The expression of fibroblast growth factor receptors during early bovine conceptus development and pharmacological analysis of their actions on trophoblast growth in vitro

Abstract

The overall aim of this work was to examine the expression profiles for fibroblast growth factor receptors (FGFRs) and describe their biological importance during bovine pre-and peri-implantation conceptus development. FGFR1 and FGFR2 mRNAs were detected at 1-, 2-, 8-cell, morula and blastocyst stages whereas FGFR3 and FGFR4 mRNAs were detected after the 8-cell stage but not earlier.The abundance of FGFR1, FGFR3, and FGFR4 mRNAs increased at the morula and blastocyst stages. Immunofluorescence microscopy detected FGFR2 and FGFR4 exclusively in trophoblast cells whereas FGFR1 and FGFR3 were detected in both trophoblast cells and inner cell mass in blastocysts. Neither transcripts for FGF10 nor its receptor (FGFR2b) were temporally related to interferon t (IFNT) transcript profile during peri-and postimplantation bovine conceptus development. A series of studies used a chemical inhibitor of FGFR kinase function (PD173074) to examine FGFR activation requirements during bovine embryo development. Exposing embryos to the inhibitor (1 mM) beginning on day 5 post-fertilization did not alter the percentage of embryos that developed into blastocysts or blastocyst cell numbers. The inhibitor did not alter the abundance of CDX2 mRNA but decreased (P!0.05) the relative abundance of IFNT mRNA in blastocysts. Exposing blastocysts to the inhibitor from days 8 to 11 post-fertilization reduced (P!0.05) the percentage of blastocysts that formed outgrowths after transfer to Matrigel-coated plates. In conclusion, each FGFR was detected in bovine embryos, and FGFR activation is needed to maximize IFNT expression and permit outgrowth formation.

Temporal regulation of fibroblast growth factors and their receptors in the endometrium and conceptus during the pre-implantation period of pregnancy in cattle

“…Ozawa et al (2013) demonstrated the presence of FGFR1, FGFR2, FGFR3 and FGFR4 in bovine embryos up to the blastocyst stage and showed that FGFR activation is needed to maximise IFNT expression and permit outgrowth formation. However, there is a paucity of data relating to posthatching stages of development, probably a reflection of the difficulty in obtaining such stages in vivo.…”

Binding of FGF2 to FGFR2 in an autocrine mode in trophectoderm cells is indispensable for mouse blastocyst formation through PKC-p38 pathway

“…FGF1, FGF2 and FGF4 have been reported to be expressed in preimplantation embryos. 20,31,32 Here, we found that the formation of expanded blastocysts was arrested only after the knockdown of FGF2 in TEs in vitro, which suggests that FGF2 is required in TEs for the development of expanded blastocysts. The knockdown of FGF2 in TEs affected the growth of expanded blastocysts, which could be reversed when the embryos were cultured with 1000 ng/ml exogenous FGF2 for 12 h. However, this rescue was completely abolished when the activation of FGF2 was eliminated by the application of an FGF2 antibody.…”

“…18,20,[31][32][33][34][35] However, the functions of these molecules in TEs that lead to the formation of the blastocyst as well as the underlying mechanism have not been elucidated. The present study has demonstrated for the first time that endogenous FGF2 secreted by TEs can regulate protein expression and distribution in TEs via the FGFR2-mediated activation of PKC and p38, which are important for the development of expanded blastocysts (Fig.…”

“…31,32 The development of expanded blastocysts was not affected when the expression of FGFR1, FGFR3 or FGFR4 was suppressed by transfection with specific siRNAs. In addition, using the double-immunofluorescence confocal technique, we found that FGF2 and FGFR2 were co-expressed in the membrane and the cytoplasm of TEs.…”

The requirement for protein kinase C delta (PRKCD) during preimplantation bovine embryo development

“…Histone H2A (H2A) was used as the reference transcript for examining changes in transcript abundance from one-cell to blastocyst stages. Previous work verified the consistency of expression for H2A between the one-cell and blastocyst stages in bovine embryos (Robert et al 2002;Ozawa et al 2013). Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was used as the reference transcript for blastocyst studies given its consistency of expression after blastocyst formation (Purcell et al 2009).…”