Contents Shallow trophoblast invasion is detrimental in human pregnancies, but represents normal endotheliochorial placentation in dogs. Factors regulating shallow trophoblast invasion into the canine decidua are not well described, but it is known that matrix metalloproteinases (MMPs) play a crucial role in trophoblast invasion in many species. Following the methods previously described for isolating human trophoblasts, canine trophoblasts were isolated using collagenase and trypsin digestions with Percoll density gradient centrifugation. In addition, placental pieces were cryopreserved prior to primary culture following methods previously described for human tissue. Expression of cytokeratin‐7, MMP2 and MMP9 was confirmed using fluorescent immunocytochemistry. Cellular morphology was similar to that reported for trophoblasts. More than 97% of the cells cultured expressed cytokeratin‐7. More cultured canine trophoblasts expressed MMP9 (54.7 ± 3.4%) compared with MMP2 (40.3 ± 1.8%) (p = 0.02). Although both MMPs were immunolocalized to the cytoplasm, MMP2 was found in large, coalescing granules, whereas MMP9 was more diffusely expressed throughout the cell. Cryogenic freezing of placental tissue prior to primary cell culture had no effect on cell proliferation (p = 0.37). This research has established a baseline for future studies investigating the canine placenta as a model for disorders of shallow trophoblast invasion in humans.

Contents Matrix metalloproteinases (MMPs), tissue inhibitors of metalloproteinases (TIMPs), vascular endothelial growth factor (VEGF)‐A, VEGF‐A receptor (Flt‐1) and KiSS‐1 receptor (KiSS‐1R) all play a role in trophoblast invasion in a number of mammalian species. However, mRNA expression of factors regulating trophoblast invasion has not been studied in dogs. Abnormal expression of these factors at the end of canine gestation may contribute to placental retention and/or subinvolution of placental sites. Therefore, we sought to determine the relative mRNA expression of these factors in canine chorioallantois tissue at 61 ± 1 day past the LH surge (pre‐term; n = 4), following elective c‐section at 64 ± 1 day past the LH surge prior to first‐stage labour (pre‐labour; n = 4) and following natural delivery (parturient; n = 3). Total RNA was isolated, real‐time RT‐PCR was performed, and relative expression was calculated using the relative quantitation (2−ΔΔCT) method. MMP‐9 mRNA expression was significantly higher in pre‐term samples compared to pre‐labour and parturient samples. The results showed no significant difference between MMP‐2, TIMP‐2, VEGF‐A and Flt‐1 mRNA expression among the three groups. KiSS‐1R mRNA was not expressed in any tissues studied. Gene expression of MMP‐9 may be related to the onset of labour, whereas MMP‐2, VEGF‐A, Flt‐1, TIMP‐2 and KiSS‐1R mRNA do not appear to play a role at the end of gestation in the dog.