American journal of physiology. Lung cellular and molecular physiology最新文献

Lung fibroblasts generate and respond to mechanical, biochemical, and matrix cues present in their microenvironment. With the advent of next-generation sequencing technologies, multiple studies describe transcriptionally unique fibroblast subpopulations in the human lung. However, limited published data suggest a loss of fibroblast native phenotypes and functions after culture ex vivo. In this study, we characterized changes in transcriptional programs of human lung mesenchyme isolated from freshly procured tissue and maintained in traditional cell culture conditions. Our results demonstrate that fibroblasts isolated and cultured in this manner adopt transcriptional programs largely distinct from those observed in vivo. To recapitulate distinct native fibroblast states in vitro, we sought to develop a screening approach to identify cues promoting native fibroblast identities. From published single-cell data, we identified unique transcriptional markers of alveolar and adventitial fibroblast subpopulations and validated the sensitivity of ELISAs for detecting changes in secreted markers of these fibroblast subpopulations. We then stimulated primary human lung fibroblasts with soluble cues known to act on fibroblasts, quantifying changes in secreted and transcriptional markers by ELISA and qPCR. Although our small pilot screen did not identify single cues capable of fully recapitulating fibroblast in vivo states, it established a system that can be expanded to broadly screen additional cues and pointed toward factors likely to be critical in developing better culture models for studying human lung fibroblast function and plasticity.NEW & NOTEWORTHY Recent studies highlight transcriptionally distinct fibroblast subpopulations in human lungs. We observed the loss of these native transcriptional programs as fresh isolated cells are maintained in traditional culture conditions. Identifying the signals defining native fibroblast identities will be pivotal to creating culture models that preserve unique subpopulations. The screening system developed here will allow the investigation of a broad selection of cues, leading to better culture models for studying human lung fibroblast function and plasticity.

The pulmonary alveolar-capillary niche is a highly specialized interface that balances gas exchange with maintenance functions and repair. Advances in single cell transcriptomics have uncovered endothelial heterogeneity, which underlies developmental angiogenesis and plastic responses to injury. Emerging evidence from a neonatal hyperoxia model highlights CAP1 to CAP2 transitions and the role of p53 in maintaining lineage fidelity. Beyond intrinsic lineage plasticity, circulating mediators such as cell-free hemoglobin drive endothelial barrier disruption through oxidative injury and lipid modification. As new signaling pathways and therapeutics targets emerge, complementary strategies are being developed at the cellular level, including adoptive transfer of mesenchymal stromal and immune cells, although mechanisms of endothelial adhesion and homing remain incompletely defined. Finally, biomechanical forces such as shear stress have become critical contextual cues for endothelial signaling, yet remain underrepresented in some experimental models. Together, these insights underscore the central role of endothelial heterogeneity, injury responses, and environmental cues in shaping pulmonary vascular health and repair, with implications for designing targeted therapies in both pediatric and adult lung disease.

Trikafta (elexacaftor/tezacaftor/ivacaftor; ETI) is approved for cystic fibrosis (CF) patients with at least one F508del mutation in the cystic fibrosis transmembrane conductance regulator (CFTR) gene or another responsive mutation based on in vitro data. However, the pharmacological effects of ETI on F508del-CFTR remain incompletely defined in vitro. To explore the mechanisms underlying Trikafta's clinical efficacy, we used primary bronchial epithelial cells from F508del homozygous patients and CFBE41o-cells expressing F508del-CFTR. We assessed CFTR maturation, turnover, chloride transport, and thermal stability under various ETI concentrations and treatment durations at physiological temperature using electrophysiology (Ussing chamber, patch-clamp) and biochemical assays. We found that ETI efficacy on F508del-CFTR is strongly influenced by both treatment duration and concentration. Reducing ETI from standard doses, i.e., E (3 µM), T (18 µM), and I (1 µM), to 33%, 11%, 3.3%, and 1.1% decreased function and maturation, but 33% retained most of the corrective effect. After 2 h of treatment, around 50% of the CFTR-dependent current was preserved, unlike in untreated cells. Notably, replacing elexacaftor with bamocaftor further improved F508del-CFTR maturation and function compared with ETI, though it did not affect the rate of current decline over time. These findings highlight the importance of optimizing ETI dose and exposure duration, as both significantly affect F508del-CFTR stability and function. The retained efficacy at reduced concentrations suggests possible individualized dosing strategies, particularly for patients experiencing adverse effects with full-dose ETI.NEW & NOTEWORTHY Our in vitro study underscores that ETI/BTI's efficacy in improving F508del-CFTR function depends on treatment concentration and duration, impacting the protein's metabolic and thermal stability. Although ETI/BTI only partially addresses F508del-CFTR's inherent thermal instability, reduced doses retained significant effectiveness. This finding supports dose optimization as a promising strategy to sustain therapeutic benefits while minimizing side effects, offering a personalized approach to treatment for individuals with cystic fibrosis experiencing adverse effects from standard dosing.

Sex appears to modulate interactions between neural mechanisms involved in regulating pulmonary ventilation during mild hypoxic exercise. Therefore, we compared pulmonary ventilation responses elicited by isolated and combined stimulation of the carotid chemoreflex and muscle mechanoreflex between males and females. Twenty-eight healthy adults (14 females) underwent four experimental manipulations: 1) normoxic rest (no stimulation), 2) hypoxic rest (carotid chemoreflex stimulation), 3) normoxic passive movement (muscle mechanoreflex stimulation), and 4) hypoxic passive movement (reflexes costimulation). Isocapnia was maintained using a rebreathing system, and hypoxia was induced by breathing 12% oxygen. Passive movement involved 30-s bouts of unilateral knee manipulation at 300°/s, with surface electromyography confirming absence of voluntary muscle contractions. In males, the pulmonary ventilation response to passive limb movement (last 10 s change vs. rest) was greater under hypoxia than normoxia (means ± SD: hypoxia = 3.6 ± 2.0 vs. normoxia = 1.6 ± 2.4 L/min; P = 0.003), whereas no difference was observed in females (hypoxia = 1.9 ± 2.4 vs. normoxia = 2.2 ± 1.5 L/min; P = 1.000). Moreover, pulmonary ventilation remained elevated in males (hypoxia = 2.7 ± 2.4 vs. normoxia = -0.1 ± 2.2; P < 0.001) but not in females (hypoxia = 0.4 ± 3.3 vs. normoxia = 0.5 ± 1.5; P = 1.000), 30 s following passive limb movement under hypoxia. These findings support a synergistic carotid chemoreflex-muscle mechanoreflex interaction in males but not in females. The persistent ventilatory elevation poststimulation indicates that short-term potentiation contributes to this synergistic interaction in males.NEW & NOTEWORTHY Pulmonary ventilation response to passive limb movement is greater under hypoxia than normoxia in males but not in females. These results support a synergistic interaction between the carotid chemoreflex and muscle mechanoreflex in males but not in females. In addition, pulmonary ventilation remains elevated in males but not in females after the cessation of passive limb movement under hypoxia, suggesting that short-term potentiation may be a mechanism mediating this synergistic reflex interaction in males.

Intravenous bolus (ivb) injection of fentanyl triggers a vagal-mediated immediate apnea and subsequent respiratory depression in anesthetized rats. This study compared the gender-dependence of these responses in conscious rats and roles of peripheral and central opioid receptors (ORs), especially µ- and mu1 opioid receptor (µ₁-ORs) in the genesis of these responses. Cardiorespiratory responses to ivb injection of fentanyl (50 µg·kg^-1) were recorded in male and female conscious rats (study I). The same protocols were performed after: naloxone (NLX) and naloxone methiodide (NLM) to systemically and peripherally antagonize ORs, respectively (study II); D-Phe-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH2 (CTAP) and methylnaltrexone (MNTX) to systemically and peripherally block opioid mu-receptors (study III); and naloxonazine (NLZ) to systemically block µ₁-ORs (study IV). ivb injection of fentanyl induced an immediate life-threatening apnea (∼1.5 min) and severe bradycardia, which was followed by cardiorespiratory depression lasting for ∼55 min with little difference between genders. NLX fully eliminated and CTAP substantially blunted all cardiorespiratory responses to fentanyl, whereas NLM and MNTX substantially minimized the immediate apnea and reduced bradycardia by ∼50% with limited impact on the subsequent cardiorespiratory depression. NLZ nearly abolished the fentanyl-evoked responses. Our results indicate that peripherally restricted OR (particularly µ₁-OR) antagonism prevents the fentanyl-induced immediate apnea, but fails to change the subsequent respiratory depression.NEW & NOTEWORTHY The cardiorespiratory responses to rapid intravenous injection of fentanyl have not been fully investigated. We demonstrate in this study that intravenous bolus injection of fentanyl triggers an immediate sustained apnea and subsequent respiratory depression without remarkable gender-difference in conscious rats. The immediate apnea is triggered by activating peripheral opioid receptors and the subsequent respiratory depression is mediated by activating central opioid receptors, in which µ1-opioid receptors play a key role.

Acute lung injury (ALI)/acute respiratory distress syndrome (ARDS) are inflammatory conditions with substantial rates of morbidity and mortality, but no effective treatments. The lack of effective treatments and unacceptably high mortality rates for ARDS are partly due to an incomplete understanding of the mechanisms that control ALI/ARDS and subsequent vascular repair. Transforming growth factors (TGFs) are a class of growth factors that regulate the vascular response to inflammation, including migration, proliferation, and differentiation of cells comprising the lung vasculature. Here we review studies that describe the impact of the TGF family on inflammatory lung injury and subsequent vascular repair and fibrosis. We highlight gaps in understanding TGF isoform-specific roles in ALI/ARDS and outline directions for future research in the field of TGF-dependent regulation of inflammatory lung injury and vascular repair. Functional roles of the TGFs have been investigated in ALI/ARDS pathogenesis and pulmonary fibrosis, with a predominance for studies showing a proinjurious and profibrotic impact of TGF-β1. Studies have also shown that TGF-α is positively associated with inflammatory lung injury and fibrosis. However, the contributions of TGF-β2 and TGF-β3 to ALI/ARDS are unclear, and the contributions of all the TGF isoforms to vascular repair after ALI/ARDS are not well characterized. Improved understanding of the regulation of inflammatory lung injury and repair by the TGFs could lead to the development of a safe and effective treatment strategy for patients with ALI/ARDS.

Invasive mechanical ventilation is a lifesaving intervention for patients with acute respiratory distress syndrome (ARDS) but it also causes ventilator-induced lung injury (VILI) that can be challenging to avoid due to interpatient and temporal heterogeneity. Thus, the aim of this study was to characterize and predict experimental VILI using readily available measures of lung function. Initially healthy (CTL) and hydrochloric acid (HCL) lung-injured mice were ventilated for 4 h at positive end-expiratory pressure (PEEP) 1, 3, or 8 cmH₂O to produce graded VILI severity as measured in lung function, alveolocapillary leak, and inflammation. Optimally protective PEEP was found to be 8 and 3 cmH₂O in the HCL and CTL groups, respectively. A novel computational model was fit to the data to investigate elastance dynamics described by the "compliance factor" (C_F), which was also used to predict VILI over 4 subsequent hours of ventilation. The model C_F is a sensitive measure of injury-induced alterations in the pressure and pressure history dependence of lung elastance that are known to correlate with recruitment and derecruitment dynamics. The C_F was then combined with PEEP and plateau pressures calculated from 10 min at the start of prolonged ventilation and used to accurately predict VILI outcomes measured 4 h later. This model outperformed other commonly used measures of injury such as driving pressure and mechanical power. The computer model provides a new tool for understanding lung dynamics and for predicting VILI. In future work, this approach could be used to guide identification of lung-protective ventilation settings.NEW & NOTEWORTHY Computer model-based analysis of lung function in healthy and lung-injured mice showed that model compliance factor (C_F) characteristics were sensitive measures of acute lung injury and ventilator-induced lung injury (VILI) severity. The [Formula: see text] Area, calculated from C_F and pressures from minutes 5-15 of ventilation, was a stronger predictor of VILI measured 4 h later than the driving pressure or mechanical power, suggesting potential utility for monitoring ventilation safety and guiding ventilator adjustments to reduce VILI.

Pulmonary fibrosis (PF) is a severe consequence of respiratory infections, characterized by excessive extracellular matrix deposition and irreversible lung architectural damage. Once considered a rare condition, PF is now increasingly recognized in the wake of viral infections, particularly among survivors of viral-induced acute respiratory distress syndrome (ARDS). The COVID-19 pandemic has highlighted in bold relief the observation that many survivors of severe viral pneumonia do not recover fully but develop chronic fibrotic changes that impair lung function. This review examines the clinical evidence and underlying mechanisms linking viral infections-COVID-19, influenza, and other respiratory viruses-to the onset of pulmonary fibrosis. By probing the mechanisms of cellular injury, immune dysregulation, and aberrant repair mechanisms, we aim to illuminate the pathways that transform an acute viral insult into a chronic, fibrotic disease.

Session III of the inaugural biennial Research Symposium on Pulmonary Injury and Repair of the Endothelium (ReSPIRE) highlighted key advancements in endothelial-inflammatory cell interactions. The work presented illustrates a growing recognition that pulmonary endothelial cell interactions and direct cross talk with inflammatory cells are integral in both health and disease in the developing and aging lung. Data presented detail the impact of targeting neutrophil- and macrophage-endothelial interactions in acute lung injury, and the role of fibroblast-endothelial inflammatory communication in interstitial pulmonary fibrosis of the aging lung. In the developing lung, the paradoxical responses of the pulmonary circulation to inflammatory cell interactions and mediators illustrate the complexities in cross talk. State-of-the-art advances in intravital microscopy have recently revealed our ability to visualize and measure the mechanotransduction involved in neutrophil migration. This review highlights these recent advances and suggests future directions for understanding pulmonary endothelial-inflammatory cell cross talk.