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Phenotypic and functional characteristics of porcine peritoneal mesothelial cells
J Ohan1, M A Gilbert, J P Brouland
1Laboratoire de Chirurgie Vasculaire et Thoracique, Clichy, France.
Abstract:
The objective of this study was to establish a technique to isolate porcine mesothelial cells (PMCs) from omental tissue and to compare them to human mesothelial cells (HMCs). The PMCs were dispersed by collagenase digestion and isolated on a Ficoll layer. Their morphologic and ultrastructural features were assessed at confluence by light and electronic microscopy, and they were characterized by immunohistochemistry using specific HMC markers. PMC proliferation was studied in the presence of growth factors platelet-derived growth factor (PDGF), epidermal growth factor (EGF) or transforming growth factors beta1, beta2, or beta3 (TGF). Fibrinolytic PMC activity was detected by zymography for tissue plasminogen activator (tPA) and by reverse zymography for plasminogen activator inhibitor-1 (PAI-1). The recalcification time of cell lysates was used to define PMC procoagulant activity, and gelatinase zymography was used to detect metalloproteinase production. At confluence, PMCs formed typical cobblestone monolayers and exhibited structural features characteristic of HMCs. Weibel Palade bodies were never seen. Specific HMC markers (HBME1, ME1, WT1) cross-reacted with PMCs. As HMCs and PMCs coexpressed cytokeratin and vimentin, and also expressed vinculin and alpha-actin. Addition of PDGF or EGF to the culture medium stimulated PMC proliferation. PMCs constitutively expressed fibrinolytic and procoagulant activity and secreted MMP9 and MMP2. The technique described in this study allows isolation of mesothelial cells from porcine omental tissue. These porcine cells exhibit a mesothelial phenotype and functional properties similar to those of HMCs. Our data warrant an evaluation of mesothelial cells as targets in several therapeutic strategies with porcine models.
Insights
Researchers developed a method to isolate porcine mesothelial cells (PMCs) from omental tissue, finding they share key characteristics and functions with human mesothelial cells (HMCs). This validates PMCs as a model for studying HMC behavior and potential therapies.
Area of Science:
- Cell Biology
- Comparative Medicine
- Tissue Engineering
Background:
- Mesothelial cells play crucial roles in serosal membrane function.
- Understanding mesothelial cell behavior is vital for developing treatments for conditions affecting serosal linings.
- Porcine models offer potential for preclinical studies due to physiological similarities with humans.
Purpose of the Study:
- To establish a reliable method for isolating porcine mesothelial cells (PMCs) from omental tissue.
- To characterize the morphology, phenotype, and functional properties of isolated PMCs.
- To compare PMCs with human mesothelial cells (HMCs) for potential use in preclinical research.
Main Methods:
- Isolation of PMCs via collagenase digestion and density gradient centrifugation (Ficoll).
- Morphological and ultrastructural assessment using light and electron microscopy.
- Immunohistochemical characterization with human mesothelial cell markers.
- Functional assays for proliferation, fibrinolytic activity, procoagulant activity, and matrix metalloproteinase production.
Main Results:
- Established a technique for isolating PMCs exhibiting a cobblestone morphology.
- PMCs expressed key mesothelial markers (HBME1, ME1, WT1, cytokeratin, vimentin, vinculin, alpha-actin), similar to HMCs.
- Porcine mesothelial cells demonstrated growth factor-responsive proliferation and constitutive fibrinolytic and procoagulant activities, secreting MMP2 and MMP9.
Conclusions:
- The described technique successfully isolates porcine mesothelial cells with characteristics comparable to human mesothelial cells.
- Porcine mesothelial cells serve as a valuable in vitro model for studying mesothelial biology and disease.
- These findings support the use of porcine models for evaluating therapeutic strategies targeting mesothelial cells.