Relationship between sperm ubiquitination and equine semen freezability

Abstract

This study aimed to assess the semen ubiquitin levels of stallions with good (GF) and poor semen freezability (PF) and to evaluate the relationship between sperm ubiquitination and sperm morphological defects. Five ejaculates from eight adult stallions (n = 40) were collected and cryopreserved. Then, the ubiquitin level in equine sperm cells was assessed by immunohistochemistry with epifluorescence microscopy, and sperm morphology was assessed by differential interference contrast microscopy. Sperm cells were classified according to the intensity (classification 1: from I to IV; I = very low ubiquitin intensity and IV = very high ubiquitin intensity) and location of ubiquitin staining (classification 2). Statistical analyses were performed using SAS software (version 9.4), and p ≤ .05 was considered significant. We observed that PF stallions showed higher percentages (p < .05) of sperm cells with high ubiquitination (11.82% of ubiquitin intensity grade I, 39.13% of ubiquitin intensity grade II, 27.25% of ubiquitin intensity grade III, and 20.67% of grade IV), while GF stallions showed higher percentages (p < .05) of sperm cells with lower staining intensity (28.52% grade I, 59.83% grade II, 7.92% grade III, and 7.02% grade IV). Furthermore, for PF stallions, 23 significant correlations were detected (p < .05) between sperm abnormalities and ubiquitin intensity in different sperm regions. Increased ubiquitination of the sperm head, midpiece, and tail was positively correlated with their respective morphological defects. We concluded that high sperm ubiquitin levels are observed in ejaculates from stallions with poor semen quality (poor freezability), and ubiquitin marking in specific cellular locations can identify sperm morphological defects.

Comprehensive Flow-Cytometric Quality Assessment of Ram Sperm Intended for Gene Banking Using Standard and Novel Fertility Biomarkers

“…According to this, specific fluorescent signal and specific protein were detected in ram testis samples using both microscopic and Western blot analyses, respectively, thus confirming the specificity of flow-cytometric analysis. Although no differences in the presence of ubiquitin on the sperm plasma membrane were found between analyzed groups of ram semen with different viability in this study, it may be important to monitor this marker because very recently, it has been reported that high ubiquitin levels in spermatozoa positively correlate with poor freezing ability of stallion semen [ 71 ]. On the other hand, significantly higher contents of AGG were observed in Group 2 compared to Group 1 ( Figure 7 ), which clearly correlates with the poor quality of semen samples in Group 2.…”

>b<Efeitos da criopreservação sobre as características da motilidade espermática (CASA) em equinos e bovinos>/b<

“…Dentre essas mudaças que podem acontecer ao espermatozoide durante a criopreservação, destacam-se o choque frio, estresse osmótico, alteração do volume celular, formação de cristais de gelo, danos oxidativos, onde estes são os principais fatores que ocasionam lesões na célula espermática decorrentes do frio, causando lesões na membrana plasmática, acrossonal, potencial mitocondrial, citoesqueleto, cromatina, podendo levar a perda da capacidade fertilizante do espermatozoide (AMAN & PICKET, 1987;PENÃ et al, 2011;KHALIL et al, 2018). Bem como, foram observadas altos níveis de ubiquitina em espermatozoides com baixa congelabilidade (LANÇONI et al, 2022).…”