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Mesothelial cell apoptosis is confirmed in vivo by morphological change in cytokeratin distribution
E Marchi1, W Liu, V C Broaddus
1Lung Biology Center, San Francisco General Hospital, San Francisco, California 94143-0854, USA.
Abstract:
Apoptosis of mesothelial cells has been demonstrated in vitro but not in vivo. To identify apoptotic pleural cells as mesothelial, we used cytokeratin as a marker and found a striking spheroid, aggregated appearance of cytokeratin in apparently apoptotic mesothelial cells. In in vitro studies, we found that the aggregated cytokeratin pattern correlated with apoptosis in primary mesothelial cells from mice, rabbits, and humans and was not seen with necrosis. In in vivo studies in mice, we then used this cytokeratin pattern to identify and quantitate apoptotic mesothelial cells. Apoptotic mesothelial cells were best harvested by pleural lavage, indicating that they were loosely adherent or nonadherent. Instillation of RPMI 1640 medium or wollastonite for 24 h induced apoptosis in 0.1 +/- 0. 1 (SE) and 1.0 +/- 0.7%, respectively, of all mesothelial cells recovered, whereas instillation of known apoptotic stimuli, crocidolite asbestos (25 microg) for 24 h or actinomycin D plus murine tumor necrosis factor-alpha for 12 h, induced apoptosis in 5. 1 +/- 0.5 and 22.4 +/- 4.5%, respectively (significantly greater than in control experiments, P < 0.05). By analysis of cytokeratin staining, mesothelial cell apoptosis has been confirmed in vivo.
Insights
This study confirms mesothelial cell apoptosis in vivo using cytokeratin staining. This novel method identifies apoptotic cells in the pleura, advancing our understanding of mesothelial cell dynamics.
Area of Science:
- Cell Biology
- Pathology
- Immunology
Background:
- Apoptosis of mesothelial cells has been observed in vitro but not confirmed in vivo.
- Identifying apoptotic pleural cells as mesothelial requires specific markers.
Purpose of the Study:
- To establish a method for identifying and quantifying apoptotic mesothelial cells in vivo.
- To confirm mesothelial cell apoptosis in vivo using a novel cytokeratin staining pattern.
Main Methods:
- Utilized cytokeratin staining to identify a unique aggregated pattern in apoptotic mesothelial cells.
- Developed pleural lavage techniques for harvesting mesothelial cells in vivo.
- Administered various agents (RPMI 1640, wollastonite, crocidolite asbestos, actinomycin D/TNF-alpha) to induce apoptosis in vivo.
Main Results:
- A striking spheroid, aggregated cytokeratin pattern was observed in apoptotic mesothelial cells, distinct from necrosis.
- Pleural lavage effectively recovered apoptotic mesothelial cells, indicating they are loosely adherent or nonadherent.
- Specific stimuli like crocidolite asbestos and actinomycin D/TNF-alpha significantly increased mesothelial cell apoptosis in vivo compared to controls.
Conclusions:
- Mesothelial cell apoptosis can be reliably identified and quantified in vivo using the described cytokeratin staining pattern.
- The findings confirm that mesothelial cells undergo apoptosis in the pleural environment.
- This research provides a crucial tool for studying pleural diseases and mesothelial cell fate.
