*4.1. Animals*

The Institutional Animal Care and Use Committee at Rutgers-New Jersey Medical School approved the studies (PROTO2018000086; October, 29, 2018). The experiments were designed to determine the role of endothelial Jag1/Notch signaling in the formation and function of uterine decidual blood vessels during early pregnancy development, prior to placentation. To delete Jag1 in the endothelium, *Cdh5-*CreERT2 *,* an endothelial specific, tamoxifen inducible *Cre* transgenic strain (gift from Ralf Adams [38]) were crossed with *Jagged1flox*/*flox* mice [39]. To validate our protocol for tamoxifen administration with the *Cdh5-CreERT2* driver, tomato reporter mice *ROSA26 tdTomato* (B6.Cg-Gt(ROSA)26Sortm14(CAG-tdTomato)Hze/J, Jackson Laboratories) were used. *Cdh5-CreERT2* mice were bred to *Jagged1flox*/*flox* mice or *ROSA26 tdTomato* mice, thereby generating *Cdh5-Cre*;*Jagged1fl*/*fl* (*Jag1*∆*EC*) mutant females and *Cdh5-CreERT2*; *ROSA26 tdTomato* reporter females. *Cdh5-CreERT2* female mice were used as controls. All strains were maintained on a C57BL/6 background.

*Jag1*∆*EC* or control females, 8–12 weeks of age, were mated to C57BL/6 males for their first pregnancy. Identification of a vaginal plug in the morning was interpreted as successful mating. Noon was designated as embryonic day (E) 0.5. At E4.5, tamoxifen (0.1mg/g body weight (Sigma, Milwakee, WI, USA) was administered by oral gavage. Since the impact of loss of endothelial Jag1/Notch signaling on ovarian function and progesterone (P4) secretion is not known, P4 replacement was performed. Two extended-release P4 capsules (15 mg P4 per capsule, 21-d release, 4-mm diameter; Innovative Research of America, Sarasota, FL, USA) were placed subcutaneously. Pregnant female mice were euthanized at E7.5 and implantation sites without myometrium were isolated.
