Plasma Membrane Integrity and Morphology of Frozen-Thawed Bull Spermatozoa Supplemented with Desalted and Lyophilized Seminal Plasma

摘要

Cryopreservation induces extensive changes in the sperm cell and the success of artificialinsemination (AI) with frozen semen implies reduction of the deleterious effects on sperm plasma membranesinduced by this technique. This study compare three methods for evaluating plasma membrane integrity offrozen-thawed Japanese Black bull spermatozoa supplemented with desalted and lyophilized seminal plasma(SP): eosin-nigrosin (EN); Propidium iodide (PI); and hypoosmotic swelling test (HOST). SP was desalted usingSephadex G-25 desalting column and lyophilized before added to semen extender at final concentrations 0, 2.5,12.5 and 25 mg/ml. Plasma and acrosomal membranes integrity as well as abnormal sperm morphology wereassessed. The results showed that the proportions of membrane-intact sperm with EN, PI and HOST were notsignificantly different and all methods failed to detect any significant effect of the added SP. Centrifugation,washing and HOST solution adversely affect acrosomal membrane. The proportions of abnormal spermatozoaidentified by Glutaraldehyde fixation was equivalent to that of EN. Overall, EN, PI and HOST have no significantdifference in identifying the proportions of membrane-intact bull spermatozoa supplemented with desalted andlyophilized SP. Subsequently, the added SP has no beneficial effect on either physical or functional integrityof the sperm membranes.