Heterogeneity of Sperm Nuclear Chromatin Structure and its Relationship to Bull Fertility1

Abstract

The relationship between sperm nuclear chromatin structure and fertility was evaluated in two groups of Holstein bulls: Group 1, 49 mature bulls, and Group 2, 18 young bulls. Fertility ratings had been estimated for Group 1 and nonreturn rates were known for Group 2. Semen samples were measured by the sperm chromatin structure assay (SCSA): sperm were treated to induce partial in situ DNA denaturation, stained with acridine orange, and evaluated by flow cytometry. Acridine orange intercalated into double-stranded DNA emits green fluorescence upon excitation with 488 nm light, and red fluorescence when associated with single-stranded DNA. An index of DNA denaturation per cell is provided by alpha-t [alpha t = red/(red + green) fluorescence]. The standard deviation (SD alpha t), coefficient of variation (CV alpha t) and proportion of cells outside the main population (COMP alpha t) of the alpha t distribution quantify the extent of denaturation for a sample. Intraclass correlations of the alpha t values were high (greater than or equal to 0.70), based on four collections obtained over several years from Group 1 bulls. Negative correlations were obtained between fertility ratings and both SD alpha t (-0.58, p less than 0.01) and COMP alpha t (-0.40, p less than 0.01) in Group 1, and between nonreturn rates and both SD alpha t (-0.65, p less than 0.01) and COMP alpha t (-0.53, p less than 0.05) in Group 2. These data suggest that the SCSA will be of value for identification of low fertility sires and poor quality semen samples.

Sperm Chromatin Structure Assay: Its Clinical Use for Detecting Sperm DNA Fragmentation in Male Infertility and Comparisons With Other Techniques

“…Much of the development and establishment of the SCSA as a meaningful biological assay for sperm nuclear integrity were conducted on rodent and livestock systems to study toxicology and fertility (Evenson et al, 1985(Evenson et al, , 1989a(Evenson et al, ,b, 1993a(Evenson et al, ,b,c, 1994Ballachey et al, 1987Ballachey et al, , 1988Evenson and Jost, 1993;Sailer et al, 1995a,b;Evenson, 1999a). One study with our laboratory showed that SCSA-defined chromatin damage in bovine spermatozoa negatively affects early embryo formation from what appear to be normally fertilized eggs (Ellington et al, unpublished).…”

“…The results from these studies showed that SCSA was highly dose-responsive to toxicants, highly repeatable, and provided meaningful biological information on sperm nuclear DNA defects. The extensive data from nonhumans (Evenson et al, 1985(Evenson et al, , 1989a(Evenson et al, ,b, 1993a(Evenson et al, ,b,c, 1994Ballachey et al, 1987Ballachey et al, , 1988Evenson and Jost, 1993; Sailer et al, 1995a,b;Evenson, 1999a), and human toxicology and fertility studies (Evenson et al, 1978(Evenson et al, , 1980a(Evenson et al, ,b, 1984(Evenson et al, , 1991Evenson and Melamed, 1983;Evenson, 1999b;Larson et al, 1999Larson et al, , 2000Larson et al, , 2001 provide This work was supported in part by grant R827019 from the US Environmental Protection Agency, grants EPS-9720642 and OSR-9452894 from the National Science Foundation, and by SCSA Diagnostics, Inc. This is South Dakota Agricultural Experiment Station publication 3273 of the journal series.…”

Detection of DNA Damage in Response to Cooling Injury in Equine Spermatozoa Using Single‐Cell Gel Electrophoresis

“…Recently, abnormalities of chromatin structure and DNA integrity in human, bovine, porcine, equine, and mouse sperm have been identified by various techniques as resulting in cellular damage induced by environmental conditions, notably by low-temperature storage of sperm (Evenson et al, 1980b;Ballachey et al, 1987;Evenson et al, 1994;Kenney et al, 1995;Sailer et al, 1996;Aravindan et al, 1997).…”

Susceptibility of Sperm Chromatin to Acid Denaturation in Situ: A Study in Endogenous Fsh-Deprived Adult Male Bonnet Monkeys (Macaca radiata)

“…Abnormal alterations in the kinetics of germ cell transformations and defective spermiogenesis have been implicated for the qualitative differences observed in sperm from FSH-deprived monkeys [I, 2, 141. Due to the condensation and compaction that sperm nuclear chromatin undergoes during spermiogenesis and epididymal maturation, the DNA of normal spermatozoa is relatively resistant to denaturation induced either by heat or acid treatment [7,8,211. Susceptibility to such denaturation treatment has been studied by measuring changes in the ratio of emitted red (R) and green (G) fluorescences by sperm nuclei following acridine orange (AO) binding to sperm DNA using either the two-parametric flow cytometric assay [5] or fluorescence microscopy [ 17,241. Increased susceptibility of sperm to acid-induced denaturation has been strongly correlated to semen quality and infertility in mammals [5].…”