Sperm morphometry and chromatin condensation in Nelore bulls of different ages and their effects on IVF

Abstract

The aim of this study was to evaluate the chromatin packing and sperm head morphometry of cryopreserved semen of Nelore bulls (Bos taurus indicus) of different ages. Furthermore, the influence of the degree of chromatin compaction on in vitro embryo production (IVP) was investigated. Forty bulls were divided into three groups: young (1.8-2 years), adult (3.5-7 years), and senile (8-14.3 years). The ejaculates were frozen according to standards established by the Artificial Insemination Center located in the Southeast of Brazil. Toluidine blue staining was used for simultaneous evaluation of the sperm chromatin and sperm head morphometry. Chromomycin A3 (CMA3) was applied to analyze sperm protamination and IVP for embryonic development. Spermatozoa of young bulls presented higher values for area (A, pixels), perimeter (P, pixels), and width (W, pixels) compared to adults and senile (young: A = 1848.5 ± 119.79, P = 10.23 ± 0.29, and W = 1.95 ± 0.1; adults: A = 1672.9 ± 104.46, P = 9.86 ± 0.33, and W = 1.81 ± 0.06; senile: A = 1723.1 ± 124.41, P = 9.97 ± 0.33, and W = 1.83 ± 0.09; P < 0.0001) and showed higher protamination deficiency when analyzed by CMA3 (young: 1.57 ± 0.76; adults: 1.09 ± 0.63, and senile: 0.90 ± 0.59; P < 0.05). Likewise, variables of sperm head size (A, P, and W) and protamination assessed by CMA3 showed negative correlation with age and positive correlation with ellipticity, evaluated by toluidine blue method (P < 0.05). Sperm head area was larger in spermatozoa presenting chromatin instabilities than spermatozoa without chromatin alteration (P < 0.0001). There was no difference in IVP when using semen with larger or smaller portions of spermatozoa with chromatin instabilities, indicating that the proportion of sperm with abnormal chromatin compaction (4%-16.15%) did not interfere with early embryonic development. From our results, it can be concluded that sperm of young Nelore bulls have larger heads compared to adults and senile due to reduced protamine content when evaluated by CMA3 and higher proportion of major sperm defects assessed by differential interference contrast microscopy.

Does sperm quality and DNA integrity differ in cryopreserved semen samples from young, adult, and aged Nellore bulls?

“…The rationale behind these investigations was that if abnormal head morphology was a characteristic of young bull spermatozoa this might be due to nuclear alterations as it is supported by several studies [31–34]. To our knowledge there is no other study that has focused on the bull sperm nuclear integrity upon aging.…”

“…These points are rather important in the bovine industry for the efficiency and the cost effectiveness of insemination programs. Regarding bovine sperm nuclear condensation, Kipper et al (2017) [34] noted that spermatozoa from young bulls were morphometrically larger compared to those from older bulls and that alterations in chromatin condensation were related to heads with greater diameters. In another study concerning a different model of agricultural interest, Martí et al (2011) [35] morphometrically evaluated the heads of sperm cells from young goats shortly after reaching sexual maturity and observed heads with larger sizes compared to those of the adult animals, indicating a possible transition period between puberty and full sperm head size maturity.…”

Morphological and morphometric characterization of domestic cat epididymal sperm

“…The description of ellipticity, elongation, rugosity and regularity parameters is fundamental for distinguishing normal‐format gametes from amorphous, round and/or very sharp shaped heads (Bergstein et al, ). The association of these data with area and perimeter values can be used to optimize seminal analysis in automated systems, as well as they subsidize the creation of clustering software that provides information about genetic characteristics, such as chromatin stability (Kipper et al, ); differentiation of sperm X and Y (Santolaria et al, ); fertility potential (Ramón, Soler, et al, ); and resistance to cryogenic process (Núñez‐Martínez, Moran, & Peña, ; Ramón, Pérez‐Guzmán, et al, ).…”

“…The description of ellipticity, elongation, rugosity and regularity parameters is fundamental for distinguishing normal-format gametes from amorphous, round and/or very sharp shaped heads (Bergstein et al, 2014). The association of these data with area and perimeter values can be used to optimize seminal analysis in automated systems, as well as they subsidize the creation of clustering software that provides information about genetic characteristics, such as chromatin stability (Kipper et al, 2017); differentiation of sperm X and Y (Santolaria et al, 2016); fertility potential (Ramón, Soler, et al, 2013); and resistance to cryogenic process ( Thus, the sperm morphologically normal of the cat can be described as an ovoid-shaped cell, being relatively longer than wide, since the ellipticity was >1 and the elongation was 21.3 ± 0.12. The rugosity value (0.14 ± 0.00) categorized a greater heterogenicity in the size of the feline sperm, since it was less than 1 and closer to 0, different from that observed in the bovine species (0.74 ± 0.001) (Santolaria et al, 2016) and caprine (0.79 ± 0.01) (Aragão et al, 2013).…”

Protein biomarkers for male artificial insemination subfertility in bovine spermatozoa

“…Thus, the detection of spermatozoa with a lack of protamines is indicative of bull AI subfertility. The research regarding Nelore cattle showed that the spermatozoa from young bulls had larger head diameters due to the lower protamination level and resultant deficiency of chromatin condensation, compared with the spermatozoa from adult bulls 72. This might account for the cause of unstable results of AI when using cryopreserved spermatozoa from young bulls.…”