PRELIMINARY CHARACTERIZATION OF A FACTOR IN HUMAN FOLLICULAR-FLUID THAT STIMULATES HUMAN SPERMATOZOA MOTION

Follicular fluid alters the physiology and behaviour of spermatozoa by increasing acrosome reaction, accelerating capacitation, attracting t he spermatozoon and enhancing vigorous motion of the cell. The objecti ve of this study was to characterize the factor(s) in human follicular fluid that causes vigorous spermatozoa motion. Follicular fluid and i ts fractions were tested for stimulation of spermatozoa motion using a standardized assay which employs a computerized digital imaging syste m. Our results show that both follicular fluid and its methanol extrac t stimulate vigorous spermatozoa motion. To determine the characterist ics of the active factor(s), the methanol extract was subjected to mol ecular weight fractionation, protease digestion, microcrystalline thin -layer chromatography (TLC) and C-18 reverse-phase high-pressure liqui d chromatography (HPLC). The spermatozoa motion stimulator in the meth anol extract was dialysable against a low molecular weight membrane (1 000 Da), insensitive to boiling and low pH (3.5) and was largely inact ivated by proteinase K digestion. The activity was detected near the s olvent front on TLC. Using reverse-phase HPLC monitored at 254 nm (UV) , the activity eluted as a single peak of activity at low methanol con centration, indicating that the activity was relatively hydrophilic. T he activity in the HPLC peak lost most, of its motion-stimulating abil ity after digestion with proteinase K. The motion stimulator could be a peptide analogous to the egg-associated peptides characterized in ec hinoderms which stimulate spermatozoa motion, respiration and chemotax is.