jueves, 21 de julio de 2016
The 6th AQUAGAMETE COST Action Training School (“Molecular basis of fish gamete quality: genomic tools”) was held in Rennes (France) from June 6th to 10th, 2016, on the Rennes 1 University campus of Beaulieu. It was organized by Dr. Julien Bobe, leader of the AQUAGAMETE Working Group 1. Techniques for evaluation of gametes quality, and Dr. Catherine Labbé. The trainers included 11 staff from the INRA Fish Physiology and Genomics lab, for both the practical and the conferences, and two trainers from Spain, Dr. Paz Herráez and Dr. Francesc Piferrer. The training course welcomed 18 trainees from 11 countries.
The Training School was focused on the Molecular basis of gamete quality and reproduction, with a strong orientation towards genomic tools. The 2h conferences spanned over sperm genetic damage and repair in fish, epigenetics and inheritance, microRNA and female fecundity, and insight into several genomic tools including the sequencing technologies, and transgenesis approaches with a special focus on CrispR/Cas9.
The practical sessions were focused on RNA extraction and quality checking, microarray analysis of fish samples in reproduction research, and sequence searching including tools presentation, primer design and phylogeny/syntheny search.
A visit of the INRA experimental fish facility allowed the trainees to discover the fully equipped behavior room, with the test chambers and the video recording system, and the rearing rooms of more than 7 freshwater species with specific photoperiod, water parameters, and temperature. The functioning of the water recycling system was explained, together with the anti-escapees system required for the transgenic fish of the facility (trout, medaka, goldfish, zebrafish). The dedication of the trainers and the motivation of the trainees ensured the success of this intense week of training.
martes, 19 de julio de 2016
La Dra. Luz Pérez realizó una estancia a finales de mayo en la Universidad de Utrecht (Campus de Uithof), en el laboratorio del Dr. Rüdiger Schulz (con ella en la foto), con quien ha estado aprendiendo a diferenciar algunos de los estados celulares de los testículos y el modo de cuantificación de estados celulares usado en este laboratorio.
El técnico Hank van der Kant le ha estado enseñando la técnica inmunohistoquímica de BrDU para ver proliferación celular, y ha podido dar un primer vistazo a los cultivos de testículo de zebrafish realizados por Diego Crespo.
La estancia fue financiada mediante una Short Term Scientific Mission de la Acción COST AQUAGAMETE.
lunes, 18 de julio de 2016
El viernes 15 nuestra compañera Marina Morini defendió su Tesis Doctoral titulada: “Molecular approaches related to the European eel (Anguilla anguilla) reproductive process”, que ha sido codirigida por Juan F. Asturiano y David S. Peñaranda, y financiada fundamentalmente por el Proyecto PRO-EEL (UE), el proyecto REPRO-TEMP (MINECO) y la Acción COST AQUAGAMETE.
El Dr. José Miguel Cerdá Reverter (del Instituto de Acuicultura de Torre de la Sal, CSIC) actuó como Presidente del Tribunal, el Dr. Ibon Cancio (de la Universidad del País Vasco/Euskal Herriko Unibertsitatea) fue el Secretario, y la Dra. Salima Aroua (de la Université du Havre, Francia) fue el tercer miembro del mismo.
Al acabar el acto de defensa, el Tribunal le concedió la máxima calificación, Sobresaliente Cum Laude. Enhorabuena Dra. Morini!
El documento final es un compendio de 4 artículos:
● Temperature modulates the testis steroidogenesis in European eel
Peñaranda et al., 2016. Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology
● Expression of nuclear and membrane estrogen receptors in the European eel throughout spermatogenesis
Morini et al., en revisión en Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology
● Nuclear and membrane progestin receptors in the European eel: characterization and expression in vivo through spermatogenesis
Morini et al., en revisión en PloS ONE
● Transcript levels of the soluble sperm factor protein phospholipase C zeta 1 (PLCζ1) increase through induced spermatogenesis in European eel
Morini et al., 2015. Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology
con una introducción y una discusión general. El pdf con el texto completo esta disponible on-line en: .
The European eel (Anguilla anguilla, L., 1758) population is in dramatic decline, so much so that this species has been listed as “Critically Endangered” on the Red List of Threatened Species, by the International Union for Conservation of Nature (IUCN). The European eel has a complex life cycle, with sexual maturation blocked in the absence of the reproductive oceanic migration, and an inability to mature in captivity without the administration of hormonal treatments. Even though experimental maturation induces gamete production of both sexes, the fertilization results in infertile eggs, unviable embryos and larvae, which die within a few days of hatching. Therefore, understanding the eel reproductive physiology during maturation is very important if we want to recover the wild eel population. Furthermore, due to its phylogenetic position, representative of a basal group of teleosts, the Elopomorphs, the Anguilla species may provide insights into ancestral regulatory physiology processes of reproduction in teleosts, the largest group of vertebrates.
In this thesis, characterization, phylogeny and synteny analyses have given us new insight into the evolutionary history of the reproductive process in vertebrates. The European eel possesses five membrane (mPRs) and two nuclear (nPR or pgrs) progestin receptors. Eel mPRs clustered in two major monophyletic groups. Phylogeny analysis of vertebrate nPRs and PLCζ1 (sperm specific protein) places both eel PLCζ1 and nPR sequences at the base of the teleost clade, which is consistent with the basal position of elopomorphs in the phylogeny of teleosts. To further resolve the origin of the duplicated eel nPRs, synteny analyses of the nPR neighboring genes in several vertebrate genomes were performed. Phylogeny and synteny analyses allowed us to propose the hypothesis that eel duplicated nPRs originated from the 3R.
In order to gain a better understanding of the role of the genes implicated in eel reproduction, analyses of their regulation during experimental maturation were carried out. The change in salinity induced parallel increases in E2 plasma and nuclear estrogen receptor expression levels, revealing a stimulatory effect of salinity on the E2 signalling pathway along the BPG axis, leading to a control of spermatogonial stem cell renewal. Brain and pituitary estrogen receptors may then mediate the stimulation of androgens and steroidogenic enzymes linked to androgen synthesis. Androgen synthesis is not dependent on temperature, but further maturation requires higher temperatures to induce a change in the steroidogenic pathway towards estrogen and progestin synthesis. This is consistent with our studies on estrogen and progestin receptors. In the testis, progestin seems to regulate meiosis through membrane and nuclear progestin receptors, and final sperm maturation seems to be controlled by both estrogen and progestin through the estrogen and progestin membrane receptors. Finally, eel sperm-specific PLCζ1 seems to have an important function in spermatozoa by inducing egg activation and temperature may play a role in its regulation, especially during the process of spermiogenesis.
This thesis attempts to evaluate the physiological function of the genes involved in eel reproduction during spermatogenesis, and demonstrates that salinity and temperature play crucial roles in the sexual maturation of the male European eel.
domingo, 17 de julio de 2016
Amin Golpour Dehsari, etudiante de doctorado de la Faculty of Fisheries and Protection of Waters de la University of South Bohemia (Vodnany, República Checa) ha realizado una estancia con nuestro grupo de investigación desde principios de uno a principios de julio, financiada con una Short Term Scientific Mission de la Acción COST AQUAGAMETE.
Durante estas semanas Amin ha aplicado el protocolo de aislamiento de espermatogonias en el testículo de otras especies de peces, ajustándolo hasta optimizarlo para la anguila europea.
Esto supone el primer paso para futuros estudios sobre la fisiología de estas células y su utilización en el control de la reproducción de esta especie.
Ha sido estupendo conseguir estos resultados en tan poco tiempo. Gracias Amin!.
martes, 5 de julio de 2016
In the 9th call, the last one in the AQUAGAMETE COST Action, 7 Short Term Scientific Missions were granted: 3 persons from Hungary travelling to the Czech Republic, Slovenia and Spain; 1 participant from Denmark visiting France; 1 person from Portugal travelling to Norway; 1 person from Greece visiting to South Africa and 1 person from Israel going to Norway.
Gook luck to everyone!
A comparison of techniques for studying oogenesis in the European eel Anguilla anguilla
Ilaria Mazzeo, Elisabetta Giorgini, Giorgia Gioacchini, Francesca Maradonna, M. Carmen Vílchez, Sylvie Baloche, Sylvie Dufour, Luz Pérez, Oliana Carnevali, Juan F. Asturiano
A multi-technique approach was used to study the changes occurring in European eel Anguilla anguilla ovaries during hormonally-induced vitellogenesis. Aside from classic techniques used to monitor the vitellogenic process, such as ovary histology, fat content analysis, sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE), and vitellogenin enzyme linked immunosorbent assay (ELISA), a new technique, Fourier Transform Infrared (FT-IR) Microspectroscopy, was used for the first time to analyze A. anguilla ovaries. The results from the different techniques provided different ways of approaching the same process. Although it is considered a time consuming approach, of all the employed techniques, histology provided the most direct evidences about vitellogenesis. SDS-PAGE and ELISA were also useful for studying vitellogenesis, whereas fat analysis cannot be used for this purpose. The FT-IR analysis provided a representative IR spectrum for each ovarian stage (PV, EV, MV and LV), demonstrating that it is a valid method able to illustrate the distribution of the oocytes within the ovary slices. The obtained chemical maps confirmed changes in lipid concentrations, and revealed their distribution within the oocytes at different maturational stages. When the results and the accuracy of the FT-IR analysis were compared to those of the traditional techniques commonly used to establish the vitellogenic stage, it became evident that FT-IR is a useful and reliable tool, with many advantages, including the fact that it requires little biological material, the costs involved are low, analysis times are short, and, last but not least, the fact that it offers the possibility of simultaneously analyzing various biocomponents of the same oocyte.