5/20/2023 0 Comments Clc genomics workbench dukeTo identify any differences in the spatial distribution of cell types in the dorsal versus ventral trachea, we compared the confocal images of epithelial cells, smooth muscles and neurons in whole-mount samples and the stereomicroscopic images of vascular networks and performed immunohistochemical analysis of BCs in tissue sections ( Fig. 2A-G). These results suggest that there are intrinsic differences in the stem cell behaviour of dorsal versus ventral BCs, or differences in the stem cell environment in the two domains. Long-term lineage tracing of BCs also showed that the large clones were located in the dorsal trachea between the cartilage and smooth muscles ( Fig. S1B). The proportion of clones in the dorsal trachea was also higher than that in the ventral trachea (72% and 28%, respectively, Fig. 1D). The results showed that, on an average, the clones in the dorsal domain (average of 4.03 cells/clone) were larger than those in the ventral domain (average of 2.61 cells/clone, Fig. 1C). In addition to the airway epithelial cells, a few myoepithelial cells were lineage-labelled after 1 year without injury ( Fig. S1A), suggesting that some of the clones in the ventral trachea could be derived from myoepithelial cells present in the SMGs. Two weeks after the treatment, we determined the number and sizes of individual clones of cells labelled with a nuclear green fluorescent protein (GFP) reporter on the dorsal half versus the ventral half of the trachea ( Fig. 1B). To determine whether there is heterogeneity in the reparative behaviour of BCs in the dorsal versus ventral trachea, we performed clonal lineage tracing of KRT5-CreER T2 Rosa-Confetti mice ( Fig. 1A). In addition, a recent study has revealed that the loss of Wnt secretion in epithelial cells affects cartilage formation from mesenchymal cells and results in the loss of FGF10 which affects the differentiation of airway epithelial cells, including BCs ( Hou et al., 2019).īrief treatment of mice with sulphur dioxide gas has been observed to cause the death of luminal cells in the trachea, and was closely followed by the proliferation of surviving BCs and their differentiation into ciliated and secretory cells ( Rock et al., 2009). Genetic disruption of cartilage or smooth muscle formation affects BC composition and differentiation in tracheal epithelium, which indicates the importance of the mesenchyme for the normal development of the tracheal epithelium ( Hines et al., 2013). Wnt5a-Ror2 signalling in mesenchymal cells regulates the elongation of the trachea through the polarisation of smooth muscle progenitors. For example, a recent study demonstrated that the mesenchyme regulates elongation along the anterior-posterior axis and the expansion of the developing trachea ( Kishimoto et al., 2018 Young et al., 2020). There is increasing evidence that supports the existence of reciprocal epithelial-mesenchymal interactions in the development of the mouse trachea. With respect to the development of airway epithelium, TRP63-positive BCs and SCGB3A2-positive secretory cells were observed in NKX2-1-positive airway epithelium from E10.5 ( Kurotani et al., 2008 Hou et al., 2019), whereas FOXJ1-positive ciliated cells were observed to form from E14.5 ( Tsao et al., 2009). Cartilage segmentation begins from E13.5 ( Young et al., 2020). Cartilage development was observed to commence at E10.5, characterised by the expression of SOX9-positive cells, whereas smooth muscle development commenced at E11.5, characterised by the expression of ACTA2 ( Hines et al., 2013). The results support the hypothesis that BCs from the dorso-ventral airways have intrinsic molecular and behavioural differences relevant to their in vivo function.ĭuring mouse development, the tracheal primordium was observed to form at E9.5 and continued to be elongated throughout the tracheal development period. Significantly, ventral BCs express Myostatin, which inhibits the growth of smooth muscle cells, and HGF, which facilitates cartilage repair. Gene ontology analysis revealed that genes expressed in dorsal BCs are enriched in wound healing while ventral BCs are enriched in response to external stimulus and immune response. Three-dimensional organoid culture of BCs demonstrated that dorsal BCs show higher colony forming efficacy to ventral BCs. Large clones were found between cartilage and smooth muscle where subpopulation of dorsal BCs exists. Lineage tracing using Krt5-CreER shows dorsal BCs produce more, larger, clones than ventral BCs. The airway is surrounded by cartilage ventrolaterally and smooth muscle dorsally. The tracheal basal cells (BCs) function as stem cells to maintain the epithelium in steady state and repair it after injury.
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