Oxidative DNA damage in lung tissue from patients with COPD is clustered in functionally significant sequences

Lung tissue from COPD patients displays oxidative DNA damage. The present study determined whether oxidative DNA damage was randomly distributed or whether it was localized in specific sequences in either the nuclear or mitochondrial genomes. The DNA damage-specific histone, gamma-H2AX, was detected immunohistochemically in alveolar wall cells in lung tissue from COPD patients but not control subjects. A PCR-based method was used to search for oxidized purine base products in selected 200 bp sequences in promoters and coding regions of the VEGF, TGF-β1, HO-1, Egr1, and β-actin genes while quantitative Southern blot analysis was used to detect oxidative damage to the mitochondrial genome in lung tissue from control subjects and COPD patients. Among the nuclear genes examined, oxidative damage was detected in only 1 sequence in lung tissue from COPD patients: the hypoxic response element (HRE) of the VEGF promoter. The content of VEGF mRNA also was reduced in COPD lung tissue. Mitochondrial DNA content was unaltered in COPD lung tissue, but there was a substantial increase in mitochondrial DNA strand breaks and/or abasic sites. These findings show that oxidative DNA damage in COPD lungs is prominent in the HRE of the VEGF promoter and in the mitochondrial genome and raise the intriguing possibility that genome and sequence-specific oxidative DNA damage could contribute to transcriptional dysregulation and cell fate decisions in COPD

Oxidative DNA damage in lung tissue from patients with COPD is clustered in functionally significant sequences

Viktor M Pastukh1, Li Zhang2, Mykhaylo V Ruchko1, Olena Gorodnya1, Gina C Bardwell1, Rubin M Tuder2, Mark N Gillespie11Department of Pharmacology and Center for Lung Biology, University of South Alabama College of Medicine, Mobile, AL, USA; 2Program in Translational Lung Research, Division of Pulmonary Sciences and Critical Care Medicine, Department of Medicine, University of Colorado at Denver, Aurora, CO, USAAbstract: Lung tissue from COPD patients displays oxidative DNA damage. The present study determined whether oxidative DNA damage was randomly distributed or whether it was localized in specific sequences in either the nuclear or mitochondrial genomes. The DNA damage-specific histone, gamma-H2AX, was detected immunohistochemically in alveolar wall cells in lung tissue from COPD patients but not control subjects. A PCR-based method was used to search for oxidized purine base products in selected 200 bp sequences in promoters and coding regions of the VEGF, TGF-β1, HO-1, Egr1, and β-actin genes while quantitative Southern blot analysis was used to detect oxidative damage to the mitochondrial genome in lung tissue from control subjects and COPD patients. Among the nuclear genes examined, oxidative damage was detected in only 1 sequence in lung tissue from COPD patients: the hypoxic response element (HRE) of the VEGF promoter. The content of VEGF mRNA also was reduced in COPD lung tissue. Mitochondrial DNA content was unaltered in COPD lung tissue, but there was a substantial increase in mitochondrial DNA strand breaks and/or abasic sites. These findings show that oxidative DNA damage in COPD lungs is prominent in the HRE of the VEGF promoter and in the mitochondrial genome and raise the intriguing possibility that genome and sequence-specific oxidative DNA damage could contribute to transcriptional dysregulation and cell fate decisions in COPD.Keywords: DNA damage, VEGF hypoxic response element, mtDNA, COP

Plasma mitochondrial DNA is elevated in obese type 2 diabetes mellitus patients and correlates positively with insulin resistance.

Cells damaged by mechanical or infectious injury release proinflammatory mitochondrial DNA (mtDNA) fragments into the circulation. We evaluated the relation between plasma levels of mtDNA fragments in obese type 2 diabetes mellitus (T2DM) patients and measures of chronic inflammation and insulin resistance. In 10 obese T2DM patients and 12 healthy control (HC) subjects, we measured levels of plasma cell-free mtDNA with quantitative real-time polymerase chain reaction, and mtDNA damage in skeletal muscle with quantitative alkaline Southern blot. Also, markers of systemic inflammation and oxidative stress in skeletal muscle were measured. Plasma levels of mtDNA fragments, mtDNA damage in skeletal muscle and plasma tumor necrosis factor α levels were greater in obese T2DM patients than HC subjects. Also, the abundance of plasma mtDNA fragments in obese T2DM patients levels positively correlated with insulin resistance. To the best of our knowledge, this is the first published evidence that elevated level of plasma mtDNA fragments is associated with mtDNA damage and oxidative stress in skeletal muscle and correlates with insulin resistance in obese T2DM patients. Plasma mtDNA may be a useful biomarker for predicting and monitoring insulin resistance in obese patients

Mitochondrial Targeted Endonuclease III DNA Repair Enzyme Protects against Ventilator Induced Lung Injury in Mice

The mitochondrial targeted DNA repair enzyme, 8-oxoguanine DNA glycosylase 1, was previously reported to protect against mitochondrial DNA (mtDNA) damage and ventilator induced lung injury (VILI). In the present study we determined whether mitochondrial targeted endonuclease III (EndoIII) which cleaves oxidized pyrimidines rather than purines from damaged DNA would also protect the lung. Minimal injury from 1 h ventilation at 40 cmH2O peak inflation pressure (PIP) was reversed by EndoIII pretreatment. Moderate lung injury due to ventilation for 2 h at 40 cmH2O PIP produced a 25-fold increase in total extravascular albumin space, a 60% increase in W/D weight ratio, and marked increases in MIP-2 and IL-6. Oxidative mtDNA damage and decreases in the total tissue glutathione (GSH) and the GSH/GSSH ratio also occurred. All of these indices of injury were attenuated by mitochondrial targeted EndoIII. Massive lung injury caused by 2 h ventilation at 50 cmH2O PIP was not attenuated by EndoIII pretreatment, but all untreated mice died prior to completing the two hour ventilation protocol, whereas all EndoIII-treated mice lived for the duration of ventilation. Thus, mitochondrial targeted DNA repair enzymes were protective against mild and moderate lung damage and they enhanced survival in the most severely injured group