Atmospheric circulation and mortality by unintentional drowning in Spain: from 1999 to 2018.

AIMS: Drowning deaths are a leading cause of unintentional deaths worldwide. Few studies have analysed the role of meteorology in drowning, and with inconclusive results. The aim of this work is to analyse the temporal and geographical distribution of deaths by accidental drowning and submersion in Spain over 20 years, and to assess the relationship between accidental drowning and main atmospheric circulation patterns. METHODS: An ecological study was performed, in which drowning and submersion mortality data from 1999 to 2018, considering demographic variables, were analysed. To study the association with atmospheric circulation we used an ERA5 reanalysis product over the whole European continent and the Climatic Research Unit Time Series (CRU TS) data set. RESULTS: The annual average rate of deaths by accidental drownings was 11.86 deaths per million of habitants in Spain. The incidence in males was four times higher than in females, and when comparing age groups, the rate in the eldest group was the highest. Unintentional drowning deaths were not equally distributed around the country; the provinces with the highest registered standardized drowning death rates were touristic waterfront provinces either in Eastern Spain or in one of the archipelagos. There was a significant relationship between accidental drowning and meteorological variables during summer months, and drowning deaths were spatially correlated with sea-level pressure over the Mediterranean basin. CONCLUSION: Although the mortality rate registered a statistically significant decreasing tendency over the studied period, our results must be taken into consideration to improve the prevention strategies in the country since most of these deaths are avoidable.

Analysis of post-transplant chimerism by using a single amplification reaction of 38 Indel polymorphic loci.

In order to detect chimerism after allogeneic hematopoietic SCT (HSCT), several methods have been developed. In this study we describe the use of a set of insertion/deletion (Indel) polymorphic loci to determine the level of donor cell engraftment. We analyzed 50 DNA samples from patients who had undergone HSCT, and also several artificial chimeric samples created by mixing different DNA specimens from non-transplanted donors in various proportions. A specific set of 38 autosomic Indel polymorphisms were analyzed. For comparison purposes, a set of 15 short tandem repeats (STRs) were analyzed using the Identifiler Plus Amplification Kit. Our results suggest that Indel-based and STR-based procedures behave similarly in most cases. However, Indel analysis may provide additional information in some cases with a small minor chimeric component or when the presence of stutter bands complicates chimerism estimation.

Missense polymorphisms of the WNT16 gene are associated with bone mass, hip geometry and fractures.

UNLABELLED: Two missense polymorphisms of WNT16 were associated with hip bone mineral density (BMD), the buckling ratio of the femoral neck, calcaneal ultrasound and hip fractures in individuals under 80 years of age. These results confirm the association of the WNT16 gene with bone mass and osteoporotic fractures. INTRODUCTION: Osteoporosis has a strong genetic component. Wnt ligands stimulate the differentiation of osteoblast precursors and play a major role in skeletal homeostasis. Therefore, the aim of this study was to explore the association of allelic variants of the WNT16 gene with BMD, other structural parameters of bone and osteoporotic hip fractures. METHODS: Six single nucleotide polymorphisms were analysed in 1,083 Caucasian individuals over 49 years of age. RESULTS: Two missense polymorphisms (rs2908004 and rs2707466) were associated with femoral neck BMD, with average differences across genotypes of 35 mg/cm(2) (p = 0.00037 and 0.0015, respectively). Likewise, the polymorphisms were associated with calcaneal quantitative ultrasound parameters (p = 0.00004 and 0.0014, respectively) and the buckling ratio, an index of cortical instability of the femoral neck (p = 0.0007 and 0.0029, respectively). Although there were no significant differences in the genotype frequency distributions between 294 patients with hip fractures and 670 controls, among the subgroup under 80 years of age, TT genotypes were underrepresented in patients with fractures (odds ratio 0.50; CI 0.27-0.94). CONCLUSION: Common missense polymorphisms of the WNT16 gene are associated with BMD at the hip, calcaneal ultrasound and the buckling ratio of the femoral neck, as well as with hip fractures in individuals under 80 years of age. Overall, these results confirm the association of the WNT16 locus with BMD identified in genome-wide association studies and support its role in determining the risk of osteoporotic fractures.

Evaluation of the sensitivity of two recently developed STR multiplexes for the analysis of chimerism after haematopoietic stem cell transplantation.

Forensic-oriented kits analysing short tandem repeat (STR) polymorphisms are widely used to determine the proportions of donor and recipient cells after haematopoietic stem cell transplantation. The sensitivity of this technology is crucial for the early detection of relapse and, in consequence, the adjustment of the treatment to enhance donor-origin haematopoiesis in transplant recipients. The objective of this study was to compare the performance of two recently developed STR multiplex kits, AmpFℓSTR(®) Identifiler(®) Plus PCR Amplification Kit (Applied Biosystems) and Investigator™ IDplex(®) (Qiagen), in the analysis of chimerism. Fifteen STR loci were amplified with both kits in 26 peripheral blood samples of transplantated patients showing chimerism. Peak amplitude threshold, detection limit (%DL), per cent donor chimerism and efficacy of each multiplex and STR were determined, and the results with both kits were compared. The %DL and the estimated per cent donor chimerism were similar with both kits. On the other hand, Identifiler(®) Plus kit allowed chimerism identification only in 24 (92%) of the 26 cases with chimerism detected by using the Investigator™ IDplex(®) when only 'type 5' allelic constellations (i.e. without potential interference by stutter peaks) were taken into account. However, IDplex(®) efficacy was somewhat lower than that of Identifiler Plus when only the most informative loci (D2S1338, D21S11, D18S51 and FGA) were considered. Therefore, although each system had some particular advantages and disadvantages, overall both STR multiplexes showed similar performance in qualitative and quantitative chimerism analysis.

Genetic uniqueness of the Waorani tribe from the Ecuadorian Amazon.

South America and especially the Amazon basin is known to be home to some of the most isolated human groups in the world. Here, we report on a study of mitochondrial DNA (mtDNA) in the Waorani from Ecuador, probably the most warlike human population known to date. Seeking to look in more depth at the characterization of the genetic diversity of this Native American tribe, molecular markers from the X and Y chromosomes were also analyzed. Only three different mtDNA haplotypes were detected among the Waorani sample. One of them, assigned to Native American haplogroup A2, accounted for more than 94% of the total diversity of the maternal gene pool. Our results for sex chromosome molecular markers failed to find close genetic kinship between individuals, further emphasizing the low genetic diversity of the mtDNA. Bearing in mind the results obtained for both the analysis of the mtDNA control region and complete mitochondrial genomes, we suggest the existence of a 'Waorani-specific' mtDNA lineage. According to current knowledge on the phylogeny of haplogroup A2, we propose that this lineage could be designated as subhaplogroup A2s. Its wide predominance among the Waorani people might have been conditioned by severe genetic drift episodes resulting from founding events, long-term isolation and a traditionally small population size most likely associated with the striking ethnography of this Amazonian community. In all, the Waorani constitute a fine example of how genetic imprint may mirror ethnopsychology and sociocultural features in human populations.

Common allelic variants of the farnesyl diphosphate synthase gene influence the response of osteoporotic women to bisphosphonates.

Farnesyl diphosphate synthase (FDPS) is necessary for osteoclast survival and activity and is considered as a major molecular target of aminobisphosphonates. Our objective was to analyze the influence of FDPS polymorphisms on bone mineral density (BMD) and the response to antiresortive drugs. Three single-nucleotide polymorphisms of FDPS were analyzed in 1186 postmenopausal women. There was only a marginally significant association of baseline hip BMD with rs11264359 alleles (P=0.043). However, among 191 women receiving antiresortive therapy, there was a very significant association between rs2297480 or rs11264359 alleles and the BMD changes after aminobisphosphonate therapy for an average period of 2.5 years (P=0.001). The genotype explained 7.2% of the variance in the BMD response. On the other hand, there was no association between the BMD changes after raloxifene therapy and any of the polymorphisms studied. These results suggest that common polymorphisms of the FDPS gene influence the response to aminobisphosphonates.

Chimerism detection by short tandem repeat analysis when donor and recipient genotypes are not known.

BACKGROUNDS: The analysis of chimerism after bone marrow transplantation by STR-PCR is frequently carried out with commercial kits designed for forensic purposes and including too many non informative STR. Furthermore, in routine clinical practice it is not uncommon to lack the pre-transplant genotype of the recipient or the donor, thus making it difficult to identify both components in the post-transplant genotype. The objective of this paper is to overcome these drawbacks by analyzing the informativity of STR markers from a perspective which can be applied whether the pretransplant genotypes are available or not, and selecting a minimum STR panel that allows an effective direct detection of chimerism. METHODS: DNA extraction, STR-PCR and fragment analysis of 15 STR in 90 donor-recipient pairs, 60 of which were part of the discovery set and 30 in a validation set. Loci were considered as informative when there were 3 or 4 different alleles in the combined genotypes of the recipient and the donor. RESULTS: The informativity varied between 41.6 and 76.6. The 4 most informative loci were D2S1338, D21S11, D18S51 and FGA. We could select a minimum set of 8 markers (D2S1338, D21S11, D18S51, FGA, VWA, D19S433, TH01 and D3S1358) that provided at least 3 informative loci in 95% of cases. CONCLUSION: This minimum STR panel may be an efficient way to detect and quantitate donor-recipient chimerism after transplantation.

Association of ACACB polymorphisms with obesity and diabetes.

Acetyl-CoA carboxylase beta, encoded by the ACAB gene, plays an important role in the oxidation of fatty acids. The aim of this study was to check the hypothesis that allelic variants of ACACB influence the risk of obesity and type 2 diabetes mellitus. Twenty five tagging single nucleotide polymorphisms (SNPs) capturing common variants of the ACACB gene were selected and analyzed in two cohorts including 1695 postmenopausal women of the general population and in 161 women with severe obesity (BMI>35). In vitro binding of transcription factors was explored by electrophoretic mobility shift assays (EMSA). T alleles at the rs2268388 locus were overrepresented in women with severe obesity (18% vs. 10% in controls; OR 1.74 [95% confidence interval 1.30-2.47]), which was statistically significant after multiple-test adjustment (p=0.0004). Likewise, T alleles at the rs2268388 locus and C alleles at the rs2239607 locus were associated with diabetes, in the discovery as well as in the replication cohorts, even after women with severe obesity were excluded (OR 3.6 and 2.8, for TT and CC homozygotes, respectively). Allelic differences in the binding affinity for nuclear proteins were revealed in vitro by EMSA and competition experiments were consistent with the binding of glucorticoid receptor and serum response factor. In conclusion, common polymorphisms of ACACB gene are associated with obesity and, independently, with type 2 diabetes in postmenopausal women, suggesting that the activity of acetyl-CoA carboxylase beta plays an important role in these disorders related to energy metabolism.

Common variations in estrogen-related genes are associated with severe large-joint osteoarthritis: a multicenter genetic and functional study.

OBJECTIVE: Several lines of evidence suggest that estrogens influence the development of osteoarthritis (OA). The aim of this study was to explore the association of two common polymorphisms within the aromatase (CYP19A1) and estrogen receptor (ER) alpha (ESR1) genes with severe OA of the lower limbs. METHODS: The rs1062033 (CYP19A1) and rs2234693 (ESR1) single nucleotide polymorphisms were genotyped in 5528 individuals (3147 patients with severe hip or knee OA, and 2381 controls) from four centres in Spain and the United Kingdom. Gene expression was measured in femoral bone samples from a group of patients. RESULTS: In the global analysis, both polymorphisms were associated with OA, but there was a significant sex interaction. The GG genotype at rs1062033 was associated with an increased risk of knee OA in women [odds ratio (OR) 1.23; P=0.04]. The CC genotype at rs2234693 tended to be associated with reduced OA risk in women (OR 0.76, P=0.028, for knee OA; OR=0.84, P=0.076 for hip OA), but with increased risk of hip OA in men (OR 1.28; P=0.029). Women with unfavourable genotypes at both loci had an OR of 1.61 for knee OA (P=0.006). The rs1062033 genotype associated with higher OA risk was also associated with reduced expression of the aromatase gene in bone. CONCLUSIONS: Common genetic variations of the aromatase and ER genes are associated with the risk of severe OA of the large joints of the lower limb in a sex-specific manner. These results are consistent with the hypothesis that estrogen activity may influence the development of large-joint OA.

Wnt pathway genes in osteoporosis and osteoarthritis: differential expression and genetic association study.

UNLABELLED: In comparison with hip fractures, increased expression of genes in the Wnt pathway and increased Wnt activity were found in bone samples and osteoblast cultures from patients with osteoarthritis, suggesting the involvement of this pathway in subchondral bone changes. No consistent differences were found in the genetic association study. INTRODUCTION: This study aims to explore the allelic variations and expression of Wnt pathway genes in patients with osteoporosis and osteoarthritis. METHODS: The expression of 86 genes was studied in bone samples and osteoblast primary cultures from patients with hip fractures and hip or knee osteoarthritis. The Wnt-related activity was assessed by measuring AXIN2 and in transfection experiments. Fifty-five SNPs of the LRP5, LRP6, FRZB, and SOST genes were analyzed in 1,128 patients. RESULTS: Several genes were differentially expressed in bone tissue, with the lowest values usually found in hip fracture and the highest in knee osteoarthritis. Overall, seven genes were consistently upregulated both in tissue samples and in cell cultures from patients with knee osteoarthritis (BCL9, FZD5, DVL2, EP300, FRZB, LRP5, and TCF7L1). The increased expression of AXIN2 and experiments of transient transfection of osteoblasts with the TOP-Flash construct confirmed the activation of Wnt signaling. Three SNPs of the LRP5 gene and one in the LRP6 gene showed marginally significant differences in allelic frequencies across the patient groups, but they did not resist multiple-test adjustment. CONCLUSIONS: Genes in the Wnt pathway are upregulated in the osteoarthritic bone, suggesting their involvement not only in cartilage distortion but also in subchondral bone changes.

Association of the 163A/G and 1181G/C osteoprotegerin polymorphism with bone mineral density.

The aim of the study was to investigate the distribution of 163 A/G osteoprotegerin gene promoter and 1181 G/C osteoprotegerin exon 1 polymorphisms in a group of women with different hormonal status and to analyze their relationship with BMD. Osteoprotegerin polymorphisms and BMD were analyzed in 332 women (69 premenopausal and 263 postmenopausal). BMD was quantified at the lumbar spine (L 2-4), femoral neck, and total hip. Genotyping for the presence of different polymorphisms was performed using the Custom Taqman ((R)) SNP Genotyping assays. There were not significant differences in BMD according to 163 A/G genotype. However, significant differences in lumbar spine BMD were found according to 1181 G/C alleles. Thus, women with CC genotype had significant higher BMD at the lumbar spine than those with GC or GG genotype. No differences were found in femoral neck and total hip BMD. In age-adjusted models, the 1181 G/C OPG polymorphism explained 2.2% of BMD variance at the spine, 0.3% at the femoral neck, and 0.9% at the total hip in the whole group. In the subgroup of premenopausal women, the polymorphism was strongly related to spine BMD, and explained 11.5% of the variance, whereas body weight explained 7.9%. The 1181 G/C polymorphism was associated with lumbar spine BMD in Spanish women. Premenopausal women with the CC genotype had a higher BMD.

Association of aromatase and estrogen receptor gene polymorphisms with hip fractures.

UNLABELLED: Two polymorphisms of the aromatase and estrogen receptor genes appeared to interact to influence the risk of hip fractures in women. INTRODUCTION: Allelic variants of the aromatase gene have been associated with bone mineral density and vertebral fractures. Our objective was to analyze the relationship between two polymorphisms of the aromatase and estrogen receptor genes and hip fractures. METHODS: We studied 498 women with hip fractures and 356 controls. A C/G polymorphism of the aromatase gene and a T/C polymorphism of the estrogen receptor alpha gene were analyzed using Taqman assays. Aromatase gene expression was determined in 43 femoral neck samples by real-time RT-PCR. RESULTS: There were no significant differences in the overall distribution of genotypes between the fracture and control groups. However, among women with a TT genotype of the estrogen receptor, the CC aromatase genotype was more frequent in women with fractures than in controls (39 vs. 23%, p = 0.009). Thus, women homozygous for T alleles of estrogen receptor and C alleles of aromatase were at increased risk of fracture (odds ratio 2.0; 95% confidence interval 1.2-3.4). The aromatase polymorphism was associated with RNA levels in bone tissue, which were three times lower in samples with a CC genotype (p = 0.009). CONCLUSIONS: These common polymorphisms of the aromatase and estrogen receptor genes appear to interact, influencing the risk of hip fractures in women.

MTHFR C677T polymorphism and osteoporotic fractures.

The C677T (rs1801133) polymorphism of MTHFR (methylenetetrahydrofolate reductase) has been associated with the risk of cardiovascular events, and also with osteoporosis in some studies. However, the results are controversial. Our objective was to determine the relationship of the polymorphism with osteoporotic fractures by means of a case-control study. C677T was analyzed in 823 subjects (365 controls, 136 with vertebral fractures and 322 with hip fracture) by using a Taqman assay. The distribution of MTHFR genotypes was similar in patients and controls. In comparison with TC/CC genotypes, the age-adjusted OR for hip fractures of the TT genotype was 1.0 (95% confidence interval 0.6-1.7) in women and 0.7 (0.3-1.8) in men. The OR for vertebral fractures was 0.8 (0.4-1.7) in women and 1.7 (0.4-6.7) in men. A meta-analysis combining these data with previous reports confirmed the lack of association between MTHFR and fractures, with an OR of 1.1 (0.7-1.9, p=0.65) for vertebral fractures and 1.2 (0.7-2.0; p=0.45) for peripheral fractures, but there was significant heterogeneity among the results of individual studies, particularly about peripheral fractures. In conclusion, the C677T polymorphism of the MTHFR gene does not appear to be associated with the overall risk of osteoporotic fractures. However, given the heterogeneity of the results of published studies, further investigations are needed to evaluate its influence in specific population subgroups.

Klotho gene polymorphism and male bone mass.

The Klotho gene codes for a protein that is thought to influence the homeostasis of several tissues, including bone, as well as the aging process. Although the mechanism of action has not been fully elucidated, some studies in women have associated Klotho allelic variants to bone mineral density (BMD). The objective of this study was to determine the relationship of a common T/G polymorphism, resulting in a phenylalanine (F) to valine (V) substitution, with male bone mass. BMD was measured by dual-energy X-ray absorptiometry in 362 Spanish men aged 19-83 years. Klotho alleles were determined by a Taqman assay. Allele frequencies were 85% and 15% for the F and V alleles, respectively. In comparison with the most common FF genotype, young and middle-aged men (age less than 53 years) with FV/VV genotypes had higher age- and body mass index-adjusted BMD at the lumbar spine (1.059 +/- 0.017 vs. 1.016 +/- 0.011 g/cm(2), P = 0.036), the hip (1.077 +/- 0.017 vs. 1.033 +/- 0.011 g/cm(2), P = 0.028), and the calcaneus (0.599 +/- 0.125 vs. 0.547 +/- 0.108 g/cm(2), P = 0.012). Klotho alleles explained about 2-4% of BMD variance. However, Klotho genotype was not associated to BMD in older men. There were no Klotho-related differences in height, body weight, calcium intake, tobacco or alcohol consumption, or serum testosterone levels. In conclusion, these results suggest that allelic variants of Klotho constitute one of the genetic factors influencing BMD in male adults.

MTHFR polymorphism and bone mineral density: meta-analysis of published studies.

The C677T (rs1801133) polymorphism of methylenetetrahydrofolate reductase (MTHFR) has been associated with bone status in some studies, but the results have been mixed. In order to have a better understanding of this issue, we performed a meta-analysis of studies about the association of the C677T polymorphism and bone mineral density (BMD). Eight studies analyzed the relationship with spine BMD. When their results were combined, individuals with TT genotype showed a small but significantly reduced BMD compared to those with TC and CC genotypes. The weighted mean difference (WMD) was 18.0 mg/cm2 (P = 0.001, 95% confidence interval [CI] 7.1-28.9), without statistical evidence for between-study heterogeneity (P = 0.28, I2 = 17%). Six studies analyzed femoral neck BMD. A test for heterogeneity was significant (P = 0.03, I2 = 56%). Individuals with TT alleles tended to have somewhat lower BMD, but the difference was not statistically significant. In random effects model, the WMD between the TT and TC/CC genotypes was 6.4 mg/cm2 (95% CI -7.8 to 21.2, P = 0.36). Total hip BMD was measured in four studies. They showed a significantly lower BMD in subjects with TT alleles: WMD 19.7 (95% CI 5.3-34.1) mg/cm2, P = 0.007, in comparison with TC/CC subjects. When we considered only studies on women, the WMD in BMD between TT and TC/CC genotypes was significant at the spine (22.1 mg/cm2, 95% CI 8.6-35.6; P = 0.001) and the femoral neck (15.5 mg/cm2, 95% CI 4.3-26.7; P = 0.007). There was no evidence for heterogeneity. The small number of studies did not allow a meaningful sex-stratified analysis of total hip BMD or a separate analysis of male data. In conclusion, the C677T polymorphism of the MTHFR gene is associated with small differences in BMD, at least in women.

Bone mass in young adults: relationship with gender, weight and genetic factors.

OBJECTIVES: To determine the relationship of the bone mass attained in young adults with anthropometric and genetic factors. DESIGN: Cross-sectional study of normal individuals. METHODS: We studied 341 healthy subjects between 22 and 45 years of age. Bone mineral density (BMD) was measured by dual-energy X-ray absorptiometry (DXA) and correlated with body weight, height and nine polymorphisms in six genes involved in sex steroid metabolism (17-hydroxylase, aromatase and 5-reductase) and activity (oestrogen receptors (ER)-alpha and -beta, and androgen receptor). RESULTS: The BMD was higher in men than in women (spine: 1.048 +/- 0.120 vs. 1.034 +/- 0.112; hip: 0.907 +/- 0.131 vs. 0.822 +/- 0.104 g cm(-2), P < 0.001). However, the difference was due, at least in part, to the larger body size in men and diminished markedly after height adjustment. There was a negative correlation between age and hip BMD. Body weight was the single most influential factor on spine and hip BMD in both sexes, explaining 8-9% of BMD variance. Amongst the genetic factors studied, a common CA repeat polymorphism in ER-beta showed a significant association with BMD in women (P = 0.03 at the spine, and 0.008 at the hip). The relationship between ER-beta genotype and BMD persisted after adjustment by body weight and age, explaining a further 2-3% of BMD variance. Allelic variants of other genes studied were not related with BMD. CONCLUSIONS: Body weight and allelic variants of ER-beta are associated with BMD in young adults.

Mutation rates at Y chromosome specific microsatellites.

A collaborative work was carried out by the Spanish and Portuguese ISFG Working Group (GEP-ISFG) to estimate Y-STR mutation rates. Seventeen Y chromosome STR loci (DYS19, DYS385, DYS389I and II, DYS390, DYS391, DYS392, DYS393, DYS437, DYS438, DYS439, DYS460, DYS461, DYS635 [GATA C4], GATA H4, and GATA A10) were analyzed in a sample of 3,026 father/son pairs. Among 27,029 allele transfers, 54 mutations were observed, with an overall mutation rate across the 17 loci of 1.998 x 10(-3) (95% CI, 1.501 x 10(-3) to 2.606 x 10(-3)). With just one exception, all of the mutations were single-step, and they were observed only once per gametogenesis. Repeat gains were more frequent than losses, longer alleles were found to be more mutable, and the mutation rate seemed to increase with the father's age. Hum Mutat 26(6), 520-528, 2005. (c) 2005 Wiley-Liss, Inc.

X-linked microsatellites in two Northern Spain populations.

The X-chromosomal microsatellites HPRTB, DXS101, ARA, DXS7423, and DXS8377 were analysed by a pentaplex PCR in an expanded population sample from Cantabria and an independent sample of unrelated individuals from the Basque Country. Allele frequencies showed similar distributions, but minor variations were found for some loci.

7-Locus Y chromosome haplotype profiling in a northern Spain population.

Allele and haplotype frequencies for 7 Y-specific STR loci (DYS19, DYS389-I, DYS389-II, DYS390, DYS391, DYS392, DYS393) had been determined in a sample of 107 unrelated males living in Cantabria, a region in northern Spain, by means of two multiplex PCRs.