A CD2-green fluorescence protein-transgenic mouse reveals very late antigen-4-dependent CD8+ lymphocyte rolling in inflamed venules

K. Singbartl, J. Thatte, M. L. Smith, K. Wethmar, K. Day, K. Ley

Research output: Contribution to journalArticle

49 Citations (Scopus)

Abstract

Intravital microscopy allows detailed analysis of leukocyte trafficking in vivo, but fails to identify the nature of leukocytes investigated. Here, we describe the development of a CD2-enhanced green fluorescence protein (EGFP)-transgenic mouse to characterize lymphocyte trafficking during inflammation in vivo. A CD2-EGFP plasmid construct including the CD2 promoter, the EGFP transgene, and the CD2 locus control region was injected into B6CBA/F1 pronuclei. EGFP+ offspring were backcrossed into C57BL/6 mice for six generations. Flow cytometry demonstrated that all peripheral blood EGFP+ cells were positive for CD2 and negative for the granulocyte Ag Ly 6-G (GR-1). EGFPhigh cells stained positive for CD2, CD3, CD8, TCR β-chain, and NK1.1 but did not express the B cell and monocyte markers CD45RA, CD19, and CD11b. In vitro stimulation assays revealed no difference in lymphocyte proliferation and IL-2 secretion between EGFP+ and EGFP- mice. Intravital microscopy of untreated or TNF-α-treated cremaster muscle venules showed EGFP+ cells in vivo, but these cells did not roll or adhere to the vessel wall. In cremaster muscle venules treated with both TNF-α and IFN-γ, EGFPhigh cells rolled, adhered, and transmigrated at a rolling velocity slightly higher (11 μm/s) than that of neutrophils (10 μm/s). Blocking α4 integrin with amAb increased rolling velocity to 24 μm/s. These findings show that CD8+ T cells roll in TNF-αIFN-γ-pretreated vessels in vivo via an α4 integrin-dependent pathway.

Original languageEnglish (US)
Pages (from-to)7520-7526
Number of pages7
JournalJournal of Immunology
Volume166
Issue number12
DOIs
StatePublished - Jun 15 2001

Fingerprint

Integrin alpha4beta1
Venules
Transgenic Mice
Fluorescence
Lymphocytes
Proteins
Abdominal Muscles
Integrins
Leukocytes
Locus Control Region
Transgenes
Inbred C57BL Mouse
Granulocytes
Interleukin-2
Monocytes
Flow Cytometry
Neutrophils
Plasmids
B-Lymphocytes
Inflammation

All Science Journal Classification (ASJC) codes

  • Immunology

Cite this

Singbartl, K. ; Thatte, J. ; Smith, M. L. ; Wethmar, K. ; Day, K. ; Ley, K. / A CD2-green fluorescence protein-transgenic mouse reveals very late antigen-4-dependent CD8+ lymphocyte rolling in inflamed venules. In: Journal of Immunology. 2001 ; Vol. 166, No. 12. pp. 7520-7526.
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A CD2-green fluorescence protein-transgenic mouse reveals very late antigen-4-dependent CD8+ lymphocyte rolling in inflamed venules. / Singbartl, K.; Thatte, J.; Smith, M. L.; Wethmar, K.; Day, K.; Ley, K.

In: Journal of Immunology, Vol. 166, No. 12, 15.06.2001, p. 7520-7526.

Research output: Contribution to journalArticle

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