Cardiovascular Research Center of the General Medical Services, Massachusetts General Hospital, Boston, Massachusetts.
Harvard Medical School, Harvard University, Cambridge, Massachusetts.
Am J Physiol Lung Cell Mol Physiol. 2022 Feb 1;322(2):L204-L223. doi: 10.1152/ajplung.00158.2021. Epub 2021 Dec 8.
During newborn lung injury, excessive activity of lysyl oxidases (LOXs) disrupts extracellular matrix (ECM) formation. Previous studies indicate that TGFβ activation in the O-injured mouse pup lung increases lysyl oxidase (LOX) expression. But how TGFβ regulates this, and whether the LOXs generate excess pulmonary aldehydes are unknown. First, we determined that O-mediated lung injury increases LOX protein expression in TGFβ-stimulated pup lung interstitial fibroblasts. This regulation appeared to be direct; this is because TGFβ treatment also increased LOX protein expression in isolated pup lung fibroblasts. Then using a fibroblast cell line, we determined that TGFβ stimulates expression at a transcriptional level via Smad2/3-dependent signaling. is translated as a pro-protein that requires secretion and extracellular cleavage before assuming amine oxidase activity and, in some cells, reuptake with nuclear localization. We found that pro-LOX is processed in the newborn mouse pup lung. Also, O-mediated injury was determined to increase pro-LOX secretion and nuclear LOX immunoreactivity particularly in areas populated with interstitial fibroblasts and exhibiting malformed ECM. Then, using molecular probes, we detected increased aldehyde levels in vivo in O-injured pup lungs, which mapped to areas of increased pro-LOX secretion in lung sections. Increased activity of LOXs plays a critical role in the aldehyde generation; an inhibitor of LOXs prevented the elevation of aldehydes in the O-injured pup lung. These results reveal new mechanisms of TGFβ and LOX in newborn lung disease and suggest that aldehyde-reactive probes might have utility in sensing the activation of LOXs in vivo during lung injury.
在新生儿肺损伤中，赖氨酰氧化酶（LOXs）的过度活跃会破坏细胞外基质（ECM）的形成。先前的研究表明，TGFβ在 O 损伤的小鼠肺中的激活会增加赖氨酰氧化酶（LOX）的表达。但是，TGFβ如何调节这一点，以及 LOX 是否会产生过多的肺醛，目前尚不清楚。首先，我们确定 O 介导的肺损伤会增加 TGFβ刺激的幼鼠肺间质成纤维细胞中的 LOX 蛋白表达。这种调节似乎是直接的；这是因为 TGFβ 处理还会增加分离的幼鼠肺成纤维细胞中的 LOX 蛋白表达。然后，我们使用成纤维细胞系确定，TGFβ通过 Smad2/3 依赖性信号转导刺激转录水平上的 表达。 被翻译为前蛋白，需要分泌和细胞外切割，然后才能发挥胺氧化酶活性，并在某些细胞中，再摄取并定位于核内。我们发现前 LOX 在新生小鼠肺中被加工。此外，我们发现 O 介导的损伤会增加前 LOX 的分泌和核 LOX 免疫反应性，特别是在富含间质成纤维细胞并表现出 ECM 畸形的区域。然后，我们使用分子探针在体内检测到 O 损伤幼鼠肺中的醛水平升高，这些区域与肺切片中前 LOX 分泌增加的区域相对应。LOXs 的活性增加在醛的生成中起着关键作用；LOXs 的抑制剂可防止 O 损伤幼鼠肺中醛的升高。这些结果揭示了 TGFβ 和 LOX 在新生儿肺疾病中的新机制，并表明醛反应性探针可能在肺损伤过程中用于体内检测 LOXs 的激活。
