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Altered proliferative response by T lymphocytes of Ly-6A (Sca-1) mice.

Stanford WL, Haque S, Alexander R, Liu X, Latour AM, Snodgrass HR, Koller BH, Flood PM - J. Exp. Med. (1997)

Bottom Line: To better understand the function of Ly-6A, we used gene targeting to produce Ly-6A mice which are healthy and have normal numbers and percentages of hematopoietic lineages.This enhanced proliferation is not due to alterations in kinetics of response, sensitivity to stimulant concentration, or cytokine production by the T cell population, and is manifest in both in vivo and in vitro T cell responses.Moreover, T cells from Ly-6A-deficient animals exhibit a prolonged proliferative response to antigen stimulation, thereby suggesting that Ly-6A acts to downmodulate lymphocyte responses.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, University of North Carolina, Chapel Hill, North Carolina 27599-7455, USA.

ABSTRACT
Ly-6A is a murine antigen which is implicated in lymphocyte activation and may be involved in activation of hematopoietic stem cells. Antibody cross-linking studies and antisense experiments have suggested that Ly-6A is a lymphocyte coactivation molecule. To better understand the function of Ly-6A, we used gene targeting to produce Ly-6A mice which are healthy and have normal numbers and percentages of hematopoietic lineages. However, T lymphocytes from Ly-6A-deficient animals proliferate at a significantly higher rate in response to antigens and mitogens than wild-type littermates. In addition, Ly-6A mutant splenocytes generate more cytotoxic T lymphocytes compared to wild-type splenocytes when cocultured with alloantigen. This enhanced proliferation is not due to alterations in kinetics of response, sensitivity to stimulant concentration, or cytokine production by the T cell population, and is manifest in both in vivo and in vitro T cell responses. Moreover, T cells from Ly-6A-deficient animals exhibit a prolonged proliferative response to antigen stimulation, thereby suggesting that Ly-6A acts to downmodulate lymphocyte responses.

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Absence of cell surface Ly-6A expression in Ly-6A−/− mice.  Splenocytes were stained with the PE-conjugated anti–Ly-6A antibody  D7 (12). Stainings of wild-type, heterozygous, and homozygous mutant  splenocytes from littermates were determined. No expression of Ly-6A  on mutant lymphocytes was detected using two other anti–Ly-6A antibodies (data not shown). 16,000 gated events were collected with the  FACScan®. Negative control staining was determined with PE-conjugated anti–rat IgG.
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Figure 2: Absence of cell surface Ly-6A expression in Ly-6A−/− mice. Splenocytes were stained with the PE-conjugated anti–Ly-6A antibody D7 (12). Stainings of wild-type, heterozygous, and homozygous mutant splenocytes from littermates were determined. No expression of Ly-6A on mutant lymphocytes was detected using two other anti–Ly-6A antibodies (data not shown). 16,000 gated events were collected with the FACScan®. Negative control staining was determined with PE-conjugated anti–rat IgG.

Mentions: Analysis of Lymphoid and Myeloid Subpopulations. Flow cytometry was used to verify the absence of Ly-6A expression in homozygous mutant mice (Fig. 2). Approximately 60% of wild-type splenocytes express Ly-6A. Splenocytes from heterozygous animals show a slight decrease in Ly-6A expression intensity; however, splenocytes from homozygotes do not stain with any of three antibodies to Ly-6A (Fig. 2). In addition, phenotypic analysis was performed to determine if the absence of Ly-6A expression in homozygous mutant mice altered the differentiation of various cell populations. Although there are minor variations between littermates, as a population Ly-6A animals (as old as 8 mo) contain normal percentages of B220, TCR-α/β, TCR-γ/δ, CD3, CD4, CD8, Mac-1, and Ly-6G (Gr-1) positive cells in the bone marrow, lymph nodes, spleen, and thymus (data not shown). In addition to Ly-6G, antibodies to other members of the Ly-6 gene family were used to determine if the lack of phenotypic changes is due to compensation by other family members. Ly-6C, Sca-2 (TSA-1), and ThB do not appear to be overexpressed in any hematopoietic tissues (data not shown).


Altered proliferative response by T lymphocytes of Ly-6A (Sca-1) mice.

Stanford WL, Haque S, Alexander R, Liu X, Latour AM, Snodgrass HR, Koller BH, Flood PM - J. Exp. Med. (1997)

Absence of cell surface Ly-6A expression in Ly-6A−/− mice.  Splenocytes were stained with the PE-conjugated anti–Ly-6A antibody  D7 (12). Stainings of wild-type, heterozygous, and homozygous mutant  splenocytes from littermates were determined. No expression of Ly-6A  on mutant lymphocytes was detected using two other anti–Ly-6A antibodies (data not shown). 16,000 gated events were collected with the  FACScan®. Negative control staining was determined with PE-conjugated anti–rat IgG.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2199024&req=5

Figure 2: Absence of cell surface Ly-6A expression in Ly-6A−/− mice. Splenocytes were stained with the PE-conjugated anti–Ly-6A antibody D7 (12). Stainings of wild-type, heterozygous, and homozygous mutant splenocytes from littermates were determined. No expression of Ly-6A on mutant lymphocytes was detected using two other anti–Ly-6A antibodies (data not shown). 16,000 gated events were collected with the FACScan®. Negative control staining was determined with PE-conjugated anti–rat IgG.
Mentions: Analysis of Lymphoid and Myeloid Subpopulations. Flow cytometry was used to verify the absence of Ly-6A expression in homozygous mutant mice (Fig. 2). Approximately 60% of wild-type splenocytes express Ly-6A. Splenocytes from heterozygous animals show a slight decrease in Ly-6A expression intensity; however, splenocytes from homozygotes do not stain with any of three antibodies to Ly-6A (Fig. 2). In addition, phenotypic analysis was performed to determine if the absence of Ly-6A expression in homozygous mutant mice altered the differentiation of various cell populations. Although there are minor variations between littermates, as a population Ly-6A animals (as old as 8 mo) contain normal percentages of B220, TCR-α/β, TCR-γ/δ, CD3, CD4, CD8, Mac-1, and Ly-6G (Gr-1) positive cells in the bone marrow, lymph nodes, spleen, and thymus (data not shown). In addition to Ly-6G, antibodies to other members of the Ly-6 gene family were used to determine if the lack of phenotypic changes is due to compensation by other family members. Ly-6C, Sca-2 (TSA-1), and ThB do not appear to be overexpressed in any hematopoietic tissues (data not shown).

Bottom Line: To better understand the function of Ly-6A, we used gene targeting to produce Ly-6A mice which are healthy and have normal numbers and percentages of hematopoietic lineages.This enhanced proliferation is not due to alterations in kinetics of response, sensitivity to stimulant concentration, or cytokine production by the T cell population, and is manifest in both in vivo and in vitro T cell responses.Moreover, T cells from Ly-6A-deficient animals exhibit a prolonged proliferative response to antigen stimulation, thereby suggesting that Ly-6A acts to downmodulate lymphocyte responses.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, University of North Carolina, Chapel Hill, North Carolina 27599-7455, USA.

ABSTRACT
Ly-6A is a murine antigen which is implicated in lymphocyte activation and may be involved in activation of hematopoietic stem cells. Antibody cross-linking studies and antisense experiments have suggested that Ly-6A is a lymphocyte coactivation molecule. To better understand the function of Ly-6A, we used gene targeting to produce Ly-6A mice which are healthy and have normal numbers and percentages of hematopoietic lineages. However, T lymphocytes from Ly-6A-deficient animals proliferate at a significantly higher rate in response to antigens and mitogens than wild-type littermates. In addition, Ly-6A mutant splenocytes generate more cytotoxic T lymphocytes compared to wild-type splenocytes when cocultured with alloantigen. This enhanced proliferation is not due to alterations in kinetics of response, sensitivity to stimulant concentration, or cytokine production by the T cell population, and is manifest in both in vivo and in vitro T cell responses. Moreover, T cells from Ly-6A-deficient animals exhibit a prolonged proliferative response to antigen stimulation, thereby suggesting that Ly-6A acts to downmodulate lymphocyte responses.

Show MeSH
Related in: MedlinePlus