The research activities of the laboratory are based on in vitro fertilization (IVF) and embryo culture as a model to study a wide variety of aspects related to reproduction in animals of zootechnical interest. Of great relevance is the study of male fertility through the evaluation of sperm physiological functional abilities during the phases of interaction with oocytes. Thanks to IVF model it is possible to study the relations between male and female gametes from the very first moments in which mutual identification and fusion takes place to generate the first embryonic cell, the zygote. The phases following fertilisation, that cover the embryonic development through the stages of morula and blastocyst, provide important information on the mechanisms leading to the creation of a new individual. These complex mechanisms, which are related to male and female qualities, contribute to the characterization of the complex parameter defined as “fertility level”.
SEMINOLOGY AND CYTOMETRY
The lab dedicated to the evaluation of sperm cells has been conducting researches on animal reproduction since 1994. The studies, based on reproductive technologies applied to selection, contributed over the years to growing insights which were fundamental for the development of the zootechnical compartment both in Lombardy and nationally. The species of interest are: cows, buffaloes, horses, pigs, sheep, goats, fish and bees. The research activities mainly focus, but are not limited to the following subjects:
· the standardisation of objective methods for the evaluation of spermatozoa in mammals and fish
· the development of innovative analytical techniques and tools for predicting the fertility of sires
· the use of sperm as a strategy in reproductive toxicology as an alternative to animal testing
· the evaluation of cell aging based on telomere length assessment
· sperm sexing after freezing/thawing for use in IVF aimed at the production of embryos of predefined sex
· the development of new technologies for sperm sexing
These last topics have involved important collaboration with the following nationally prestigious Institutes: The “Scuola Normale Superiore di Pisa”, the Italian Institute of Technology, the National Institute of Applied Optics of the National Research Council (CNR), the Institute of Microelectronic and Microsystems of the CNR, and Novaetech, the spin-off of the National Institute of Astrophysics. These collaborations contributed to the creation of patents.
In its complexity, the final goal of the lab is to implement efficient services through continuous and close interchange between service and research needs so as to guarantee the ever-growing information flow towards the users about the research innovations. Some of the results obtained over the years have directly influenced the breeding Italian industry and have contributed to the definition of the process of official control of frozen sperm in Italy (CUS), carried out by the Institute Spallanzani on behalf of MiPAAF. This is realised by qualifying the algorithms for the comparison of the sperm quality ascertained by ISILS and the quality certificated by the semen production centres.
The laboratory is equipped with modern instruments for objective semen analyses, i.e. CASA systems for computerized analysis of moving images, phase-contrast microscopes, NIC and DIC microscopes by Nomarsky for the evaluation of cell morphology and cytofluorimeters for the qualitative and quantitative evaluation of cell status.
The laboratory of Cryobiology cooperates synergistically with the laboratory of Seminology. The cryopreservation allows to keep the cells in a “latent” state at a very low temperature (-196°C). This technology is particularly relevant for the implementation of germplasm banks, which are a “reservoir” of genetic variability. This is strategic for genetic improvement of various species and for planning of crossbreeding in the context of conservation programs for endangered species. Sperm cryopreservation is accomplished either using the standard or alternative methods based on multi thermal-gradient. The laboratory utilizes programmable freezers and freezing curves which are adapted to the specific requirements of each species. In this respect, the lab of Cryobiology is equipped with innovative instruments for sperm analysis and is committed to implement new protocols also oriented to species which are not traditionally selected for reproduction based on fertility criteria and sperm propensity to freezing.
Over the years the research activities helped to perfect the protocols for freezing fish sperm, some of which contributed to the creation of patents.