Team:Montpellier/Demonstrate
Proof of concept : analyse of spermtozoa motility
Introduction
Antimicrobial peptides (AMPs) have a great affinity for spermatozoa membranes. They are composed of anionic phospholipids such as phosphatidylglycerol and phosphatidylserine in the plasma membrane. Therefore AMP have to ability to inhibit motility of spermatozoa and prevent the fertilization. Antisperm antibodies are antibodies that can fix on specific parts of spermatozoa (YLP12 antigen) and inhibit their motility. The exact mechanism for this motility inhibition is not known yet. We quantified the action of the AMPs and antibodies on the motility of spermatozoa. This is our proof of concept to test the anticonceptional efficiency of the molecules produced by our bacteria.
Use of mice sperm
The animals that we are using are males Mus musculus, 8 weeks old. They are used by the team of Thomas Robert, at the IGF of Montpellier, that is working on the genetic factors that control the integrity of the meiotic cell division. For our study, the epididymis was taken to collect the sperm of the animal. This tissue is not used by the team of Thomas Robert. The epididymis is squeezed to take the sperm out of it. Following extraction, the sperm is stored in BWW buffer for the experiments.
We are using the spermatozoa from the mice’s epididymis cauda because the maturation of spermatozoa is more advanced in this region of the epididymis and thus represent in the best way fully mature spermatozoa while still being easy to extract. For our experiments there were two cases. Experiments could be done right after the dissection and extraction of the sperm, else we did the experiment one or two days after the dissection. In the first case we put the sperm in Biggers-Whitten-Whittingham medium *insert BWW protocol link*. Otherwise we used a conservation method, which allows to conserve spermatozoa and their motility during 72 hours. *insert link to preservation protocol*
Measurements
We carried out different types of measurements, including their controls. Negative controls allow us to quantify the motility of spermatozoa alone, while positive controls measure the efficiency and the concentration needed to completely stop the motility of spermatozoa of a commercial molecule. (figure 1)
| Type of experiment | What we put in the device |
|---|---|
| Only spermatozoa | Negative control |
| Spermatozoa + commercial spermicides | Positive control |
| Spermatozoa + lysed bacteria | Control the efficiency of the molecules produce by the bacteria |
| Spermatozoa + Bacteria supernatant | Control the secretion |