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Identification of a committed T cell precursor population in adult human peripheral blood.

Bruno L, Res P, Dessing M, Cella M, Spits H - J. Exp. Med. (1997)

Bottom Line: At a molecular level, they express the pre-T cell receptor alpha (pT alpha) gene, CD3-gamma, CD-delta and CD-epsilon, and RAG-1 recombination enzyme and have initiated rearrangements in the T cell receptor (TCR)-beta locus (D-J).Moreover, low levels of CD3 epsilon protein, but not of TCR-beta chain, can be detected in their cytoplasm.Our results suggest that CD4+CD3- cells identified in peripheral blood are different from CD3-CD4+CD8- thymocytes and may contain precursors of an extrathymic T cell differentiation pathway.

View Article: PubMed Central - PubMed

Affiliation: Basel Institute for Immunology, Switzerland.

ABSTRACT
Here, we report data concerning the discovery in adult human peripheral blood of a precursor cell population able to differentiate into CD4+CD3+ alpha beta + mature T cells. These cells, which represent 0.1-0.5% of total peripheral blood mononuclear cells (PBMC), express substantial levels of CD4, but lack CD3 surface expression. At a molecular level, they express the pre-T cell receptor alpha (pT alpha) gene, CD3-gamma, CD-delta and CD-epsilon, and RAG-1 recombination enzyme and have initiated rearrangements in the T cell receptor (TCR)-beta locus (D-J). Moreover, low levels of CD3 epsilon protein, but not of TCR-beta chain, can be detected in their cytoplasm. Our results suggest that CD4+CD3- cells identified in peripheral blood are different from CD3-CD4+CD8- thymocytes and may contain precursors of an extrathymic T cell differentiation pathway.

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(A) CD3-γ, CD3-ε,  and CD3-δ transcripts are expressed in pTα+ cells. RT–PCR  analysis with primers specific for  three components (γ, ε, and δ)  of the CD3 signaling molecule  was performed on CD4+  CD3−CD14− cell population.  The T cell line MOLT4 and water were used as positive and  negative controls, respectively.  (B) CD3-ε molecule, but not  TCR β chain, is present in the  cytoplasm of pTα+ cells. Cytoplasmic staining was performed  with a TCR-β–specific (top right  and left) or a CD3-ε–specific (bottom right and left) mAbs on a T  cell clone (right top and bottom)  and on CD4+CD3−CD14−  sorted PBMC (left top and bottom). In the case of TCR-β cytoplasmic staining, mouse anti– human IgG1 was subsequently  used. Dotted histogram lines  represent negative controls that  were the following: for β-cytoplasmic, mouse anti–human  IgG1 alone and for CD3-cytoplasmic, CD20–FITC.
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Figure 5: (A) CD3-γ, CD3-ε, and CD3-δ transcripts are expressed in pTα+ cells. RT–PCR analysis with primers specific for three components (γ, ε, and δ) of the CD3 signaling molecule was performed on CD4+ CD3−CD14− cell population. The T cell line MOLT4 and water were used as positive and negative controls, respectively. (B) CD3-ε molecule, but not TCR β chain, is present in the cytoplasm of pTα+ cells. Cytoplasmic staining was performed with a TCR-β–specific (top right and left) or a CD3-ε–specific (bottom right and left) mAbs on a T cell clone (right top and bottom) and on CD4+CD3−CD14− sorted PBMC (left top and bottom). In the case of TCR-β cytoplasmic staining, mouse anti– human IgG1 was subsequently used. Dotted histogram lines represent negative controls that were the following: for β-cytoplasmic, mouse anti–human IgG1 alone and for CD3-cytoplasmic, CD20–FITC.

Mentions: Although the pTα+ peripheral blood cell population does not express CD3 on the cell surface, as assessed by FACS® analysis, we investigated whether transcripts for the different components of this TCR-associated signalling molecule could be detected. For this purpose, cDNA from CD4+CD3−CD14− sorted cells was amplified with three different sets of primers specific for CD3-γ, CD3-ε, and CD3-δ transcripts, respectively. Analysis of the PCR products showed that CD4+CD3− CD14− cells expressed all three CD3 components, as can be observed in the T cell line MOLT 4 (Fig. 5 A); CD3-γ was found to be expressed in low amount, because it appeared barely detectable on agarose gels after staining with ethidium bromide; nonetheless, the presence of the message was readily detectable after hybridization with a specific internal probe of the blotted PCR products (data not shown).


Identification of a committed T cell precursor population in adult human peripheral blood.

Bruno L, Res P, Dessing M, Cella M, Spits H - J. Exp. Med. (1997)

(A) CD3-γ, CD3-ε,  and CD3-δ transcripts are expressed in pTα+ cells. RT–PCR  analysis with primers specific for  three components (γ, ε, and δ)  of the CD3 signaling molecule  was performed on CD4+  CD3−CD14− cell population.  The T cell line MOLT4 and water were used as positive and  negative controls, respectively.  (B) CD3-ε molecule, but not  TCR β chain, is present in the  cytoplasm of pTα+ cells. Cytoplasmic staining was performed  with a TCR-β–specific (top right  and left) or a CD3-ε–specific (bottom right and left) mAbs on a T  cell clone (right top and bottom)  and on CD4+CD3−CD14−  sorted PBMC (left top and bottom). In the case of TCR-β cytoplasmic staining, mouse anti– human IgG1 was subsequently  used. Dotted histogram lines  represent negative controls that  were the following: for β-cytoplasmic, mouse anti–human  IgG1 alone and for CD3-cytoplasmic, CD20–FITC.
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Related In: Results  -  Collection

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Figure 5: (A) CD3-γ, CD3-ε, and CD3-δ transcripts are expressed in pTα+ cells. RT–PCR analysis with primers specific for three components (γ, ε, and δ) of the CD3 signaling molecule was performed on CD4+ CD3−CD14− cell population. The T cell line MOLT4 and water were used as positive and negative controls, respectively. (B) CD3-ε molecule, but not TCR β chain, is present in the cytoplasm of pTα+ cells. Cytoplasmic staining was performed with a TCR-β–specific (top right and left) or a CD3-ε–specific (bottom right and left) mAbs on a T cell clone (right top and bottom) and on CD4+CD3−CD14− sorted PBMC (left top and bottom). In the case of TCR-β cytoplasmic staining, mouse anti– human IgG1 was subsequently used. Dotted histogram lines represent negative controls that were the following: for β-cytoplasmic, mouse anti–human IgG1 alone and for CD3-cytoplasmic, CD20–FITC.
Mentions: Although the pTα+ peripheral blood cell population does not express CD3 on the cell surface, as assessed by FACS® analysis, we investigated whether transcripts for the different components of this TCR-associated signalling molecule could be detected. For this purpose, cDNA from CD4+CD3−CD14− sorted cells was amplified with three different sets of primers specific for CD3-γ, CD3-ε, and CD3-δ transcripts, respectively. Analysis of the PCR products showed that CD4+CD3− CD14− cells expressed all three CD3 components, as can be observed in the T cell line MOLT 4 (Fig. 5 A); CD3-γ was found to be expressed in low amount, because it appeared barely detectable on agarose gels after staining with ethidium bromide; nonetheless, the presence of the message was readily detectable after hybridization with a specific internal probe of the blotted PCR products (data not shown).

Bottom Line: At a molecular level, they express the pre-T cell receptor alpha (pT alpha) gene, CD3-gamma, CD-delta and CD-epsilon, and RAG-1 recombination enzyme and have initiated rearrangements in the T cell receptor (TCR)-beta locus (D-J).Moreover, low levels of CD3 epsilon protein, but not of TCR-beta chain, can be detected in their cytoplasm.Our results suggest that CD4+CD3- cells identified in peripheral blood are different from CD3-CD4+CD8- thymocytes and may contain precursors of an extrathymic T cell differentiation pathway.

View Article: PubMed Central - PubMed

Affiliation: Basel Institute for Immunology, Switzerland.

ABSTRACT
Here, we report data concerning the discovery in adult human peripheral blood of a precursor cell population able to differentiate into CD4+CD3+ alpha beta + mature T cells. These cells, which represent 0.1-0.5% of total peripheral blood mononuclear cells (PBMC), express substantial levels of CD4, but lack CD3 surface expression. At a molecular level, they express the pre-T cell receptor alpha (pT alpha) gene, CD3-gamma, CD-delta and CD-epsilon, and RAG-1 recombination enzyme and have initiated rearrangements in the T cell receptor (TCR)-beta locus (D-J). Moreover, low levels of CD3 epsilon protein, but not of TCR-beta chain, can be detected in their cytoplasm. Our results suggest that CD4+CD3- cells identified in peripheral blood are different from CD3-CD4+CD8- thymocytes and may contain precursors of an extrathymic T cell differentiation pathway.

Show MeSH