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A protocol for phenotypic detection and characterization of vascular cells of different origins in a lung neovascularization model in rodents

Nature Protocols volume 3, pages 388–397 (2008)Cite this article

Abstract

The goal of many current studies of neovascularization is to define the phenotype of vascular cell populations of different origins and to determine how such cells promote assembly of vascular channel. Here, we describe a protocol to immunophenotype vascular cells by high-resolution imaging and by fluorescence-activated flow cytometry in an in vivo rodent model of pulmonary microvascular remodeling. Analysis of cells by this combined approach will characterize their phenotype, quantify their number and identify their role in the assembly of vascular channels.

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Figure 1: Quantification of gold particle density using image processing and analysis.
Figure 2: Representative flow cytometry plots using FlowJo software to analyze cells from lung tissue or from rat peripheral blood.
Figure 3: Inverted images of PDGF-Rβ+ vascular-associated cells in the rat lung after breathing high oxygen.
Figure 4: Schematic of vessel wall thickening by precursor cells in the rat after high oxygen (left); values for gold-labeled antigenic sites quantified by macro for each cell population at each time point (right).

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References

  1. Jones, R. Ultrastructural analysis of contractile cell development in lung microvessels in hyperoxic pulmonary hypertension. Fibroblasts and intermediate cells selectively reorganize nonmuscular segments. Am. J. Pathol. 141, 1491–1505 (1992).

    CAS  PubMed  PubMed Central  Google Scholar 

  2. Jones, R.C., Capen, D., Petersen, B., Jain, R.K. & Duda, D.G. A protocol for a lung neovascularization model in rodents. Nat. Protoc. 3, 378–387 (2008).

    Article  CAS  Google Scholar 

  3. Jones, R.C. & Jacobson, M. Angiogenesis in the hypertensive lung: response to ambient oxygen tension. Cell Tissue Res. 300, 263–284 (2000).

    Article  CAS  Google Scholar 

  4. Jones, R., Capen, D., Jacobson, M. & Munn, L. PDGF and microvessel wall remodeling in adult rat lung: imaging PDGF-AA and PDGF-Rα molecules in progenitor smooth muscle cells developing in experimental pulmonary hypertension. Cell Tissue Res. 326, 759–769 (2006).

    Article  CAS  Google Scholar 

  5. Jones, R., Capen, D. & Jacobson, M. PDGF and microvessel wall remodeling in adult lung: imaging PDGF-Rβ and PDGF-BB molecules in progenitor smooth muscle cells developing in pulmonary hypertension. Ultrastruct. Pathol. 30, 267–281 (2006).

    Article  Google Scholar 

  6. Duda, D.G. et al. Evidence for incorporation of bone marrow-derived endothelial cells into perfused blood vessels in tumors. Blood 107, 2774–2776 (2006).

    Article  CAS  Google Scholar 

  7. Jones, R., Jacobson, M. & Steudel, W. Alpha-smooth-muscle actin and microvascular precursor smooth-muscle cells in pulmonary hypertension. Am. J. Respir. Cell Mol. Biol. 20, 582–594 (1999).

    Article  CAS  Google Scholar 

  8. Jones, R., Steudel, W., White, S., Jacobson, M. & Low, R. Microvessel precursor smooth muscle cells express head-inserted smooth muscle myosin heavy chain (SM-B) isoform in hyperoxic pulmonary hypertension. Cell Tissue Res. 295, 453–465 (1999).

    Article  CAS  Google Scholar 

  9. De Palma, M. et al. Tie2 identifies a hematopoietic lineage of proangiogenic monocytes required for tumor vessel formation and a mesenchymal population of pericyte progenitors. Cancer Cell 8, 211–226 (2005).

    Article  CAS  Google Scholar 

  10. De Palma, M., Venneri, M.A., Roca, C. & Naldini, L. Targeting exogenous genes to tumor angiogenesis by transplantation of genetically modified hematopoietic stem cells. Nat. Med. 9, 789–795 (2003).

    Article  CAS  Google Scholar 

  11. Duda, D.G. et al. Differential transplantability of tumor-associated stromal cells. Cancer Res. 64, 5920–5924 (2004).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

Supported by NIH R01HL 070866 (to RJ), NIH R01CA 115767 (to RKJ) and by a Career Development Award to DGD (from the American Association for Cancer Research-Genentech BioOncology).

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Authors and Affiliations

  1. Department of Anesthesia and Critical Care, Massachusetts General Hospital and Harvard Medical School, 55 Fruit Street, Boston, 02114, Massachusetts, USA

    Rosemary C Jones & Diane E Capen

  2. Center for Regenerative Medicine, Massachusetts General Hospital and Harvard Medical School, 55 Fruit Street, Boston, 02114, Massachusetts, USA

    Kenneth S Cohen

  3. Department of Medicine, Section of Hematology/Oncology, University of Chicago, 5841 South Maryland Avenue, Chicago, 60637, Illinois, USA

    Kenneth S Cohen

  4. Steele Laboratory for Tumor Biology, Massachusetts General Hospital and Harvard Medical School, 55 Fruit Street, Boston, 02114, Massachusetts, USA

    Lance L Munn, Rakesh K Jain & Dan G Duda

Authors
  1. Rosemary C Jones
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  2. Diane E Capen
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  3. Kenneth S Cohen
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  4. Lance L Munn
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  5. Rakesh K Jain
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  6. Dan G Duda
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Correspondence to Rosemary C Jones.

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Jones, R., Capen, D., Cohen, K. et al. A protocol for phenotypic detection and characterization of vascular cells of different origins in a lung neovascularization model in rodents. Nat Protoc 3, 388–397 (2008). https://doi.org/10.1038/nprot.2007.537

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