Dynamic mRNA Expression Analysis of the Secondary Palatal Morphogenesis in Miniature Pigs

Abstract

Normal mammalian palatogenesis is a complex process that requires the occurrence of a tightly regulated series of specific and sequentially regulated cellular events. Cleft lip/palate (CLP), the most frequent craniofacial malformation birth defects, may occur if any of these events undergo abnormal interference. Such defects not only affect the patients, but also pose a financial risk for the families. In our recent study, the miniature pig was shown to be a valuable alternative large animal model for exploring human palate development by histology. However, few reports exist in the literature to document gene expression and function during swine palatogenesis. To better understand the genetic regulation of palate development, an mRNA expression profiling analysis was performed on miniature pigs, Sus scrofa. Five key developmental stages of miniature pigs from embryonic days (E) 30–50 were selected for transcriptome sequencing. Gene expression profiles in different palate development stages of miniature pigs were identified. Nine hundred twenty significant differentially expressed genes were identified, and the functional characteristics of these genes were determined by gene ontology (GO) function and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. Some of these genes were associated with HH (hedgehog), WNT (wingless-type mouse mammary tumor virus integration site family), and MAPK (mitogen-activated protein kinase) signaling, etc., which were shown in the literature to affect palate development, while some genes, such as HIP (hedgehog interacting protein), WNT16, MAPK10, and LAMC2 (laminin subunit gamma 2), were additions to the current understanding of palate development. The present study provided a comprehensive analysis for understanding the dynamic gene regulation during palate development and provided potential ideas and resources to further study normal palate development and the etiology of cleft palate.

Transcriptomic analysis of the upper lip and primary palate development in mice

“…Signaling pathways play important roles in multi-organ development and congenital malformations. The Wnt, MAPK, Hippo, and TGF-beta signaling pathways are involved in the development of the lip and/or palate and CL/P (Zhu et al, 2012;Liu et al, 2019a;Reynolds et al, 2019). In our results, the PI3K-Akt and Hippo signaling pathways also intensified from E10.5 to E12.5.…”

“…Further analysis of the signal-gene network revealed highly upregulated DEGs associated with these significant pathways, including Tgfb2, Wnt16, Fgf18, Wnt3a, Col6a1, and Inhba, providing a series of candidate genes for normal development or CL/P. Among these genes, Tgfb2, Wnt16, and Inhba are important for palatogenesis (Zhu et al, 2012;Liu et al, 2019a). Fgf18 and Wnt3a were associated with non-syndromic cleft lip with or without palate (NSCL/P) (Riley et al, 2007;Reynolds et al, 2019).…”

“…Excessive intake of retinoic acid (RA) could cause cleft palate and PI3K-Akt signaling was related to the action of RA, implying the possible influence of PI3K-Akt signaling on palate development (Hu et al, 2013). MAPK signaling affected palate development in animal studies (Liu et al, 2019a). Differences in DNA methylation have been reported in many candidate genes contributing to non-syndromic cleft upper lip with or without cleft palate (NSCL/ P), meanwhile, common differential methylation was also found in genes correlated with the Hippo signaling pathway, indicating an association between this pathway and NSCL/P (Young et al, 2021).…”