Standardization of European eel (Anguilla anguilla) sperm motility evaluation by CASA software

[EN] The development of powerful computer-assisted sperm analysis software has made kinetic studies of spermatozoa possible. This system has been used and validated for several species, but some technical questions have emerged regarding fish sample evaluations (i.e., frame rate, sperm dilution, chamber model, time of analysis, magnification lens, etc.). In the present study, we have evaluated the effects of different procedural and biological settings with the aim to correctly measure sperm quality parameters of the European eel. The use of different chambers did not affect the sperm motility parameters. However, regarding lens magnification, 10x was the most accurate lens, showing the least variation in the acquired data. Similarly, the frame rate setting resulted in a dramatic effect in some sperm kinetic parameters, primarily in terms of curvilinear velocity; we therefore recommend using the camera's highest available frame rate setting. Finally, the reduction in sperm motility over postactivation times suggests that sperm analysis should be performed within the first 60 seconds after activation of the European eel sperm. In conclusion, some protocol variables of sperm analysis by computer-assisted sperm analysis software can affect the measurement of eel sperm quality parameters, and should be considered before directly comparing results obtained by different laboratories. Moreover, because marine fish species show relatively similar features of sperm kinetic parameters, these results could be considered in the evaluation of the motility of sperm from other fish species. (C) 2013 Elsevier Inc. All rights reserved.This study was funded from the European Community's 7th Framework Programme under the Theme 2 "Food, Agriculture and Fisheries, and Biotechnology," grant agreement 245257 (Pro-Eel), and Generalitat Valenciana (ACOMP/2011/229). D.S.P. and P.C.F.C. have postdoctoral grants from UPV (CE-01-10) and PAC-EMBRAPA, respectively. I.M. and V.G. have predoctoral grants from Generalitat Valenciana and Spanish MICINN, respectively. The authors thank the Proiser R&D, S.L. team, who performed the task of fractioning the original sequences for the experiment described in section 3.2.Gallego Albiach, V.; Carneiro, PCF.; Mazzeo, I.; Vilchez Olivencia, MC.; Peñaranda, D.; Soler, C.; Pérez Igualada, LM.... (2013). Standardization of European eel (Anguilla anguilla) sperm motility evaluation by CASA software. Theriogenology. 79(7):1034-1040. https://doi.org/10.1016/j.theriogenology.2013.01.019S1034104079

Study of the effects of thermal regime and alternative hormonal treatments on the reproductive performance of European eel males (Anguilla anguilla) during induced sexual maturation

[EN] Since 1960, the European eel (Anguilla anguilla) has suffered a dramatic reduction in natural stocks. Breeding in captivity is considered an alternative, but obtaining high quality sperm seems basic on this regard. The main objective of this study was to assess the effects of three thermal regimes (two of them variable: T10 and T15; and one of them constant: T20) and three hormonal treatments with different hormones (hCG, hCGrec and PSMG) on the induction of maturation in European eel males. In the case of the thermal regimes, our results demonstrated that the onset and progression of spermiation are strongly influenced, and perhaps closely regulated, by water temperature. T20 demonstrated the best results in all the sperm parameters (volume, density, motility, kinetic features, etc.) throughout most weeks of treatment, becoming a reliable and productive method for inducing spermiation in this species. In the case of hormonal treatments, the onset and progression of spermiation in European eel males were influenced by the hormone used. In this respect, hCGrec produced the best results in all the sperm parameters including volume, density, motility, kinetic features, etc., throughout most weeks of treatment, thus becoming an effective alternative treatment to the standard hCG treatment used to induce spermiation in eel species. Moreover, hCGrec gave rise to the best economical profitability, making it possible to obtain good quality sperm samples at a lower price than by using the other two hormonal treatments. © 2012 Elsevier B.V. All rights reserved Guardar / Salir Siguiente >Funded by the European Community's 7th Framework Programme under the Theme 2 "Food, Agriculture and Fisheries, and Biotechnology", grant agreement no. 245257 (PRO-EEL) and Generalitat Valenciana (ACOMP/2012/086). Victor Gallego, Ilaria Mazzeo and M. Carmen Vilchez have predoctoral grants from the Spanish Ministry of Science and Innovation (MICINN), Generalitat Valenciana, and UPV PAID Programme (2011-S2-02-6521), respectively. Paulo C.F. Carneiro had post-doc grants from PAC-EMBRAPA and UPV PAID Programme (PAID-02-11). David S. Penaranda was supported by a contract co-financed by MICINN and UPV (PTA2011-4948-I).Gallego Albiach, V.; Mazzeo, I.; Vilchez Olivencia, MC.; Peñaranda, D.; Carneiro, PCF.; Pérez Igualada, LM.; Asturiano Nemesio, JF. (2012). Study of the effects of thermal regime and alternative hormonal treatments on the reproductive performance of European eel males (Anguilla anguilla) during induced sexual maturation. Aquaculture. 354:7-16. https://doi.org/10.1016/j.aquaculture.2012.04.041S71635

Exploring correlations between sex steroids and fatty acids and their potential roles in the induced maturation of the male European eel

[EN] The present study was undertaken to evaluate the correlations between the fatty acids in the liver and testis and the plasma levels of the hormonal steroids used during eel spermatogenesis, in order to clarify the physiological roles fatty acids play in the spermatogenetic process. The stages of testis development (S1-S5) were assessed by histological observations in order to classify the different phases of hormonally-induced spermatogenesis and evaluate the possible relationships between the hormones and fatty acids in each stage. The highest plasma levels of 17 beta-estradiol (E2), testosterone (T) and 11-ketotestosterone (11KT) were found in S1, when spermatogonial proliferation occurs. A correlation was found between 17 alpha-20 beta-dihydroxy-4-pregnen-3- one (DHP) levels and some fatty acids during the proliferation and growing phases (S1-2), suggesting that DHP might modulate lipid metabolism in the liver during early spermatogenesis. The DHP levels increased significantly during the growing phase (S2) and remained at high levels throughout the subsequent development stages (S3-S5). Similar to results found in mammals, our results show that in the eel there are regulatory mechanisms, including eicosapentaenoic acid (20:5-n3, EPA) and docosahexaenoic acid (22:6-n3, DHA), which act as modulators in the synthesis of androgens, particularly during the final phase of sperm maturation. Our results suggest that the fact that EPA, ARA and DHA concentrations in the eel testis remain constant/stable during spermiation could be related to the subsequent union of the spermatozoa and the egg. The findings from this research provide new insights for further studies about the possible effect of steroids on desaturase activity and highlight the importance of the effect of lipid metabolism during male eel spermatogenesis. (C) 2014 Elsevier B.V. All rights reserved.Funded by the European Community's 7th Framework Programme under the Theme 2 "Food, Agriculture and Fisheries, and Biotechnology", grant agreement no. 245257 (PRO-EEL). D. S. P. had a contract co-financed by MICINN and UPV (PTA2011-4948-I) and received a Shortterm Scientific Mission grant from COST Office (Food and Agriculture COST Action FA1205: AQUAGAMETE) to carry out the steroids analyses in Norway.Baeza Ariño, R.; Peñaranda, D.; Vilchez Olivencia, MC.; Tveiten, H.; Pérez Igualada, LM.; Asturiano Nemesio, JF. (2015). Exploring correlations between sex steroids and fatty acids and their potential roles in the induced maturation of the male European eel. Aquaculture. 435:328-335. https://doi.org/10.1016/j.aquaculture.2014.10.016S32833543

Specific recombinant gonadotropins induce for the first time in European eel (Anguilla anguilla) full spermiating fish from immature males

Trabajo presentado en la 22nd International Conference of Zoology, celebrado en Okinawa (Japón), del 14 al 19 de noviembre de 2016In teleosts, recombinant gonadotropins (rGTHs), both in vitro and in vivo, have been able to induce the steroidogenesis and gonad development, however the in vivo results have been variable. In vivo, specific rGTH stimulated estradiol (E2) production in female of species such as orange-spotted grouper (Epinehelus coioides) or rosy bitterling (Rhodeus ocellatus), achieving ovulation in the latter, in that study however, already sexually matured females were used. In males, rGTHs were also able to induce the androgenesis process e.g. in zebrafish (Danio rerio) and European sea bass (Dicentrarchus labrax). Generally, the hormonal treatment was not able to induce the spermiation in vivo, with the exception of goldfish (Carassius auratus) and European sea bass, where the treated fish were already sexually matured at the beginning of the study. In Japanese eel (Anguilla japonica), recombinant FSH stimulated the in vitro production of E2 and testosterone in oocytes, but only at vitellogenic. In vivo, rGTHs did not stimulate an increase in the gonadosomatic index in females, but some induction of vitellogenesis was observed. In males, rGTHs expressed by baculovirus, yeasts or Drosophila cell lines were able to induce androgenesis in vitro. A complete spermatogenesis, with the presence of some spermatozoa in the testis, was achieved with rGTHs expressed by baculovirus. In vivo, Japanese eel and goldfish rGTHs, expressed by baculovirus, stimulated a complete spermatogenesis in male eels, but spermiation has never been achieved. Therefore, in the current work, the aim was to check the functionality in vivo of specific rGTHs in European eel. European eel males (n=72; 100.1±1.9 g) from a local fish farm were distributed in four 150-L aquaria and progressively adapted to sea water and 20 ºC. Single-chain recombinant FSH and LH were obtained by transfection of a mammalian cell line with further partial purification and up-concentration (Rara Avis Biotec S.L.; Valencia, Spain). In a first experiment, males were submitted to five hormonal treatments: one group was treated during 9 weeks with constant doses of recombinant European eel follicle-stimulating hormone (aarFSH; 4 µg/fish), a second group received constant doses of recombinant European eel luteinizing hormone (aarLH; 2 µg/fish) also during 9 weeks, while other three groups were injected with different combinations of constant or changing doses of both aarGTHs. All treatments stimulated the androgen synthesis, but this increase was more pronounced in those fish treated with a combination of both aarGTHs. Unlike aarLH or aarFSH alone were able to induce spermiating males, the fish treated with constant doses of aarFSH and increasing ones of aarLH registered the best results, with 20% of motile spermatozoa, stimulating spermiation for first time in a teleost from immature fish. In order to improve the sperm quality, a second experiment was performed. Immature males received three constant aarFSH doses (2.8, 1.4 or 0.7 µg/fish) and increasing doses of aarLH (every 3 weeks; 1, 2, 6 µg/fish). All treatments induced spermiating males, however the best sperm quality (¿50% motile spermatozoa) was observed in males treated with the highest aarFSH dose. In conclusion, these new specific recombinant gonadotropins have demonstrated their capacity to induce the spermatogenesis and spermiation in vivo in a teleost fish, the European eel.Peer reviewe

Specific recombinant gonadotropins induce spermatogenesis and spermiation by first time in a teleost fish, the European EEL

Trabajo presentado en el 10º Congreso de la Asociación Ibérica de Endocrinología Comparada (AIEC), celebrado en Castellón (España), del 23 al 25 de septiembre de 2015Eel recombinant gonadotropins have induced spermatogenesis in vitro, but had limited effect in vivo. New specific single-chain recombinant gonadotropins (Rara Avis Biotec S.L.) were tested treating European eel males with rFSH and rLH. Males received rFSH in three doses (2.8, 1.4 and 0.7 ¿g/fish; high, medium and low treatments) during 3 weeks. Later, an increasing rLH dose (every 3 weeks; 1, 2, 6 ¿g/fish) was combined with rFSH. Three males/treatment were sacrificed every 3 weeks. Blood and testis samples were taken for steroids and histological analyses and GSI was calculated. Sperm quality was determined by CASA software. rFSH by itself was able to induce the spermatogenesis (week 3), causing higher androgen levels and development until the spermatogonia 2 stage in the high rFSH group. The rLH (week 6) promoted further maturation to spermatocyte 2 stage in the medium and high groups, but no gonadal progression was observed in the low group. Higher T levels were reached in all the groups, and the biggest 11KT values in the rFSH high group. A second increase of rLH (week 9) induced further maturation to spermatid-spermatozoa 1 and spermatocyte 1 stages in the high and low groups, respectively. The medium group showed no further maturation (spermatocyte 1-2 stages). The highest dose of rLH (week 12) induced the most advanced stage (spermatozoa 2), and significant GSI increase in all the groups. A progressive 11KT (high group) and T (all treatments) decrease was observed from week 9 to 12. All treatments induced spermiating males, however, the best sperm quality with ?50% motile cells and volumes ~0.4 ml was observed in males treated with the highest rFSH dose and a progressive increase of rLH treatment. On the other hand, a 20% of nonresponders were observed and the sperm quality was variable. So, further experiments are required to improve these treatments.Funded by IMPRESS (Marie Sklodowska-Curie Actions; Grant agreement nº: 642893) and MINECO (REPRO-TEMP; AGL2013-41646-R).Peer reviewe

Neuroblastoma in Spain : Linking the national clinical database and epidemiological registries - A study by the Joint Action on Rare Cancers

Altres ajuts: Ministerio de Sanidad; Universitat de València; Sociedad Española de Oncología Pediátrica; Fundación de Oncología Infantil Enriqueta Villavecchia.Purpose: Linkage between clinical databases and population-based cancer registries may serve to evaluate European Reference Networks' (ERNs) activity, by monitoring the proportion of patients benefiting from these and their impact on survival at a population level. To test this, a study targeting neuroblastoma (Nb) was conducted in Spain by the European Joint Action on Rare Cancers. Material and methods: Subjects: Nb cases, incident 1999-2017, aged < 15 years. Linkage included: Spanish Neuroblastoma Clinical Database (NbCDB) (1217 cases); Spanish Registry of Childhood Tumours (RETI) (1514 cases); and 10 regional population-based registries (RPBCRs) which cover 33% of the childhood population (332 cases). Linkage was semiautomatic. We estimated completeness, incidence, contribution, deficit, and 5-year survival in the databases and specific subsets. Results: National completeness estimates for RETI and NbCDB were 91% and 72% respectively, using the Spanish RPBCRs on International Incidence of Childhood Cancer (https://iicc.iarc.fr/) as reference. RPBCRs' specific contribution was 1.6%. Linkage required manual crossover in 54% of the semiautomatic matches. Five-year survival was 74% (0-14 years) and 90% (0-18 months). Conclusions: All three databases were incomplete as regards Spain as a whole and should therefore be combined to achieve full childhood cancer registration. A unique personal patient identifier could facilitate such linkage. Most children have access to Nb clinical trials. Consolidated interconnections between the national registry and clinical registries (including ERNs and paediatric oncology clinical groups) should be established to evaluate outcomes

Effect of thermal regime on fatty acid dynamics in male European eels (Anguilla anguilla) during hormonally-induced spermatogenesis

[EN] Little is known about the role of fat and fatty acids in European eel spermatogenesis. The aim of this research was to study the changes in fat content and to carry out a quantitative analysis of the fatty acid composition of the muscle, liver and gonad of European male eels during hormonally induced sexual maturation. Three different thermal regimes were used (two variable: T10 and T15; and one constant: T20) to replicate the changes in temperature that these fish experience during their transoceanic reproductive migration. Spermatogenesis was reached earlier in treatment T20, suggesting that spermatogenesis in the European eel is closely regulated by water temperature. Although eels lose body mass due to the period of fasting that accompanies gonadal growth, no significant changes were found in the fat and fatty acid content of the muscle during the experimental period. With regard to the liver, the levels of palmitic, eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids increased significantly at the start of the spermiation process in all the thermal treatments. In the testis, levels of EPA, arachidonic acid (ARA) and DHA remained constant during the maturation process whereas the levels of the rest of the fatty acids decreased significantly. The stability of the ARA and EPA levels in the testis may have a physiological significance, whereas the stability of the DHA levels may have a structural significance. The results suggest that the progression of spermiation is influenced by water temperature and demonstrate the importance of EPA, ARA and DHA in European eel reproduction. This study demonstrates that complementary research focusing on the lipid composition of commercial diets could improve the sperm quality of this species.Funded by the European Community's 7th Framework Programme under the Theme 2 "Food, Agriculture and Fisheries, and Biotechnology", grant agreement no. 245257 (PRO-EEL). Victor Gallego, Ilaria Mazzeo and M. Carmen Vilchez had predoctoral grants from the Spanish Ministry of Science and Innovation (MICINN), Generalitat Valenciana, and UPV PAID Programme (2011-S2-02-6521), respectively. David S. Penaranda was supported by a contract co-financed by MICINN and UPV (PTA2011-4948-I). Rosa Baeza was supported by contract financed by PRO-EEL. The authors want to thank Vicente Javier Moya Salvador for his technical assistance with gas chromatography analyses.Baeza Ariño, R.; Mazzeo, I.; Vilchez Olivencia, MC.; Gallego Albiach, V.; Peñaranda, D.; Pérez Igualada, LM.; Asturiano Nemesio, JF. (2014). Effect of thermal regime on fatty acid dynamics in male European eels (Anguilla anguilla) during hormonally-induced spermatogenesis. Aquaculture. 430:86-97. https://doi.org/10.1016/j.aquaculture.2014.03.045869743

International eDelphi Study to Reach Consensus on the Methotrexate Dosing Regimen in Patients With Psoriasis.

A clear dosing regimen for methotrexate in psoriasis is lacking, and this might lead to a suboptimal treatment. Because methotrexate is affordable and globally available, a uniform dosing regimen could potentially optimize the treatment of patients with psoriasis worldwide. To reach international consensus among psoriasis experts on a uniform dosing regimen for treatment with methotrexate in adult and pediatric patients with psoriasis and identify potential future research topics. Between September 2020 and March 2021, a survey study with a modified eDelphi procedure that was developed and distributed by the Amsterdam University Medical Center and completed by 180 participants worldwide (55 [30.6%] resided in non-Western countries) was conducted in 3 rounds. The proposals on which no consensus was reached were discussed in a conference meeting (June 2021). Participants voted on 21 proposals with a 9-point scale (1-3 disagree, 4-6 neither agree nor disagree, 7-9 agree) and were recruited through the Skin Inflammation and Psoriasis International Network and European Academy of Dermatology and Venereology in June 2020. Apart from being a dermatologist/dermatology resident, there were no specific criteria for participation in the survey. The participants worked mainly at a university hospital (97 [53.9%]) and were experienced in treating patients with psoriasis with methotrexate (163 [91.6%] had more than 10 years of experience). In a survey with eDelphi procedure, we tried to reach consensus on 21 proposals. Consensus was defined as less than 15% voting disagree (1-3). For the consensus meeting, consensus was defined as less than 30% voting disagree. Of 251 participants, 180 (71.7%) completed all 3 survey rounds, and 58 participants (23.1%) joined the conference meeting. Consensus was achieved on 11 proposals in round 1, 3 proposals in round 2, and 2 proposals in round 3. In the consensus meeting, consensus was achieved on 4 proposals. More research is needed, especially for the proposals on folic acid and the dosing of methotrexate for treating subpopulations such as children and vulnerable patients. In this eDelphi consensus study, consensus was reached on 20 of 21 proposals involving methotrexate dosing in patients with psoriasis. This consensus may potentially be used to harmonize the treatment with methotrexate in patients with psoriasis