized within a 6-cM interval on bovine chromosome 13 in the Montbéliarde breed.

Caprine-like Generalized Hypoplasia Syndrome (or SHGC) is a new hereditary disorder described in the Montbéliarde breed. We report here the characterization of this new disease, based on the visual examination of animals affected by SHGC, and on physiological and biochemical studies undertaken on samples of both SHGC and normal animals. Biological samples for more than 150 affected calves and their parents have been collected over the past 4 years within the framework of the Bovine Genetic Disease Observatory. First, pedigree analyses showed that the mode of inheritance is most probably autosomal recessive. Then, a genome scan with 113 animals and 140 microsatellite markers revealed a single locus within a 35-cM region on bovine chromosome 13. Genotypes of 261 animals for 18 new microsatellite markers from the region confirmed the localization of the disorder to a 6-cM interval. Finally, based on the analysis of haplotypes in 463 Montbéliarde sires, we estimated the frequency of the SHGC mutated allele in the population and could propose a strategy for the systematic eradication of this disorder in the near future.

Characterization of a balanced reciprocal translocation, rcp(9;11)(q27;q11) in cattle.

Cytogenetic analysis of a phenotypically normal young bull from Marchigiana breed revealed the presence of an abnormal karyotype. The observation of longer and smaller chromosomes than BTA1 and BTA29, respectively in all metaphases suggested the presence of a reciprocal translocation. RBG-banding confirmed this hypothesis revealing the involvement of BTA9 and BTA11. FISH analyses using cattle-specific BAC clones (474A12 and 293G09 for BTA9; 035D03 for BTA11) identified rcp(9;11)(q27;q11) in the two regions affected. Moreover analyses performed on both parents established the 'de novo' origin of the anomaly. Comparison with human homologue sequences (HSA6q24.3-->q25.3 for BTA9q27 and HSA2q11.1-->q12.1 for BTA11q11) revealed that both breakpoint regions are gene rich as up to date at least 200 genes have been localized in these regions. Thus, further analyses are required to identify the sequences disrupted by the breakpoints and to verify their consequences on rcp carrier phenotype.

A new case of reciprocal translocation in a young bull: rcp(11;21)(q28;q12).

Routine cytogenetic investigations of the Chianina cattle (BTA) breed revealed the presence of longer and smaller chromosomes than the largest (BTA1) and smallest (BTA29) chromosomes in the cells of a young, normal-looking bull used for reproduction. Application of both RBA-banding and Ag-NOR techniques, as well as the use of the FISH technique and specific molecular markers of both BTA11 (IL1B, ASS and LGB) and BTA21 (SERPINA and D21S45) established that these two abnormal chromosomes were the product of a reciprocal translocation between BTA11 and BTA21. Both der(11) and der(21) were C-band positive and the chromosome regions affected were rcp(11;21)(q28;q12). The young bull had a normal body conformation, including external genitalia, normal levels of testosterone (as in the control) and non-detectable levels of both 17 beta-estradiol and progesterone (as in the control). The animal never showed libido in the presence of both males and females in oestrus. After slaughter at 18 months, histological evaluation revealed normal organized testes, seminiferous tubules and epididymis but with poor proliferative germ cells consisting mainly of spermatogonia, middle pachytene spermatocytes and early spermatids with late spermatids and spermatozoa being very rare.

Characterization of the DGAT1 K232A and variable number of tandem repeat polymorphisms in French dairy cattle.

A quantitative trait locus (QTL) underlying different milk production traits has been identified with a high significance threshold value in the genomic region containing the acylCoA:diacylglycerol acyltransferase (DGAT1) gene, in the 3 main French dairy cattle breeds: French Holstein, Normande, and Montbéliarde. Previous studies have confirmed that the K232A polymorphism in DGAT1 is responsible for a major QTL underlying several milk production traits in Holstein dairy cattle and several other bovine breeds. In this study, we estimate the frequency of the 2 alternative alleles, K and A, of the K232A polymorphism in French Holstein, Normande, and Montbéliarde breeds. Although the K allele segregates in French Holstein and Normande breeds with a similar effect on production traits, the existence of additional mutations contributing to the observed QTL effect is strongly suggested in both breeds by the existence of sires heterozygous at the QTL but homozygous at the K232A polymorphism. One allele at a variable number of tandem repeats (VNTR) locus in the 5' noncoding region of DGAT1 has been recently proposed as a putative causative variant. In our study, this marker was found to present a high mutation rate of 0.8% per gamete and per generation, making the allele diversity observed compatible with that expected under neutrality. Moreover, among the sires homozygous at the K232A polymorphism, no allele at the VNTR can fully explain their QTL status. Finally, no allele at the VNTR was found to be significantly associated with the fat percentage variation in the 3 breeds simultaneously after correction for the effect of the K232A polymorphism. Therefore, our results suggest the existence of at least one other causative polymorphism not yet described. Because the A allele is nearly fixed in the Montbéliarde breed, this breed represents an interesting model to identify and confirm other mutations that have a strong effect on milk production traits.

Genomic structure and an alternative transcript of bovine mitochondrial glycerol-3-phosphate acyltransferase gene (GPAM).

GPAM maps in BTA26q22, where several QTLs affecting milk production, milk fat and protein content have been mapped. On the basis of the QTL location, the GPAM gene could be considered a good candidate gene for the mentioned traits. Glycerol-3-phosphate acyltransferase mitochondrial (GPAM) is the enzyme that catalyses the initial and committed step of glycerolipid synthesis and, therefore, it is a potential site for triacylglycerol synthesis regulation. In this study, the structure of the cDNA and the genomic DNA of the bovine GPAM gene were determined and the expression of its mRNA was studied. The cDNA of the gene was cloned by RT-PCR, 5' and 3' rapid amplification of cDNA ends. The GPAM mRNA sequence contains a 2,475-bp coding region and a 3,689-bp 3' UTR. Its ORF encoded for an 825-amino acid protein and has an 89% homology with the coding regions of previously characterized mouse and human GPAM genes. The predicted amino acid sequence had an 89 and 93% similarity with mouse and human GPAM proteins, respectively. Using a 5' RACE strategy, two different 5' UTRs were cloned. Northern blot analysis confirmed the presence of two different transcripts. Adipose tissues and lung had the highest levels of GPAM mRNA expression, whereas it was barely detectable in liver. This expression pattern differs with those of non-ruminant animals where liver is one of the tissues with higher GPAM mRNA expression level.

Sperm nuclei analysis of 1/29 Robertsonian translocation carrier bulls using fluorescence in situ hybridization.

In 1964, Gustavsson and Rockborn first described the 1/29 Robertsonian translocation in cattle. Since then, several studies have demonstrated the negative effect of this particular chromosomal rearrangement on the fertility of carrier animals. During the last decade, meiotic segregation patterns have been studied on human males carrying balanced translocations using FISH on decondensed sperm nuclei. In this work, we have applied the 'Sperm-FISH' technique to determine the chromosomal content of spermatozoa from two bulls heterozygous for the 1/29 translocation and one normal bull (control). 5425 and 2702 sperm nuclei were scored, respectively, for the two heterozygous bulls, using whole chromosome painting probes of chromosomes 1 and 29. Very similar proportions of normal (or balanced) spermatozoa resulting from alternate segregation were observed (97.42% and 96.78%). For both heterozygous bulls, the proportions of nullisomic and disomic spermatozoa did not follow the theoretical 1:1 ratio. Indeed, proportions of nullisomic spermatozoa were higher than those of disomic sperma tozoa (1.40% vs 0.09% (bull 1) and 1.29% vs 0.15% (bull 2) for BTA1, and 0.65% vs 0.40% (bull 1) and 1.11% vs 0.63% (bull 2) for BTA29). The average frequencies of disomic and diploid spermatozoa in the normal bull were 0.11% and 0.05%, respectively.

Isolation, mapping and identification of SNPs for four genes (ACP6, CGN, ANXA9, SLC27A3) from a bovine QTL region on BTA3.

On the basis of fine mapping of a quantitative trait loci region of BTA3 for milk fat content, an examination of the comparative map between cattle and human indicates that the annexin 9 protein gene (ANXA9) and the fatty acid transport protein type 3 gene (SLC27A3) are two strong candidate genes. The objective of the present study is to isolate, map and characterize these genes and identify polymorphisms that could be further utilized in linkage or association studies. Furthermore, two new genes which are in the same region, cingulin protein gene (CGN) and lysophosphatidic acid phosphatase protein gene (ACP6) were studied. DNA fragments (869, 1778, 1933 and 2618 bp) corresponding to partial sequences of ACP6,CGN,ANXA9 and SLC27A3 genes were isolated. Direct sequencing of PCR products amplified from different cattle breeds revealed 1, 4, 4 and 2 SNPs for ACP6, CGN,ANXA9 and SLC27A3, respectively. For ANXA9 one SNP was located in exon 5 (A-->G 951) resulting in an amino acid change from histidine to arginine. Finally, ACP6,CGN,ANXA9 and SLC27A3 genes were located on chromosome 3 between ILSTS096 and BMS819 markers, in a region in which quantitative trait loci (QTL) for several milk traits have been described.

Comparative FISH mapping of mucin 1, transmembrane (MUC1) among cattle, river buffalo, sheep and goat chromosomes: comparison between bovine chromosome 3 and human chromosome 1.

Four bovine BAC clones (0494F01, 0069D07, 0060B06, and 0306A12) containing MUC1, as confirmed by mapping MUC1 on a RH3000 radiation hybrid panel, were hybridised on R-banded chromosomes of cattle (BTA), river buffalo (BBU), sheep (OAR) and goat (CHI). MUC1 was FISH-mapped on BTA3q13, BBU6q13, OAR1p13 and CHI3q13 and both chromosomes and chromosome bands were homoeologous confirming the high degree of chromosome homoeologies among bovids and adding more information on the pericentromeric regions of these species' chromosomes. Indeed, MUC1 was more precisely assigned to BTA3 and assigned for the first time to BBU6, OAR1p and CHI3. Moreover, detailed and improved cytogenetic maps of BTA3, CHI3, OAR1p and BBU6 are shown and compared with HSA1.

Genomic structure and alternative transcript of bovine fatty acid synthase gene (FASN): comparative analysis of the FASN gene between monogastric and ruminant species.

Fatty acid synthesis differs considerably between monogastric and ruminant species. Fatty acid synthase (FASN) plays a central role in de novo lipogenesis in mammals. FASN has seven active sites which help to catalyse all the reaction steps in the conversion of acetyl-CoA and malonyl-CoA to palmitate. In this work, the bovine fatty acid synthase gene (FASN) was cloned, characterized and compared to the human and rat orthologs. Comparative analysis reveals evolutionarily conserved exon regions and gene flanking sequences. Analysis of the DNA sequence in the 5' flanking region of the FASN bovine gene revealed a potential TATA box, CAAT box and 5 Sp1 binding sites located in a CpG island. RT-PCR and Western blot analysis showed that FASN expression was higher in brain, testis and adipose tissue than in liver and heart. The longer form of the FASN cDNA includes a 7,542-bp sequence which encodes a protein with 2,513 amino acids. An alternative transcript was discovered in bovine and ovine tissues devoid of part of exon 9. The removal of part of exon 9 by post-transcriptional splicing causes a frameshift in the open reading frame and results in a premature termination codon. We hypothesize that in ruminants, FASN may be regulated by the ratio between the two transcripts. The small transcript is mostly produced in tissues with low fatty acid synthesis.

Mapping quantitative trait loci for milk production and health of dairy cattle in a large outbred pedigree.

Quantitative trait loci (QTL) affecting milk production and health of dairy cattle were mapped in a very large Holstein granddaughter design. The analysis included 1794 sons of 14 sires and 206 genetic markers distributed across all 29 autosomes and flanking an estimated 2497 autosomal cM using Kosambi's mapping function. All families were analyzed jointly with least-squares (LS) and variance components (VC) methods. A total of 6 QTL exceeding approximate experiment-wise significance thresholds, 24 QTL exceeding suggestive thresholds, and 34 QTL exceeding chromosome-wise thresholds were identified. Significance thresholds were determined via data permutation (for LS analysis) and chi-square distribution (for VC analysis). The average bootstrap confidence interval for the experiment-wise significant QTL was 48 cM. Some chromosomes harbored QTL affecting several traits, and these were always in coupling phase, defined by consistency with genetic correlations among traits. Chromosome 17 likely harbors 2 QTL affecting milk yield, and some other chromosomes showed some evidence for 2 linked QTL affecting the same trait. In each of these cases, the 2 QTL were in repulsion phase in those families appearing to be heterozygous for both QTL, a finding which supports the build-up of linkage disequilibrium due to selection.

Development of a comprehensive comparative radiation hybrid map of bovine chromosome 7 (BTA7) versus human chromosomes 1 (HSA1), 5 (HSA5) and 19 (HSA19).

In this study, we present a comprehensive 3,000-rad radiation hybrid (RH) map of bovine chromosome 7 (BTA7) with 108 markers including 54 genes or ESTs. For 52 of them, a human ortholog sequence was found either on HSA1 (one gene), HSA5 (31 genes) or HSA19 (19 genes and one non-annotated sequence) confirming previously described syntenies. Moreover, in order to refine boundaries of blocks of conserved synteny, nine new genes were mapped to the bovine genome on the basis of their localization on the human genome: six on BTA7 and originating from HSA1 (TRIM17), HSA5 (MAN2A1, LMNB1, SIAT8D and FLJ1159) and HSA19 (VAV1), and the three others (AP3B1, APC and CCNG1) on BTA10. The available draft of the human genome sequence allowed us to present a detailed picture of the distribution of conserved synteny segments between man and cattle. Finally, the INRA bovine BAC library was screened for most of the BTA7 markers considered in this study to provide anchors for the bovine physical map.

Mapping of 195 genes in cattle and updated comparative map with man, mouse, rat and pig.

Our on-going goal is to improve and update the comparative genome organization between cattle and man but also among the most detailed mammalian species genomes i.e. cattle, mouse, rat and pig. In this work, we localized 195 genes in cattle and checked all human/bovine non-concordant localizations found in the literature. Next, we compiled all the genes mapped in cattle, goat, sheep and pig (2,166) for which the human ortholog with its chromosomal position is known, added corresponding data in mouse and rat, and ordered the genes relatively to the human genome sequence. We estimate that our compilation provides bovine mapping information for about 89% of the human autosomes. Thus, a near complete, overall and detailed picture of the number, distribution and extent of bovine conserved syntenies (regardless of gene order) on human R-banded autosomes is proposed as well as a comparison with mouse, rat and pig genomes.