IMMUNOLOCALIZATION OF HEAT SHOCK PROTEIN 70 (Hsp70) IN BOAR SPERMATOZOA

Hsp70 has been identified in human (Miller et al., Hum Reprod 7:637, 1992) and bovine (Kamaruddin et al., Andrologia 36:327, 2004) ejaculates as well as in extracts from boar sperm (Huang et al., Anim Reprod Sci 63:231, 2000). The aim of this study was to determine Hsp 70 presence and distribution in ejaculated, capacitated and acrosome-reacted boar spermatozoa; the fate of this protein after fertilization was also studied. In fixed-permeabilized fresh semen 92.3±1.8% of cells showed a triangular-shaped immunoreactivity in the equatorial segment of the head. The localization of fluorescent signal underwent several rearrangements after capacitation, becoming particularly evident in the equatorial line in 72.6±5.3% of cells, and in a sub-equatorial band in 88.9±2.3% of sperm after induced acrosome reaction (AR). The proportion of unfixed cells showing the fluorescent signal significantly (P<0.05) increased after AR. No differences in Hsp70 amount between fresh, capacitated and reacted sperm were observed by Western blot. A loss of Hsp immunoreactivity in demembranated spermatozoa was recorded. Perivitelline sperm cells after IVF showed the same pattern as AR-induced cells; after sperm incorporation into the oocyte Hsp positivity became fuzzy and disappeared during sperm decondensation. Our results indicate that Hsp 70 undergoes a redistribution along sperm membrane and is probably translocated from the inner to the outer leaflet after capacitation and acrosome reaction.