Laser-Based Microscopic Approaches: Application to Cell Signaling in Environmental Lung Disease

Taatjes, Douglas J.; Palmer, Cathy J.; Pantano, Cristen; Hoffmann, Scott; Cummins, Andrew; Mossman, Brooke T.

doi:10.2144/01314rv01

Cited by 22 publications

(14 citation statements)

References 26 publications

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“…LCM is a powerful tool that has been used in several pulmonary studies (3,4,10,34). Several investigators have combined LCM with microarray analysis to study gene expression patterns in specific pulmonary cell types or structures (4,25,34).…”

Section: Discussionmentioning

confidence: 99%

“…Several investigators have combined LCM with microarray analysis to study gene expression patterns in specific pulmonary cell types or structures (4,25,34). The methods for accomplishing this always include amplification of the RNA isolated from LCM-dissected samples because the amount of RNA that can be recovered from the LCM samples is very small (22).…”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Galectin-1 in secondary alveolar septae of neonatal mouse lung

Foster¹,

Goss²,

George³

et al. 2006

American Journal of Physiology-Lung Cellular and Molecular Phys

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Galectin-1 in secondary alveolar septae of neonatal mouse lung

Foster¹,

Goss²,

George³

et al. 2006

American Journal of Physiology-Lung Cellular and Molecular Phys

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“…LSC was performed with C10 cells on coverslips using a laser scanning cytometer with WinCyte 3.3 data analysis software (CompuCyte, Cambridge, MA) as described previously (19). To evaluate cells for expression of BrdUrd and cyclin D1, the primary contouring parameter was set on orange-red fluorescence of PI-stained nuclei with detector gain voltages set for a maximum of 75% saturation for the brightest pixel event scanned.…”

Section: Methodsmentioning

confidence: 99%

The Duration of Nuclear Extracellular Signal-Regulated Kinase 1 and 2 Signaling during Cell Cycle Reentry Distinguishes Proliferation from Apoptosis in Response to Asbestos

et al. 2004

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Asbestos exposure causes activation of extracellular signal-regulated kinases 1 and 2 (ERK1/2) in lung epithelial cells, the targets of asbestosassociated lung carcinomas. The functional significance of ERK1/2 activation in pulmonary epithelial and mesothelial cells is unclear. Using serum-stimulated mouse alveolar type II epithelial cells as a model for cell cycle reentry, we show that the duration of phospho-ERK1/2 in the nucleus determines cell fate in response to crocidolite asbestos. In response to 10% serum, a proliferative stimulus, phosphorylated ERK1/2 initially accumulated in the nucleus, and reduction of nuclear phospho-ERK1/2 after 2 to 4 hours was followed by expression of cyclin D1 and S-phase entry. Low levels of asbestos (<0.5 g/cm 2 ) promoted S-phase entry in low (2%) serum through an epidermal growth factor receptor-dependent pathway but did not promote cell cycle progression or induce apoptosis in the presence of high (10%) serum-containing medium. Higher levels of asbestos (1.0 to 5.0 g/cm 2 ) prolonged the localization of phospho-ERK1/2 in the nucleus in the presence of high serum, impeded S-phase entry, and induced apoptosis in a dose-dependent manner. Immunofluorescence microscopy indicated that the duration of signaling by phospho-ERK1/2 in the nucleus was predictive of cell fate at any concentration of asbestos. After 8 hours of exposure, cells with nuclear phospho-ERK1/2 also were positive for nuclear localization of apoptosis-inducing factor (AIF), an early event in apoptosis. In contrast, asbestos-exposed cells that displayed cytoplasmic phospho-ERK1/2 at 8 hours expressed cyclin D1 and proceeded to S phase. Our studies show that prolonged localization of phospho-ERK1/2 in the nucleus is incompatible with expression of cyclin D1 and is predictive of asbestos-associated cell death by AIF, thereby providing an approach for determining cell fate in asbestos-induced tumorigenesis.

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“…The primary contouring parameter used to detect and quantify cells was set on the red fluorescence emitted by the propidium iodide as previously described. 5,18 Detector gain voltages were set so that the brightest pixels scanned were no more than 75% of saturated values. The threshold contour was set within the nucleus at a distance one-third from the nuclear border, using the instrument's scan data display function.…”

Section: Cell Immunostaining and Lscmentioning

confidence: 99%

Persistent Localization of Activated Extracellular Signal-Regulated Kinases (ERK1/2) Is Epithelial Cell-Specific in an Inhalation Model of Asbestosis

Cummins

Palmer

Mossman

et al. 2003

The American Journal of Pathology

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Asbestos fibers up-regulate the extracellular signal-regulated kinase (ERK1/2) pathway in mesothelial and pulmonary epithelial cells in vitro, but the cell-type expression patterns and intracellular localization of activated, ie, phosphorylated, ERK in the lung after inhalation of asbestos are unclear. C57/BL6 mice were exposed to 7-mg/m 3 air of crocidolite asbestos for 5 and 30 days, the times required for the development of epithelial cell hyperplasia and fibrotic lesions, respectively. Exposure to asbestos caused striking increases in both unphosphorylated and phosphorylated ERK (p-ERK), which were most marked at 30 days and co-localized in bronchiolar and alveolar epithelial cells using an antibody to cytokeratin. Alveolar macrophages, detected with an anti-macrophage antibody, did not express p-ERK. p-ERK was localized at the apical cell surface of bronchiolar and alveolar type II epithelial cells exposed to asbestos fibers, and was most marked in areas of epithelial hyperplasia in association with fibrotic lesions. Because translocation of p-ERK to the nucleus is associated with activation of early response genes and transcription factors, laser scanning cytometry was used to determine the kinetics of activation and nuclear translocation of p-ERK in an alveolar type II epithelial cell line in vitro after exposure to asbestos or the ERK stimuli, epidermal growth factor, or H 2 O 2 . Results showed that cytoplasmic to nuclear translocation of p-ERK occurred in a protracted manner in cells exposed to asbestos

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Laser-Based Microscopic Approaches: Application to Cell Signaling in Environmental Lung Disease

Cited by 22 publications

References 26 publications

Galectin-1 in secondary alveolar septae of neonatal mouse lung

Galectin-1 in secondary alveolar septae of neonatal mouse lung

The Duration of Nuclear Extracellular Signal-Regulated Kinase 1 and 2 Signaling during Cell Cycle Reentry Distinguishes Proliferation from Apoptosis in Response to Asbestos

Persistent Localization of Activated Extracellular Signal-Regulated Kinases (ERK1/2) Is Epithelial Cell-Specific in an Inhalation Model of Asbestosis

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