Colombo/Luvoni

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Martina Colombo graduated cum laude in Veterinary Biotechnology Sciences at the University of Milan in October 2017. In 2016, she got a scholarship to carry out her master’s thesis experiments at the Universidade Estadual Paulista – Jaboticabal, Brasil – where she spent 5 months under the supervision of Professor Maricy Apparicio. She acquired scientific skills concerning in vitro cell culture in two and three-dimensional systems, in vitro embryo production and gametes cryopreservation. Professor Gaia Cecilia Luvoni supervised her master’s thesis entitled “Embryo development in 3D culture of low competence vitrified feline oocytes”.

 

 

 

 

 

 

 

 

 

Gaia Cecilia Luvoni graduated in 1986 and got a PhD in reproduction in 1992 at the Faculty of Veterinary Medicine, University of Milan, Italy. She conducted research on in vitro embryo production in the U.S. at the Louisiana State University in 1990, and at the University of Georgia in 1994. In 2002 she became a Diplomate of the European College of Animal Reproduction (ECAR) and currently she is Full Professor in Obstetrics, Gynecology and Reproductive Biotechnologies at the University of Milan and responsible of the Canine Semen Bank Vetogene (http://www.vetogene.com). She is Past President of the EVSSAR (European Society for Small Animal Reproduction) and ECAR Board member. Her main scientific interests are Small Animal Theriogenology and Biotechnologies for Assisted Reproduction, which include in vitro embryo production, cryopreservation of male, female gametes and ovarian tissue, and ultrasonographic foetal biometry in dogs and cats. The results have been published in more than 190 paper and proceedings.

 

Title:  Microenvironment factors promoting carnivore oocyte quality

An ideal culture system for oocytes has not been found yet, as they often fail to develop into embryos or to do so at satisfactory rates in vitro.

The follicle, with its physical and chemical features, promotes the oocyte developmental competence in vivo, and to improve oocyte outcomes in vitro, researchers are directing their efforts towards the re-creation of a microenvironment able to mimic the physiological ovarian conditions.

The physical environment where the oocyte grows and develops has a three-dimensional and dynamic architecture, with the presence of follicular cells around the gamete, whereas the chemical factors in the follicular fluid include hormones and regulatory molecules, also secreted by surrounding cells.

Therefore, in this project, focused on the study of factors affecting oocyte developmental competence in vitro, several physical or chemical modifications of the extra and intracellular environments will be experimented to ameliorate the in vitro milieu, making it more similar to the in vivo conditions.

Two-dimensional systems, traditionally used in cell culture, will be compared with three-dimensional and microfluidic microenvironments. The addition of granulosa cells to these systems could help to further mimic the ovarian physical organization.

In a following phase of the project, paracrine factors secreted by granulosa cells or oocytes themselves will be investigated, and their synthetic forms will be supplemented to the culture medium to avoid the use of companion cells, characterized by an intrinsic variability.

In addition, manipulation of the intracellular environment will be investigated through the microinjection into the oocytes of compounds able, for instance, to stimulate the signaling cascade that regulates meiosis resumption and further development of female gametes, as cyclin molecules.

The domestic cat and the dog will be used as models, since the availability of gonads from routine ovariectomy and orchiectomy allows to obtain oocytes and spermatozoa to design and test protocols that could later be applied in other species, including humans and many endangered wild animals, improving their reproductive ability and contributing to fertility and biodiversity preservation.

In conclusion, in this project, the evaluation of in vitro outcomes of carnivore oocytes in various physicochemical conditions will be performed. The data collected will be useful to design a species-specific culture system that should be able to allow not only the development of high quality oocytes, but also of low competence gametes, such as the cryopreserved ones.

 

Figure 1. Feline oocytes cultured in 3D microcapsules

 

Figure 2. In vitro-derived feline embryos