S. Shariatzadeh1, M. G. Martin2, J. Dunn1 1Stanford University, Division Of Pediatric Surgery, Department Of Surgery, Palo Alto, CA, USA 2University Of California – Los Angeles, Division Of Gastroenterology And Nutrition, Department Of Pediatrics, Los Angeles, CA, USA
Introduction:
The formation of the circular muscle layer (CM) during embryonic intestine development exerts a circumferential force within the intestinal tube, which is indispensable for the subsequent formation of the longitudinal muscle layer (LM). Yes-associated protein (YAP), the primary downstream effector of the Hippo pathway, acts as mechanosensitive transcriptional coactivator and its translocation from the cytoplasm to the nucleus can influence cell fate. Herein, we aim to characterize the localization of YAP within each intestinal muscle layer in response to circumferential force utilizing a partial obstruction mouse model that provides a non-dissipative force over time.
Methods:
5-mm tubes were placed around the terminal ileum of C57BL/6 mice that were euthanized at post-operation days (POD) 1-7. Dilated ileum proximal to the tube was harvested and compared with the normal bowel of untreated mice. The thickness of CM and LM was measured. RNA sequencing was performed to profile the transcriptome adaptations. The proliferation, cell size, and YAP subcellular localization of each muscle layer were analyzed using immunofluorescence.
Results:
The lumen of the ileum dilated over time, and the muscularis thickness gradually increased after an initial decrease on POD3. The maximal decrease in thickness was observed in the LM on POD3, suggestive of maximal mechanosensation. The number and size of muscle cells significantly increased, indicating hyperplastic and hypertrophic adaptation. The maximum proliferation was observed on POD 5, with a larger increase in the LM compared to the CM. RNA-seq analysis identified the upregulation of Hippo pathway at POD 3 (fig. a). Total YAP expression escalated, reaching 5 and 3 times higher in LM and CM on POD5, respectively. Furthermore, a significant temporal increase in the nuclear to cytoplasmic YAP ratio was observed, with the maximum ratio observed in LM on POD 3 (fig. b).
Conclusion:
The study reveals that the intestinal muscle layers can proliferate in response to mechanical force due to obstruction, with the longitudinal layer being particularly well-suited for circumferential mechanosensation. These findings suggest that Hippo pathway and spatiotemporal translocation of YAP could play a role in regulating the intestinal smooth muscle cell’s proliferation and size. It provides insight into understanding the mechanisms underlying gastrointestinal wall disorders.
