Ischemia-reperfusion injury–induced pulmonary mitochondrial damage

doi:10.1016/j.healun.2011.02.001

The Journal of Heart and Lung Transplantation

Volume 30, Issue 7, July 2011, Pages 811-818

https://doi.org/10.1016/j.healun.2011.02.001 Get rights and content

Background

Mitochondrial dysfunction is a key factor in solid organ ischemia-reperfusion (IR) injury. Impaired mitochondrial integrity predisposes to cellular energy depletion, free radical generation, and cell death. This study analyzed mitochondrial damage induced by warm pulmonary IR.

Methods

Anesthetized Wistar rats received mechanical ventilation. Pulmonary clamping was followed by reperfusion to generate IR injury. Rats were subjected to control, sham, and to 2 study group conditions: 30 minutes of ischemia without reperfusion (IR30/0), or ischemia followed by 60 minutes of reperfusion (IR30/60). Pulmonary edema was quantified by wet/dry-weight ratio. Polarography determined activities of respiratory chain complexes. Mitochondrial viability was detected by using Ca²⁺-induced swelling, and integrity by citrate synthase assay. Enzyme-linked immunosorbent assay determined cytochrome C content. Mitochondrial membrane potential (ΔΨm) stability was analyzed by flow cytometry using JC1, inflammation by myeloperoxidase (MPO) activity, and matrix-metalloproteinase-9 (MMP-9) activity by gel zymography, respectively.

Results

In IR30/60 rats, tissue water content was elevated from 80.6 % (sham) to 86.9%. After ischemia, ΔΨm showed hyperpolarization and rapid decline after uncoupling compared with controls. IR, but not ischemia alone, impaired respiratory chain function complexes I, II and III (p < 0.05). Mitochondrial viability (p < 0.001) and integrity (p < 0.01) was impaired after ischemia and IR, followed by mitochondrial cytochrome C loss (p < 0.05). Increased activation of MPO (p < 0.01) and MMP-9 (p < 0.001) was induced by reperfusion after ischemia.

Conclusions

Ischemia-related ΔΨm hyper-polarization induces reperfusion-associated mitochondrial respiratory chain dysfunction in parallel with tissue inflammation and degradation. Controlling ΔΨm during ischemia might reduce IR injury.

Section snippets

Materials and methods

Differential citrate synthase assay was applied to analyze for mitochondrial membrane instability. Mitochondrial viability was assessed using photometric Ca²⁺-induced swelling assay. Comprehensive mitochondrial respiratory complex function analysis was achieved by polarographic and ΔΨm stability measurements, using fluorescent-activated cell sorting (FACS), respectively. We assessed tissue damage beyond mitochondrial injury by determination of tissue water content as a sensitive marker for

Development of pulmonary edema after ischemia was dependent on reperfusion

Tissue water content was 82.0% in the control group and 80.6% in the sham group. Ischemia alone did not alter tissue water content (79.6%), but ischemia followed by reperfusion (IR30/60) elevated tissue water content up to 86.9%. Inter-group differences reached significance (p < 0.01, Kruskal-Wallis test). Compared with sham tissue, water content differed significantly in IR30/60 (p < 0.01, Dunn's).

Mitochondrial respiratory chain function was impaired in warm pulmonary IR

Ischemia, followed by reperfusion, impairs state 2 and state 3 respiration of pulmonary

Discussion

In this study investigating IR-associated pulmonary mitochondrial injury, we show that respiratory chain dysfunction, tissue degradation, and inflammation occur during reperfusion. IR-induced alterations in mitochondrial membrane potential, demonstrating a statistical tendency, are already detectable after ischemia, before reperfusion. The data presented in this study provide a comprehensive image of mitochondrial injury during warm pulmonary IR.

Effects on pulmonary mitochondria were measured

Disclosure statement

This work was funded by a grant of the Interdisciplinary Center for Clinical Research of the University Hospital Würzburg (IZKF project A-132N 2010; http://www.izkf.uni-wuerzburg.de/).

This article contains work of the medical thesis of Jakob Wiedemann (cand. med.).

None of the authors has a financial relationship with a commercial entity that has an interest in the subject of the presented manuscript or other conflicts of interest to disclose.

The authors thank Renate Wahn and Heidi Linss for

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Original pre-clinical scienceIschemia-reperfusion injury–induced pulmonary mitochondrial damage

Background

Methods

Results

Conclusions

Section snippets

Materials and methods

Development of pulmonary edema after ischemia was dependent on reperfusion

Mitochondrial respiratory chain function was impaired in warm pulmonary IR

Discussion

Disclosure statement

Ann Thorac Surg

J Heart Lung Transplant

J Mol Cell Cardiol

Exp Cell Res

J Biol Chem

Cell

Life Sciences

Metabolism

Neurobiol Dis

Brain Res

Neurochem Int

Original pre-clinical science
Ischemia-reperfusion injury–induced pulmonary mitochondrial damage