Affiliations of authors: D. E. Hallahan (Departments of Radiation Oncology and Biomedical Engineering), L. Geng (Department of Radiation Oncology), and Y. Shyr (Biostatistics), Vanderbilt University, Nashville, TN.
Correspondence to: Dennis E. Hallahan, M.D., Vanderbilt Department of Radiation Oncology, 1301 22nd Ave. South, B-902, The Vanderbilt Clinic, Nashville, TN 372325671 (e-mail: dennis.hallahan{at}mcmail.vanderbilt.edu).
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ABSTRACT |
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INTRODUCTION |
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Radiation induces inflammation in the lung by inducing the expression of two cell adhesion proteins, E-selectin and intercellular adhesion molecule 1 (ICAM-1), on endothelial cells of the microvasculature (13,14). Leukocytes subsequently adhere to the endothelial cells via these adhesion molecules (14,15). Mice genetically engineered to lack the ICAM-1 gene (i.e., ICAM-1-/- or knockout mice) show no inflammatory cell infiltration of their lungs after thoracic irradiation (16,17). Therefore, the ICAM-1 knockout mouse is an excellent animal model in which to study the role of inflammation in the pathogenesis of impaired compliance in the irradiated lung.
We have previously shown that C57BL/6 mice that lack ICAM-1 gene expression do not develop pulmonary inflammation following thoracic irradiation, whereas C57BL/6 mice that express wild-type levels of ICAM-1 show inflammation beginning within 4 weeks of irradiation (16). Here we determined the effects of ICAM-1 gene expression on pulmonary compliance and fibrosis in the lungs of mice subjected to thoracic irradiation.
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MATERIALS AND METHODS |
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Six-week-old C57BL/6 mice of both sexes with (ICAM-1-/-) or without (ICAM-1+/+) a homozygous null mutation in the ICAM-1 gene (The Jackson Laboratory, Bar Harbor, ME) were subjected to thoracic irradiation as previously described (16,17). Briefly, the mice were first anesthetized by intraperitoneal injection with a mixture of ketamine (44 mg/kg) and xylazine (15 mg/kg) in normal saline. The thorax of each mouse was then irradiated using a GE Maxitron orthovoltage generator (General Electric, Milwaukee, WI) operated at 250 kV and equipped with an aluminum filter to give a dose rate of 2 Gy/minute. Age-matched control (mock-irradiated) mice were similarly anesthetized and placed into the Maxitron irradiation vault but were not irradiated (i.e., 0 Gy group). Thoracic irradiation was verified by developing a piece of radiography film that had been placed under each mouse before irradiation. The dose to the thorax was verified by thermal luminescence detectors. After irradiation, the mice were returned to their cages, observed until they began to move, and then housed in a barrier facility until plethysmography was performed or respiratory compromise was observed as described below. Animal care was in accord with institutional guidelines.
Determination of the Lethal Dose of Thoracic Irradiation
Six-week-old ICAM-1+/+ or ICAM-1-/- mice were subjected to thoracic irradiation as described above. Ten mice of each genotype were randomly assigned to receive a total dose of 12.5, 14, 16, 17, or 18 Gy or mock irradiation (0 Gy). The mice were subsequently housed in a barrier facility for 18 months during which time they were monitored twice weekly for signs of respiratory distress, as defined by hypomotility, hunched backs, and tachypnea. Mice with respiratory distress were euthanized by an intraperitoneal injection of phenobarbital. The lungs of euthanized mice were dissected, fixed, and sectioned for microscopic verification of lung injury without pneumonia as described below.
To determine whether radiation-induced pulmonary fibrosis and respiratory compromise were time-dependent, ten 6-week-old ICAM-1+/+ or ICAM-1-/- mice were randomly assigned to receive mock irradiation or a total thoracic dose of 12.5, 14, 16, 17, or 18 Gy. The mice were observed twice weekly for signs of hypomotility, hunched backs, and tachypnea. Nine months after receiving thoracic or mock irradiation, all mice were sacrificed and their lung histology was studied. Mice with respiratory distress were euthanized and their lungs were sectioned to detect radiation-induced lung injury.
Plethysmography
Groups of five mice were randomly assigned to receive thoracic irradiation at a dose of 14 Gy or mock irradiation (0 Gy). At 10 weeks after actual or mock irradiation, the mice were anesthetized by intraperitoneal injection with a mixture of ketamine (44 mg/kg) and xylazine (15 mg/kg) in normal saline, and plethysmography was performed as previously described (18,19). Briefly, a tracheostomy was surgically created in each mouse, and the trachea was intubated with a 12-mm-long, 22-gauge metal cannula. The mice were mechanically ventilated (tidal volume = 8 mL/kg; frequency = 140 breaths/minute) using a volume-controlled rodent ventilator (Harvard Apparatus, South Natick, MA). Changes in plethysmographic pressure, measured relative to a Whiter reference chamber by means of a differential pressure transducer (Honeywell Microswitch, Freeport, IL), reflected changes in lung volume (18). Lung volume and pressure signals were recorded digitally, and dynamic compliance, in mL/cm H2O, was calculated, breath by breath, using Mouse PRC software (Lakeshore Technologies, Chicago, IL). This experiment was repeated once.
Lung Histology
We studied the pulmonary histology of mice that underwent plethysmography, that showed respiratory distress, or that survived for 12 months after irradiation. Whole lungs were dissected and fixed in 10% formalin (Histoprep; Fisher Scientific, Suwanee, GA) for 24 hours. Lungs were dehydrated and embedded in paraffin. Lungs were then sectioned into 5-µm-thick sections, mounted on glass slides, and stained.
We used Masson's trichrome stain to detect fibrosis as previously described (20). Briefly, tissue sections were rinsed in absolute alcohol followed by 95% alcohol and distilled water. Bouin's fixative was heated to 60 °C and dropped onto slides containing lung tissue sections, and the slides were then incubated for 3 minutes at 60 °C. Sections were then cooled to room temperature, rinsed in running tap water for 5 minutes, and then rinsed once with distilled water. The sections were incubated in Weigert's iron hematoxylin solution for 7 minutes at room temperature and then rinsed with distilled water. The sections were then incubated with Biebrich scarlet-acid fuchsin solution for 30 seconds followed by a distilled water rinse. Sections were placed in a phosphotungstic acid solution for 105 seconds and then rinsed in distilled water. Sections were then placed in Aniline blue solution (Sigma Chemical Co., St. Louis, MO) for 3 minutes followed by a distilled water rinse. The sections were placed in 1% acetic acid for 1 minute, followed by 95% alcohol, two changes of absolute alcohol, and two changes of xylene. The sections were mounted under coverslips with a xylene-soluble media (Sigma Chemical Co.). Randomly selected microscopic fields (at x400 magnification) were photographed, and staining was quantified using ImagePro software (Media Cybernetics, Des Moines, IA).
Quantitation of Inflammatory Cell Infiltration
Inflammatory cell infiltration into the lungs of irradiated and mock-irradiated mice (10 mice per group) was quantified by indirect immunofluorescence microscopy using anti-leukocyte-common antigen (anti-LCA) antibody (Pharmingen, San Diego, CA) to detect the presence of LCA in lung sections, as previously described (16). We counted the number of LCA-stained cells within 10 randomly selected fields of slides viewed at x400. The fluorescence intensity of stained tissue was then quantified with NIH Image software.
Assessment of Type III Collagen Deposition in Lung Sections
Histologic sections of lungs from ICAM-1-/- and ICAM-1+/+ mice that had survived 18 months after thoracic irradiation were examined for evidence of fibrosis by staining with a goat anti-mouse collagen III immunoglobulin G (IgG) (Chemicon International, Temecula, CA). The lung sections were first rehydrated and incubated in 3% H2O2 for 1 hour at room temperature to quench endogenous peroxidases. The sections were then incubated in 5% goat serum in phosphate-buffered saline (PBS) for 1 hour at 37 °C to block nonspecific binding. Goat anti-mouse collagen III IgG (20 µg/mL) was added to the blocking buffer, and the sections were incubated for 60 minutes at 37 °C. The sections were then washed with blocking buffer at 37 °C, and biotin-conjugated donkey anti-goat IgG (Accurate Chemicals, Westbury, NY) at 20 µg/mL in blocking buffer was added to the sections and incubated for 40 minutes at 37 °C. The sections were then washed, and ABC Reagent (Vector Laboratories, Inc., Burlingame, CA) was added in PBS, followed by reagents A and B for 30 minutes at room temperature, according to the manufacturer's instructions (Vector Laboratories, Inc.). Sections were washed, peroxidase substrate was added, and the sections were incubated at room temperature for 10 minutes. The sections were then dehydrated in a graded alcohol series followed by xylene and mounted under coverslips.
Determination of Hydroxyproline Content of Lung
Groups of 10 ICAM-1+/+ or ICAM-1-/- mice were randomly assigned to receive mock irradiation (0 Gy) or thoracic irradiation with 14, 16, or 18 Gy. The mice were euthanized at 10 weeks, 9 months, and 18 months after mock or actual irradiation, and their lungs were removed, washed in PBS, and weighed. The total collagen content of each right lung was determined using a colorimetric assay to measure lung hydroxyproline content as previously described (21,22). Briefly, a minced aliquot of lung tissue was hydrolyzed overnight at 110 °C in 800 µL of 6 N HCl. To 200 µL of that hydrolysate were added 100 µL of 0.02% methyl red and 20 µL of 0.04% bromthymol blue. The sample volume was adjusted to 2 mL with 0.5x assay buffer (0.024 M citric acid, 0.02 M acetic acid, 0.088 M sodium acetate, 0.085 M NaOH), and the pH was adjusted to 6.57.0. One milliliter of chloramine T solution (Sigma Chemical Co.) was added to each sample; the mixture was then incubated at room temperature for 20 minutes, after which 1 mL of dimethyl benzaldehyde solution (Sigma Chemical Co.) was added followed by incubation at 60 °C for 15 minutes. We measured the absorbance of each sample at 550 nm. Whole-lung hydroxyproline values, expressed as micrograms of hydroxyproline per lung, were determined by normalizing the hydroxyproline values obtained from this colorimetric assay of the minced lung aliquots to the wet weight of the whole right lung. Values were corrected for total lung wet weight.
Statistical Analysis
We used a general linear model to test for associations between the numbers of leukocytes present in the lungs, dynamic compliances (mL/cm H2O), ICAM-1 genotype (wild type versus homozygous null), and the doses of irradiation the mice received (0, 14, 16, and 18 Gy). The general linear model (logistic regression analysis) was also used to test the association between the respiratory insufficiency (yes/no), ICAM-1 genotype (wild type versus homozygous null), and the doses of irradiation the mice received (12.5, 14, 16, 17, and 18 Gy). We applied the Bonferroni method to adjust the overall statistical significance levels to 5% for the multiple comparisons in this study. All statistical tests were two-sided, and differences were considered statistically significant for P<.05. SAS software version 8.1 (SAS Institute Inc., Cary, NC) was used for all statistical analyses.
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RESULTS |
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Pulmonary inflammation precedes radiation-induced fibrosis. To determine whether there is a causal relationship between radiation-induced pneumonitis and subsequent fibrosis, we studied these processes in ICAM-1-/- mice, which do not display radiation-induced pulmonary inflammation. Anti-LCA antibody binds to all leukocytes and provides the means of quantifying neutrophils, lymphocytes, and monocytes within the irradiated lung. As shown in Fig. 1, at 5 weeks after mock irradiation, the lungs of ICAM-1-/- mice had statistically significantly fewer LCA-positive cells than the lungs of ICAM-1+/+ mice. For example, the mean numbers of LCA-positive cells/field in lung sections from ICAM-1+/+ mice subjected to 0, 14, 16, and 18 Gy of radiation were 13 LCA-positive cells/field, 50 LCA-positive cells/field, 54 LCA-positive cells/field, and 61 LCA-positive cells/field, respectively. These values were statistically significantly higher than the mean numbers of LCA-positive cells/field in lung sections from ICAM-1-/- mice subjected to 0 Gy (four LCA-positive cells/field), 14 Gy (six LCA-positive cells/field), 16 Gy (eight LCA-positive cells/field), and 18 Gy (seven LCA-positive cells/field) of radiation, respectively (P<.001 for each; difference in mean number of LCA-positive cells/field between ICAM-1+/+ mice and ICAM-1-/- mice subjected to mock irradiation [0 Gy] = nine cells/field [95% CI = seven cells/field to 11 cells/field], between ICAM-1+/+ mice and ICAM-1-/- mice subjected to 14 Gy = 44 cells/field [95% CI = 31 cells/field to 57 cells/field], between ICAM-1+/+ mice and ICAM-1-/- mice subjected to 16 Gy = 46 cells/field [95% CI = 34 cells/field to 58 cells/field], and between ICAM-1+/+ mice and ICAM-1-/- mice subjected to 18 Gy = 54 cells/field [95% CI = 43 cells/field to 65 cells/field]). LCA-positive cells were present in both the perivascular and alveolar spaces of the lungs of all irradiated ICAM-1+/+ mice.
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Diminished ventilation and perfusion are two aspects of impaired lung function that are a consequence of thoracic irradiation. The onset of impaired ventilation occurs within weeks of irradiation; thus early pathologic events, such as cell loss, edema, diminished surfactant, and inflammation, have been implicated in radiation-induced impairment of lung function. One physiologic measure of these events is dynamic pulmonary compliance, which is a function of lung volume, surface tension, and the elasticity of lung parenchyma. We therefore evaluated dynamic pulmonary compliance, as measured by changes in plethysmographic pressure, in ICAM-1+/+ and ICAM-1-/- mice 10 weeks after they had received 14, 16, and 18 Gy of thoracic irradiation to determine whether the absence of inflammation in irradiated lung had an impact on dynamic pulmonary compliance.
At 10 weeks after mock irradiation, ICAM-1+/+ mice had a mean dynamic pulmonary compliance of 0.054 mL/cm H2O and ICAM-1-/- mice had a mean dynamic pulmonary compliance of 0.051 mL/cm H2O. Ten weeks after receiving 14 Gy of thoracic irradiation, ICAM-1+/+ mice had a mean dynamic pulmonary compliance of 0.043 mL/cm H2O, which was statistically significantly lower than that of the mock-irradiated ICAM-1+/+ mice (P<.001; difference in mean dynamic pulmonary compliance between mock-irradiated and irradiated ICAM-1+/+ mice = 0.011 mL/cm H2O [95% CI = 0.006 mL/cm H2O to 0.016 mL/cm H2O]) (Fig. 2). By contrast, at 10 weeks after receiving 14 Gy of thoracic irradiation, the ICAM-1-/- mice had a mean dynamic pulmonary compliance of 0.049 mL/cm H2O, which was not statistically significantly different from that of the mock-irradiated ICAM-1-/- mice (P = .17; difference in mean dynamic pulmonary compliance between mock-irradiated and irradiated ICAM-1-/- mice = 0.002 mL/cm H2O [95% CI = 0.001 mL/cm H2O to 0.004 mL/cm H2O]) (Fig. 2
).
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To determine whether the radiation-induced reduction in pulmonary compliance we observed was associated with respiratory distress, mice were monitored for signs of tachypnea, hunched backs, and hypomotilitythree indicators of respiratory distresstwice weekly for 18 months after receiving thoracic irradiation with 12.5, 14, 16, 17, and 18 Gy. These mice began to display signs of respiratory distress 6 months after thoracic irradiation. We compared the incidence of respiratory distress in ICAM-1-/- mice with that in ICAM-1+/+ mice that had received the same dose of radiation. At 18 months after irradiation, 20% (95% CI = 0% to 44%) of the ICAM-1+/+ mice that received a dose of 14 Gy, 50% (95% CI = 19% to 81%) of the ICAM-1+/+ mice that received a dose of 16 Gy, and 80% (95% CI = 56% to 100%) of the 10 ICAM-1+/+ mice that received a dose of 18 Gy displayed respiratory distress (Fig. 3, A). By contrast, at this same time point, 0% (95% CI = 0% to 31%) of the ICAM-1-/- mice that received a dose of 14 Gy, 20% (95% CI = 0% to 44%) of the ICAM-1-/- mice that received a dose of 16 Gy, and 50% (95% CI = 19% to 81%) of the ICAM-1-/- mice that received a dose of 18 Gy displayed respiratory distress (Fig. 3, A
). The overall incidence of respiratory distress at 18 months after thoracic irradiation was statistically significantly lower in all groups of irradiated ICAM-1-/- mice than it was in all groups of irradiated ICAM-1+/+ mice (overall P = .0036, odds ratio [OR] = 0.19; 95% CI = 0.06 to 0.58).
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To determine whether the difference in incidence of respiratory distress in response to irradiation between ICAM-1+/+ and ICAM-1-/- mice was time-dependent, we repeated this experiment and monitored the mice over a shorter time period for signs of respiratory distress (Fig. 3, B). At 9 months after irradiation, 10% (95% CI = 0% to 29%) of the ICAM-1+/+ mice that received a dose of 14 Gy and 40% (95% CI = 10% to 70%) of the ICAM-1+/+ mice that received a dose of 18 Gy displayed tachypnea and hypomotility (Fig. 3, B
). By contrast, at this same time point, 0% (95% CI = 0% to 31%) of the ICAM-1-/- mice that received a dose of 14 Gy, 10% (95% CI = 0% to 29%) of the ICAM-1-/- mice that received a dose of 16 Gy, and 20% (95% CI = 0% to 44%) of the ICAM-1-/- mice that received a dose of 18 Gy developed tachypnea and hypomotility (Fig. 3, B
). Although the overall incidence of respiratory distress at 9 months after thoracic irradiation was lower in all groups of irradiated ICAM-1-/- mice than it was in all groups of irradiated ICAM-1+/+ mice, that difference was not statistically significant (overall P = .07; general linear model). These findings indicate that the ICAM-1 null mutation may attenuate radiation-induced respiratory distress, but that it does not entirely eliminate radiation-induced pulmonary injury. Moreover, the onset of respiratory insufficiency was time-dependent in both the ICAM-1+/+ and ICAM-1-/- mice.
Mouse Lung Histology Following Actual or Mock Thoracic Irradiation
To determine whether the wild-type ICAM-1 genotype was associated with radiation-induced alveolar thickening and pulmonary fibrosis, we studied histologic sections of lungs from ICAM-1-/- and ICAM-1+/+ mice that had survived for 12 months after receiving 16 Gy of thoracic irradiation. Lung sections were stained with Masson's trichrome stain to visualize collagen deposition. Representative micrographs of stained lung tissue are shown in Fig. 4. The lungs of all mice irradiated with 16 Gy displayed prominent staining around the large blood vessels that was consistent with fibrosis. The lungs of ICAM-1+/+ mice subjected to 16 Gy showed more alveolar thickening and collagen deposition than the lungs of the mock-irradiated control mice (Fig. 4, A
). In comparison, ICAM-1-/- mice subjected to 16 Gy showed no increase in alveolar septal wall thickening and no increase in Masson's trichrome staining as compared with mock-irradiated mice (Fig. 4, C
).
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Collagen Deposition in the Lungs of Irradiated Mice
We used an antibody to collagen III to stain lung sections prepared from ICAM-1+/+ and ICAM-1-/- mice at various times after thoracic irradiation to determine the patterns of collagen deposition in response to radiation-induced pulmonary injury. The lungs of irradiated ICAM-1+/+ mice had higher collagen deposition, primarily in the perivascular regions, 12 months after irradiation with 16 Gy than did the lungs of mock-irradiated mice and ICAM-1-/- mice at 12 months after irradiation (Fig. 5).
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DISCUSSION |
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Previous studies (13) have failed to clarify the role of inflammation in the development of pulmonary fibrosis. For example, C57BL/6 mice, which are prone to radiation-induced fibrosis, develop less inflammation than C3H mice, which are prone to radiation-induced inflammation. Conversely, the inflammation-prone mice develop less fibrosis than the fibrosis-prone mice. Glucocorticoids reduce inflammation but not subsequent fibrosis in the lungs of irradiated rats (23). However, steroids have many nonspecific effects that influence lung physiology (24). The purpose of the present study was to specifically study the role of ICAM-1 in radiation-induced lung injury.
We found that hydroxyproline content and collagen stainingtwo markers of fibrosisincreased in the lungs of ICAM-1+/+ mice following thoracic irradiation. These findings indicate that fibrosis may account for the diminished pulmonary compliance observed in these mice following thoracic irradiation. The role of fibrosis in radiation-induced reduction in pulmonary compliance is also supported by results from previous studies (5,25,26).
Fibrosis may develop in response to inflammation (12). Leukocytes release cytokines, such as tumor necrosis factor- (TNF-
) and transforming growth factor-
(TGF-
), which stimulate collagen production. Fibrosis occurs in foci that surround a nidus of inflammatory cell infiltration, thus implicating inflammatory cells in both the early (pneumonitic) and the late (fibrotic) phases of radiation-induced lung injury (12). However, inflammatory cells are not the only source of TGF-
and TNF-
, and the elimination of inflammation does not entirely prevent the reduced pulmonary compliance and respiratory compromise, as shown in this study. Although ICAM-1 knockout mice serve as a model to study the role of inflammation (27) in the pathogenesis of treatment-related pulmonary fibrosis, this model has limitations. For example, injection of an ICAM-1-blocking antibody into ICAM-1+/+ mice is more efficient than a homozygous knockout of the ICAM-1 gene at preventing tissue injury, indicating that other cell adhesion molecules, such as P-selectin, may also contribute to inflammation (28). This finding suggests that redundant mechanisms of inflammatory cell trafficking may compensate for the effects of a null mutation in the ICAM-1 gene.
In the present study, the reduction in dynamic pulmonary compliance occurred 10 weeks after irradiation of wild-type mice, even though pulmonary inflammation occurred at 5 weeks. This late progression to impaired pulmonary compliance may indicate that radiation-induced injury is multifactorial and related to inflammation, edema, and diminished surfactant. This speculation is supported by the finding that irradiated ICAM-1-/- mice also have diminished pulmonary compliance, albeit to a lesser extent than wild-type mice. We propose that the ICAM-1 null mutation reduces the inflammatory component of lung injury but has little effect on the cytotoxic effects of irradiation. This hypothesis is supported by a recent study (29) that showed that endothelial cell cytotoxicity is the predom-inant event in the pathophysiology of radiation-induced organ injury.
The present study shows that the incidence of tachypnea and hypomotility increases between 9 and 18 months after thoracic irradiation in both ICAM-1+/+ and ICAM-1-/- mice. The radiation doses required to induce a 50% incidence of respiratory distress at 18 months were 18 Gy in ICAM -/- mice but only 16 Gy in ICAM-1+/+ mice. Taken together with the findings that mice bearing the ICAM-1 null mutation showed essentially no radiation-induced pulmonary fibrosis but had attenuated impairment of pulmonary compliance, these results suggest that ICAM-1 expression contributes to the pathogenesis of treatment-related lung injury. ICAM-1 is required for radiation-induced pneumonitis, which supports the link between inflammation and collagen deposition in the irradiated lung. These findings suggest that agents that block ICAM-1 function or expression should be studied for their effects on the prevention of radiation-induced pulmonary fibrosis. In addition, other means of protecting lungs from the cytotoxic effects of ionizing radiation should be considered.
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NOTES |
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REFERENCES |
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1 Down J, Steel G. The expression of early and late damage after thoracic irradiation: a comparison between CBA and C57B1 mice. Radiat Res 1983;96:60310.[Medline]
2 Franko A, Sharplin J, Ward W, Taylor J. Evidence for two patterns of inheritance of sensitivity to induction of lung fibrosis in mice by radiation, one of which involves two genes. Radiat Res 1996;146:6874.[Medline]
3 Franko AJ, Sharplin J, Ward WF, Hinz JM. The genetic basis of strain-dependent differences in the early phase of radiation injury in mouse lung. Radiat Res 1991;126:34956.[Medline]
4 Ward W, Sharplin J, Franko A, Hinz J. Radiation-induced pulmonary endothelial dysfunction and hydroxyproline accumulation in four strains of mice. Radiat Res 1989;120:11320.[Medline]
5 Travis EL. The sequence of histological changes in mouse lungs after single doses of x-rays. Int J Radiat Oncol Biol Phys 1980;6:3457.[Medline]
6 Penney DP, Rubin P. Specific early fine structural changes in the lung following irradiation. Int J Radiat Oncol Biol Phys 1977;2:112332.[Medline]
7 Hopewell J, Calvo W, Jaenke R, Reinhold H, Robbins M, Whitehouse E. Microvasculature and radiation damage. In: Hopewell J, editor. Recent results in cancer research. Vol 130. Berlin (Germany): Springer-Verlag; 1993. p. 153.
8 Steinberg F, Quabeck K, Rehn B, Kraus R, Mohnke M, Costabel U, et al. Lung effects after TBI of mice and bone marrow transplant patients: comparison of experimental and clinical data. In: Hopewell J, editor. Recent results in cancer research. Vol 130. Berlin (Germany): Springer-Verlag; 1993. p. 133143.
9 Ward HE, Kemsley L, Davies L, Holecek M, Berend N. The pulmonary response to sublethal thoracic irradiation in the rat. Radiat Res 1993;136:1521.[Medline]
10 Travis EL. The sequence of histological changes in mouse lungs after single doses of x-rays. Int J Radiat Oncol Biol Phys 1980;6:3457.[Medline]
11 Graham MM, Evans ML, Dahlen, DD, Mahler PA, Rasey JS. Pharmacological alteration of the lung vascular response to radiation. Int J Radiat Oncol Biol Phys 1990;19:32939.[Medline]
12 Franko AJ, Sharplin J, Ghahary A, Barcellos-Hoff MH. Immunohistochemical localization of TGF-B and TNF in the lungs after irradiation. Radiat Res 1997;147:24557.[Medline]
13 Hallahan DE, Clark ET, Kuchibhotla J, Gewertz B, Collins T. E-selectin gene induction by ionizing radiation is independent of cytokine induction. Biochem Biophys Res Commun 1995;217:78495.[Medline]
14 Hallahan DE, Kuchibholta J, Wyble C. Cell adhesion molecules mediate radiation-induced leukocyte adhesion to the vascular endothelium. Cancer Res 1996;56:51506.[Abstract]
15 Hallahan DE, Kuchibhotla J, Wyble C, Sialyl LX. Mimetics attenuate E-selectin-mediated leukocyte adhesion to irradiated human endothelial cells. Radiat Res 1997;147:417.[Medline]
16
Hallahan D, Virudachalam S. ICAM-1 null mutation abrogates the inflammatory response to ionizing radiation. Proc Natl Acad Sci U S A 1997;94:643237.
17 Hallahan DE, Virudachalam S. Ionizing radiation mediates expression of cell adhesion molecules in distinct histological patterns within the lung. Cancer Res 1997;57:20969.[Abstract]
18
Padrid P, Mathur M, Li X, Herrmann K, Qin Y, Cattamanchi A, et al. CTLA41g inhibits airway eosinophilia and hyperresponsiveness by regulating the development of Th1/Th2 subsets in a murine model of asthma. Am J Respir Cell Mol Biol 1998;18:45362.
19 Solway J, Hershenson MB. Structural and functional abnormalities of the airways of hyperoxia-exposed immature rats. Chest 1995;107(3 Suppl):89S93S.[Medline]
20 Dileto CL, Travis EL. Fibroblast radiosensitivity in vitro and lung fibrosis in vivo: comparison between a fibrosis-prone and fibrosis-resistant mouse strain. Radiat Res 1996;146:617.[Medline]
21 Woessner JF Jr. The determination of hydroxyproline in tissue and protein samples containing small proportions of this imino acid. Arch Biochem Biophys 1961;93:440.
22 Haschek WM, Klein-Szanto AJ, Last JA, Reiser KM, Witschi H. Long-term morphologic and biochemical features of experimentally induced lung fibrosis in the mouse. Lab Invest 1982;46:4389.[Medline]
23 Ward HE, Kemsley L, Davies L, Holecek M, Berend N, et al. The effects of steroids on radiation-induced lung disease in the rat. Radiat Res 1993;136:228.[Medline]
24 Gross NJ. Radiation pneumonitis in mice. Some effects of corticosteroids on mortality and pulmonary physiology. J Clin Invest 1980;66:50410.[Medline]
25 Travis EL, Down JD, Holmes SJ, Hobson B. Radiation pneumonitis and fibrosis in mouse lung assayed by respiratory frequency and histology. Radiat Res 1980;84:13343.[Medline]
26 Travis EL, Vojnovic B, Davies EE, Hirst DG. A plethysmographic method for measuring function in locally irradiated mouse lung. Br J Radiol 1979;52:6774.[Medline]
27
Lloyd CM, Gonzalo JA, Salant DJ, Just J, Gutierrez-Ramos JC. Intercellular adhesion molecule-1 deficiency prolongs survival and protects against the development of pulmonary inflammation during murine lupus. J Clin Invest 1997;100:96371.
28 Doerschuk CM, Quinlan WM, Doyle NA, Bullard DC, Vestweber D, Jones ML, et al. The role of P-selectin and ICAM-1 in acute lung injury as determined using blocking antibodies and mutant mice. J Immunol 1996;157:460914.[Abstract]
29
Paris F, Fuks Z, Kang A, Capodieci P, Juan G, Ehleiter D, et al. Endothelial apoptosis as the primary lesion initiating intestinal radiation damage in mice. Science 2001;293:2937.
Manuscript received July 10, 2001; revised February 22, 2002; accepted March 14, 2002.
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