In vitro decidualisation of canine uterine stromal cells

Abstract

BackgroundThe uterine response to the presence of embryos is poorly understood in the domestic dog (Canis familiaris). The intimate embryo-maternal cross-talk, which begins following the hatching of blastocysts and embryo attachment leads to strong structural and functional remodelling of the uterus. A part of this process is decidualisation, comprising morphological and biochemical changes that result in formation of maternal stroma-derived decidual cells. These are an integral part of the canine placenta materna, which together with the maternal vascular endothelium are the only cells of the canine endotheliochorial placenta able to resist trophoblast invasion. These cells are also the only ones within the canine placenta expressing the progesterone receptor (PGR). Understanding the decidualisation process thus appears essential for understanding canine reproductive physiology.MethodsHere, we investigated the capability of canine uterine stromal cells to decidualise in vitro, thereby serving as a canine model of decidualisation. A dbcAMP-mediated approach was chosen during a time course of 24 - 72 h. Tissue material from six (n = 6) healthy, dioestric bitches was used (approximately 2 weeks after ovulation). Cells were characterized by differential staining, nearly 100 % of which were vimentin-positive. Scanning and transmission electron microscope analyses were applied, and morphological changes were recorded with a live cell imaging microscope. Expression of several decidualisation markers was investigated.ResultsThe in vitro cultured stromal cells acquired characteristics of decidual cells when incubated with 0.5 mM dbcAMP for 72 h. Their shape changed from elongated to rounded, while ultrastructural analysis revealed higher numbers of mitochondria and secretory follicles, and an increased proliferation rate. Elevated expression levels of IGF1, IGF2, PRLR and ERα were observed in decidualised cells; PRL and ERβ remained mostly below the detection limit, while PGR remained unaffected. The expression of smooth muscle α actin (αSMA), another decidualisation marker, was strongly induced. Among prostaglandin system members, levels of COX2 (PTGS2) and of PGE2-synthase (PTGES) were upregulated. Expression of the PGE2 receptors, PTGER2 and PTGER4, was clearly detectable.ConclusionAn in vitro decidualisation model with canine uterine stromal cells was successfully established, allowing future, more detailed studies to be undertaken on the underlying molecular and endocrine mechanisms of canine decidualisation.Electronic supplementary materialThe online version of this article (doi:10.1186/s12958-015-0066-4) contains supplementary material, which is available to authorized users.

Decidualization of the canine uterus: From early until late gestational in vivo morphological observations, and functional characterization of immortalized canine uterine stromal cell lines

“…Consequently, there is emerging evidence identifying PGE2 as an important factor involved in canine decidualization. This assumption is further strengthened by our recent observations from studies investigating the capability of canine uterine stromal cells to undergo decidualization in vitro (Kautz et al., ). Utilizing the cAMP‐mediated approach with stromal cells isolated from naturally oestrogen‐primed uteri from early dioestrus dogs, PTGES was strongly induced during the course of in vitro decidualization (Kautz et al., ).…”

“…This assumption is further strengthened by our recent observations from studies investigating the capability of canine uterine stromal cells to undergo decidualization in vitro (Kautz et al., ). Utilizing the cAMP‐mediated approach with stromal cells isolated from naturally oestrogen‐primed uteri from early dioestrus dogs, PTGES was strongly induced during the course of in vitro decidualization (Kautz et al., ). The cAMP‐based protocol was chosen for that study due to its strong decidualization potential as shown previously (Brar et al., ; Gellersen & Brosens, ; Tamura et al., ).…”

The study of endometrium at gestational days 5 and 15 in dairy goats (Capra hircus)

“…Endometrial receptivity has become one of the research hotspots in recent years, and accumulated evidence suggests that hormones (Young 2013), osteopontin (OPN) (Talbi et al 2006), vascular endothelial growth factor (VEGF) (Lai et al 2015), leukaemia-inhibitory factor (LIF) (Miravet-Valenciano et al 2015), prolactin (PRL) (Kautz et al 2015) and other factors participate in the preparation of endometrial receptivity. In addition, intensive research work has been performed to better understand the regulation of the endometrium and its clinical implications to improve implantation in human, and many genes have been reported to influence endometrial development (Strowitzki et al 2006).…”

“…Specifically, the endometrial stromal cells (ESC) undergo a marked rearrangement of the intracellular architecture and begin to accumulate glycogen, initiating the secretion of various proteins, growth factors, and cytokines, such as PRL, which are induced in ESC during decidualization and are therefore recognized as a marker of decidualization (Gellersen and Brosens 2003). In contrast to human and rodent models, however, canine decidualized cells exhibited very low expression of PRL (Kautz et al 2015). PRL-protein was mainly expressed in the cytoplasm of LE and GE, and in a small amount in SC in dairy goats, notably, there was no difference in LE and GE, but a remarkable increase was found in SC on D15 compared to D5.…”

Differential expression and activity of matrix metalloproteinases 2 and 9 in canine early placenta

“…Canine placentation is more invasive than that of the cat, despite their similarities. Absent or scarce decidual vimentin‐positive cells, well described in cat but unsteadily identified in the dog, might partially explain its higher invasiveness (Barbeito, Fernández, Gimeno, & Portiansky, ; Fernández, Barbeito, Portiansky, & Gimeno, ; Fernández et al., ; Kautz et al., ; Walter & Schönkypl, ). Furthermore, several cellular and matrix components participate in the invasive function of the early canine placenta.…”