ABSTRACT
The mechanisms underlying male infertility are poorly understood. Most mammalian spermatozoa have two centrioles: the typical barrel-shaped proximal centriole (PC) and the atypical fan-like distal centriole (DC) connected to the axoneme (Ax). These structures are essential for fertility. However, the relationship between centriole quality and subfertility (reduced fertility) is not well established. Here, we tested the hypothesis that assessing sperm centriole quality can identify cattle subfertility. By comparing sperm from 25 fertile and 6 subfertile bulls, all with normal semen analyses, we found that unexplained subfertility and lower sire conception rates (pregnancy rate from artificial insemination in cattle) correlate with abnormal centriolar biomarker distribution. Fluorescence-based Ratiometric Analysis of Sperm Centrioles (FRAC) found only four fertile bulls (4/25, 16%) had positive FRAC tests (having one or more mean FRAC ratios outside of the distribution range in a group's high-quality sperm population), whereas all of the subfertile bulls (6/6, 100%) had positive FRAC tests (P = 0.00008). The most sensitive biomarker was acetylated tubulin, which had a novel labeling pattern between the DC and Ax. These data suggest that FRAC and acetylated tubulin labeling can identify bull subfertility that remains undetected by current methods and may provide insight into a novel mechanism of subfertility.
Subject(s)
Centrioles , Infertility, Male , Humans , Pregnancy , Female , Male , Cattle , Animals , Pilot Projects , Tubulin , Semen , Insemination, Artificial/veterinary , Infertility, Male/diagnosis , Infertility, Male/veterinary , Fertility , Spermatozoa , Biomarkers , MammalsABSTRACT
Centrosomes play an important role in the microtubule organization of a cell. The sperm's specialized centrosome consists of the canonical barrel-shaped proximal centriole, the funnel-shaped distal centriole, and the pericentriolar material known as striated columns (or segmented columns). Here, we examined the localization of the centriole proteins CEP135 and CP110 in cattle and human spermatozoa. In canonical centrioles, CP110 is a centriole tip protein that controls cilia formation, while CEP135 is a structural protein essential for constructing the centriole. In contrast, we found antibodies recognizing CEP135 and CP110 label near the proximal and distal centrioles at the expected location of the striated columns and capitulum in cattle and humans in an antibody and species-specific way. These findings provide a pathway to understanding the unique functions of spermatozoan centrosome.
ABSTRACT
Sperm cells are transcriptionally and translationally silent. Therefore, they may use one of the remaining mechanisms to respond to stimuli in their environment, the post-translational modification of their proteins. Here we examined three post-translational modifications, acetylation, glutamylation, and glycylation of the protein tubulin in human and cattle sperm. Tubulin is the monomer that makes up microtubules, and microtubules constitute the core component of both the sperm centrioles and the axoneme. We found that the sperm of both species were labeled by antibodies against acetylated tubulin and glutamylated tubulin.
ABSTRACT
A large proportion of infertility and miscarriage causes are unknown. One potential cause is a defective sperm centriole, a subcellular structure essential for sperm motility and embryonic development. Yet, the extent to which centriolar maladies contribute to male infertility is unknown due to the lack of a convenient way to assess centriole quality. We developed a robust, location-based, ratiometric assay to overcome this roadblock, the Fluorescence-based Ratiometric Assessment of Centrioles (FRAC). We performed a case series study with semen samples from 33 patients, separated using differential gradient centrifugation into higher-grade (pellet) and lower-grade (interface) sperm fractions. Using a reference population of higher-grade sperm from infertile men with morphologically standard sperm, we found that 79% of higher-grade sperm of infertile men with substandard sperm morphology have suboptimal centrioles (P = 0.0005). Moreover, tubulin labeling of the sperm distal centriole correlates negatively with age (P = 0.004, R = -0.66). These findings suggest that FRAC is a sensitive method and that patient age and sperm morphology are associated with centriole quality.
ABSTRACT
Infertility is a devastating experience for both partners as they try to conceive. Historically, when a couple could not conceive, the woman has carried the stigma of infertility; however, men and women are just as likely to contribute to the couple's infertility. With the development of assisted reproductive technology (ART), the treatment burden for male and unexplained infertility has fallen mainly on women. Equalizing this burden requires reviving research on male infertility to both improve treatment options and enable natural conception. Despite many scientific efforts, infertility in men due to sperm dysfunction is mainly diagnosed by a semen analysis. The semen analysis is limited as it only examines general sperm properties such as concentration, motility, and morphology. A diagnosis of male infertility rarely includes an assessment of internal sperm components such as DNA, which is well documented to have an impact on infertility, or other components such as RNA and centrioles, which are beginning to be adopted. Assessment of these components is not typically included in current diagnostic testing because available treatments are limited. Recent research has expanded our understanding of sperm biology and suggests that these components may also contribute to the failure to achieve pregnancy. Understanding the sperm's internal components, and how they contribute to male infertility, would provide avenues for new therapies that are based on treating men directly for male infertility, which may enable less invasive treatments and even natural conception.