Positive SOSTDC1 staining was pre dominantly localized in the peripheral epithelium dur ing the pseudoglandular stage and persisted into the canalicular stage. Nega tive controls for in situ hybridization in the developing human lung were shown in Figure 5M P. All the in situ hybridization data obtained in this study showed that expression of canonical WNT/ B CATENIN Dasatinib Sigma signaling components was mainly loca lized in the bronchial and alveolar epithelium in em bryonic human lung tissues, although some of the components were found to be expressed at low levels in the surrounding mesenchyme. Moreover, the expres sion patterns of canonical WNT/B CATENIN signaling components detected by in situ hybridization were found to be in accordance with those detected by qRT PCR.
Activity of canonical WNT/B CATENIN signals in the developing human lung To further assess canonical WNT signaling activities in the developing human lung, CHIR 99021 was used to activate WNT/B CATENIN signaling cascades through in vitro exposure of human lung Inhibitors,Modulators,Libraries explants to 0, 5 and 10 uM CHIR 99021 for 72 h. As pre sented in Figure 6A and B, Western blot analysis showed B CATENIN expression in 15 W human lung explants decreased significantly from newly isolated to in vitro cultured for 72 h. However, B CATENIN expres sion Inhibitors,Modulators,Libraries nearly increased back to newly isolated levels in human lung explants at 15 W treated with 5 uM CHIR 99021 for 72 h, but decreased again after treatment of 10 uM CHIR 99021.
Meanwhile, Inhibitors,Modulators,Libraries qRT PCR analyses showed increased expression of transcription factors and target genes in human lung explants exposed to 5 uM CHIR 99021 compared with tissues exposed to 0 uM CHIR 99021, the levels of which were also higher than in the 10 uM CHIR 99021 groups. We then performed histological analysis. The human lung explants cultured in vitro for 72 h showed enlarged lung tubes and differentiation of Inhibitors,Modulators,Libraries the airway epi thelial cells around the tubes from short columnar to cu boidal morphology. However, parallel explant tissues treated with 5 uM CHIR 99021 for 72 h differentiated back to short columnar cells, with slighter changes between cu boidal and short columnar shape observed in the groups exposed to 5 uM CHIR 99021. In contrast, no obvious differences were observed in the amount of lung tubes formed in the control and CHIR 99021 treated explant tissues.
Discussion Crucial roles of canonical WNT/B CATENIN signaling components in the developing human lung This Inhibitors,Modulators,Libraries study described the mRNA Bicalutamide expression of canonical WNT/B CATENIN signaling components at the tissue level during human lung development at 7 W, 12 W, 17 W and 21 W. Most of the canonical WNT signaling components were detected at 7 W and increased to high levels at 17 W followed by a decrease at 21 W using quantitative real time qRT PCR.