Increased distension stimulates distal capillary growth as well as expression of specific angiogenesis genes in fetal mouse lungs.

Tracheal occlusion (TO) performed surgically in utero near the end of gestation causes a rapid increase in the distension of future airspaces, resulting in accelerated lung development. The authors hypothesize that TO stimulates microvascular growth concomitant with a rapid increase in the expression of genes implicated in angiogenesis. Mouse fetuses underwent in utero surgery (TO or sham-TO surgery) at 16.5 days of gestation, whereupon development was allowed to continue for a further 1 or 24 hours. Microvascular changes were assessed by immunohistochemical staining of fetal lung sections for platelet endothelial cell adhesion molecule-1. Levels of vascular endothelial growth factor-A (VEGF-A; isoforms 120, 164 and 188), VEGF receptors 1 and 2 (VEGFR-1 and -2), angiopoietins 1 and 2, and Tie2 mRNAs were determined by quantitative real-time polymerase chain reaction (PCR). The authors observed more intercapillary interconnection, less isolated capillaries, and a more extended capillary network inside septa of lungs that underwent 24 h of TO versus sham-TO. Moreover, the authors observed a significant increase in mRNA levels of VEGF 188 and VEGFR-1 as early as 1 hour following TO and of VEGFR-1 and angiopoietin 1 after 24 hours. Together, these results suggest that surgically applied stretching quickly enhances the expression of specific angiogenesis and vessel maintenance genes, which seems to result in the maturation and organization of a more extensive and complex capillary network.

Identification of cellular processes that are rapidly modulated in response to tracheal occlusion within mice lungs.

Lung development progresses through a process reliant on mechanical cell stretch. However, this process is not well defined at the molecular level. Our goal was to globally analyze the transcriptome of fetal mouse lungs following short periods of tracheal occlusion (TO) to identify cellular processes that are rapidly modulated in response to intraluminal stretch increase. Serial analysis of gene expression (SAGE) was used to examine the global transcriptomic response of mouse prealveolar stage lungs to in vivo TO at 1 and 3 h. SAGE results were extended by histo- and immunochemical examination. Based on the 97 TO-modulated transcripts identified, our results further point out that continuous stretch in developing lungs leads directly to rapid and highly specific phenotypic modifications in a significant proportion of pulmonary cells. We conclude that intraluminal stretch increase during prealveolar stage of lung development induces a critical transition of pulmonary cells phenotype in which there is an increase in alpha-smooth muscle actin (alpha-SMA)-containing cells along with a relative decrease in lipid-containing cells.

Short-term response to tracheal occlusion during perinatal lung development in mice.

Tracheal occlusion (TO) is known to stimulate lung growth. The aim of this research is to investigate early cellular responses to TO during perinatal growth in order to identify cellular targets in fetal mouse lungs that respond rapidly to surgically induced stretch. TO, or sham-TO, surgery was performed at 16.5 gestational days in fetal mice. Cellular activation, proliferation, and apoptosis were assessed on lung tissue sections harvested at various time points within the first 24 hours after the surgery. Lung tissue from unoperated fetuses and newborn mice served as controls to establish the pattern of cellular proliferation and apoptosis during normal lung development. When compared with sham-TO, TO induces a significantly higher expression of pulmonary c-Fos mRNA within 1 hour after surgery. When compared with sham-TO and unoperated controls, TO induces a rapid (1-hour) increase in proliferating cell nuclear antigen (PCNA) expression within differentiated epithelial airways. In contrast, a significant increase in the apoptotic index of mesenchymal cells from TO lungs was not observed before 24 hours when compared with sham-TO and controls. The data demonstrate that in vivo TO induces an immediate cellular response and that stretch both primarily and significantly accelerates epithelial cell proliferation and mesenchymal cell apoptosis. Moreover, we conclude that TO reduces the gestational time required to reach the natural peaks of proliferation and apoptosis associated with normal lung development, thus resulting in an apparent stimulation of lung growth.

In vivo tracheal occlusion in fetal mice induces rapid lung development without affecting surfactant protein C expression.

Fetal tracheal occlusion (TO) reverses lung hypoplasia by inducing rapid lung growth. Although increases in lung size accompanied by increased numbers of alveoli and capillaries have been reported, effects of TO on lung development have not been formally assessed. In the present study, the objective was to verify our prediction that the main effect of TO would be to accelerate fetal lung development. We have developed and characterized a new fetal mouse model of TO to best realize this goal. At embryonic day 16.5, pregnant CD1 mice were operated under general anesthesia. One fetus per dam was selected to undergo surgical TO with a surgical clip or a sham operation. The fetuses were delivered 24 or 36 h postsurgery. The maturation of lung parenchyma, evaluated by counting the generations of alveolar saccules from the terminal bronchiole to the pleura, was significantly accelerated in the TO group with a complexity of the gas exchange region comparable with postnatal days 1 and 3 after 24 or 36 h of TO. Cellular proliferation and apoptosis peaks, assessed by immunohistochemistry directed against PCNA and the active form of caspase-3, were significantly increased 24 h after surgery in the TO group compared with the sham group. However, in situ hybridization showed no significant difference in the density of type II pneumocytes expressing surfactant protein C mRNA. Our results show that brief TO during late gestation in fetal mice induces accelerated lung development with minimal effects on surfactant protein C mRNA expression.