title: The bronchiolar epithelium as a prominent source of pro-inflammatory cytokines after lung irradiation.
abstract: OBJECTIVE
  To study in detail the temporal and spatial release of the pro-inflammatory cytokines tumor necrosis factor alpha, interleukin (IL)-1alpha, and IL-6 in the lung tissue of C57BL/6 mice after thoracic irradiation with 12 Gy.
  
  
  METHODS
  C57BL/6J mice were exposed to either sham irradiation or a single fraction of 12 Gy delivered to the thorax. Treated and sham-irradiated control mice were killed at 0.5 h, 1 h, 3 h, 6 h, 12 h, 24 h, 48 h, 72 h, 1 week, 2 weeks, 4 weeks, 8 weeks, 16 weeks, and 24 weeks post-irradiation (p.i.). Real-time multiplex reverse transcriptase polymerase chain reaction was established to evaluate the relative messenger RNA (mRNA) expression of TNF-alpha, IL-1alpha, and IL-6 in the lung tissue of the mice (compared with nonirradiated lung tissue). Immunohistochemical detection methods (alkaline phosphatase anti-alkaline phosphatase, avidin-biotin-complex [ABC]) and automated image analysis were used to quantify the protein expression of TNF-alpha, IL-1alpha, and IL-6 in the lung tissue (percentage of the positively stained area).
  
  
  RESULTS
  Radiation-induced release of the pro-inflammatory cytokines TNF-alpha, IL-1alpha, and IL-6 in the lung tissue was detectable within the first hours after thoracic irradiation. We observed statistically significant up-regulations for TNF-alpha at 1 h p.i. on mRNA (4.99 +/- 1.60) and at 6 h p.i. on protein level (7.23% +/- 1.67%), for IL-1alpha at 6 h p.i. on mRNA (11.03 +/- 0.77) and at 12 h p.i. on protein level (27.58% +/- 11.06%), for IL-6 at 6 h p.i. on mRNA (6.0 +/- 3.76) and at 12 h p.i. on protein level (7.12% +/- 1.93%). With immunohistochemistry, we could clearly demonstrate that the bronchiolar epithelium is the most prominent source of these inflammatory cytokines in the first hours after lung irradiation. During the stage of acute pneumonitis, the bronchiolar epithelium, as well as inflammatory cells in the lung interstitium, produced high amounts of TNF-alpha (with the maximal value at 4 weeks p.i.: 9.47% +/- 1.78%), IL-1alpha (with the peak value at 8 weeks p.i.: 14.76% +/- 7.77%), and IL-6 (with the peak value at 8 weeks p.i.: 4.28% +/- 1.33%).
  
  
  CONCLUSIONS
  In the present study we have clearly demonstrated the immediate expression of the pro-inflammatory cytokines TNF-alpha, IL-1alpha, and IL-6 in the bronchiolar epithelium in the first hours after lung irradiation. A second, long-lasting release of these cytokines by the bronchiolar and alveolar epithelium, as well as by inflammatory cells, was observed at the onset of acute pneumonitis. Therefore, we postulate that lung irradiation causes immediate epithelial reaction, with the bronchiolar epithelium becoming a significant source of pro-inflammatory cytokines capable of promoting inflammation through recruitment and activation of inflammatory cells.
authors.affiliation: Department of Radiotherapy and Radiooncology, Saarland University, Homburg/Saar, Germany. claudia.ruebe@uniklinik-saarland.de
authors.forename: Claudia E
authors.initials: CE
authors.lastname: Rübe
authors.forename: Daniela
authors.initials: D
authors.lastname: Uthe
authors.forename: Falk
authors.initials: F
authors.lastname: Wilfert
authors.forename: Daniela
authors.initials: D
authors.lastname: Ludwig
authors.forename: Kunyu
authors.initials: K
authors.lastname: Yang
authors.forename: Jochem
authors.initials: J
authors.lastname: König
authors.forename: Jan
authors.initials: J
authors.lastname: Palm
authors.forename: Andreas
authors.initials: A
authors.lastname: Schuck
authors.forename: Normann
authors.initials: N
authors.lastname: Willich
authors.forename: Klaus
authors.initials: K
authors.lastname: Remberger
authors.forename: Christian
authors.initials: C
authors.lastname: Rübe
mesh_terms: D000818:Animals; D001980:Bronchi / Q000378:metabolism* / Q000528:radiation effects*; D004848:Epithelium / Q000378:metabolism / Q000528:radiation effects; D005260:Female; D007375:Interleukin-1 / Q000378:metabolism*; D015850:Interleukin-6 / Q000378:metabolism*; D008168:Lung / Q000378:metabolism / Q000528:radiation effects*; D051379:Mice; D008810:Mice, Inbred C57BL; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D011829:Radiation Dosage; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*
chemical_list: D007375:Interleukin-1; D015850:Interleukin-6; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha