Major Proteins of Bovine Seminal Plasma and High-Density Lipoprotein Induce Cholesterol Efflux from Epididymal Sperm1

Abstract

One of the hypotheses to explain the mechanism of capacitation involves the loss of sperm membrane cholesterol. Here, we studied whether or not the major proteins of bovine seminal plasma designated as BSP-A1, -A2, -A3, and -30-kDa (collectively called BSP proteins), which are implicated in sperm capacitation, induce cholesterol efflux. When epididymal sperm were labeled with [3H]cholesterol and incubated with bovine seminal plasma (0.05-2%) or BSP proteins (20-120 microg/ml) for 8 h, the sperm lost [3H]cholesterol (3.6-fold and 3-fold, respectively). The same results in the presence of BSP-A1/-A2 were obtained (3.5-fold) by direct determination of cholesterol on unlabeled epididymal sperm. Analysis of efflux particles by ultracentrifugation on a sucrose gradient revealed a single symmetrical peak of radioactivity at 1.14 g/ml. Immunoblotting of the fractions obtained from size-exclusion chromatography of the efflux particles showed that a portion of the BSP proteins were associated with [3H]cholesterol. Heparin (12 microg/ml) alone did not stimulate cholesterol efflux. In contrast, high-density lipoprotein (HDL, 100 microg/ml) alone stimulated cholesterol efflux up to 3.1-fold after 8 h. When labeled epididymal sperm were preincubated for 20 min with BSP-A1/-A2 (120 microg/ml), washed, and incubated with HDL (100 microg/ml) for 8 h, the total cholesterol efflux of the sperm suspension was 51.8 +/- 5.0% compared to 39.3 +/- 1.2% when HDL alone was used. These results indicate that BSP proteins and HDL play an important role in the sperm sterol efflux that occurs during capacitation. Furthermore, the heparin-induced sperm capacitation did not involve the efflux of sperm membrane cholesterol.

Immunohistochemical Localization of Sperm‐Preserving Proteins in the Ram Reproductive Tract

“…The homology was found from amino acid 19-34 (CVFPFTYYDDRHFDCT) with 49-64 of PDC-109 (CVFPFVYRNRKHFDCT), which is the FN2 domain (Greube et al, 2001). PDC-109 has been reported to stabilize the sperm membrane in a first step (Greube et al, 2001) and subsequently to participate in the female tract in capacitation by releasing cholesterol and binding high-density lipoprotein and heparin (Thérien et al, 1998(Thérien et al, , 2001Gwathmey et al, 2003). However, the P20 fragment was not homologous with any reported protein (Barrios et al, 2005).…”

Sperm surface changes and physiological consequences induced by sperm handling and storage

“…The main molecules of functional dominance in bull seminal plasma are BSP proteins of the fibronectin type II family, which bind tightly to choline phospholipids on the sperm membrane upon ejaculation (Manjunath et al 2009). This initially stabilises the sperm membrane but extended association causes efflux of cholesterol in a dose-and time-dependent manner (Therien et al 1998. As BSP proteins can also bind high-density lipoprotein and heparin-like glycosaminoglycans (Gwathmey et al 2003(Gwathmey et al , 2006) -known capacitation factors in follicular and oviductal fluids -in vivo, they may act as regulators of capacitation (Lane et al 1999).…”

Functional aspects of seminal plasma and sperm proteins and their potential as molecularmarkers of fertility

“…This outcome may be the result of BSP1-induced damage on sperm, given the exposure period during fertilization (18 h). In agreement with such results, studies show that excess of BSP proteins and time exposure are harmful to cryopreserved sperm because of membrane destabilization and excessive phospholipid and cholesterol efflux (Therien et al, 1995;1998;Manjunath and Therien, 2002). Also, an earlier investigation claimed that the content of BSP5 in accessory sex gland fluid has a quadratic association with bull fertility (Moura et al, 2006), suggesting that too much BSP is detrimental to sperm physiology and/or embryo development.…”