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In situ pulmonary vascular perfusion for improved recovery of pulmonary intravascular macrophages
A A Fowler1, P D Carey, C J Walsh
1Department of Internal Medicine, Medical College of Virginia, Richmond 23298.
Abstract:
The microcirculation contains mononuclear phagocytes, with features characteristic of macrophages, adhered to luminal capillary surfaces by intercellular adhesion plaques. These pulmonary intravascular macrophages may play an important role in regulating lung vascular tone and capillary permeability, and may modulate capillary endothelial cell growth and replication by the secretion of soluble mediators (i.e., arachidonate metabolites, cytokines). This study describes a technique which utilizes in situ lung perfusion to remove intravascular macrophages in large numbers from the microcirculation of porcine lung (n = 26). This technique yielded 3.8 +/- 0.5 x 10(8) (mean +/- SEM) mononuclear cells which were highly phagocytic toward particulate carbon (phagocytic index, 80 +/- 6%). Harvested mononuclear phagocytes reestablished intercellular adhesion plaques when placed on small vessel porcine pulmonary artery endothelial cell monolayers and exhibited histochemical characteristics typical of monocyte/macrophage lineage cells. Mononuclear cells obtained from lung microcirculation displayed size heterogeneity varying from 10.4 to 16.5 microns in diameter. Both large and small cell populations phagocytosed particulate carbon. Morphometric studies performed on collagenase-treated lung demonstrated that in situ perfusion removed significant numbers of intravascular macrophages in lung capillaries. The technique described permits the rapid removal of anchored mononuclear phagocytes from lung capillaries with minimal postmortem delay.
Insights
Researchers developed a new method to isolate pulmonary intravascular macrophages from pig lungs. This technique efficiently removes these lung immune cells for further study of their role in lung health and disease.
Area of Science:
- Pulmonary immunology
- Vascular biology
- Cellular biology
Background:
- The lung microcirculation harbors mononuclear phagocytes, resembling macrophages, attached to capillary surfaces.
- These pulmonary intravascular macrophages are implicated in regulating lung vascular tone, capillary permeability, and endothelial cell growth via secreted mediators.
Purpose of the Study:
- To describe a novel technique for the efficient removal of intravascular macrophages from the porcine lung microcirculation.
- To characterize the isolated cells and validate the efficacy of the perfusion method.
Main Methods:
- In situ lung perfusion of porcine lungs (n=26) to isolate intravascular macrophages.
- Phagocytosis assays using particulate carbon to assess cell function.
- Histochemical analysis and morphometric studies to characterize cell lineage and removal efficacy.
Main Results:
- The technique yielded a large number of mononuclear cells (3.8 ± 0.5 x 10^8) with high phagocytic activity (80 ± 6%).
- Isolated cells re-formed adhesion plaques on endothelial cells and showed monocyte/macrophage characteristics.
- Morphometric analysis confirmed significant removal of intravascular macrophages from lung capillaries.
Conclusions:
- The described in situ lung perfusion technique effectively isolates large quantities of functional intravascular macrophages from the lung microcirculation.
- This method allows for rapid collection of these cells with minimal delay, facilitating further research into their biological roles.