Why is follicular fluid essential for the proper functioning of sperm during fertilization?

The fertilization process can be ruined if enzymes from the fluids that accompany the oocytes and sperm cause DNA degradation. A team from the Autonomous University of Madrid (UAM) has found evidence that the follicular fluid, the liquid that bathes the oocytes, works to prevent this calamity.

In our species, fertilization is the result of the fusion of two specialized cells, called gametes: the oocyte provided by the woman and the sperm generated by the man.

Both cell types are accompanied by two liquid substances: follicular fluid (FF), which accompanies the oocytes; and the seminal plasma (PS), in which the spermatozoa are immersed. A few years ago it was shown that FF is essential for the proper functioning of sperm during fertilization. In fact, some assisted reproduction clinics use it as a good preservative of sperm quality in the samples used in pregnancy tests.

What does the follicular fluid have to perform such an important function?

A recent work led by the Department of Biology of the Autonomous University of Madrid (UAM) —and which had the collaboration of Halotech DNA (a biotechnology company located in the Madrid Science Park), the Ginemed assisted reproduction clinic (Seville) and a researcher from the University of Queensland (Australia)—has delved into this question.

The results, published in Reproductive BioMedicine Online , represent a step forward in understanding how fertilization works in molecular terms and, therefore, in improving sperm handling systems in assisted reproduction clinics.

^{Sample of spermatozoa not protected by the follicular fluid. In this case, the size of the DNA halos is much smaller, as a consequence of the destruction generated by the activity of the DNases of the seminal plasma / UAM itself.}

Experiments with purified genomic DNA

The team, led by UAM researcher Jaime Gosálvez, had already verified that FF has a protective effect on the integrity of the sperm genome. But the quality of the genetic content of a sperm can be compromised, among other things, by the activity of enzymes that are capable of destroying DNA. These enzymes, the DNases, are present in both the FF (inactive) and the PS (active).

“On this occasion we have carried out a series of experiments with purified genomic DNA where it has been mixed with FF, with PS, or also with FF together with PS to determine whether or not the DNA was degraded by the activity of the DNases present in both media. ”, describe the authors.

“In the first place —they detail— we have confirmed that FF does not degrade DNA (a reflection of the fact that its DNases are not active). In contrast, PS DNases do destroy DNA. And, what is new, the DNA remains intact in samples incubated with the two media (FF+PS). In fact, the degree of protection is linked to the relative concentration of the FF. The obvious conclusion is that there is 'something' in the FF that is capable of inactivating the DNases of the PS”.

a chelating agent

Probably, according to the results, that 'something' is a chelating agent; a chemical compound that sequesters ions and alters the functionality of other molecules. “So if we add EDTA (a well-known chelating agent) to PS, the DNA is not digested. That is, EDTA reproduces the result obtained with the mixture of FF and PS”.

Finally, the researchers also verified that if increasing amounts of calcium or magnesium are added, the effect of this chelating agent is reversed. In such cases, the DNA is digested in the samples with PS. So the DNases present in PS need calcium or magnesium to achieve DNA destruction, and FF provides a chelator to nullify that activity.

In short, during the fertilization process, and when FF and PS intermingle, the protective role of FF is essential. The consequence is to prevent the genomes that provide oocytes and sperm from suffering attacks that would surely compromise reproductive success.

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Bibliographic reference:

Bartolomé, J., Claver Romeo, S., Dorado-Silva, M., García de la Vega, C., López, C., Sánchez-Martín, P., Johnston, S., Gosálvez, J. 2021. DNase activity in human seminal plasma and follicular fluid and its inhibition by follicular fluid chelating agents . Reproductive BioMedicine Online 43(6): 1079-1086. doi: 10.1016/j.rbmo.2021.09.015

More scientific culture in UAM Gazette .

Top image: Sample of spermatozoa protected by the follicular fluid. The nuclei, where the genome is found, are intact and display large halos of DNA (colored in fuchsia) / UAM