Decrease, GLI1/2 target gene expression leads to Apoptosis

The development of the face occurs early in embryogenesis and involves precise coordination of multiple tissues. The oropharyngeal membrane appears early in the 4^th week of gestation and gives rise to the frontonasal process and the 1^st pharyngeal arch. The frontonasal process is derived from the neural crest and in turn gives rise to two medial nasal process and two lateral nasal processes that later fuse and form the intermaxillary process. The pharyngeal arch is derived from mesoderm and the neural crest. It gives rise to two mandibular process and two maxillary processes (Som and Naidich 2013). These processes are comprised of mesenchymal cells from neural crest migration and the craniopharyngeal ectoderm and are coated in an epithelium (Ferguson 1988). The upper lip is formed during weeks 5-7 when the maxillary processes grow towards the midline and fuse intermaxillary process that have formed the philtrum and columella (Warbrick 1960, Kim, Park et al. 2004). The palate develops between week 6-12 from a median palatine process and a pair of lateral palatine processes. The primary palate is formed from the posterior extension of the intermaxillary process. The lateral palatine processes arise as medial mesenchymal processes from both maxillary processes. These processes initially grow inferiorly until the tongue is pulled downwards by the elongation of the maxilla and mandible. Once above the tongue, the lateral processes grow medially until they make contact and fuse (Som and Naidich 2014). For normal facial development and growth coordinated multivariate signaling is required. For example, retinoic acid, BMP, FGF, and SHH signal together to control facial growth (Liu, Rooker et al. 2010). SHH is an important modulator of epithelial-mesenchyme interaction (EMi) during development. SHH has been shown to regulate growth and formation of the palatal shelves prior to elevation and fusion (Rice, Connor et al. 2006). During development, SHH ligand is secreted by the epithelium into the underlying mesenchyme. This causes a gradient of signaling where mesenchyme proximal to the epithelium is exposed to higher concentrations of SHH than more distal cells (Cohen, Kicheva et al. 2015). Disruption of SHH during critical windows of development is believed to work in an EMi dependent, but epithelial-mesenchyme transition (Emt) independent manner. OFCs caused by disruption to SHH are believed to be due to a decrease in cellular proliferation and an increase in apoptosis leading to a decrease in tissue outgrowth and the failure of the facial processes to meet and fuse (Lipinski, Song et al. 2010, Heyne, Melberg et al. 2015). This increase is apoptosis is believed to be due to a decrease in GLI1/2 target gene expression.   

Pubmed was used as the primary database for evidence collection. Searches are organized by the date and search terms used in the supplementary table. Search results were initially screened through review of the title and abstract for potential for data relating GLI1 target gene expression and apoptosis. Each selected publication and its’ data were then examined to determine if support or lack thereof existed for this KER. Papers that did not show any data relating to this KER were discarded. The search terms used are organized below in Table 1.

The quantitative understanding of this relationship is low. No studies were found to exist to address dose response or time-scale data. Further work is needed to address these questions and create a better understanding of this relationship.

The relationship is biologically plausible in human, but to date no specific experiments have addressed this question. The SHH pathway is well understood to be fundamental to proper embryonic development and that aberrant SHH signaling during embryonic development can cause birth defects including orofacial clefts (OFCs). For this reason, this KER is applicable to the embryonic stage with a high level of confidence. 

Ahlgren, S. C., V. Thakur and M. Bronner-Fraser (2002). "Sonic hedgehog rescues cranial neural crest from cell death induced by ethanol exposure." Proc Natl Acad Sci U S A 99(16): 10476-10481.

Cohen, M., A. Kicheva, A. Ribeiro, R. Blassberg, K. M. Page, C. P. Barnes and J. Briscoe (2015). "Ptch1 and Gli regulate Shh signalling dynamics via multiple mechanisms." Nature Communications 6(1): 6709.

Ferguson, M. W. (1988). "Palate development." Development 103 Suppl: 41-60.

Heyne, G. W., C. G. Melberg, P. Doroodchi, K. F. Parins, H. W. Kietzman, J. L. Everson, L. J. Ansen-Wilson and R. J. Lipinski (2015). "Definition of critical periods for Hedgehog pathway antagonist-induced holoprosencephaly, cleft lip, and cleft palate." PLoS One 10(3): e0120517.

Kim, C. H., H. W. Park, K. Kim and J. H. Yoon (2004). "Early development of the nose in human embryos: a stereomicroscopic and histologic analysis." Laryngoscope 114(10): 1791-1800.

Lipinski, R. J., C. Song, K. K. Sulik, J. L. Everson, J. J. Gipp, D. Yan, W. Bushman and I. J. Rowland (2010). "Cleft lip and palate results from Hedgehog signaling antagonism in the mouse: Phenotypic characterization and clinical implications." Birth Defects Res A Clin Mol Teratol 88(4): 232-240.

Liu, B., S. M. Rooker and J. A. Helms (2010). "Molecular control of facial morphology." Semin Cell Dev Biol 21(3): 309-313.

Rice, R., E. Connor and D. P. C. Rice (2006). "Expression patterns of Hedgehog signalling pathway members during mouse palate development." Gene Expression Patterns 6(2): 206-212.

Som, P. M. and T. P. Naidich (2013). "Illustrated review of the embryology and development of the facial region, part 1: Early face and lateral nasal cavities." AJNR Am J Neuroradiol 34(12): 2233-2240.

Som, P. M. and T. P. Naidich (2014). "Illustrated review of the embryology and development of the facial region, part 2: Late development of the fetal face and changes in the face from the newborn to adulthood." AJNR Am J Neuroradiol 35(1): 10-18.

Wang, Q., H. Kurosaka, M. Kikuchi, A. Nakaya, P. A. Trainor and T. Yamashiro (2019). "Perturbed development of cranial neural crest cells in association with reduced sonic hedgehog signaling underlies the pathogenesis of retinoic-acid-induced cleft palate." Dis Model Mech 12(10).

Warbrick, J. G. (1960). "The early development of the nasal cavity and upper lip in the human embryo." J Anat 94(Pt 3): 351-362.