Lung inflation with hydrogen during the cold ischemia phase decreases lung graft injury in rats

Abstract

Hydrogen has antioxidant and anti-inflammatory effects on lung ischemia-reperfusion injury when it is inhaled by donor or/and recipient. This study examined the effects of lung inflation with 3% hydrogen during the cold ischemia phase on lung graft function in rats. The donor lung was inflated with 3% hydrogen, 40% oxygen, and 57% nitrogen at 5 mL/kg, and the gas was replaced every 20 min during the cold ischemia phase for 2 h. In the control group, the donor lung was inflated with 40% oxygen and 60% nitrogen at 5 mL/kg. The recipient was euthanized 2 h after orthotropic lung transplantation. The hydrogen concentration in the donor lung during the cold ischemia phase was 1.99-3%. The oxygenation indices in the arterial blood and pulmonary vein blood were improved in the hydrogen group. The inflammation response indices, including lung W/D ratio, the myeloperoxidase activity in the grafts, and the levels of IL-8 and TNF-α in serum, were significantly lower in the hydrogen group (5.2 ± 0.8, 0.76 ± 0.32 U/g, 340 ± 84 pg/mL, and 405 ± 115 pg/mL, respectively) than those in the control group (6.5 ± 0.7, 1.1 ± 0.5 U/g, 443 ± 94 pg/mL, and 657 ± 96 pg/mL, respectively (P < 0.05), and the oxidative stress indices, including the superoxide dismutase activity and the level of malonaldehyde in lung grafts were improved after hydrogen application. Furthermore, the lung injury score determined by histopathology, the cell apoptotic index, and the caspase-3 protein expression in lung grafts were decreased after hydrogen treatment, and the static pressure-volume curve of lung graft was improved by hydrogen inflation. In conclusion, lung inflation with 3% hydrogen during the cold ischemia phase alleviated lung graft injury and improved graft function.

Keywords: Hydrogen; donor; ischemia reperfusion injury; lung transplantation.

Figure 1

Histological analysis of lung grafts (n = 6). Paraformaldehyde-fixed sections of lung grafts were stained with eosin and hematoxylin, and all pictures represent 10× original magnifications. (a) Sham group; (b) control group; (c) hydrogen group; (d) lung injury score for each criterion. The lung injury score for each criterion in the control group was higher than the sham group (P < 0.05), and the hydrogen group was lower than the control group, but the difference was not statistically significant. *P < 0.05 versus sham group; ^#P < 0.05 versus control group. (A color version of this figure is available in the online journal.)

Figure 2

Alveolar epithelial cell apoptosis in lung grafts by TUNEL staining (n = 6). (a) Sham group; (b) control group; (c) hydrogen group. Apoptosis of the cell was represented by brown-yellow staining in the nuclei (arrows in b and c). All pictures represent 40× original magnifications. The number of apoptotic cells (AI) in the control group (52 ± 11) and hydrogen group (34 ± 9) was higher than the sham group (11 ± 3) (P < 0.05), and the hydrogen group was significantly lower than the control group (P < 0.05). *P < 0.05 versus sham group; ^#P < 0.05 versus control group. (A color version of this figure is available in the online journal.)

Figure 3

Caspase-3 protein expression of alveolar epithelial cell in lung grafts (n = 6). (a) Sham group; (b) control group; (c) hydrogen group. The presence of brown granules in the cytoplasm indicated a positive cell (arrows in b and c). All pictures represent 40× original magnifications. The immunohistochemical score (IHS) in the control group (6.2 ± 2.0) and hydrogen group (4.0 ± 1.8) was higher than the sham group (1.3 ± 0.4) (P < 0.05), and the IHS of caspase-3 in the hydrogen group was lower than the control group (P < 0.05). *P < 0.05 versus sham group; ^#P < 0.05 versus control group. (A color version of this figure is available in the online journal.)

Figure 4

The static P–V curve of the lungs (n = 8). Values were means. SD bars were omitted for clarity. *P < 0.05 versus sham group; ^#P < 0.05 versus control group