Recent evidences (Matwee et al., Mol Hum Reprod 7:829, 2001) suggest that heat shock protein 70 (Hsp70) could exert an important effect during bovine gamete interaction. As Hsp70 has been identified in extracts from boar semen (Huang et al., Anim Reprod Sci 63:231, 2000), the aim of this study was to examine the possible role of this protein during in vitro fertilization in pig. In vitro matured oocytes were fertilized in presence of 2, 5 and 10 µg/ml of anti-Hsp70 monoclonal antiserum or 10 µg/ml mouse myeloma ascites IgG1 (MOPC21). After 20 h the oocytes were fixed and stained with lacmoid to evaluate the nuclear status. The presence of anti Hsp70 antibody induced a clear dose-dependent inhibition of in vitro fertilization: the fertilization rate was in fact significantly lower (P<0.05) in the presence of 10μg/ml of anti-Hsp70 as compared to control (49.8±6.6 vs 83.6±4.2%). The inhibition is unlikely due to non specific binding, since the exposure to MOPC21 did not induce similar effects (fertilization rate 84.5±3.4%). Moreover, the addition of a FITC-conjugated sheep anti-mouse antibody to spermatozoa collected from IVF wells with 10μg/ml of anti-Hsp70 antibody and spotted onto glass slides, induced a thick post-equatorial band green fluorescence, thus confirming the binding in vivo of anti-Hsp70 antibodies to sperm cells. These results strongly suggest that Hsp70 could play an important role in sperm-egg interaction during in vitro fertilization in swine species.
EFFECT OF ANTI-HEAT SHOCK PROTEIN 70 (Hsp 70) ANTIBODY ON PIG IN VITRO FERTILIZATION
SPINACI, MARCELLA;VOLPE, SARA;DE AMBROGI, MARCO;TAMANINI, CARLO;SEREN, ERALDO;GALEATI, GIOVANNA
2005
Abstract
Recent evidences (Matwee et al., Mol Hum Reprod 7:829, 2001) suggest that heat shock protein 70 (Hsp70) could exert an important effect during bovine gamete interaction. As Hsp70 has been identified in extracts from boar semen (Huang et al., Anim Reprod Sci 63:231, 2000), the aim of this study was to examine the possible role of this protein during in vitro fertilization in pig. In vitro matured oocytes were fertilized in presence of 2, 5 and 10 µg/ml of anti-Hsp70 monoclonal antiserum or 10 µg/ml mouse myeloma ascites IgG1 (MOPC21). After 20 h the oocytes were fixed and stained with lacmoid to evaluate the nuclear status. The presence of anti Hsp70 antibody induced a clear dose-dependent inhibition of in vitro fertilization: the fertilization rate was in fact significantly lower (P<0.05) in the presence of 10μg/ml of anti-Hsp70 as compared to control (49.8±6.6 vs 83.6±4.2%). The inhibition is unlikely due to non specific binding, since the exposure to MOPC21 did not induce similar effects (fertilization rate 84.5±3.4%). Moreover, the addition of a FITC-conjugated sheep anti-mouse antibody to spermatozoa collected from IVF wells with 10μg/ml of anti-Hsp70 antibody and spotted onto glass slides, induced a thick post-equatorial band green fluorescence, thus confirming the binding in vivo of anti-Hsp70 antibodies to sperm cells. These results strongly suggest that Hsp70 could play an important role in sperm-egg interaction during in vitro fertilization in swine species.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
