*2.7. EC-Specific Loss of Jag1 Does Not Impact CD31*<sup>+</sup> *EC Content in Pregnant Uteri at E7.5*

To better understand the effect of loss of *Jag1* on decidual vasculature, flow cytometry was done to quantify the proportion of CD31+CD45<sup>−</sup> ECs in the implantation site and myometrium of *Jag1*∆*EC* and control pregnancies at E7.5 (Figure 8A). Analysis of the uteri from nonpregnant C57BL/6 mice and pregnant C57BL/6 mice showed that an increase CD31+CD45<sup>−</sup> EC content was the highest in the decidua of pregnant C57BL/6 mice at E7.5 (Figure 8B). Quantification of the percentages of CD31+CD45<sup>−</sup> ECs in the decidua and myometrium of E7.5 *Jag1*∆*EC* and *Cdh5-CreERT2* control pregnancies revealed no significant differences (Figure 8C). Thus, despite loss of *Jag1* in ECs, the overall percentage of decidual CD31<sup>+</sup> ECs is similar in *Jag1*∆*EC* and control pregnancies. *2.7. EC-Specific Loss of Jag1 Does Not Impact CD31+ EC Content in Pregnant Uteri at E7.5*  To better understand the effect of loss of *Jag1* on decidual vasculature, flow cytometry was done to quantify the proportion of CD31+CD45- ECs in the implantation site and myometrium of *Jag1*Δ*EC* and control pregnancies at E7.5 (Figure 8A). Analysis of the uteri from nonpregnant C57BL/6 mice and pregnant C57BL/6 mice showed that an increase CD31+CD45- EC content was the highest in the decidua of pregnant C57BL/6 mice at E7.5 (Figure 8B). Quantification of the percentages of CD31+CD45- ECs in the decidua and myometrium of E7.5 *Jag1*Δ*EC* and *Cdh5-CreERT2* control pregnancies revealed no significant differences (Figure 8C). Thus, despite loss of *Jag1* in ECs, the overall percentage of decidual CD31+ ECs is similar in *Jag1*Δ*EC* and control pregnancies.

**Figure 8.** EC-specific loss of *Jag1* does not impact the frequency of CD31+ ECs in implantation sites. (**A**) Representative dot plot of flow cytometry (FCM) data showing gating strategy to identify CD31+ CD45– EC populations in implantation sites from wild-type C57BL/6 mice at E7.5. (**B**) Results of FCM analysis to assess CD31+CD45 cell populations in uteri from nonpregnant C57BL/6 mice, and in the myometrium and implantation sites from pregnant C57BL/6 mice at E7.5. The percentage of CD31+CD45- ECs is significantly higher in implantation sites compared to the myometrium or nonpregnant uteri. (**C**) Results of FCM analysis to determine the percentage of CD31+CD45- ECs in the myometrium and implantation sites of *Cdh5-CreERT2* control and *Jag1ΔEC* pregnancies at E7.5. The percentage of CD31+CD45- ECs in the myometrium and implantation sites of *Cdh5-CreERT2* control and *Jag1ΔEC* pregnancies is similar. IS = implantation site; myo = myometrium; NP = nonpregnant; each **Figure 8.** EC-specific loss of *Jag1* does not impact the frequency of CD31<sup>+</sup> ECs in implantation sites. (**A**) Representative dot plot of flow cytometry (FCM) data showing gating strategy to identify CD31<sup>+</sup> CD45– EC populations in implantation sites from wild-type C57BL/6 mice at E7.5. (**B**) Results of FCM analysis to assess CD31+CD45<sup>−</sup> cell populations in uteri from nonpregnant C57BL/6 mice, and in the myometrium and implantation sites from pregnant C57BL/6 mice at E7.5. The percentage of CD31+CD45<sup>−</sup> ECs is significantly higher in implantation sites compared to the myometrium or nonpregnant uteri. (**C**) Results of FCM analysis to determine the percentage of CD31+CD45<sup>−</sup> ECs in the myometrium and implantation sites of *Cdh5-CreERT2* control and *Jag1*∆*EC* pregnancies at E7.5. The percentage of CD31+CD45<sup>−</sup> ECs in the myometrium and implantation sites of *Cdh5-CreERT2* control and *Jag1*∆*EC* pregnancies is similar. IS = implantation site; myo = myometrium; NP = nonpregnant; each group contains *n* = 3–5 individual mice. Data shown as median + IQR; \* *p* < 0.05.

The recruitment and differentiation of mural cells, into pericytes and vascular smooth muscle

Δ

*EC Pregnancies* 

*EC* mutant and *Cdh5-CreERT2* control pregnancies were stained

group contains *n* = 3–5 individual mice. Data shown as median + IQR; \* *p* < 0.05.

Δ

for expression of CD31 and mural cell markers, PDGFRβ, NG2, and SMA (Figure S3).

*2.8. Vasculature Is Similar in Implantation Sites of Control and Jag1*

content, implantation sites from *Jag1*
