Automated analysis of toxicant-induced changes in rat sperm head morphometry☆

Abstract An automated sperm morphometry analysis (ASMA) instrument was developed to obtain measurements of toxicant-induced changes in rat sperm head morphometry. 1,3-dinitrobenzene (1,3-DNB), a testicular toxicant known to affect sperm parameters, was used. Twelve-week-old Sprague-Dawley rats were allocated to a control (C) and to two 1,3-DNB treatment groups (T1 = 15 mg/kg; T2 = 25 mg/kg). 1,3-DNB was administered as a single dose by gavage, and animals were sacrificed 22 days after exposure. Sperm were collected, and morphology smears were made by a standard method. One hundred sperm heads were digitized on each slide, and 8 metric measurements were automatically reported. All measurements tended to decrease in a dose-dependent manner with increasing doses of 1,3-DNB. All values for total width (W a ) and interior width (W e ) were significantly different from one another. Wa/L was significantly larger in the control than in T1 or T2, and symmetry (S = W b /W a ) was significantly smaller in the control than in T1 or T2. Multivariate cluster analysis revealed three subpopulations that were also visually distinct. Subpopulation no. 1 was normal, based on published descriptions of normal rat sperm; subpopulation no. 2 was abnormal with a flattened curvature and a normal length; subpopulation no. 3 was abnormal with a foreshortened length and a flattened curvature. T1 and T2 contained significantly more sperm from subpopulation no. 2 and no. 3 than C(T1 = 22% and T2 = 34% vs. C = 8% by cluster analysis). C had 93% normal sperm, while the treatments had 78% and 66%, respectively. Based on these findings, we conclude that ASMA is an efficient and accurate method of sperm morphometry analysis that may improve our ability to detect sperm biomarkers of reproductive health.