Cardiac conduction system malformations in heterotaxy result from dysregulated Pitx2 expression
Kunihiko Joo, Ryohei Matsuoka, Keiko Kitajima, Kenta Yashiro, Akira Shiose, Ryuji Tominaga, Michael M. Shen, Shinya Oki, Chikara Meno
Kunihiko Joo, Ryohei Matsuoka, Keiko Kitajima, Kenta Yashiro, Akira Shiose, Ryuji Tominaga, Michael M. Shen, Shinya Oki, Chikara Meno
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Research Article Cardiology Development

Cardiac conduction system malformations in heterotaxy result from dysregulated Pitx2 expression

Abstract

The cardiac conduction system (CCS) develops asymmetrically along the body axes. In heterotaxy syndrome — resulting from aberrant left-right axis formation — atrial and atrioventricular conduction defects can cause life-threatening arrhythmias. However, the developmental mechanisms regulating the atrioventricular conduction system (AVCS) disposition and integrity remain unclear. To investigate the etiology of AVCS malformations in laterality defects, we analyzed CCS development and function in mouse mutants for Cryptic and Lefty1, which are key regulators of Pitx2 in the left-right axis formation. Cryptic–/– embryos exhibited bilateral sinoatrial nodes and an ectopic anterior AV node and bundle accompanied by reduced Pitx2 expression. In contrast, Lefty1–/– embryos showed a hypoplastic sinoatrial node and AV node–bundle dissociation with ectopic Pitx2 expression. Single-cell transcriptomic analysis of Pitx2–/– hearts revealed expansion of AV node and bundle populations, consistent with a repressive role of Pitx2 in AVCS specification. Genetic lineage tracing indicated that Pitx2-expressing cells from the left lateral plate mesoderm populate cranioventral cardiac regions, where AVCS development is suppressed. Together, these findings clarify how global left-right axis information is locally integrated to shape AVCS disposition and integrity, providing a mechanistic model for AVCS abnormalities in laterality-associated congenital heart disease.

Authors

Kunihiko Joo, Ryohei Matsuoka, Keiko Kitajima, Kenta Yashiro, Akira Shiose, Ryuji Tominaga, Michael M. Shen, Shinya Oki, Chikara Meno

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Figure 1

Abnormal cardiac conduction system in the hearts of Cryptic–/– embryos at E18.5.

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Abnormal cardiac conduction system in the hearts of Cryptic–/– embryos a...
(A–J) Representative images of 3D-reconstructed hearts (A and F) and their original images of in situ hybridization with Hcn4 probes (B–E and G–J) in WT (A–E) (n = 6) and Cryptic–/– (F–J) embryos (n = 4) at E18.5. Upper panels in A and F are ventral views showing the lumen of heart chambers and great vessels, whereas lower panels are right ventral views showing Hcn4 expression in pseudocolors (SA node head, dark green; SA node tail, pale green; venous valves and sinus horn, yellow; AV ring, light blue; AV node and cranioventral nodule, blue; AV bundle and septal branch, red; bundle branches, orange). The levels for each transverse section are shown in the lower panels. (K and L) Right craniodorsal views of the base showing Hcn4 expression in the WT (K) and Cryptic–/– (L) hearts presented in A and F, respectively. (M–P) Expression of Tbx3 (M and O), Myl7 (N), and Gja5 (P) was detected in adjacent transverse sections of a Cryptic–/– heart (different from F), corresponding to the level shown in H. Scale bars: 200 μm. aAVB, (anterior) atrioventricular bundle; aAVN, (anterior) atrioventricular node; Ao, aorta; AVR, atrioventricular ring; cvN, cranioventral nodule; DMP, dorsal mesenchymal protrusion–derived tissue; LA, left atrium; (L/R)BB, (left/right) bundle branch; (L/R)SCV, (left/right) superior caval vein; LSH, left sinus horn; LV, left ventricle; (M)RA, (morphologically) right atrium; PA, pulmonary artery; PV, pulmonary vein; RV, right ventricle; SAN, sinoatrial node; VS, ventricular septum; VV, venous valve.