Melatonin for the first in vitro production of Iberian deer embryos from dead animals

    IREC scientists produce Iberian deer embryos for the first time vitro from germ cells of dead females, being the only ones in the world that produce embryos of this deer subspecies.


    Reproduction is an essential phenomenon for the survival of species, which is why reproductive biotechnologies, better known as assisted reproductive techniques, they have a essential role in conserving biodiversity. In addition, in the case of animal species that constitute a direct resource for humans, such as livestock or hunting, reproductive biotechnologies offer improvement alternatives for their management, use and well-being.

    As the largest phytophagous of our fauna and the queen of big game hunting in Spain, the Iberian deer (Cervus elaphus hispanicus) It is a good example of a wild species whose conservation and management can benefit from reproductive biotechnologies, since, on the one hand, they provide the possibility of introducing improvements in the quality of the trophies; and on the other, they make it possible to ensure the genetic variability of the most intensively managed populations, avoiding the appearance of inbreeding problems.

     

    For wild species such as the Iberian deer (Cervus elaphus hispanicus), the obtaining, conservation and transfer of embryos on the one hand would allow a faster genetic improvement and the conservation of unique individuals for the maintenance of biodiversity.

    However, the development of assisted reproductive techniques, such as embryo production vitro, is a real challenge for scientists when it comes to wild species. Although this type of biotechnologies are usually highly developed in domestic species, it is difficult to extrapolate them to other less studied species such as deer. In addition, the number of samples necessary to be able to fine-tune these techniques is high and, generally, the animals are usually far from the laboratories, further complicating, if possible, the preservation of the samples until they are used.

    An alternative for the development of these reproductive biotechnologies is use samples from dead animals. Thus, for example, embryos can be produced vitro from oocytes –female germ cells whose maturation give rise to the ovules– extracted from the ovaries of dead females, which offers the possibility of obtaining a large amount of material ready to be used and fine-tuning the protocols.

    The problem is that, after the death of the animal, the ovaries stop receiving oxygen, and a consequence of the lack of oxygen is the production of free radicals and the generation of conditions of oxidative stress, a disorder that can affect oocytes causing their degradation. The consequences of the lack of oxygen can be very rapid, so avoiding this damage from the moment of sampling is essential for the viability of the oocytes.

    A new investigation led by doctors Irene Sánchez-Ajofrín and Ana Josefa Soler, from the Animal Reproduction Area of ​​the Research Group in Health and Biotechnology (SaBio) of the Instituto de Investigación en Recursos Cinegéticos (IREC – CSIC, UCLM, JCCM), has found the key to solve these limitations adding melatonin, a potent antioxidant, into the transport medium of the ovaries, preventing the damage that occurs from their collection to their processing in the laboratory.

     

    Oocyte status: viable oocyte (1A), oocyte undergoing the first phases of apoptosis or cell death (1B) and nonviable oocytes (1C and 1D); oocytes with different levels of oxidative stress (2A, high stress level; 2B, low stress level) and oocytes with different glutathione or GSH levels (2C, high GSH level; 2D, low GSH level).

    As shown by the results of this work, the maturation of oocytes from Iberian deer ovaries that were transported in a medium with melatonin and their subsequent fertilization resulted in a higher rate of embryo production vitro in blastocyst state –8 day embryos–.

    Everything indicates that melatonin has a positive effect on the regulation of oxidative stress in oocytes, inducing the synthesis of glutathione (GSH), the most important endogenous antioxidant of cells, thus facilitating its preservation in good condition during transport to the laboratory and subsequently stimulating the development of viable embryos vitro.

    This work represents the first time that Iberian deer embryos are obtained vitro from germ cells of dead females, making the laboratory of the Animal Reproduction Area of ​​the SaBio Group of the IREC the only one in the world that produces embryos of this subspecies. In addition, its results will imply the improvement of the protocols for the production vitro of embryos of other threatened deer species, facilitating their conservation.

    The scientific publication of this research is available at: