Involvement of the Notch signaling pathway in vascular development has been demonstrated by both gain-and loss-of-function mutations in humans, mice, and zebrafish. In zebrafish, Notch signaling is required for arterial identity by suppressing the venous fate in developing artery cells. In mice, the Notch4 receptor and the Delta-like 4 (Dll4) ligand are specifically expressed in arterial endothelial cells, suggesting a similar role. Here we show that the Dll4 ligand alone is required in a dosage-sensitive manner for normal arterial patterning in development. This implicates Dll4 as the specific mammalian endothelial ligand for autocrine endothelial Notch signaling, and suggests that Dll4 may be a suitable target for intervention in arterial angiogenesis.
In the present study we examined whether LPA can be synthesized and act during in vitro maturation of bovine cumulus oocyte complexes (COCs). We found transcription of genes coding for enzymes of LPA synthesis pathway (ATX and PLA2) and of LPA receptors (LPAR 1–4) in bovine oocytes and cumulus cells, following in vitro maturation. COCs were matured in vitro in presence or absence of LPA (10−5 M) for 24 h. Supplementation of maturation medium with LPA increased mRNA abundance of FST and GDF9 in oocytes and decreased mRNA abundance of CTSs in cumulus cells. Additionally, oocytes stimulated with LPA had higher transcription levels of BCL2 and lower transcription levels of BAX resulting in the significantly lower BAX/BCL2 ratio. Blastocyst rates on day 7 were similar in the control and the LPA-stimulated COCs. Our study demonstrates for the first time that bovine COCs are a potential source and target of LPA action. We postulate that LPA exerts an autocrine and/or paracrine signaling, through several LPARs, between the oocyte and cumulus cells. LPA supplementation of maturation medium improves COC quality, and although this was not translated into an enhanced in vitro development until the blastocyst stage, improved oocyte competence may be relevant for subsequent in vivo survival.
Lysophosphatidic acid (LPA) is a known cell signaling lipid mediator in reproductive tissues. In the cow, LPA is involved in luteal and early pregnancy maintenance. Here, we evaluated the presence and role of LPA in bovine early embryonic development. In relevant aspects, bovine embryos reflect more closely the scenario occurring in human embryos than the mouse model. Transcription of mRNA and protein expression of enzymes involved in LPA synthesis (ATX and cPLA2) and of LPA receptors (LPAR1–4) were detected in Days 5 and 8 in vitro produced embryos. Embryonic LPA production into culture medium was also detected at both stages of development. Supplementation of culture medium with LPA (10−5 M) between Days 2 and 8 had no effect on embryo yield and quality and on blastocyst relative mRNA abundance of genes involved in prostaglandin synthesis (PTGS2, PGES, and PGFS) and steroidogenesis (3β
HSD). However, LPA treatment affected transcription levels of embryo quality markers, decreasing BAX (apoptotic) and increasing BCL2 (antiapoptotic) and IGF2R (growth marker) gene transcription levels. Blastocyst transcription of OCT4 (pluripotency marker) was not affected by LPA stimulation. In conclusion, LPA is an early bovine embryonic autocrine/paracrine signaling mediator, and LPA action may be relevant in early embryo-maternal interactions leading to embryonic survival.
The role of progesterone (P(4)) and prostaglandins (PGs) in bovine early embryonic development and embryo-maternal crosstalk is almost unknown. Here, the in vitro steroidogenic (P(4)) and prostanoid (PGE(2) and PGF(2α)) interactions between bovine embryos and luteal cells (LC) were evaluated. In two experiments, embryos (n = 1.900) were either co-cultured with LC or cultured alone, from days 2 to 7 (day 0 = in vitro insemination). LC were also cultured alone, and medium was used as a control, all groups being cultured either with or without oil overlay of culture medium. Oil overlay of culture medium significantly decreased the amount of P(4), but not of PGE(2) and PGF(2α) measured in culture medium. Embryos and LC had transcripts of genes coding for enzymes of the PGs (PTGS2, PGES, and PGFS) and P(4) (StAR, P450scc, and 3β-HSD) synthesis pathways, and produced P(4), PGF(2α), and PGE(2) into culture medium. Co-culture with LC exerted an embryotrophic effect, significantly increasing blastocyst yield and quality. This indicates a possible direct effect of LC in early embryo development. Embryos did not exert a luteotrophic effect upon LC. This may indicate that early embryos (until day 7) probably do not exert influence in LC main function. It is suggested that production of P(4), PGE(2), and PGF(2α) by early embryos may be associated to autocrine signaling leading to events in development and to paracrine signaling in the endometrium leading to local uterine receptivity.
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