Transmural pressure is a master regulator of airway branching morphogenesis

Mechanical forces are increasingly recognized to regulate morphogenesis, but how this is accomplished in the context of the multiple tissues present within a developing organ remains unclear. Here we use bioengineered "microfluidic chest cavities" to precisely control the mechanical environment of the fetal lung. We show that transmural pressure controls airway branching morphogenesis and regulates the frequency of airway smooth muscle contraction. Next-generation sequencing analysis shows that lungs held at higher pressure are more mature than lungs held at lower pressure. Timelapse imaging reveals that branching events are synchronized across distant locations within the lung, and are preceded by long-duration waves of airway smooth muscle contraction. Higher transmural pressure decreases the interval between systemic smooth muscle contractions and increases the rate of morphogenesis of the airway epithelium. These data reveal that the mechanical properties of the microenvironment instruct crosstalk between tissues to control the rate of development of the embryonic lung. (i) embryonic mouse lungs at E12.5 were cultured under low or high pressure for 48 hours prior to RNA extraction or (ii) embryonic mouse lungs were isolated from pregnant mice at E12.5, E13.5 and E14.5 prior to RNA extraction机械力对形态发生的调控作用已日益受到学界重视，但在发育器官内存在多种组织的情境下，这一调控过程的具体实现机制仍未明确。本研究借助生物工程构建的"微流控胸腔（microfluidic chest cavities）"，实现对胎肺机械微环境的精准操控。研究证实，跨壁压（transmural pressure）可调控气道分支形态发生（airway branching morphogenesis），并调节气道平滑肌收缩（airway smooth muscle contraction）的频率。下一代测序（next-generation sequencing）分析显示，处于较高跨壁压环境的肺脏，其成熟度高于低压力组的肺脏。延时成像（timelapse imaging）结果表明，肺内不同远端位点的分支事件存在同步性，且该过程此前会出现持续时间较长的气道平滑肌收缩波。更高的跨壁压会缩短全身性平滑肌收缩的间隔时长，并加快气道上皮（airway epithelium）的形态发生速率。上述研究结果表明，微环境的机械特性可通过调控组织间串扰（crosstalk between tissues），进而控制胚胎肺的发育速率。(i) 于胚胎发育第12.5天（E12.5）获取的小鼠胚胎肺脏，在低压力或高压力环境下培养48小时后再进行RNA提取；(ii) 于妊娠小鼠的胚胎发育第12.5天、13.5天及14.5天分离小鼠胚胎肺脏，随后进行RNA提取