Figure 3
Figure 3. Deletion of Smad4 by Vav-Cre has no influence on hematopoietic progenitor generation and homeostasis. (A) Representative FACS analysis of EYFP fluorescence in the gated subpopulations (blue boxes) of E10.5 Tie2-Cre;ROSA-EYFP (upper) and E10.5 Vav-Cre;ROSA-EYFP (lower) AGM regions, respectively. The ROSA-EYFP littermate without the Cre transgene serves as the negative controls (red lines). (B) Whole mount immunostaining of c-Kit (red) and CD31 (green) at E10.5. Confocal images of the aorta region of Vav-Cre;Smad4fl/fl and control embryos are shown (three-dimensional reconstructions with totally 120 μm thickness). Higher magnification images of representative clusters are shown to the right, respectively. Scale bars: 40 μm. (C) Representative FACS analysis of E10.5 AGM cells for CD31+c-Kithigh expression (left). The graph to the right shows the percentages of the gated populations (n = 3). (D) Representative FACS analysis of EYFP fluorescence in the CD31+c-Kithigh and CD41lowc-Kit+ subpopulations of the E11.5 Vav-Cre;ROSA-EYFP AGM region. The ROSA-EYFP littermate without the Cre transgene serves as the negative controls (red lines). (E) Representative FACS analysis of E11.5 AGM cells for CD31+c-Kithigh and CD41lowc-Kit+ expression (left). The graphs to the right show the percentages of the gated populations (n = 3). (F) Representative FACS analysis of EYFP fluorescence in the hematopoietic progenitors (CD45+c-Kit+) and hematopoietic cells (CD45+) of the E12.5 Vav-Cre;ROSA-EYFP fetal liver. The ROSA-EYFP littermate without the Cre transgene serves as the negative controls (red lines). (G) Graphs showing the frequencies of hematopoietic progenitors and hematopoietic cells in the E12.5 fetal liver of Vav-Cre;Smad4fl/fl and control embryos (n = 3). (H) Graph showing the number of hematopoietic colonies in the E11.5 AGM region of Vav-Cre;Smad4fl/fl and control embryos (n = 3). (I) Graphs showing the number of hematopoietic colonies in the E12.5 fetal liver of Vav-Cre;Smad4fl/fl and control embryos (n = 3). Data are mean ± standard deviation. DA, dorsal aorta; NS, not significant.

Deletion of Smad4 by Vav-Cre has no influence on hematopoietic progenitor generation and homeostasis. (A) Representative FACS analysis of EYFP fluorescence in the gated subpopulations (blue boxes) of E10.5 Tie2-Cre;ROSA-EYFP (upper) and E10.5 Vav-Cre;ROSA-EYFP (lower) AGM regions, respectively. The ROSA-EYFP littermate without the Cre transgene serves as the negative controls (red lines). (B) Whole mount immunostaining of c-Kit (red) and CD31 (green) at E10.5. Confocal images of the aorta region of Vav-Cre;Smad4fl/fl and control embryos are shown (three-dimensional reconstructions with totally 120 μm thickness). Higher magnification images of representative clusters are shown to the right, respectively. Scale bars: 40 μm. (C) Representative FACS analysis of E10.5 AGM cells for CD31+c-Kithigh expression (left). The graph to the right shows the percentages of the gated populations (n = 3). (D) Representative FACS analysis of EYFP fluorescence in the CD31+c-Kithigh and CD41lowc-Kit+ subpopulations of the E11.5 Vav-Cre;ROSA-EYFP AGM region. The ROSA-EYFP littermate without the Cre transgene serves as the negative controls (red lines). (E) Representative FACS analysis of E11.5 AGM cells for CD31+c-Kithigh and CD41lowc-Kit+ expression (left). The graphs to the right show the percentages of the gated populations (n = 3). (F) Representative FACS analysis of EYFP fluorescence in the hematopoietic progenitors (CD45+c-Kit+) and hematopoietic cells (CD45+) of the E12.5 Vav-Cre;ROSA-EYFP fetal liver. The ROSA-EYFP littermate without the Cre transgene serves as the negative controls (red lines). (G) Graphs showing the frequencies of hematopoietic progenitors and hematopoietic cells in the E12.5 fetal liver of Vav-Cre;Smad4fl/fl and control embryos (n = 3). (H) Graph showing the number of hematopoietic colonies in the E11.5 AGM region of Vav-Cre;Smad4fl/fl and control embryos (n = 3). (I) Graphs showing the number of hematopoietic colonies in the E12.5 fetal liver of Vav-Cre;Smad4fl/fl and control embryos (n = 3). Data are mean ± standard deviation. DA, dorsal aorta; NS, not significant.

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