Background
In newborns with an anorectal malformation (ARM), the anal opening is absent or mal-positioned. ARMs are relatively common (1:~5,000 newborns) and often remain associated with poor quality of life after surgical correction. Functionally more effective treatments require better knowledge of the development of both unaffected and malformed structures. Previous studies in our group demonstrated that between 5-7 weeks of development the ventral part of the cloaca is a growth zone, whereas the dorsal part suffers from regression. This study aimed to detect if the dorsal cloaca continues to regress during early fetal development.

Methods
3D-reconstructions from serial sections of 16 human embryos between 7-20 weeks of development were generated with Amira and Cinema software. Landmarks are positions that remain identifiable over successive time points in development. Landmarks near the cloaca were 1) the distal end of the smooth muscle coat of the hindgut, 2) the deep end of the anal canal, 3) the urethral and anal sphincters, 4) the pudendal nerve, and 5) the pubococcygeal line.

Results
At 7.5 weeks, the hindgut epithelium separated into an upper region with, and a lower region without goblet cells. The lower region formed vertical folds (columns of Morgagni), and became contiguous with the deep end of the anal canal at 10 weeks. This finding showed that the dorsal cloaca, initially located between Morgagni’s columns and the anal canal, had disappeared. The anal canal was covered by a thick layer of stratified squamous epithelium layer, whereas its surrounding skin had a much thinner squamous lining.

Conclusion
This study provides a 3D-topographic atlas of the fetal development of the ano-rectum. None of its developmental features can be linked to the spectrum of ARMs, indicating that the malformation arises as a consequence of cloacal remodeling during its septation.