Lentiviral tracking of vascular differentiation in bone marrow progenitor cells

Jeffrey Schmeckpeper, Yasuhiro Ikeda, Arun H.S. Kumar, Pat Metharom, Stephen J. Russell, Noel M. Caplice

Research output: Contribution to journalArticle

17 Scopus citations

Abstract

Lentiviral vectors encoding for identifiable marker genes controlled by lineage-specific promoters can be used to track differentiation of bone marrow progenitors into endothelial cells and/or smooth muscle cells. Human VE-Cadherin and Smoothelin-B promoters were cloned into a self-inactivating lentiviral vector (HR-VECad and HR-SMTHB) and used to drive expression of green fluorescent protein (eGFP). These constructs demonstrated specific promoter activity in mature endothelial and smooth muscle cells respectively in vitro. Lin- bone marrow progenitor cells (Lin- BMCs) in culture were used to test vector ability to track vascular differentiation. HR-VECad transduced Lin- BMCs were plated on collagen-coated slides and grown in endothelial media, while HR-SMTHB transduced Lin- BMCs were cultured on fibronectin-coated slides and grown in smooth muscle media. For in vivo differentiation assessment, lentiviral transduced Lin- BMCs resuspended in Matrigel were injected subcutaneously into C57BL/6J mice. Explants were evaluated for eGFP expression. Lin- BMCs grown in endothelial differentiation media produced groups of polygonal endothelial-like cells by days 16-21. When transduced with HR-VECad vector, these expressed eGFP in distinct cells within the colony by days 18-21, and coexpressed VE-Cadherin and eNOS. Lin- BMCs grown in smooth muscle differentiation media produced spindle-shaped cells between days 10-14 in culture. When transduced with the HR-SMTHB vector, these cells showed eGFP expression at ∼12 days, which increased over time and coexpressed αSMA, calponin and myosin heavy chain. Within Matrigel plugs containing HR-VECad transduced cells, eGFP+ constituted 0.4±0.2% of total cells. In contrast, within Matrigel plugs containing HR-SMTHB transduced cells, eGFP+ cells constituted 0.2±0.1% of total cells. These data demonstrate the feasibility of selectively marking BMC populations for cell fate determination.

Original languageEnglish (US)
Pages (from-to)169-176
Number of pages8
JournalDifferentiation
Volume78
Issue number2-3
DOIs
StatePublished - Sep 1 2009

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Keywords

  • Differentiation
  • Lentiviral vectors
  • Lin bone marrow
  • Smoothelin
  • VE-Cadherin

ASJC Scopus subject areas

  • Molecular Biology
  • Developmental Biology
  • Cell Biology
  • Cancer Research

Cite this

Schmeckpeper, J., Ikeda, Y., Kumar, A. H. S., Metharom, P., Russell, S. J., & Caplice, N. M. (2009). Lentiviral tracking of vascular differentiation in bone marrow progenitor cells. Differentiation, 78(2-3), 169-176. https://doi.org/10.1016/j.diff.2009.01.002