Viscous Cervical Environment-on-a-Chip for Selecting High-Quality Sperm from Human Semen

When ejaculated sperm travels through the vagina to the uterus, mucus secreted by the cervical canal generally filters out sperm having low motility and poor morphology. To investigate this selection principle in vivo, we developed a microfluidic sperm-sorting chip with a viscous medium (polyvinylpyrrolidone: PVP) to imitate the biophysical environment mimic system of the human cervical canal. The material property of the PVP solution was tuned to the range of viscosities of cervical mucus using micro-viscometry. The selection of high-quality human sperm was experimentally evaluated in vitro and theoretically analyzed by the convection-diffusion mechanism. The convection flow is shown to be dominant at low viscosity of the medium used in the sperm-sorting chip when seeded with raw semen; hence, the raw semen containing sperm and debris convectively flow together with suppressed relative dispersions. Also, it was observed that the sperm selected via the chip not only had high motilities but also normal morphologies and high DNA integrity. Therefore, the biomimetic sperm-sorting chip with PVP medium is expected to improve male fertility by enabling the selection of high-quality sperm as well as uncovering pathways and regulatory mechanisms involved in sperm transport through the female reproductive tract for egg fertilization.

P–069 microfluidic sperm sorting vs density gradient to yield sperm with reduced DFI for patients undergoing IVF-ICSI

Study question To compare the effect of sperm preparation methods on the DFI of semen sample for patients undergoing ICSI. Summary answer On comparing the results, microfluidic sperm sorting yielded sperms with significantly less DFI as compared to density gradient method of sperm preparation. What is known already The DNA integrity of the sperm plays an important role to ensure formation of good quality embryos with increased potential of fertilization, growth and ultimately implantation.. Centrifugation has shown to add stress to the sperm and leading to DNA damage, therefore there is a need to develop techniques of sperm preparation which help in retrieving as many sperms with intact DNA from the unprocessed sample as possible. Microfludic is fluid dynamic based technique of sperm preparation. in this study, we evaluated if microfluidic sperm sorter can recover motile sperm with better DNA integrity compared to density gradient preparation method. Study design, size, duration Prospective randomized study conducted in 80 patients undergoing IVF-ICSI with normal semen parameters (based WHO criteria 2010). DFI was done using Sperm Chromatin Dispersion (SCD) test in split semen samples prepared by microfluidic sperm sorter and density gradient method. Sperm morphology and motility were also recorded and evaluated based on the WHO 2010 criteria. Participants/materials, setting, methods Semen parameters of the sample were assessed by microscopic examination. DFI of each unprocessed sample was carried out using SCD test, following that the sample was split and sperm preparation was done using microfluidic sperm sorter and density gradient. the recovered sperm were tested for DFI and the results were compared. Main results and the role of chance Mean DFI in unprocessed semen samples was 23%. the analysis of split semen samples post preparation showed that the DFI was significantly reduced with the use of microfluidic sperm sorter (mean DFI 0.6%) as compared to density gradient (mean DFI 9%). Limitations, reasons for caution A major limitation of the microfluidic sperm sorter is the use sperm concentration and motility of the semen sample. In oligospermic and asthenospermic samples, density gradient is the preferred method of preparation. Lack of data showing improvement in clinical outcomes with reduced DFI is also a major limitation. Wider implications of the findings: Microfluidics has shown to significantly reduce the DFI of the semen sample, it requires no extra equipment and cost and is relatively easy to pick up. Density gradient method of sperm preparation continues to be the preferred method due to its versatility and recovery of good quality sperm. Trial registration number Not applicable

P–235 Are morphokinetics parameters and profitability of an assisted reproduction cycle related to sperm selection technique? Microfluidic sperm sorter chip versus magnetic activated cell sorting

Study question Does microfluidic sperm sorter offer any biological improvement over magnetic activated cell sorting (MACS)? Summary answer Microfluidic approach for selecting high profile spermatozoa is as good as magnetic activated cell sorting in terms of morphokinetic, fertilization and good quality blastocyst rate. What is known already Microfluidic sperm sorter chip is a method for select non-fragmented DNA sperm. We know that the use of these non-fragmented DNA sperms can improve the clinical results and the useful blastocyst rate. As several studies have shown previously, embryos morphokinetics parameters are affected by culture medium, ovarian stimulation, oxygen tension, origin of the oocytes, or the age of the patient, this is why we wanted to compare if the cleavage times using time lapse technology, are different depending on the sperm selection method used to select sperm with the non-fragmented DNA. Study design, size, duration Prospective and observational study performed between May 2019 to January 2021 in IVI Madrid and IVI Sevilla. Seminal samples from couples participating in the study were divided into two aliquots; each of them was processed according to one of the study methods. 53 couples were included in the study. Half of the oocyte from each donor were microinjected with sperm selected through MACS (n = 281) and the other half through a microfluidic device (n = 275). Participants/materials, setting, methods These oocytes were microinjected with both types of sperm samples and incubated in EmbryoScope. Cellular events studied in this study included cellular divisions until blastocyst stage, appearance and fading of some cellular structures and the duration of the first, second and third cellular cycle (cc1, cc2 and cc3) as well as their synchrony (S1, S2, S3). Data were exported from the EmbryoViewer data base. We perform an ANOVA statistical analysis to analyze the data. Main results and the role of chance No significant differences between both sperm selection methods were found regarding the time of cell division (from T2 to Tblstocyst), the cellular cycles duration (cc1, cc2 and cc3) or the synchrony of the cellular cycles divisions (s1, s2 and s3). However, a clear trend towards statistical significance has been found in both duration of cc2 (p = 0.052) being longer in MACS embryos than in microfluidic sperm sorting embryos, and in the expansion of the blastocyst, which occurs earlier in embryos that come from MACS than in those that come from microfluidic sperm sorting (p = 0.097). These two events could indicate a better embryo cleavage dynamic in the case of MACS embryos, with a better blastocyst expandability and the necessary time to carry out all the biological events that must occur in the cc2. However, significant difference was found in the direct cleavage from 1 to 3 cells embryo stage, which is one of the adverse events that more affects embryo implantation, being higher in microfluidic sperm sorting group (p = 0,037). Finally, the fertilization rate (73.1% vs 76.9%) and the good quality blastocyst rate (53.7% vs 56.5%) were higher in MACS embryos than in microfluidic sperm sorting embryos, although no significant differences were found. Limitations, reasons for caution This study has been performed in donated oocytes, so these results may not be extrapolated to other groups of assisted reproduction patients. However, more data are needed to draw firm conclusions. Furthermore, it´s crucial to increase the sample size to check if the trends founded reach statistical significance. Wider implications of the findings: Microfluidic sorting of unprocessed semen in un unselected population is as efficacious as magnetic activated cell sorting according embryo morphokinetic, fertilization rate and useful blastocyst rate. Microfluidic sperm sorting does not show clinical advantage over MACS considering this data collection. Trial registration number NCT04061484