Role and mechanism of signal pathway mediated by Toll-like receptor 9-myeloid differentiation factor 88 in alveolar macrophages in ventilator-induced lung injury in rats

Abstract

To investigate the role of Toll-like receptor 9 (TLR9)-myeloid differentiation factor 88 (MyD88) signal pathway in alveolar macrophages in ventilator-induced lung injury (VILI). 30 adult male Sprague-Dawley (SD) rats were randomly assigned to three groups (with 10 rats in each group). Group A was the control group, with spontaneous respiration after tracheostomy. Rats in group B received mechanical ventilation for 4 hours with normal tidal volume (VT) 7 ml/kg after tracheostomy, and group C rats received mechanical ventilation with VT 40 ml/kg for 4 hours. After termination of ventilation, examination with transmission electron microscopy was performed to observe the ultrastructure changes in alveolar epithelial cell type II (AECII) of the lung. Lung wet/dry ratios (W/D) and total protein concentration, the concentration of interleukins (IL-6 and IL-1β) in bronchoalveolar lavage fluid (BALF) were determined. The protein and mRNA expressions of TLR9, MyD88 and nuclear factor-ΚB (NF-ΚB) in alveolar macrophages were assayed by Western Blot and real-time reverse transcription-polymerase chain reaction (RT-PCR). The ultrastructure of AECII in the group A and group B was almost normal, whereas the chromatin of the nuclei, the lamellar corpuscles in the cytoplasm, the cell membrane and the microvilli of the AECII in the group C showed injurious changes in various degrees. When the group C was compared with the group A and the group B, it was shown that the W/D ratios (5.54±0.17 vs. 4.58±0.17, 4.69±0.16) and total protein concentration (6.33±0.61 g/L vs. 0.45±0.05 g/L, 0.47±0.04 g/L), IL-6 (1.989±0.103 μg/L vs. 1.033±0.061 μg/L, 1.010±0.069 μg/L) and IL-1β (2.79±0.25 ng/L vs. 1.05±0.15 ng/L, 1.23±0.22 ng/L) in BALF, the protein expressions of TLR9, MyD88 and NF-ΚB [TLR9 (A value): 0.770±0.042 vs. 0.300±0.027, 0.310±0.037; MyD88 (A value): 0.950±0.091 vs. 0.560±0.082, 0.580±0.084; NF-ΚB(A value): 1.020±0.076 vs. 0.740±0.052, 0.700±0.076] in alveolar macrophages were all increased significantly, and all of which showed significant difference (P<0.05 or P<0.01). The mRNA levels of TLR9, MyD88 and NF-ΚB in alveolar macrophages in the group B were (1.13±0.32), (1.18±0.33), and (1.11±0.22) folds of those of the group A, respectively, but there were no significant differences (all P>0.05). While the mRNA levels of TLR9, MyD88 and NF-ΚB of alveolar macrophages in the group C were (8.66±0.69), (6.41±0.53) and (5.29±0.71) folds of those of the group A, respectively, and all of them showed significant difference (all P<0.01). TLR9-MyD88 signaling in alveolar macrophages plays a role in pathogenesis of VILI.