Nectin-2-Null Spermatozoa Display Impaired Binding to Zona-Intact Mouse Oocytes
Zona-intact mouse oocytes were recovered from ovulated CD1 females 15 h post-hCG administration and incubated with 5 X 105 nectin-2wt/LacZ or nectin-2LacZ/LacZ sperm per milliliter (Fig. 4). We observed a 6-fold reduction in the binding capacity of nectin-2LacZ/LacZ spermatozoa to oocytes. An average of 40.8 nectin-2LacZ/LacZ spermatozoa bound per oocyte, but only 6.7 r^e^1^^^-2wt/LacZ spermatozoa were found to bind per oocyte (Fig. 4).
Nectin-2LacZ/LacZ Sperm Bind to Oolemma Normally But Fail to Penetrate
The SPA using zona-free hamster oocytes is a well-established test that has been used extensively to assess the penetration capacity of a wide variety of mammalian sperm. Removing the zona pellucida sidesteps the requirement for the species-specific sperm-zona binding reaction, and it allows one to evaluate sperm/oolemma binding and fusion. buy ventolin inhaler
FIG. 4. Reduction of nectin-2LacZ/LacZ spermatozoa binding to zona-intact mouse oocytes. A) Six sets of approximately 20 zona-intact mouse oocytes (61 oocytes/genotype) were incubated with spermatozoa of three nectin-2wt/LacZ and three nectin-2LacZ/LacZ males, respectively. Bound spermatozoa were stained with Hoechst 33258 DNA stain and counted under an inverted epifluorescence microscope. Binding of nectin-2LacZ/LacZ spermatozoa to mouse zona pellucida was found to be reduced 6-fold compared to nectin-2wt/LacZ controls (P < 0.0001). B-E) Representative phase-contrast image of nectin-2LacZ/LacZ (B and C) and nectin-2wt/LacZ (D and E) spermatozoa bound to oocytes. Magnification X600 (B-D).