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Quantitative assessment of the specific CD4+ T lymphocyte proliferative response in bovine herpesvirus 1 immune
M Denis1, M J Kaashoek, J T van Oirschot
1Faculty of Veterinary Medicine, University of Liège, Belgium.
Insights
Quantifying bovine herpesvirus 1 (BHV-1) specific CD4+ T cell proliferation in cattle is crucial for vaccine development. A refined limiting dilution method accurately measures this immune response without IL-2 interference.
Area of Science:
- Immunology
- Veterinary Virology
Background:
- Bovine herpesvirus 1 (BHV-1) infection poses significant challenges in cattle.
- Assessing cell-mediated immunity, particularly T cell responses, is vital for evaluating BHV-1 vaccine efficacy.
- Existing methods for quantifying specific T cell proliferation in cattle are often variable and lack precision.
Purpose of the Study:
- To develop a sensitive and quantitative method for measuring CD4+ T cell proliferation specific to BHV-1 in cattle.
- To identify the key cellular players involved in antigen-specific T cell responses to BHV-1.
- To optimize assay conditions for accurate enumeration of BHV-1 specific T cells.
Main Methods:
- Peripheral blood mononuclear cells (PBMCs) from immune cattle were used.
- Proliferative assays were performed with and without BHV-1 antigen.
- Negative selection was employed to isolate specific cell populations.
- Limiting dilution assays were optimized, with and without interleukin-2 (IL-2), to quantify antigen-specific T cells.
Main Results:
- CD4+ T cells were identified as the limiting cell type for BHV-1 antigen-induced proliferation.
- B cells, gamma delta T cells, and CD8+ T cells were not significantly involved in the specific response.
- Interleukin-2 (IL-2) addition led to non-specific proliferation, masking the antigen-specific response.
- A method was established to quantify BHV-1 specific CD4+ T cells by comparing proliferation in the presence and absence of antigen, without IL-2.
Conclusions:
- The developed limiting dilution assay provides a precise and sensitive method for quantifying BHV-1 specific CD4+ T cell immune responses in cattle.
- This method is valuable for assessing the efficacy of BHV-1 vaccine candidates.
- Understanding the role of CD4+ T cells and optimizing assay conditions are key for accurate immunological assessments in veterinary medicine.
Abstract:
We quantified the CD4+ T cell proliferation specific for bovine herpesvirus 1 (BHV-1) in peripheral blood mononuclear cells from cattle. The stimulation index as detected in proliferative assays performed in the presence of BHV-1 antigen is highly variable in immune cattle. By using proliferative assays performed after negative selection we showed that, as expected, CD4+ T cells were the limiting cell type for antigen-induced proliferation. Neither B, gamma delta T nor CD8+ cells seemed to be involved. The limiting dilution method was established to obtain quantitative estimations, namely frequencies of specific T cells. When limiting dilution cultures were supplemented with interleukin-2 (IL-2), an IL-2 induced unspecific cell proliferation masked the specific T cell proliferation. Natural killer cells were not the major cell type involved, but CD4+ lymphocytes themselves seemed to respond to IL-2 irrespective of the presence of antigen. When cultures were performed without addition of IL-2, the frequency of BHV-1 specific proliferative T cells could be obtained by the difference between the frequency of proliferating cells calculated in the presence and absence of antigen. The method provides a sensitive and quantitative means to measure the T cell immune response to BHV-1 vaccine candidates.