Fig. 5: Tbx2 and Tbx3 are required together for aortic arch artery and arterial branching from the aortic arch.

a–c Intracardiac ink injection of control (a) (n = 3) and Wnt1-Cre;Tbx2 ^f/f;Tbx3 ^f/f conditional null mutant embryos at E15.5 (b, c) (n = 3). Panel b shows a double Tbx2/Tbx3 cKO embryo with an aberrant retro-esophageal right subclavian artery (ARSA). d–l Hematotoxin and Eosin staining on traverse sections of control (d, f) (n = 18) and Tbx2/Tbx3 cKO mutant embryos (g–l) (n = 9) at E15.5. Scale bars: 300 μm. Panels g and h show a Tbx2/3 cKO embryo with ARSA and panels j and k show a Tbx2/3 cKO embryo with a normal right subclavian artery. Panel i shows a Tbx2/3 cKO embryo with aortic arch hypoplasia and panel l shows a Tbx2/3 cKO embryo with a normal aortic arch as compared with control embryos (f). m Table of the cardiovascular defects in Tbx2/Tbx3 conditional mutants. ARSA was identified in 5/13 Tbx2/3 cKO embryos: one after intracardiac ink injection, three after H&E histology, and one by anatomic analysis after micro-dissection. Aortic arch hypoplasia was found in 2/8 Tbx2/3 cKO embryos after H&E histology. n–q Immunofluorescence on coronal sections of controls at E11.5 (n) (n = 8) and Tbx2/3 cKO embryos (o–q) (n = 8), at the level of the pharyngeal arch arteries for GFP and ACTA2 expression. Nuclei are stained with DAPI. Scale bars: 100 μm. The bottom panels are a high magnification of the dashed regions in n–q. Note the strong reduction of ACTA2 expression, ranging from almost total absence to low-level expression, in the right and left fourth PAAs in Tbx2/3 conditional mutant embryos compared to control embryos (white arrowheads). RSA right subclavian artery, RCC right common carotid, LCC left common carotid, BA brachiocephalic artery, Ao aorta, AoA aortic arch, LSA left subclavian artery, PT pulmonary trunk, E esophagus, PAA pharyngeal arch artery.