High-Altitude Hypoxic Preconditioning Attenuates Lipopolysaccharide-Induced Lung Injury and is Associated with...

High-altitude hypoxic preconditioning (HAP) is an adaptive state induced by sustained exposure to hypobaric hypoxia. In addition to improving tolerance to hypoxia itself, HAP may also modify organ responses to subsequent inflammatory stress. The alveolar-capillary barrier is essential for maintaining pulmonary structural integrity, fluid balance, and gas exchange, and its disruption is a central event in inflammatory lung injury. However, whether HAP-related pulmonary protection is associated with preservation of the alveolar-capillary barrier and coordinated changes in barrier-related cells remains unclear. This study aimed to investigate the effects of HAP on alveolar-capillary barrier preservation under lipopolysaccharide (LPS) challenge and to explore the associated cellular transcriptional characteristics.

A rat model of HAP was established by exposure to hypobaric hypoxia for 4 weeks. Rats were assigned to four groups: normoxic control (C), normoxia+LPS (CL), HAP control (H), and HAP+LPS (HL). Gross lung appearance, lung dry/wet ratio, histopathology, inflammatory and oxidative stress markers, and arterial blood gas parameters were evaluated. Single-cell RNA sequencing was performed in the CL and HL groups to assess transcriptional changes in endothelial cells, epithelial cells, and macrophages. Transmission electron microscopy and immunofluorescence staining for VE-cadherin and E-cadherin were used to evaluate barrier ultrastructure and junctional integrity.

Compared with the CL group, the HL group showed reduced pulmonary edema, less exudation and congestion, milder histological damage, and better oxygenation under LPS challenge. Lactate elevation was attenuated and metabolic changes were less pronounced in the HL group. Single-cell transcriptomic analysis showed distinct transcriptional characteristics in barrier-related cells after HAP. Endothelial cells in the HL group exhibited increased expression of genes related to cell junctions and cytoskeletal organization, epithelial cells exhibited upregulated expression of genes associated with cell membrane structure, cell adhesion, and alveolar fluid clearance, accompanied by an increased proportion of AT1 cells and a decreased proportion of tuft cells. Macrophages tended to express more repair-related genes. In contrast, the CL group showed more stress-, injury-, inflammation-, and interferon-related transcriptional features. Ultrastructural examination demonstrated better preservation of endothelial and alveolar epithelial integrity in the HL group. Immunofluorescence analysis further showed stronger and more continuous VE-cadherin and E-cadherin signals in the HL group than in the CL group.

HAP was associated with better preservation of the alveolar-capillary barrier under LPS challenge, as reflected by reduced pulmonary edema, improved oxygenation, stronger junctional protein signals, and better preserved barrier structure. This barrier-protective effect was accompanied by coordinated transcriptional changes in endothelial cells, epithelial cells, and macrophages.