IV3à(NeuGcà2-8NeuGc)-Gg4Cer Is Restricted to CD4+ T Cells Producing Interleukin-2 and a Small Population of Mature Thymocytes in Mice

Kyoko Nakamura,Hidenori Suzuki,Yoshio Hirabayashi,Akemi Suzuki

doi:10.1074/jbc.270.8.3876

Kyoko Nakamura, Hidenori Suzuki + Show 2 more

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https://doi.org/10.1074/jbc.270.8.3876

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Abstract

Monoclonal antibody YK-3 was established by immunization with IV3 alpha (NeuGc alpha 2-8NeuGc)-Gg4Cer (GD1c-(NeuGc-NeuGc-)), and its epitope was determined to be NeuGc alpha 2-8NeuGc alpha 2-3Gal beta 1. Thin layer chromatography immunostaining with YK-3 detected only GD1c-(NeuGc-NeuGc-) among the gangliosides of mouse thymocytes and splenocytes. Immunohistochemical staining with YK-3 visualized the medulla of mouse thymus and T cell-dependent areas of mouse spleen and mesenteric lymph nodes. Two-color flow cytometry demonstrated that GD1c (NeuGc-NeuGc-) was expressed on a quarter of CD3+ mature thymocytes and strongly expressed on three quarters of CD4+ T cells in the spleen, lymph nodes, and peripheral blood but not on CD8+ T cells or B cells. GD1c (NeuGc-NeuGc-)-positive cells and negative cells were separated by panning with YK-3 on Petri dishes into adherent and nonadherent fractions. Following stimulation with concanavalin A, adherent cells, predominantly GD1c (NeuGc-NeuGc-)+, produced more interleukin-2 (IL-2) and markedly less interleukin-4 (IL-4) than nonadherent cells. This conclusion is supported by data obtained by lysis of cells by YK-3 and complement. These data indicate that the cell surface expression of GD1c (NeuGc-NeuGc-) is restricted to a small number of mature thymocytes and a subset of CD4+ T cells, which produce abundant IL-2 and very little IL-4, suggesting that GD1c(NeuGc-NeuGc-) is an excellent marker for mouse naive T or T helper 1-like cells in vivo.

Highlights

From the Wepartment of Membrane Biochemistry and §Deparlment of Cardiovascular Research, Tokyo Metropolitan Institute of Medical Science, Honkomagome, Bunhyo-ku, Tokyo, 113, Japan and the 1\Laboratory for Glyco Cell Biology, Frontier Research Program, The Institute of Chemical and Physical Research, Wako-shi, Saitama, 351-01, Japan
Using monoclonal antibody YK-3, we demonstrated that the expression ofGndNeuGc-NeuGc-) is limited to a small number of mature CD3+ thymocytes and a subset of CD4+ T cells, which produce abundant IL-2 and little IL-4
GDIc(NeuGc-NeuGc-) should be an excellent marker for mouse naive T or T helper 1 (Thl)-like cells in vivo. It is still a matter of controversy how naive T helper cells differentiate into Thl or T helper 2 (Th2) cells, there appear to be at least two distinguishable T helper subsets in vitro

Summary

To whom correspondence should be addressed

Dept. of Membrane Biochemistry, Tokyo Metropolitan lnst. of Medical Science, 18-22, Honkomagome3-chome,Bunkyo-ku, Tokyo113,Japan. (NeuGca2-8NeuGc)-Gg4Cer (GDlc(NeuGc-NeuGc-» (Table I) in mouse thymocytes [15]. These gangliosides are all synthesized by extension of Gg4Cer, and activation of the biosynthetic pathways for Gg4Cer is a unique characteristic of mouse immune tissues. In mature or stimulated T cells, which contrasts with the evidence that gangliosides synthesized from GM1 restricted to B lymphocytes [18]. In the course of our studies on gangliosides of mouse lymphocytes, we noted that GDlc(NeuGc-NeuGc-) is a major disialoganglioside in thymocytes and occurs in splenocytes as well.

The abbreviations used are

EXPERIMENTAL PROCEDURES

Findings

DISCUSSION