| 1. |
- Cruz, Raquel, et al.
(författare)
-
Novel genes and sex differences in COVID-19 severity
- 2022
-
Ingår i: Human Molecular Genetics. - : Oxford University Press. - 0964-6906 .- 1460-2083. ; 31:22, s. 3789-3806
-
Tidskriftsartikel (refereegranskat)abstract
- Here, we describe the results of a genome-wide study conducted in 11 939 coronavirus disease 2019 (COVID-19) positive cases with an extensive clinical information that were recruited from 34 hospitals across Spain (SCOURGE consortium). In sex-disaggregated genome-wide association studies for COVID-19 hospitalization, genome-wide significance (P < 5 × 10−8) was crossed for variants in 3p21.31 and 21q22.11 loci only among males (P = 1.3 × 10−22 and P = 8.1 × 10−12, respectively), and for variants in 9q21.32 near TLE1 only among females (P = 4.4 × 10−8). In a second phase, results were combined with an independent Spanish cohort (1598 COVID-19 cases and 1068 population controls), revealing in the overall analysis two novel risk loci in 9p13.3 and 19q13.12, with fine-mapping prioritized variants functionally associated with AQP3 (P = 2.7 × 10−8) and ARHGAP33 (P = 1.3 × 10−8), respectively. The meta-analysis of both phases with four European studies stratified by sex from the Host Genetics Initiative (HGI) confirmed the association of the 3p21.31 and 21q22.11 loci predominantly in males and replicated a recently reported variant in 11p13 (ELF5, P = 4.1 × 10−8). Six of the COVID-19 HGI discovered loci were replicated and an HGI-based genetic risk score predicted the severity strata in SCOURGE. We also found more SNP-heritability and larger heritability differences by age (<60 or ≥60 years) among males than among females. Parallel genome-wide screening of inbreeding depression in SCOURGE also showed an effect of homozygosity in COVID-19 hospitalization and severity and this effect was stronger among older males. In summary, new candidate genes for COVID-19 severity and evidence supporting genetic disparities among sexes are provided.
|
|
| 2. |
- Ademuyiwa, Adesoji O., et al.
(författare)
-
Determinants of morbidity and mortality following emergency abdominal surgery in children in low-income and middle-income countries
- 2016
-
Ingår i: BMJ Global Health. - : BMJ Publishing Group Ltd. - 2059-7908. ; 1:4
-
Tidskriftsartikel (refereegranskat)abstract
- Background: Child health is a key priority on the global health agenda, yet the provision of essential and emergency surgery in children is patchy in resource-poor regions. This study was aimed to determine the mortality risk for emergency abdominal paediatric surgery in low-income countries globally.Methods: Multicentre, international, prospective, cohort study. Self-selected surgical units performing emergency abdominal surgery submitted prespecified data for consecutive children aged <16 years during a 2-week period between July and December 2014. The United Nation's Human Development Index (HDI) was used to stratify countries. The main outcome measure was 30-day postoperative mortality, analysed by multilevel logistic regression.Results: This study included 1409 patients from 253 centres in 43 countries; 282 children were under 2 years of age. Among them, 265 (18.8%) were from low-HDI, 450 (31.9%) from middle-HDI and 694 (49.3%) from high-HDI countries. The most common operations performed were appendectomy, small bowel resection, pyloromyotomy and correction of intussusception. After adjustment for patient and hospital risk factors, child mortality at 30 days was significantly higher in low-HDI (adjusted OR 7.14 (95% CI 2.52 to 20.23), p<0.001) and middle-HDI (4.42 (1.44 to 13.56), p=0.009) countries compared with high-HDI countries, translating to 40 excess deaths per 1000 procedures performed.Conclusions: Adjusted mortality in children following emergency abdominal surgery may be as high as 7 times greater in low-HDI and middle-HDI countries compared with high-HDI countries. Effective provision of emergency essential surgery should be a key priority for global child health agendas.
|
|
| 3. |
- Ferrando, Carlos, et al.
(författare)
-
Effects of oxygen on post-surgical infections during an individualised perioperative open-lung ventilatory strategy : a randomised controlled trial
- 2020
-
Ingår i: British Journal of Anaesthesia. - : ELSEVIER SCI LTD. - 0007-0912 .- 1471-6771. ; 124:1, s. 110-120
-
Tidskriftsartikel (refereegranskat)abstract
- Background: We aimed to examine whether using a high fraction of inspired oxygen (FIO2) in the context of an individualised intra- and postoperative open-lung ventilation approach could decrease surgical site infection (SSI) in patients scheduled for abdominal surgery. Methods: We performed a multicentre, randomised controlled clinical trial in a network of 21 university hospitals from June 6, 2017 to July 19, 2018. Patients undergoing abdominal surgery were randomly assigned to receive a high (0.80) or conventional (0.3) FIO2 during the intraoperative period and during the first 3 postoperative hours. All patients were mechanically ventilated with an open-lung strategy, which included recruitment manoeuvres and individualised positive end-expiratory pressure for the best respiratory-system compliance, and individualised continuous postoperative airway pressure for adequate peripheral oxyhaemoglobin saturation. The primary outcome was the prevalence of SSI within the first 7 postoperative days. The secondary outcomes were composites of systemic complications, length of intensive care and hospital stay, and 6-month mortality. Results: We enrolled 740 subjects: 371 in the high FIO2 group and 369 in the low FIO2 group. Data from 717 subjects were available for final analysis. The rate of SSI during the first postoperative week did not differ between high (8.9%) and low (9.4%) FIO2 groups (relative risk [RR]: 0.94; 95% confidence interval [CI]: 0.59-1.50; P=0.90]). Secondary outcomes, such as atelectasis (7.7% vs 9.8%; RR: 0.77; 95% CI: 0.48-1.25; P=0.38) and myocardial ischaemia (0.6% [n=2] vs 0% [n=0]; P=0.47) did not differ between groups. Conclusions: An oxygenation strategy using high FIO2 compared with conventional FIO2 did not reduce postoperative SSIs in abdominal surgery. No differences in secondary outcomes or adverse events were found.
|
|
| 4. |
- Kehoe, Laura, et al.
(författare)
-
Make EU trade with Brazil sustainable
- 2019
-
Ingår i: Science. - : American Association for the Advancement of Science (AAAS). - 0036-8075 .- 1095-9203. ; 364:6438, s. 341-
-
Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)
|
|
| 5. |
- Alvarez-Rodríguez, Manuel, et al.
(författare)
-
Brown bear sperm double freezing : Effect of elapsed time and use of PureSperm(®) gradient between freeze-thaw cycles.
- 2013
-
Ingår i: Cryobiology. - : Elsevier. - 0011-2240 .- 1090-2392. ; 67:3, s. 339-346
-
Tidskriftsartikel (refereegranskat)abstract
- The use of sexed spermatozoa has great potential to captive population management in endangered wildlife. The problem is that the sex-sorting facility is a long distance from the semen collection place and to overcome this difficulty two freeze-thaw cycles may be necessary. In this study, effects of refreezing on brown bear electroejaculated spermatozoa were analyzed. We carried out two experiments: (1) to assess the effects of the two freezing-thawing cycles on sperm quality and to analyze three different elapsed times between freezing-thawing cycles (30, 90 and 180 min), and (2) to analyze the use of PureSperm between freezing-thawing cycles to select a more motile and viable sperm subpopulation which better survived first freezing. The motility, viability and undamaged acrosomes were significantly reduced after the second thawing respect to first thawing into each elapsed time group, but the elapsed times did not significantly affect the viability and acrosome status although motility was damaged. Our results with the PureSperm gradient showed higher values of viability in freezability of select sample (pellet) respect to the rest of the groups and it also showed a significant decrease in the number of acrosome damaged. In summary, the double freezing of bear semen selected by gradient centrifugation is qualitatively efficient, and thus could be useful to carry out a sex-sorting of frozen-thawed bear spermatozoa before to send the cryopreserved sample to a biobank. Given the low recovery of spermatozoa after applying a selection gradient, further studies will be needed to increase the recovery rate without damaging of the cell quality.
|
|
| 6. |
- Alvarez-Rodríguez, Manuel, et al.
(författare)
-
The antioxidant effects of soybean lecithin- or low-density lipoprotein-based extenders for the cryopreservation of brown-bear (Ursus arctos) spermatozoa.
- 2013
-
Ingår i: Reproduction, Fertility and Development. - : CSIRO Publishing. - 1031-3613 .- 1448-5990. ; 25:8, s. 1185-1193
-
Tidskriftsartikel (refereegranskat)abstract
- Egg yolk low-density lipoproteins (LDL) and soybean lecithin were evaluated as replacements for egg yolk in extenders used for the cryopreservation of brown-bear spermatozoa. The motility, viability and acrosomal status of post-thawed spermatozoa were analysed, and an egg-yolk extender was used as a control. The total antioxidant capacity of these extenders was tested. Soybean lecithin showed an effect that was dependent on the soybean concentration (2%, 3.5% and 5%) and source (Type A: 24% L-α-phosphatidylcholine, and Type B: 14-23% L-α-phosphatidylcholine). Only semen cryopreserved with 5% Type A soybean exhibited a sperm motility similar to that of semen cryopreserved in egg-yolk-based extender after thawing, although the sperm viability and acrosome status were not as high. Semen frozen in an extender containing LDL (10-15%) exhibited improved sperm viability in comparison with the control, but sperm motility was lower. The LDL-based extender exhibited a higher anti-oxidant activity than the egg-yolk extender and soy lecithin-based extenders. The extenders with higher anti-oxidant activity showed improvements in frozen sperm viability but lower semen motility. These results indicate that soybean lecithin did not have the same protective effect as egg yolk during the freezing of brown-bear spermatozoa but suggest that LDL (10-15%) could be a useful substitute for egg yolk in these extenders.
|
|
| 7. |
- Anel-López, Luis, et al.
(författare)
-
Reduced glutathione and Trolox (vitamin E) as extender supplements in cryopreservation of red deer epididymal spermatozoa
- 2012
-
Ingår i: Animal Reproduction Science. - : Elsevier. - 0378-4320 .- 1873-2232. ; 135:1-4, s. 37-46
-
Tidskriftsartikel (refereegranskat)abstract
- The use of assisted reproductive techniques in cervids is increasing as the commercial use of these species increase. We have tested the suitability of the antioxidants Trolox and reduced glutathione (GSH) for freezing red deer epididymal spermatozoa, aiming at improving post-thawing quality. Samples from 19 stags were frozen in a TES-Tris-fructose extender (20% egg yolk, 8% glycerol), with 1 or 5 mM of antioxidant. Motility (CASA), lipoperoxidation (malondialdehyde -MDA- production), membrane status, mitochondrial activity, acrosomal status (flow cytometry) and chromatin status (SCSA: %DFI and %HDS; flow cytometry) were assessed after thawing and after 6 h at 39°C. There were few differences between treatments after thawing, with Trolox reducing MDA production in a dose-response manner. After the incubation, sperm quality decreased and %DFI increased moderately, with no change for MDA. GSH improved motility, kinematic parameters and mitochondrial status, with a slight increase in %HDS. GSH 5 mM also increased moderately MDA production and %DFI, possibly due to enhanced metabolic activity and reducing power. Trolox maintained MDA low, but was detrimental to sperm quality. Trolox might not be appropriate for the cryopreservation of red deer epididymal spermatozoa, at least at the millimolar range. GSH results are promising, especially regarding motility improvement after the post-thawing incubation, and should be selected for future fertility trials.
|
|
| 8. |
- de Paz, Paulino, et al.
(författare)
-
Optimization of glycerol concentration and freezing rate in the cryopreservation of ejaculate from brown bear (Ursus arctos).
- 2012
-
Ingår i: Reproduction in domestic animals. - : Blackwell Verlag. - 0936-6768 .- 1439-0531. ; 47:1, s. 105-112
-
Tidskriftsartikel (refereegranskat)abstract
- In order to establish a semen bank for the endangered Cantabrian brown bear, we tested five glycerol concentrations and three freezing rates for electroejaculated semen. Electroejaculation was performed on nine males. Semen was diluted in TES-Tris-Fructose (20% egg yolk, 2% EDTA, 1% Equex) with 2%, 4%, 6%, 8% or 10% glycerol and frozen at -10, -20 or -40°C/min. Before and after cryopreservation, samples were analysed for motility (CASA), viability and acrosomal status (flow cytometry). Pre-freezing results showed that glycerol concentration had no significant effect on total motility or progressive motility, but it significantly decreased VCL, ALH, viability and acrosomal status (p < 0.05). After thawing, sperm motility was higher at extender with 4%, 6% and 8% glycerol, but only at 4% and 6% glycerol for viability and acrosomal status. For 4% and 6% glycerol, freezing rates did not have significant effects. The curve fitting gave an estimate of the optimal glycerol concentration, with all the optimal values for every parameter between 6% and 7% glycerol falling. We propose using 6% glycerol and a freezing velocity of -20°C/min for freezing brown bear ejaculated spermatozoa.
|
|
| 9. |
- Lacalle, Estibaliz, et al.
(författare)
-
Bicarbonate-Triggered In Vitro Capacitation of Boar Spermatozoa Conveys an Increased Relative Abundance of the Canonical Transient Receptor Potential Cation (TRPC) Channels 3, 4, 6 and 7 and of CatSper-gamma Subunit mRNA Transcripts
- 2022
-
Ingår i: Animals. - : MDPI. - 2076-2615. ; 12:8
-
Tidskriftsartikel (refereegranskat)abstract
- Simple Summary The detection of sub-fertile boars has been a difficult task, and despite their prevalence being low, its impact is very significant because it implies economic drawbacks for artificial insemination (AI) centers and farms. Unfortunately, some crucial reproductive processes fall beyond the routine analysis performed in the porcine model, such as sperm capacitation, which is a necessary event for fertilization. A synergistic action of bicarbonate (HCO3-) with calcium (Ca2+) is needed to achieve capacitation. The transport of Ca2+ is mediated by CatSper channels and Canonical Transient Potential Channels (TRPC). We quantified mRNA transcripts of different subunits of CatSper (beta, gamma and delta) and TRPC (1, 3, 4, 6 and 7) before and after in vitro capacitation by HCO3- ions. Our results showed that in vitro capacitation using HCO3- increases the relative abundance of mRNA transcripts of almost all subunits of Ca2+ channels, except CatSper-delta and TRPC1, which were significantly reduced. More studies are needed to elucidate the specific roles of the TRPC channels at a physiological and functional level. Sperm capacitation is a stepwise complex biochemical process towards fertilization. It includes a crucial early calcium (Ca2+) transport mediated by CatSper channels and Canonical Transient Potential Channels (TRPC). We studied the relative abundance of mRNA transcripts changes of the CatSper beta, gamma and delta subunits and TRPC-channels 1, 3, 4, 6 and 7 in pig spermatozoa, after triggering in vitro capacitation by bicarbonate ions at levels present in vivo at the fertilization site. For this purpose, we analyzedfive5 ejaculate pools (from three fertile adult boars) before (control-fresh samples) and after in vitro exposure to capacitation conditions (37 mM NaHCO3, 2.25 mM CaCl2, 2 mM caffeine, 0.5% bovine serum albumin and 310 mM lactose) at 38 degrees C, 5% CO2 for 30 min. In vitro capacitation using bicarbonate elicits an increase in the relative abundance of mRNA transcripts of almost all studied Ca2+ channels, except CatSper-delta and TRPC1 (significantly reduced). These findings open new avenues of research to identify the specific role of each channel in boar sperm capacitation and elucidate the physiological meaning of the changes on sperm mRNA cargo.
|
|
| 10. |
- Naghavi, Mohsen, et al.
(författare)
-
Global, regional, and national age-sex specific all-cause and cause-specific mortality for 240 causes of death, 1990-2013: a systematic analysis for the Global Burden of Disease Study 2013
- 2015
-
Ingår i: The Lancet. - 1474-547X .- 0140-6736. ; 385:9963, s. 117-171
-
Tidskriftsartikel (refereegranskat)abstract
- Background Up-to-date evidence on levels and trends for age-sex-specifi c all-cause and cause-specifi c mortality is essential for the formation of global, regional, and national health policies. In the Global Burden of Disease Study 2013 (GBD 2013) we estimated yearly deaths for 188 countries between 1990, and 2013. We used the results to assess whether there is epidemiological convergence across countries. Methods We estimated age-sex-specifi c all-cause mortality using the GBD 2010 methods with some refinements to improve accuracy applied to an updated database of vital registration, survey, and census data. We generally estimated cause of death as in the GBD 2010. Key improvements included the addition of more recent vital registration data for 72 countries, an updated verbal autopsy literature review, two new and detailed data systems for China, and more detail for Mexico, UK, Turkey, and Russia. We improved statistical models for garbage code redistribution. We used six different modelling strategies across the 240 causes; cause of death ensemble modelling (CODEm) was the dominant strategy for causes with sufficient information. Trends for Alzheimer's disease and other dementias were informed by meta-regression of prevalence studies. For pathogen-specifi c causes of diarrhoea and lower respiratory infections we used a counterfactual approach. We computed two measures of convergence (inequality) across countries: the average relative difference across all pairs of countries (Gini coefficient) and the average absolute difference across countries. To summarise broad findings, we used multiple decrement life-tables to decompose probabilities of death from birth to exact age 15 years, from exact age 15 years to exact age 50 years, and from exact age 50 years to exact age 75 years, and life expectancy at birth into major causes. For all quantities reported, we computed 95% uncertainty intervals (UIs). We constrained cause-specific fractions within each age-sex-country-year group to sum to all-cause mortality based on draws from the uncertainty distributions. Findings Global life expectancy for both sexes increased from 65.3 years (UI 65.0-65.6) in 1990, to 71.5 years (UI 71.0-71.9) in 2013, while the number of deaths increased from 47.5 million (UI 46.8-48.2) to 54.9 million (UI 53.6-56.3) over the same interval. Global progress masked variation by age and sex: for children, average absolute diff erences between countries decreased but relative diff erences increased. For women aged 25-39 years and older than 75 years and for men aged 20-49 years and 65 years and older, both absolute and relative diff erences increased. Decomposition of global and regional life expectancy showed the prominent role of reductions in age-standardised death rates for cardiovascular diseases and cancers in high-income regions, and reductions in child deaths from diarrhoea, lower respiratory infections, and neonatal causes in low-income regions. HIV/AIDS reduced life expectancy in southern sub-Saharan Africa. For most communicable causes of death both numbers of deaths and age-standardised death rates fell whereas for most non-communicable causes, demographic shifts have increased numbers of deaths but decreased age-standardised death rates. Global deaths from injury increased by 10.7%, from 4.3 million deaths in 1990 to 4.8 million in 2013; but age-standardised rates declined over the same period by 21%. For some causes of more than 100 000 deaths per year in 2013, age-standardised death rates increased between 1990 and 2013, including HIV/AIDS, pancreatic cancer, atrial fibrillation and flutter, drug use disorders, diabetes, chronic kidney disease, and sickle-cell anaemias. Diarrhoeal diseases, lower respiratory infections, neonatal causes, and malaria are still in the top five causes of death in children younger than 5 years. The most important pathogens are rotavirus for diarrhoea and pneumococcus for lower respiratory infections. Country-specific probabilities of death over three phases of life were substantially varied between and within regions. Interpretation For most countries, the general pattern of reductions in age-sex specifi c mortality has been associated with a progressive shift towards a larger share of the remaining deaths caused by non-communicable disease and injuries. Assessing epidemiological convergence across countries depends on whether an absolute or relative measure of inequality is used. Nevertheless, age-standardised death rates for seven substantial causes are increasing, suggesting the potential for reversals in some countries. Important gaps exist in the empirical data for cause of death estimates for some countries; for example, no national data for India are available for the past decade.
|
|
| 11. |
- Nicolas, Manuel, et al.
(författare)
-
Evaluation of the qualitative and quantitative effectiveness of three media of centrifugation (Maxifreeze, Cushion Fluid Equine, and PureSperm 100) in preparation of fresh or frozen-thawed brown bear spermatozoa.
- 2012
-
Ingår i: Theriogenology. - : Elsevier. - 0093-691X .- 1879-3231. ; 77:6, s. 1119-1128
-
Tidskriftsartikel (refereegranskat)abstract
- Centrifugation is a crucial procedure in sperm cryopreservation protocols of brown bear (Ursus arctos), because the semen must be processed to increase sperm concentration and/or clean urine-contaminated samples. The efficacy of three media for centrifugation (Maxifreeze [IMV technologies, L'Aigle, France], Cushion Fluid Equine (Minitübe, Tiefenbach, Germany), and PureSperm [Nidacon, Gothenburg, Sweden]) on the quality of bear spermatozoa was evaluated. In experiment one, two cushioned media used for protecting against mechanical stress during centrifugation were analyzed. In experiment two, a density gradient based on PureSperm was assessed in relation to the maximum retrieval and the quality of fresh spermatozoa, and the freezability of the spermatozoa selected in this density gradient was studied in experiment three. Finally, the selection of frozen-thawed sperm using PureSperm was analyzed in experiment four. Our results indicate that the use of dense isotonic cushion solutions (Maxifreeze, Cushion Fluid Equine) in centrifugation did not improve the quality of recovered spermatozoa compared with standard centrifugation. However, a density gradient prepared with PureSperm improved the quality of spermatozoa in fresh semen and frozen-thawed semen, but the spermatozoa selected from the fresh sample with this density gradient did not show a better resistance to freezing with this density gradient in comparison with the control sample.
|
|
| 12. |
- Alvarez-Rodriguez, Manuel, et al.
(författare)
-
Chicken seminal fluid lacks CD9-and CD44-bearing extracellular vesicles
- 2020
-
Ingår i: Reproduction in domestic animals. - : Wiley-VCH Verlagsgesellschaft. - 0936-6768 .- 1439-0531. ; 55:3, s. 293-300
-
Tidskriftsartikel (refereegranskat)abstract
- The avian seminal fluid (SF) is a protein-rich fluid, derived from the testis, the rudimentary epididymis and, finally, from the cloacal gland. The SF interacts with spermatozoa and the inner cell lining of the female genital tract, to modulate sperm functions and female immune responsiveness. Its complex proteome might either be free or linked to extracellular vesicles (EVs) as it is the case in mammals, where EVs depict the tetraspanin CD9; and where those EVs derived from the epididymis (epididymosomes) also present the receptor CD44. In the present study, sperm-free SF from Red Jungle Fowl, White Leghorn and an advanced intercross (AIL, 12th generation) were studied using flow cytometry of the membrane marker tetraspanin CD9, Western blotting of the membrane receptor CD44 and electron microscopy in non-enriched (whole SF) or enriched fractions obtained by precipitation using a commercial kit (Total Exosome Precipitation Solution). Neither CD9- nor CD44 could be detected, and the ultrastructure confirmed the relative absence of EVs, raising the possibility that avian SF interacts differently with the female genitalia as compared to the seminal plasma of mammals.
|
|
| 13. |
- Alvarez-Rodriguez, Manuel, et al.
(författare)
-
Does the Act of Copulation per se, without Considering Seminal Deposition, Change the Expression of Genes in the Porcine Female Genital Tract?
- 2020
-
Ingår i: International Journal of Molecular Sciences. - : MDPI. - 1661-6596 .- 1422-0067. ; 21:15
-
Tidskriftsartikel (refereegranskat)abstract
- Semen-through its specific sperm and seminal plasma (SP) constituents-induces changes of gene expression in the internal genital tract of pigs, particularly in the functional sperm reservoir at the utero-tubal junction (UTJ). Although seminal effects are similarly elicited by artificial insemination (AI), major changes in gene expression are registered after natural mating, a fact suggesting the act of copulation induces per se changes in genes that AI does not affect. The present study explored which pathways were solely influenced by copulation, affecting the differential expression of genes (DEGs) of the pre/peri-ovulatory genital tract (cervix, distal uterus, proximal uterus and UTJ) of estrus sows, 24 h after various procedures were performed to compare natural mating with AI of semen (control 1), sperm-free SP harvested from the sperm-peak fraction (control 2), sperm-free SP harvested from the whole ejaculate (control 3) or saline-extender BTS (control 4), using a microarray chip (GeneChip(R)porcine gene 1.0 st array). Genes related to neuroendocrine responses (ADRA1,ADRA2,GABRB2,CACNB2), smooth muscle contractility (WNT7A), angiogenesis and vascular remodeling (poFUT1,NTN4) were, among others, overrepresented with distal and proximal uterine segments exhibiting the highest number of DEGs. The findings provide novel evidence that relevant transcriptomic changes in the porcine female reproductive tract occur in direct response to the specific act of copulation, being semen-independent.
|
|
| 14. |
- Alvarez-Rodriguez, Manuel, et al.
(författare)
-
Effect of colloid (Androcoll-Bear, Percoll, and PureSperm) selection on the freezability of brown bear (Ursus arctos) sperm
- 2016
-
Ingår i: Theriogenology. - : ELSEVIER SCIENCE INC. - 0093-691X .- 1879-3231. ; 85:6, s. 1097-1105
-
Tidskriftsartikel (refereegranskat)abstract
- The development of a species-specific conservation protocol that involves artificial insemination with frozen semen needs to validate an effective methodology for freezing semen. Colloid centrifugation has been suggested and widely applied as an effective tool for selecting animal spermatozoa for artificial breeding. The objective of the present study was to compare different methods of centrifugation, single layer using Androcoll-Bear and Percoll and double layer using PureSperm 100 (in two different discontinuous gradients 40%-80% and 45%-90%), for the selection of fresh brown bear sperm samples. In the before freezing group, all selected samples showed a higher progressive motility and viability (except Percoll for motility 43.0 +/- 5.3 [P < 0.051); all colloids except PureSperm 45/90% rendered samples with fewer damaged acrosomes. In the after thawing group, all tested centrifugation colloids showed a good capacity to decrease the number of damaged acrosomes. Furthermore, PureSperm treatment (45/90%) resulted in an increase in apoptotic-like changes not only immediately after thawing but also after the incubation test, leading us to suggest that this gradient could induce some kind of deleterious effects on the sperm samples. On the other hand, PureSperm treatment (40/80%) yielded a quality preservation capacity similar to Androcoll-Bear in number of damaged acrosomes, different relative to the control (control, 5.3 +/- 0.6; PureSperm 80, 2.0 +/- 0.3; Androcoll, 2.1 +/- 0.9 [P < 0.051) but a decrease in the number of viable spermatozoa recovered after thawing relative to the control (control, 21.2 +/- 3.1; PureSperm 80, 13.7 +/- 2.7 [P < 0.051). In conclusion, Androcoll-Bear constitutes a useful tool for handling of brown bear ejaculates owing to its simple handling and procedure with a reliable sperm selection and freezability. This colloid yielded an improvement in several sperm parameters in brown bear frozen-thawed semen; the selected spermatozoa of fresh samples with this colloid showed a better resistance to freezing compared with the control sample not only for motility but also for viability. (C) 2016 Elsevier Inc. All rights reserved.
|
|
| 15. |
- Alvarez-Rodriguez, Manuel, et al.
(författare)
-
Effect of length of time post-mortem on quality and freezing capacity of Cantabric chamois (Rupicapra pyrenaica parva) epididymal spermatozoa
- 2018
-
Ingår i: Animal Reproduction Science. - : ELSEVIER SCIENCE BV. - 0378-4320 .- 1873-2232. ; 198, s. 184-192
-
Tidskriftsartikel (refereegranskat)abstract
- Genome Resource Banks are keystones in the ex-situ conservation of wild species. Post-mortem (PM) collection of epididymal spermatozoa is an opportunistic and valuable source of germplasm, the time from the death of the animal limits its use. Seeking to improve germplasm preservation strategies for the chamois (Rupicapra sp.), the effect of PM time on epididymal sperm quality and freezability was studied using the Cantabrian chamois. Samples were classified according to PM collection time, up to 216 h (refrigerated), and cryopreserved (Tris-citric acid-fructose, 430 mOsm/kg, 15% egg yolk, 8% glycerol; freezing at - 20 degrees C/min). Sperm quality was assessed after recovery and post-thawing (motility by CASA, HOS test, abnormal forms, cytoplasmic droplets, and viability and acrosomal damage by flow cytometry). The sperm mass pH and osmolality showed a positive correlation with time. Total sperm motility dropped after 2 days PM, with progressivity and sperm velocities remained similar up to 3 days PM. Sperm freezability was acceptable, with the post-thawing HOST, motility, progressivity, VAP, VCL, VSL and BCF negatively correlating with PM time. Overall, chamois epidydimal samples were not adequate for preservation after 6 days PM. Freezability capacity could make these spermatozoa suitable for specific ART even if kept refrigerated for several days PM.
|
|
| 16. |
- Alvarez-Rodriguez, Manuel, 1983-, et al.
(författare)
-
Exogenous Individual Lecithin-Phospholipids (Phosphatidylcholine and Phosphatidylglycerol) Cannot Prevent the Oxidative Stress Imposed by Cryopreservation of Boar Sperm.
- 2017
-
Ingår i: Journal of veterinary medicine and surgery. - : imedpub. - 2574-2868. ; 1:1
-
Tidskriftsartikel (refereegranskat)abstract
- Objective: Despite the use of high proportions of the chemically undefined lipoprotein/phospholipid-rich egg-yolk in extenders, boar sperm are highly sensitive to cooling, which induces ROS generation and disrupts the plasma membrane.Here, we studied whether replacement of hen egg-yolk by commercially defined lecithin phospholipids, derived from egg (LPGE: phosphatidyl glycerol, LPCE: phosphatidyl choline) or soybean (LPCS: phosphatidyl choline), could individually ameliorate such oxidative effects during cryopreservation of ejaculated (sperm rich fraction, SRF) or of cauda-epididymal sperm, retrieved post-mortem from the same males.Methods: A conventional extender (lactose buffer, with 20% egg-yolk, 0.5% OEP and 3% glycerol) was used as control. Cryodamage was assessed as loss of sperm motility, membrane and acrosome intactness, early membrane destabilization changes, mitochondrial potential, superoxide and ROS production, to finally determine lipid peroxidation (LPO) using specific probes.Results and conclusion: In general, the exogenous phospholipids assayed were unable of maintaining neither sperm motility nor viability post-thaw compared to controls, owing to increased ROS production and lipid peroxidation. In our study, mitochondrial superoxide production resulted in very high levels for all groups, whereas both ROS production and lipid peroxidation were reduced in the control group, containing emulsified hen egg yolk. Further studies using various dosage and combination of LPCS should be followed for their eventual protective effect.Keywords: Cryodamage; Sperm; Boar; Mitochondrial activation; Mitochondrial superoxide; ROS production; Lipid peroxidation
|
|
| 17. |
- Alvarez-Rodriguez, Manuel, et al.
(författare)
-
Exosomes in specific fractions of the boar ejaculate contain CD44: A marker for epididymosomes?
- 2019
-
Ingår i: Theriogenology. - : ELSEVIER SCIENCE INC. - 0093-691X .- 1879-3231. ; 140, s. 143-152
-
Tidskriftsartikel (refereegranskat)abstract
- Seminal plasma (SP) is a complex fluid containing proteins, peptides, enzymes, hormones as well as extracellular vesicles (EVs). The SP interacts with spermatozoa and the inner cell lining of the female genital tract, adsorbing proteins and exosomes that modulate sperm functions and female immune responsiveness. In the present study, boar sperm-free SP was studied using flow cytometry (FC) after membrane tetraspanins (CD9, CD63 and CD81) and membrane receptor CD44 marking of non-enriched (whole SP) or gradient fractions enriched through two-step discontinuous KBr-density-gradient ultracentrifugation, in whole ejaculate or in selected ejaculate fractions. The results, evaluated by transmission electron microscopy, confirmed the presence of exosomes in all fractions of the pig SP. Noteworthy, these pig SP-exosomes were CD44-bearing when analysed by FC, with bands detected by western blotting (WB) at the expected 85 kD size. The two-step discontinuous KBr-density-gradient ultracentrifugation enriched the population of exosomes in two specific gradient fractions, indicating exosomes (either prostasomes or epididymosomes) could be separated from low-density lipoprotein (LDL) but they co-sediment with the high-density lipoprotein (HDL)-bearing fraction. The findings pave for the selective isolation of exosomes in functional studies of their function when interacting with spermatozoa, the oocyte and/or the female genitalia, including hyaluronan-CD44 interplay. (C) 2019 Elsevier Inc. All rights reserved.
|
|
| 18. |
- Alvarez-Rodriguez, Manuel, 1983-, et al.
(författare)
-
Expression of Immune Regulatory Genes in the Porcine Internal Genital Tract Is Differentially Triggered by Spermatozoa and Seminal Plasma
- 2019
-
Ingår i: International Journal of Molecular Sciences. - : MDPI. - 1661-6596 .- 1422-0067. ; 20:3
-
Tidskriftsartikel (refereegranskat)abstract
- Mating or cervical deposition of spermatozoa or seminal plasma (SP) modifies the expression of genes affecting local immune defense processes at the oviductal sperm reservoir in animals with internal fertilization, frequently by down-regulation. Such responses may occur alongside sperm transport to or even beyond the reservoir. Here, immune-related gene expression was explored with cDNA microarrays on porcine cervix-to-infundibulum tissues, pre-/peri-ovulation. Samples were collected 24 h post-mating or cervical deposition of sperm-peak spermatozoa or SP (from the sperm-peak fraction or the whole ejaculate). All treatments of this interventional study affected gene expression. The concerted action of spermatozoa and SP down-regulated chemokine and cytokine (P00031), interferon-gamma signaling (P00035), and JAK/STAT (P00038) pathways in segments up to the sperm reservoir (utero-tubal junction (UTJ)/isthmus). Spermatozoa in the vanguard sperm-peak fraction (P1-AI), uniquely displayed an up-regulatory effect on these pathways in the ampulla and infundibulum. Sperm-free SP, on the other hand, did not lead to major effects on gene expression, despite the clinical notion that SP mitigates reactivity by the female immune system after mating or artificial insemination.
|
|
| 19. |
- Alvarez-Rodriguez, Manuel, et al.
(författare)
-
Hyaluronan improves neither the long-term storage nor the cryosurvival of liquid-stored CD44-bearing Al boar spermatozoa
- 2018
-
Ingår i: Journal of reproduction and development. - : SOCIETY REPRODUCTION & DEVELOPMENT-SRD. - 0916-8818 .- 1348-4400. ; 64:4, s. 351-360
-
Tidskriftsartikel (refereegranskat)abstract
- Hyaluronan (hyaluronic acid, HA) apparently improves sperm survival in vitro and in vivo (oviduct), maintaining sperm motility and inducing capacitation, but not acrosome exocytosis, either by direct action as a macromolecule or via CD44 membrane receptors. This study explored ejaculated, liquid-extended pig spermatozoa to ascertain (i) the presence (Western blotting) and specific location (immunocytochemistry) of the CD44 receptor, using a specific monoclonal commercial antibody; (ii) whether the CD44 receptor changed location when exposed to bicarbonate, a capacitating trigger, in vitro; and (iii) whether the addition of HA, of molecular size comparable to that produced in the oviduct sperm reservoir (0.0625 to 2.0 mg/ml; 0 HA: control), to semen extenders would improve sperm liquid storage in vitro or cryosurvival post freezing. Variables tested were sperm velocity and progressive motility (Qualisperm (TM)), sperm viability and acrosome status, membrane integrity and early destabilization, mitochondrial activation, and superoxide production (flow cytometry). The CD44 receptor presence in ejaculated, liquid-stored AI boar spermatozoa, as confirmed by a porcine-specific monoclonal antibody, maintained its membrane location under in vitro capacitation-inducing conditions. HA exposure to 24-, 48-, or 72-h liquid-stored (17-20 degrees C) spermatozoa lowered sperm velocity in membrane-intact spermatozoa, but increased mitochondrial superoxide production. Finally, HA addition during cooling did not improve cryosurvival but did increase mitochondrial activation and membrane destabilization in surviving cells. These results confirm the existence of a CD44 receptor in pig spermatozoa, but the usefulness of adding HA for long-term storage or cryopreservation of liquid-stored, extended boar semen remains in question, thereby warranting further non-empirical analyses of HA-sperm membrane interactions.
|
|
| 20. |
- Alvarez-Rodriguez, Manuel, et al.
(författare)
-
Mating modifies the expression of crucial oxidative-reductive transcripts in the pig oviductal sperm reservoir: is the female ensuring sperm survival?
- 2023
-
Ingår i: Frontiers in Endocrinology. - : FRONTIERS MEDIA SA. - 1664-2392. ; 14
-
Tidskriftsartikel (refereegranskat)abstract
- BackgroundMating induces large changes in the female genital tract, warranting female homeostasis and immune preparation for pregnancy, including the preservation of crucial oxidative status among its pathways. Being highly susceptible to oxidative stress, sperm survival and preserved function depend on the seminal plasma, a protection that is removed during sperm handling but also after mating when spermatozoa enter the oviduct. Therefore, it is pertinent to consider that the female sperm reservoir takes up this protection, providing a suitable environment for sperm viability. These aspects have not been explored despite the increasing strategies in modulating the female status through diet control and nutritional supplementation. AimsTo test the hypothesis that mating modifies the expression of crucial oxidative-reductive transcripts across the entire pig female genital tract (cervix to infundibulum) and, particularly in the sperm reservoir at the utero-tubal junction, before ovulation, a period dominated by estrogen stimulation of ovarian as well as of seminal origin. MethodsThe differential expression of estrogen (ER) and progesterone (PR) receptors and of 59 oxidative-reductive transcripts were studied using a species-specific microarray platform, in specific segments of the peri-ovulatory sow reproductive tract in response to mating. ResultsMating induced changes along the entire tract, with a conspicuous downregulation of both ER and PR and an upregulation of superoxide dismutase 1 (SOD1), glutaredoxin (GLRX3), and peroxiredoxin 1 and 3 (PRDX1, PRDX3), among other NADH Dehydrogenase Ubiquinone Flavoproteins, in the distal uterus segment. These changes perhaps helped prevent oxidative stress in the area adjacent to the sperm reservoir at the utero-tubal junction. Concomitantly, there were a downregulation of catalase (CAT) and NADH dehydrogenase (ubiquinone) oxidoreductases 1 beta subcomplex, subunit 1 (NDUFB1) in the utero-tubal junction alongside an overall downregulation of CAT, SOD1, and PRDX3 in the ampullar and infundibulum segments. ConclusionsNatural mating is an inducer of changes in the expression of female genes commanding antioxidant enzymes relevant for sperm survival during sperm transport, under predominant estrogen influence through the bloodstream and semen. The findings could contribute to the design of new therapeutics for the female to improve oxidative-reductive balance.
|
|
| 21. |
- Alvarez-Rodriguez, Manuel, 1983-
(författare)
-
Mejora y evolución de los protocolos de congelación de eyaculados de oso pardo (Ursus arctos)
- 2012
-
Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- La situación crítica del oso pardo (Ursus arctos) en España plantea la necesidad de aplicar diferentes estrategias de conservación, dentro de las cuales tiene especial relevancia la creación de un banco de recursos genéticos. La eficacia de esta herramienta depende, entre otros factores, de la adaptación específica de los protocolos de congelación espermática ya existentes y que permitan el almacenamiento eficaz de los espermatozoides del oso pardo en un banco de germoplasma. Este trabajo nos permite concluir la utilidad de la selección entre ciclos con el fin de mejorar la calidad de los espermatozoides sometidos a recongelación
|
|
| 22. |
- Alvarez-Rodriguez, Manuel, et al.
(författare)
-
MicroRNA expression in specific segments of the pig periovulatory internal genital tract is differentially regulated by semen or by seminal plasma
- 2024
-
Ingår i: Research in Veterinary Science. - : ELSEVIER SCI LTD. - 0034-5288 .- 1532-2661. ; 168
-
Tidskriftsartikel (refereegranskat)abstract
- microRNAs play pivotal roles during mammalian reproduction, including the cross-talk between gametes, embryos and the maternal genital tract. Mating induces changes in the expression of mRNA transcripts in the female, but whether miRNAs are involved remains to be elucidated. In the current study, we mapped 181 miRNAs in the porcine peri-ovulatory female reproductive tract: Cervix (Cvx), distal and proximal uterus (Dist-Ut, ProxUt), Utero-tubal-junction (UTJ), isthmus (Isth), ampulla (Amp), and infundibulum (Inf) when exposed to semen (natural mating (NM) or artificial insemination (AI-P1)) or to infusions of sperm-free seminal plasma (SP): the first 10 mL of the sperm rich fraction (SP-P1) or the entire ejaculate (SP-E). Among the most interesting findings, NM decreased mir-671, implicated in uterine development and pregnancy loss prior to embryo implantation, in Cvx, Dist-UT, Prox-UT, Isth, and Inf, while it increased in Amp. NM and SP-E induced the downregulation of miRlet7A-1 (Dist-UT, Prox-UT), a regulator of immunity during pregnancy. miR-34C-1, a regulator of endometrial receptivity gene expression, was increased in Dist-UT, UTJ and Amp (NM), in Prox-UT (AI-P1), and in Amp (SPP1). miR-296, a modulator of the inflammatory response and apoptosis, was upregulated in the UTJ (all treatments). NM elicited the highest miRNA activity in the sperm reservoir (UTJ), suggesting that key-regulators such as miR-34c or miR-296 may modulate the metabolic processes linked to the adequate preparation for gamete encounter in the oviduct. Our results suggest that SP should be maintained in AI to warrant miRNA regulation within the female genital tract for reproductive success.
|
|
| 23. |
- Alvarez-Rodriguez, Manuel, et al.
(författare)
-
mRNA expression of oxidative-reductive proteins in boars with documented different fertility can identify relevant prognostic biomarkers
- 2021
-
Ingår i: Research in Veterinary Science. - : Elsevier Science Ltd. - 0034-5288 .- 1532-2661. ; 141, s. 195-202
-
Tidskriftsartikel (refereegranskat)abstract
- Oxidative stress unbalance is a major factor causing impairment of sperm function and, ultimately, sperm death. In this study, we identified transcriptomic and proteomic markers for oxidative-related protectors from the generation of reactive oxygen species (ROS) in spermatozoa from breeding boars with documented high- or lowfertility. Particular attention was paid to glutathione peroxidases, and to transcripts related to DNA stabilization and compaction, as protamine and transition proteins. mRNA cargo analysis was performed using porcinespecific micro-arrays (GeneChip (R) miRNA 4.0 and GeneChip (R) Porcine Gene 1.0 ST) and qPCR validation. Differences between fertility-classed boars were ample among biomarkers; some upregulated only at protein level (catalase (CAT), superoxide dismutase 1 (SOD1) and glutathione proteins), or only at the mRNA level (ATOX1, Antioxidant Protein 1). In addition, protamines 2 and 3, essential for sperm DNA condensation and also transition proteins 1 and 2 (TNP1 and TNP2), required during histone-to-protamine replacement, were overexpressed in spermatozoa from high-fertile boars. This up-regulation seems concerted to reduce DNA accessibility to ROS attack, protecting the DNA. The upregulated intracellular phospholipid hydroperoxide glutathione peroxidase (GPx4), in high-fertile boars at mRNA level, can be considered a most relevant biomarker for fertility disclosure during sperm evaluation.
|
|
| 24. |
- Alvarez-Rodriguez, Manuel, et al.
(författare)
-
Quality of frozen-thawed semen in brown bear is not affected by timing of glycerol addition.
- 2011
-
Ingår i: Theriogenology. - : Elsevier. - 0093-691X .- 1879-3231. ; 75:8, s. 1561-1565
-
Tidskriftsartikel (refereegranskat)abstract
- We have tested several freezing protocols for brown bear semen, modifying the time when glycerol was added (before and after cooling to 5 °C). No differences were found among protocols, indicating a good tolerance of brown bear semen to glycerol. This finding indicates that freezing protocols for brown bear semen could be modified to fit practical solutions which would facilitate preparation of the seminal samples in the field with the addition of glycerol at ambient temperature.
|
|
| 25. |
- Alvarez-Rodríguez, Manuel, et al.
(författare)
-
The addition of heat shock protein HSPA8 to cryoprotective media improves the survival of brown bear (Ursus arctos) spermatozoa during chilling and after cryopreservation.
- 2013
-
Ingår i: Theriogenology. - : Elsevier. - 0093-691X .- 1879-3231. ; 79:3, s. 541-550
-
Tidskriftsartikel (refereegranskat)abstract
- The Cantabrian brown bear survives as a small remnant population in northern Spain and semen cryopreservation for future artificial insemination is one of the measures being implemented for conservation of this species. As part of this program we investigated the value of adding heat shock protein A8 (HSPA8) to media (N-[Tris(hydroxymethyl)methyl]-2-aminoethanesulfonic acid-TRIS-fructose with 20% egg yolk) used for chilling and cryopreserving the spermatozoa. Semen samples from eight brown bears were obtained by electroejaculation during the breeding season. In experiment 1, we tested three concentrations of HSPA8 (0.5, 1, and 5 μg/mL) to determine whether sperm motility (computer assisted sperm analysis system) and sperm survival could be improved during refrigeration (5 °C) up to 48 hours. Results showed that sperm viability (test with propidium iodide) was improved by the addition of 0.5 and 5 μg/mL HSPA8. In experiment 2, HSPA8 was added to the cryopreservation media (6% final glycerol concentration) before the freezing process. Though there were no differences in sperm viability immediately after thawing (analyses to 0 hours), plasma membrane permeability (test with YO-PRO-1) was significantly lower by the presence of HSPA8 (1 μg/mL) and acrosomal damage (test with peanut agglutinin-fluorescein isothiocyanate conjugate) was reduced by higher concentrations of HSPA8 (1 and 5 μg/mL) (analyses after thermal stress test incubating over 2 hours to 37 °C). In experiment 3, results of a simple progression test carried out through artificial mucus (hyaluronic acid 4 mg/mL) showed a significant decrease in the total number of sperm able to swim a distance of 0.5 to 2 cm through a capillary tube for all HSPA8-based extenders. Nevertheless, the distance traveled by the vanguard spermatozoa, which represent a highly motile subpopulation, was restored by the inclusion of 1 and 5 μg/mL HSPA8 in the cryopreservation media. Thus, the HSPA8 addition to extender improves the quality of brown bear (Ursus arctos) sperm during chilling (viability) and after cryopreservation (number of sperm with damaged acrosomes and "apoptotic-like" changes).
|
|
