Characterization of organic cation/carnitine transporter family in human sperm
Section snippets
Materials and methods
Sperm preparation and analysis. All studies were performed with the approval of the Institutional Review Board of the Hospital for Sick Children, Toronto. Residual unused portions of semen samples were obtained, with informed consent, from men, with normal semen parameters, who were donating samples for artificial insemination at the Sunnybrook and Women’s College Hospital Fertility Centre (Toronto, Canada). Samples were examined and selected according to World Health Organization [41]
Western blot of OCTN1, OCTN2, and OCTN3
To date, three proteins have been shown to transport l-carnitine, namely OCTN1, OCTN2, and OCTN3. Importantly, we have demonstrated the expression of all three, each with an expected molecular weight of ∼63 kDa, in human sperm by Western blot analysis using our anti-murine transporter-specific antibodies (Fig. 1).
Carnitine uptake in sperm
Carnitine uptake studies in human sperm from normal control samples support the presence of a high-affinity transport process with an apparent Km of 3.39 ± 1.16 μM and a maximal velocity
Acknowledgements
This work was supported in part by a grant from the Physicians’ Services Incorporated Foundation of Ontario and the Heart and Stroke Foundation of Ontario NA # 4964. We thank Nelson Palomaris, Domingo Molingbayan, Jennifer Willis, and Dr. Wei Qiu for their helpful assistance.
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