CHARATERIZATION OF OVIDUCTAL PROTEINS THAT PARTICIPATE IN THE SELECTION OF SPERM

VANZETTI AGUSTIN; MELINA FAGGI; JUAN MANUEL TEIJEIRO

Congreso; XXV Jornadas de la Sociedad Argentina de Biología; 2023

In previous works, weidentified to DMBT1 and ANXA2 as oviductal proteins that interact with sperm. IsolatedDMBT1 reduce viability and motility in vitro and thus, involved innegative sperm selection, while ANXA2 may be involved in oviductal reservoirformation. The objective of this work was to evaluate the transcriptional andtranslational expression of DMBT1 and ANXA2, the presence of isoforms andsubcellular locations of both proteins in oviductal epithelium through theporcine estrous cycle. Also, to analyze the effect of no isolated DMBT1 presentin different subcellular fractions on sperm physiology. Oviducts wereclassified according its estrous cycle stage and processed to obtain oviductalfluid (OF), oviductosomes (OVS) and plasma membrane of epithelial cells (MEC). Transcriptionaland translational expression were assayed by RT-qPCR and Western blot, respectively.Sperm were incubated in capacitating medium supplemented with OVS (mOVS), OF (mOF)or MEC (mMEC) corresponding to each estrous stage. The specificity of theeffect of DMBT1 present in each fraction was evaluated by blocking itsinteraction with sperm using anti-DMBT1 antibody and pre-immune serum ascontrol. The results show that expression DMBT1 and ANXA2 are regulated at transcriptionallevel through the estrous cycle (p < 0.05) but only DMBT1 is regulated attranslational level (p < 0.05). Both proteins were detected in OVS, OF andMEC and the presence of alternative transcripts and putative protein isoforms forANXA2 and DMBT1 were also demonstrated. Greater viability was found in mOF ofluteal stage and in mFO and mOVS of follicular stage all supplemented with theanti-DMBT1 antibody (p > 0.05), indicating an effective blocking effect ofthe antibody in these fractions. Also, greater motility in mOF of luteal stagesupplemented with antibody was found. These results are in line with previous hypothesisof the role of DMBT1 in negative selection by reducing viability and motilityof sperm