P-098 Cryopreservation process deregulates the expression of pathways important for fertility in human spermatozoa

Abstract Study question Does cryopreservation impact on transcriptome of conventional frozen human spermatozoa? Summary answer The sperm alters pathways important for fertility, although the upregulation some genes can compensate harmful effects freezing. What is known already Sperm a widely used procedure storing gametes later use, preserving fertility in patients before gonadotoxic treatments or surgery, and donation programs. technique cause damage to through induction DNA alterations, fragmentation oxidation, altered motility, mitochondrial activity morphology. Transcriptomic alterations have been reported cryopreserved from various animal species, but no experimental data are available effect transcriptome-wide profile sperm. design, size, duration Semen samples were obtained 20 normospermic men April May 2022. Each sample was divided two aliquots. From one aliquot total RNA immediately extracted. second slowly after week storage liquid nitrogen A 13 paired passed quality control analyzed randomization into 4 pools, each 6 patients. Participants/materials, setting, methods who participated this study had median age 35.0 years (range: 29.0-46.0). extracted by miRNeasy Micro Kit (Qiagen), High Sensitivity 2200 Tape Station system (Agilent), amplified Ovation Pico WTA SystemV2 (Nugen), labeled hybridized GE 4x44K v2 microarrays (Agilent). Significance Analysis Microarray (SAM) performed using limma hgug4112a.db samr packages R/BioConductor. Main results role chance expression profiles significantly differed those fresh 219 down-regulated 28 up-regulated unique transcripts. gene ontology analysis disclosed that downregulates involved motility mitochondria function (CABS1, SPATA19) correct organization midpiece flagellum cytoskeleton (ACTL7A, AKAP4, C9ORF24, CAPZB, CCIN, IQCG, ODF2, SPATA6, SPATA6L, TBC1D21), fertilization PLCZ1, PRSS37, TUBGCP3), early embryo development (AGT, CLMN, DCC, DHRS3, EGFLAM, FAM20A, FREM1, GPI, HEMGN, KRTDAP, IFT172, MICAL2, MLF1, NF2, PGRMC2, PHACTR1, POPDC3, RO60, ROBO1, RORA, SGCA, SPRR2D, TCF4, TNC, TNNI3), oxidant detoxification (TXNDC29) repair (BRIP1, ERCC6, TRIP12), calcium ion binding homeostasis (AMY1C, ANO1, ANO2, ASGR1, CABS1, CPNE9, KCNIP2, ITPR3, PKD2L1, SELENOK, SYN3, TNNI3). Upregulated enriched related negative regulation response (ALOX15B, CD74, RPL10, SNAI1, THBS1), cellular FOSB, HLA-DQB1, S100A4, transcription (CD74, HLA-DRB1, MAMLD1, RASD1, TCF15, TLE1), protein stabilization (CD74), ubiquitination (BAG1, DCAF6, HERPUD2, KLHL7, MARCHF8, PSMA6, RNF133, SERGEF, TRIP12, UBE2DNL, UBL3, UBQLN3, ZNRF3) ubiquitin ligase (GPI, HSPA1L, PRKAR2A, STX8). Limitations, reasons caution Data validation larger cohort quantitative PCR would be useful, we applied stringent criteria selection (FDR = 0). Further research should undertaken experimentally test whether length cryostorage any Wider implications findings Specific deregulated cryopreservation, accordance with morphology, molecular integrity literature. Functional enrichment analyses also revealed changes cryoinjury. This finding noteworthy safety issue banking i.e., preservation, gamete donation. Trial registration number not applicable