The H₂S-generating enzymes cystathionine β-synthase and cystathionine γ-lyase play a role in vascular development during normal lung alveolarization

Abstract

The gasotransmitter hydrogen sulfide (H2S) is emerging as a mediator of lung physiology and disease. Recent studies revealed that H2S administration limited perturbations to lung structure in experimental animal models of bronchopulmonary dysplasia (BPD), partially restoring alveolarization, limiting pulmonary hypertension, limiting inflammation, and promoting epithelial repair. No studies have addressed roles for endogenous H2S in lung development. H2S is endogenously generated by cystathionine β-synthase (Cbs) and cystathionine γ-lyase (Cth). We demonstrate here that the expression of Cbs and Cth in mouse lungs is dynamically regulated during lung alveolarization and that alveolarization is blunted in Cbs(-/-) and Cth(-/-) mouse pups, where a 50% reduction in the total number of alveoli was observed, without any impact on septal thickness. Laser-capture microdissection and immunofluorescence staining indicated that Cbs and Cth were expressed in the airway epithelium and lung vessels. Loss of Cbs and Cth led to a 100-500% increase in the muscularization of small- and medium-sized lung vessels, which was accompanied by increased vessel wall thickness, and an apparent decrease in lung vascular supply. Ablation of Cbs expression using small interfering RNA or pharmacological inhibition of Cth using propargylglycine in lung endothelial cells limited angiogenic capacity, causing a 30-40% decrease in tube length and a 50% decrease in number of tubes formed. In contrast, exogenous administration of H2S with GYY4137 promoted endothelial tube formation. These data confirm a key role for the H2S-generating enzymes Cbs and Cth in pulmonary vascular development and homeostasis and in lung alveolarization.

Functional and Molecular Insights of Hydrogen Sulfide Signaling and Protein Sulfhydration

“…Exogenous administration of H2S with GYY4137 promoted endothelial tube formation. These data confirm a key role for the H2S-generating enzymes CBS and CSE in pulmonary vascular development and homeostasis and in lung alveolarization [84]. The blood-brain barrier (BBB) integrity can be maintained by the cross-talk of endothelial cells, junction proteins, and neurogliovascular network and a recent study has shown that miR29b regulates BBB dysfunction by regulating BBB permeability through DNMT3b.…”

Stereological monitoring of mouse lung alveolarization from the early postnatal period to adulthood

“…The practical conduct of isolating, fixing, and sectioning lungs for design-based stereology has been described in extensive detail (38,44). The application and validation of this approach to neonatal mouse lungs have been described by our group (26,28,41). This approach, which is also applied in the present study to adult (mice aged up to 22 mo) mouse lungs, is based on pioneering work addressing the analysis of lung growth and lung structure in adult mice, rats, and humans (15, 35-38, 44, 51, 52).…”

“…Analyses were performed using the NewCast PLUS computerassisted stereology system (Visiopharm, Hoersholm, Denmark). Parameters analyzed included the mean linear intercept, alveolar septal wall thickness, total surface area, as well as alveolar number and alveolar density, as described previously (26,28,33…”

H2S biosynthesis and catabolism: new insights from molecular studies

“…This particular line is useful for studying the in vivo role of elevated levels of homocysteine in the aetiology of cardiovascular diseases and was developed in the lab of Dr Nobuyo Maeda at the University of North Carolina at Chapel Hill. A number of researchers have utilised this mouse model to determine the functional role of H 2 S in colitis [253] for the role of H 2 S in alveolarization [254] and in the prevention of hyperhomocysteinemia associated chronic renal failure [255], however, studies are limited primarily due to the high mortality rates in offspring. In the case of research using CSE knockout (CSE-KO) animals, this model is more widely reported in the literature.…”