Piña Jeremie Oliver, Raju Resmi, Stipano Evan, Myo Aye Chan, Wang Ziyi, Ono Mitsuaki, Chattaraj Parna, Furukawa Masae, D'Souza Rena N
Section on Craniofacial Genetic Disorders, Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD), National Institutes of Health (NIH), Bethesda, MD, USA.
Graduate School of Medicine Dentistry and Pharmaceutical Sciences, Department of Molecular Biology and Biochemistry, Okayama University, Okayama, Japan.
Sci Rep. 2025 Aug 13;15(1):29639. doi: 10.1038/s41598-025-14807-1.
Despite advances in understanding the morphological disruptions that lead to defects in palate formation, the precise perturbations within the signaling microenvironment of palatal clefts remain poorly understood. To explore in greater depth the genomic basis of palatal clefts, we designed and implemented the first single cell spatial RNA-sequencing study in a cleft palate model, utilizing the Pax9 murine model at multiple developmental timepoints, which exhibits a consistent cleft palate defect. Visium HD, an emerging platform for true single-cell resolution spatially resolved transcriptomics, was employed using custom bins of 2 × 2 μm spatial gene expression data. Validation of spatial gene expression was then validated using custom designed Xenium In Situ mRNA spatial profiling and RNAscope Multiplex assays. Functional enrichment analysis revealed a palate cell-specific perturbation in Wnt signaling effector function in tandem with disrupted expression of extracellular matrix genes in developing mesenchyme. As a key step toward laying the framework for identifying key molecular targets these data can be used for translational studies aimed at developing effective therapies for human palatal clefts.
尽管在理解导致腭裂形成的形态学破坏方面取得了进展，但腭裂信号微环境内的确切扰动仍知之甚少。为了更深入地探索腭裂的基因组基础，我们在腭裂模型中设计并实施了第一项单细胞空间RNA测序研究，在多个发育时间点利用Pax9小鼠模型，该模型表现出一致的腭裂缺陷。Visium HD是一个新兴的用于真正单细胞分辨率空间分辨转录组学的平台，使用2×2μm空间基因表达数据的自定义区域进行研究。然后使用定制设计的Xenium原位mRNA空间分析和RNAscope多重分析对空间基因表达进行验证。功能富集分析揭示了腭细胞特异性的Wnt信号效应功能扰动，同时发育中的间充质细胞外基质基因表达也受到破坏。作为为确定关键分子靶点奠定框架的关键一步，这些数据可用于旨在开发治疗人类腭裂有效疗法的转化研究。
