[Effect of mechanical stretch preconditioning on pathological stretch-induced activation of γ-aminobutyric acid signaling pathway in human type II alveolar epithelial cells].

OBJECTIVE

To evaluate the effect of mechanical stretch preconditioning on pathological stretch-induced activation of γ-aminobutyric acid (GABA) signaling pathway in human type II alveolar epithelial cells (AEC II).

METHODS

AEC II cell line (A549 cells) cultured in vitro were divided into control group (group C), pathological stretch group (group P1) and mechanical stretch preconditioning group (group P2). In group C, A549 cells were cultured routinely. In group P1, A549 cells were exposed to 20% cyclic stretch for 6 hours. In group P2, A549 cells were exposed to 5% cyclic stretch for 60 minutes, and then exposed to 20% cyclic stretch for 6 hours. The cells were harvested for determination of the cell viability by methyl thiazolyl tetrazolium assay, lactate dehydrogeuase (LDH) release was determined by colorimetric method, the levels of interleukin (IL-1β and IL-6) and tumor necrosis factor-α (TNF-α) were determined by enzyme linked immunosorbent assay (ELISA), the mRNA expressions of IL-1β, IL-6 and TNF-α were determined by reverse transcription-polymerase chain reaction (RT-PCR), and the protein expressions of glutamic acid decarboxylase (GAD) and γ-aminobutyric acid A receptor (GABAR) were determined by Western Blot.

RESULTS

Compared with group C, the cell viability of group P1 was significantly decreased (A value: 0.196 ± 0.071 vs. 0.886±0.107), the release rate of LDH was significantly increased [(12.3±2.4)% vs. (1.9±0.5)%]; the contents and mRNA expressions of IL-1β, IL-6 and TNF-α in cell culture medium were significantly increased [IL-1β (ng/L): 138.6±19.7 vs. 32.7±7.4, IL-6 (ng/L): 196.5±31.7 vs. 55.4±13.8, TNF-α (ng/L): 111.3±21.8 vs. 20.8±7.6; IL-1β mRNA (2): 2.79±0.44 vs. 0.83±0.12, IL-6 mRNA (2): 1.99±0.25 vs. 0.56±0.11, TNF-α mRNA (2): 2.54±0.37 vs. 0.72±0.09]; the protein expressions of GAD and GABAR were significantly decreased [GAD (gray value): 0.38±0.12 vs. 1.75±0.45, GABAR (gray value): 0.29±0.09 vs. 1.68±0.39; all P < 0.05]. Compared with group P1, the cell viability of group P2 was significantly increased (A value: 0.523±0.132 vs. 0.196±0.071), the release rate of LDH was significantly decreased [(6.9±1.7)% vs. (12.3±2.4)%]; the contents and mRNA expressions of IL-1β, IL-6 and TNF-α in cell culture medium were significantly decreased [IL-1β (ng/L): 79.2±11.6 vs. 138.6±19.7, IL-6 (ng/L): 89.6±15.6 vs. 196.5±31.7, TNF-α (ng/L): 55.9±11.4 vs. 111.3±21.8; IL-1β mRNA (2): 1.92±0.36 vs. 2.79±0.44, IL-6 mRNA (2): 1.09±0.18 vs. 1.99±0.25, TNF-α mRNA (2): 1.77±0.25 vs. 2.54±0.37]; the protein expressions of GAD and GABAR were significantly increased [GAD (gray value): 1.26±0.33 vs. 0.38±0.12, GABAR (gray value): 1.04±0.15 vs. 0.29±0.09; all P < 0.05].

CONCLUSIONS

The mechanism by which mechanical stretch preconditioning attenuates pathological stretch-induced injury in human AECII is related to the activation of GABA signaling pathway.