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VCAM-1 influences lymphocyte proliferation and cytokine production during mixed lymphocyte responses
N W Lukacs1, R M Strieter, H L Evanoff
1Department of Pathology, University of Michigan Medical School, Ann Arbor 48197.
Insights
Vascular Cell Adhesion Molecule-1 (VCAM-1) plays a key role in allogeneic immune responses. Inhibiting VCAM-1 significantly reduces T cell proliferation and cytokine production during mixed lymphocyte reactions (MLR).
Area of Science:
- Immunology
- Cellular and Molecular Immunology
- Transplantation Immunology
Background:
- The allogeneic mixed lymphocyte reaction (MLR) is a model for studying immune responses to foreign cells.
- Adhesion molecules are known to be involved in MLR, but the specific role of VCAM-1 was not fully understood.
Purpose of the Study:
- To investigate the role of VCAM-1 in the generation of allogeneic MLR.
- To determine the impact of VCAM-1 on T lymphocyte activation, proliferation, and cytokine production in MLR.
Main Methods:
- Reverse transcription-polymerase chain reaction (RT-PCR) to detect VCAM-1 mRNA expression.
- Immunolocalization to identify VCAM-1 on specific cell types.
- Inhibition assays using anti-VCAM-1 monoclonal antibodies (mAbs).
- Measurement of cytokine production (IL-2, TNF-alpha, MIP-1 alpha).
Main Results:
- VCAM-1 mRNA expression peaked between 12 and 48 hours during MLR.
- VCAM-1 was localized on adherent mononuclear phagocytes.
- Anti-VCAM-1 mAbs inhibited MLR proliferation by over 70%, even when added as late as Day 2.
- Anti-VCAM-1 significantly inhibited IL-2, TNF-alpha, and MIP-1 alpha production.
Conclusions:
- VCAM-1 is expressed during allogeneic MLR and contributes significantly to the immune response.
- Targeting VCAM-1 may be a therapeutic strategy to modulate T cell activation and cytokine production in allospecific immune responses and allograft rejection.
Abstract:
The allogeneic mixed lymphocyte reaction (MLR) is regarded as an effective model for examining the events which occur during an allospecific immune response. Numerous studies have delineated the role of adherence molecule interactions during the MLR response. In the present study we have identified VCAM-1 as having a contribution to the generation of an allogeneic MLR response. These findings may have broad implications in vivo during antigen-specific and allograft rejection events. RT-PCR analysis was initially used to examine whether VCAM-1 mRNA expression was observed during MLR responses and demonstrated peak expression between 12 and 48 hr of culture. Immunolocalization of VCAM-1 on adherent mononuclear phagocytes, but not nonadherent lymphocytes, from MLR cultures verified its expression during this response. Addition of anti-VCAM-1 mAbs to MLR assays inhibited the proliferative response by over 70%, while addition of anti-VCAM-1 as late as Day 2 of the assay allowed significant inhibition of the proliferative response. This correlated with peak expression of VCAM-1 mRNA observed as late as 48 hr in RT-PCR analyses. In further studies, anti-VCAM-1 significantly inhibited peak expression of IL-2 on Days 3 and 4, while TNF-alpha production was diminished at 30 min and 1, 96, and 120 hr of culture, compared to control cultures. The production of macrophage inflammatory protein-1 alpha (MIP-1 alpha), a chemotactic cytokine which has an important role in vivo for the recruitment of leukocytes to a site of inflammation, was also significantly inhibited during peak production on Days 4 and 5 of the MLR assay. This study demonstrates novel findings of VCAM-1 expression during an allogeneic MLR response. The expression of VCAM-1 may have important implications during allospecific immune responses for the activation and proliferation of T lymphocytes as well as cytokine production.