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. 2012 Sep 7;425(4):873-9.
doi: 10.1016/j.bbrc.2012.08.005. Epub 2012 Aug 7.

Profiling Molecular Changes Induced by Hydrogen Treatment of Lung Allografts Prior to Procurement

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Free PMC article

Profiling Molecular Changes Induced by Hydrogen Treatment of Lung Allografts Prior to Procurement

Yugo Tanaka et al. Biochem Biophys Res Commun. .
Free PMC article

Abstract

Background: We previously demonstrated that donor treatment with inhaled hydrogen protects lung grafts from cold ischemia/reperfusion (I/R) injury during lung transplantation. To elucidate the mechanisms underlying hydrogen's protective effects, we conducted a gene array analysis to identify changes in gene expression associated with hydrogen treatment.

Methods: Donor rats were exposed to mechanical ventilation with 98% oxygen and 2% nitrogen or 2% hydrogen for 3 h before harvest; lung grafts were stored for 4h in cold Perfadex. Affymetrix gene array analysis of mRNA transcripts was performed on the lung tissue prior to implantation.

Results: Pretreatment of donor lungs with hydrogen altered the expression of 229 genes represented on the array (182 upregulated; 47 downregulated). Hydrogen treatment induced several lung surfactant-related genes, ATP synthase genes and stress-response genes. The intracellular surfactant pool, tissue adenosine triphosphate (ATP) levels and heat shock protein 70 (HSP70) expression increased in the hydrogen-treated grafts. Hydrogen treatment also induced the transcription factors C/EBPα and C/EBPβ, which are known regulators of surfactant-related genes.

Conclusion: Donor ventilation with hydrogen significantly increases expression of surfactant-related molecules, ATP synthases and stress-response molecules in lung grafts. The induction of these molecules may underlie hydrogen's protective effects against I/R injury during transplantation.

Figures

Fig. 1
Fig. 1
Quantitative RT-PCR for (A) CC16, (B) SP-A, (C) SCGB3A2, (D) HSD11β1 and (E) PLUNC (n = 4 for each group; **p < 0.01 and *p < 0.05). (F) Representative immunohistochemistry for CC16 in the lung grafts before transplantation. Western blots for (G) CC16, (H) PLUNC, (I) C/EBPα, (J) C/EBP β, and β-actin on protein extracts from lung grafts taken after 3 h of mechanical ventilation and 4 h cold storage. The images are representative of 3 independent experiments.
Fig. 2
Fig. 2
Transmission electron microscopy of lung grafts. Cell structure of alveolar epithelial type II cells in hydrogen-treated and nitrogen-treated grafts. Lamellar bodies (white arrowheads) contain intracellular surfactant.
Fig. 3
Fig. 3
Quantitative RT-PCR for (A) ATP5b, (B) ATP5o, (C) ATP5j and (D) ATP5 l (n = 4 for each group; **p < 0.01 and *p < 0.05). (E) Lung ATP content prior to implantation. (n = 4 per group *p < 0.05). Quantitative RT-PCR for (F) HSPA5 and (G) DUSP1 (n = 4 for each group; **p < 0.01 and *p < 0.05). (H) Western blots for HSP70 and β-actin on protein extracts from lung grafts taken after 3 h of mechanical ventilation and 4 h of cold storage. (I) Western blots for phosphorylated (p) and total (t) p38, ERK1/2 and JNK on protein extracts from lung grafts taken 2 h after reperfusion. The images are representative of 3 independent experiments (n = 3 for each group).

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