5 to 17. five is usually observed in cell cultures. Levels of Crh, Crhbp, Crhr1, Crhr2b, Pomc, Mc2r, and Nr3c1 mRNAs had been compared involving fibroblast enriched cell cultures, epithelial cell enriched cell cultures, and total lungs. Fetal sex was not considered in these experiments for the reason that only Crhbp showed a sex difference in expression by QPCR in complete lungs, whereas no sex difference was observed for any of these genes by in situ hybridization. Messenger RNA levels of Krt18 and Vim, which are epithelial and mesenchymal markers, respectively, confirmed cell enrichments. For every analyzed gene, ratios corresponding to mRNA levels in epithelial cell enriched cultures over these in fibroblast enriched cultures are presented in Figure 4D. Interestingly, Crh mRNA was not or barely detected in cell cultures.
On GD 15. five and 17. 5, Crhr1 mRNA was detected in mesenchymal cells, whereas the transcript was not or barely detected in epithelial cells. On selleck chemicals GD 15. 5, Crhr2b expression level was greater in mesenchymal cells than in epithelial cells, even though on GD 17. five the opposite predicament was observed. Crhr2b mRNA was not detected in all samples. Crhr1 and Crhr2b mRNA levels had been reduce in cell cultures than in whole fetal lungs, similarly to Crh. The expression of Crhbp was higher in epithe lial cells than in fibroblasts on GD 15. five and 17. five, even though Pomc, Mc2r, and Nr3c1 are preferentially expressed in fibroblasts on GD 15. 5 and in epithelial cells on GD 17. 5. Expression levels of these last four genes have been similar between cell cultures and total lung pools.
Expression of Cyp21a1 and Cyp11b1 in fetal mouse lungs and fetal lung major cell cultures Cyp21a1 and Cyp11b1 encode the final two steroidogenic enzymes involved in de novo corticosterone synthesis. Cyp11b1 expression was previously observed on GD 15. five, but not on GD 16. 5, 17. 5, and 18. 5 inside the fetal lung. There was no obtainable data on expression of Cyp21a1 CP724714 soon after GD 15. five. We detected Cyp21a1 mRNA in fetal lung pools and in lung explants obtained on GD 17. 5. Interestingly, Cyp21a1 mRNA was detected at a great deal greater levels in main cell cul tures than in non incubated whole lung samples and lung explants. Indeed, Cyp21a1 mRNA levels had been 15 and 40 fold higher in fibroblast and in epithelial cell enriched cultures, respectively, than in lung explants, and were larger in epithelial fractions than in fibroblast fractions on each GD 15.
5 and 17. five. In agreement with preceding data, Cyp11b1 mRNA was not detected in lung explants obtained on GD 17. five within the present study. On the other hand, Cyp11b1 mRNA was surprisingly detected at relatively high levels in cell cultures, as Cyp21a1. The potential effect of reciprocal exposure of epithelial and mesenchymal enriched cell cultures to their respec tive secreted factors on gene expression of Crh, Crhr1, Crhr2b, Cyp21a1, and Cyp11b1 was addressed.