title: Postconditioning with inhaled carbon monoxide counteracts apoptosis and neuroinflammation in the ischemic rat retina.
abstract: OBJECTIVE
  Ischemia and reperfusion injury (I/R) of neuronal structures and organs is associated with increased morbidity and mortality due to neuronal cell death. We hypothesized that inhalation of carbon monoxide (CO) after I/R injury ('postconditioning') would protect retinal ganglion cells (RGC).
  
  
  METHODS
  Retinal I/R injury was performed in Sprague-Dawley rats (n = 8) by increasing ocular pressure (120 mmHg, 1 h). Rats inhaled room air or CO (250 ppm) for 1 h immediately following ischemia or with 1.5 and 3 h latency. Retinal tissue was harvested to analyze Bcl-2, Bax, Caspase-3, HO-1 expression and phosphorylation of the nuclear transcription factor (NF)-κB, p38 and ERK-1/2 MAPK. NF-κB activation was determined and inhibition of ERK-1/2 was performed using PD98059 (2 mg/kg). Densities of fluorogold prelabeled RGC were analyzed 7 days after injury. Microglia, macrophage and Müller cell activation and proliferation were evaluated by Iba-1, GFAP and Ki-67 staining.
  
  
  RESULTS
  Inhalation of CO after I/R inhibited Bax and Caspase-3 expression (Bax: 1.9 ± 0.3 vs. 1.4 ± 0.2, p = 0.028; caspase-3: 2.0 ± 0.2 vs. 1.5 ± 0.1, p = 0.007; mean ± S.D., fold induction at 12 h), while expression of Bcl-2 was induced (1.2 ± 0.2 vs. 1.6 ± 0.2, p = 0.001; mean ± S.D., fold induction at 12 h). CO postconditioning suppressed retinal p38 phosphorylation (p = 0.023 at 24 h) and induced the phosphorylation of ERK-1/2 (p<0.001 at 24 h). CO postconditioning inhibited the expression of HO-1. The activation of NF-κB, microglia and Müller cells was potently inhibited by CO as well as immigration of proliferative microglia and macrophages into the retina. CO protected I/R-injured RGC with a therapeutic window at least up to 3 h (n = 8; RGC/mm(2); mean ± S.D.: 1255 ± 327 I/R only vs. 1956 ± 157 immediate CO treatment, vs. 1830 ± 109 1.5 h time lag and vs. 1626 ± 122 3 h time lag; p<0.001). Inhibition of ERK-1/2 did not counteract the CO effects (RGC/mm(2): 1956 ± 157 vs. 1931 ± 124, mean ± S.D., p = 0.799).
  
  
  CONCLUSIONS
  Inhaled CO, administered after retinal ischemic injury, protects RGC through its strong anti-apoptotic and anti-inflammatory effects.
authors.affiliation: Department of Anesthesiology and Critical Care Medicine, University Medical Center Freiburg im Breisgau, Germany.
authors.forename: Nils
authors.initials: N
authors.lastname: Schallner
authors.forename: Matthias
authors.initials: M
authors.lastname: Fuchs
authors.forename: Christian I
authors.initials: CI
authors.lastname: Schwer
authors.forename: Torsten
authors.initials: T
authors.lastname: Loop
authors.forename: Hartmut
authors.initials: H
authors.lastname: Buerkle
authors.forename: Wolf Alexander
authors.initials: WA
authors.lastname: Lagrèze
authors.forename: Christian
authors.initials: C
authors.lastname: van Oterendorp
authors.forename: Julia
authors.initials: J
authors.lastname: Biermann
authors.forename: Ulrich
authors.initials: U
authors.lastname: Goebel
mesh_terms: D000280:Administration, Inhalation; D000818:Animals; D000893:Anti-Inflammatory Agents / Q000008:administration & dosage*; D017209:Apoptosis / Q000187:drug effects*; D002248:Carbon Monoxide / Q000008:administration & dosage*; D053148:Caspase 3 / Q000235:genetics / Q000378:metabolism; D002465:Cell Movement / Q000187:drug effects; D004789:Enzyme Activation; D005260:Female; D015870:Gene Expression / Q000187:drug effects; D051547:Heme Oxygenase-1 / Q000235:genetics / Q000378:metabolism; D007511:Ischemia / Q000188:drug therapy; D008297:Male; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D009457:Neuroglia / Q000187:drug effects / Q000502:physiology; D018696:Neuroprotective Agents / Q000008:administration & dosage*; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D015427:Reperfusion Injury / Q000188:drug therapy*; D012160:Retina / Q000187:drug effects / Q000473:pathology; D012165:Retinal Ganglion Cells / Q000187:drug effects / Q000473:pathology; D012171:Retinal Vessels / Q000473:pathology; D012173:Retinitis / Q000517:prevention & control*; D051028:bcl-2-Associated X Protein / Q000235:genetics / Q000378:metabolism
chemical_list: D000893:Anti-Inflammatory Agents; C494828:Bax protein, rat; D016328:NF-kappa B; D018696:Neuroprotective Agents; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D002248:Carbon Monoxide; D051547:Heme Oxygenase-1; D020928:Mitogen-Activated Protein Kinases; C505410:Casp3 protein, rat; D053148:Caspase 3