Enhanced Hsp70 Expression Protects against Acute Lung Injury by Modulating Apoptotic Pathways

Abstract

The Acute respiratory distress syndrome (ARDS) is a highly lethal inflammatory lung disorder. Apoptosis plays a key role in its pathogenesis. We showed that an adenovirus expressing the 70 kDa heat shock protein Hsp70 (AdHSP) protected against sepsis-induced lung injury. In this study we tested the hypothesis that AdHSP attenuates apoptosis in sepsis-induced lung injury.Sepsis was induced in rats via cecal ligation and double puncture (2CLP). At the time of 2CLP PBS, AdHSP or AdGFP (an adenoviral vector expressing green fluorescent protein) were injected into the tracheas of septic rats. 48 hours later, lungs were isolated. One lung was fixed for TUNEL staining and immunohistochemistry. The other was homogenized to isolate cytosolic and nuclear protein. Immunoblotting, gel filtration and co-immunoprecipitation were performed in these extracts. In separate experiments MLE-12 cells were incubated with medium, AdHSP or AdGFP. Cells were stimulated with TNFα. Cytosolic and nuclear proteins were isolated. These were subjected to immunoblotting, co- immunoprecipitation and a caspase-3 activity assay.TUNEL assay demonstrated that AdHSP reduced alveolar cell apoptosis. This was confirmed by immunohistochemical detection of caspase 3 abundance. In lung isolated from septic animals, immunoblotting, co-immunoprecipitation and gel filtration studies revealed an increase in cytoplasmic complexes containing caspases 3, 8 and 9. AdHSP disrupted these complexes.We propose that Hsp70 impairs apoptotic cellular pathways via interactions with caspases. Disruption of large complexes resulted in stabilization of lower molecular weight complexes, thereby, reducing nuclear caspase-3. Prevention of apoptosis in lung injury may preserve alveolar cells and aid in recovery.

Stimulation of Brain AMP-Activated Protein Kinase Attenuates Inflammation and Acute Lung Injury in Sepsis

“…The histopathological features of lung injury in septic mice demonstrated here are resembled in the clinical condition of patients with sepsis (41). Other studies have indicated that apoptosis plays an important role on progression of sepsis (42,43). Oberholzer and colleagues suggested that targeting signaling pathways that lead to apoptosis would represent a new therapeutic target for the patient with sepsis or other related clinical conditions (34).…”

Protective Effect of Naringenin against Lipopolysaccharide-Induced Acute Lung Injury in Rats

“…It was reported that HSP70 significantly decreased the inflammatory biomarkers, impaired apoptotic pathways via interactions with caspases, and improved outcome after septic and shock models of ALI in rats [34,35] . In this study, LPS caused a minor insignificant upregulation of HSP70 in lung tissues.…”

Hyperthermia Promotes and Prevents Respiratory Epithelial Apoptosis through Distinct Mechanisms

“…Bid cleavage was observed within 30 minutes of treatment with TNF-a at 42 8 C, but was not detected up to 2 hours after TNF-a treatment in 37 8 C cells. Exposure to 42 8 C without TNF-a also stimulated Bid cleavage, although the activation was delayed and reduced compared with the TNFa-treated cells (compare lanes 11-14 with lanes [15][16][17][18].…”

“…HSP70 has been shown to block apoptosis by binding to Apaf-1, thereby preventing its recruitment to the apoptosome (17). More recently, Aschkenasy and colleagues (18) showed that adenoviral transfer of HSP70 via intratracheal instillation in mice subsequently subjected to cecal ligation and puncture reduced lung injury and lung cell apoptosis. These authors also showed that HSP70 transfer to mouse lung epithelial (MLE) cell line 12 (MLE12) cells reduced TNF-a-induced apoptosis.…”