Gene expression and protein distribution of leptin and its receptor in bovine oocytes and preattachment embryos produced in vitro.

Many investigations point out the important role of leptin during the preimplantation development. Transcripts for the leptin gene (LEP) and its receptor (LEPR) have been identified in several tissues related to reproduction (e.g. ovaries, testis and oviduct) in both human and mouse. This work shows for the first time the expression and distribution patterns of LEP and LEPR in bovine oocytes and in vitro-produced embryos. Gene expression was analysed by reverse transcription PCR and real-time PCR, and the proteins were localised by immunostaining. This study included immature and mature oocytes, zygotes, two-, four-, eight- to 16-cell embryos, morulae and blastocysts and the LEP transcript was identified throughout all stages of bovine preimplantation development. However, mRNA for the LEPR gene was detected at all stages, excluding four-cell embryos. Expression of both LEP and LEPR genes was reduced at the eight- to 16-cell stage. This in addition to the absence of LEPR mRNA in four-blastomere embryos may suggest that maternally derived transcripts degenerate towards the eight- to 16-cell stage coinciding with embryonic genome activation at eight- to 16-cell stage and subsequent appearance of embryonic mRNA. Immunofluorescent staining demonstrated that LEP and LEPR proteins form a spherical rim beneath the oolemma. After maturation, however, the proteins became evenly distributed within the cytoplasm. In two- to eight-cell embryos, fluorescence was observed in the apical surface of the blastomeres, and from 10- to 16-cell stage in the apical region of outer blastomeres. This pattern persisted to the blastocyst stage, leading to LEP and LEPR distribution within trophoblast cells, but not in the inner cell mass. These results support previous findings on polar distribution of proteins within mammalian oocytes and embryos, as well as suggests leptin's potential role during early mammalian development and implantation.