Premium
Characterization of secretory proteins from cultured cauda epididymal cells that significantly sustain bovine sperm motility in vitro
Author(s) -
ReyesMoreno Carlos,
Boilard Mathieu,
Sullivan Robert,
Sirard MarcAndre
Publication year - 2002
Publication title -
molecular reproduction and development
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.745
H-Index - 105
eISSN - 1098-2795
pISSN - 1040-452X
DOI - 10.1002/mrd.10192
Subject(s) - sperm , biology , motility , andrology , epididymis , sperm motility , in vitro , incubation , protamine , microbiology and biotechnology , biochemistry , medicine , botany , heparin
Epididymis provides a safe environment in which stored‐spermatozoa could survive for days before ejaculation. In vitro studies suggested that epididymal proteins seem to be implicated in sperm survival during coincubation with cultured epididymal cells. This study was basically designed to confirm if secretory proteins from bovine epididymal cell cultures provide sperm protection against rapid loss of sperm motility in vitro. Bovine spermatozoa were incubated in conditioned media (CM), which were prepared from cultured cauda epididymal cell (CEC). Motion parameters were recorded using a computer‐assisted sperm analyzer. Sperm‐free protein extracts from CM were fractionated by ultrafiltration through a 10‐kDa cut off membrane. A significantly positive effect on sperm motility was observed when spermatozoa were incubated in CM (54 ± 4%) and CM >10 kDa (57 ± 4%) compared to CM <10‐kDa fraction (30 ± 3%) or fresh media (34 ± 3%), after a 6‐hr incubation period. This beneficial effect on sperm motility was abolished when the CM >10‐kDa fraction was heat‐treated at 100°C for 10 min. The CM >10 kDa fraction provides factors that remained active even though spermatozoa were washed twice after a 2‐hr preincubation period. To identify potential beneficial factors, bovine spermatozoa were incubated with radiolabeled proteins obtained using 35 S‐methionine in culture medium. SDS–PAGE analysis of proteins extracted from CM‐preincubated spermatozoa revealed the presence of a 42‐kDa protein strongly associated to the sperm surface. This 42‐kDa spot was trypsin‐digested and identified by matrix‐assisted laser desorption ionization‐time of flight (MALDI‐TOF) as a protein homologue to a 35‐kDa bovine estrogen‐sulfotransferase. This protein can play a role in epididymal biology and sperm function. Taken together, these results suggest that specific epididymal proteins can be implicated in the sperm protection in vitro, and can be characterized in our cell culture system. Mol. Reprod. Dev. 63: 500–509, 2002. © 2002 Wiley‐Liss, Inc.