Effects of delayed inhalation therapy on apoptosis-related proteins and late remodeling after mixed-insult lung injury in rats

Abstract Introduction: Acute lung injury (ALI) remains a critical condition associated with high mortality despite advances in intensive care. Its pathogenesis involves diffuse bronchoalveolar damage that frequently progresses to fibrotic remodeling of lung tissue. Dysregulated apoptosis of pulmonary parenchymal cells represents a key mechanism linking inflammation to fibrosis and is largely controlled by the balance between proapoptotic and antiapoptotic Bcl-2 family proteins, including Bax and Bcl-xL, as well as activation of caspase-3. Modulation of these pathways represents a promising therapeutic strategy. Aims To evaluate the effects of inhalation therapy with camostat mesylate, methylprednisolone, and enoxaparin sodium on the tissue content of apoptosis-related proteins (caspase-3, Bax, and Bcl-xL) and lung morphology in a rat model of experimental acute lung injury. Material and methods Acute lung injury was induced in male Wistar rats (n = 45) by a combination of aspiration bronchopneumonia and systemic/intratracheal lipopolysaccharide administration. From day 5 to day 21, animals received inhalation therapy with the studied agents or saline (placebo control). On day 21, lung tissue was examined using histological and immunohistochemical analyses to assess caspase-3–positive cells. Tissue levels of caspase-3, Bax (monomer and dimer), and Bcl-xL were quantified by immunoblotting. Results Placebo-treated rats demonstrated pronounced fibrotic remodeling (carnification) accompanied by a high proportion of caspase-3–positive macrophages and fibroblasts (35–75%). This was associated with a marked increase in active caspase-3 (9.3-fold) and Bax monomer (2.8-fold), along with a significant decrease in Bcl-xL (3.2-fold) compared with intact controls (p < 0.05). All inhalation therapies significantly attenuated fibrotic changes, reduced the number of caspase-3–positive cells, and decreased tissue levels of active caspase-3 (1.3–2.1-fold), Bax monomer (1.3–1.9-fold), and Bax dimer (1.3–2.4-fold), while restoring Bcl-xL expression (p < 0.05). Methylprednisolone exerted the strongest effect on Bax dimer reduction, camostat mesylate most effectively increased Bcl-xL levels, and enoxaparin sodium showed the greatest suppression of active caspase-3. Immunohistochemically, methylprednisolone predominantly reduced apoptosis of macrophages and type II alveolocytes, enoxaparin targeted fibroblasts, whereas camostat demonstrated a more uniform antiapoptotic effect. Conclusions Experimental acute lung injury is characterized by excessive apoptosis driven by Bax /Bcl-xL imbalance and caspase-3 activation. Inhalation therapy with camostat mesylate, methylprednisolone, and enoxaparin sodium effectively corrected these alterations through complementary mechanisms, attenuating fibrotic remodeling. These findings support the potential clinical relevance of combined or targeted antiapoptotic strategies in acute lung injury.