Rapid assessment of in vitro expanded human regulatory T cell function

Area of Science:

• Immunology
• Cellular Biology

Background:

• Human regulatory T cells (Treg) are crucial for suppressing autoreactive immune responses.
• Treg are phenotypically identified by CD4, CD25, CD127(lo/⁻), and FoxP3 expression, with CD45RA differentiating naïve and induced populations.
• Current functional assessment of Treg relies on lengthy 3-5 day proliferation assays.

Purpose of the Study:

• To develop and evaluate a short-term functional assay for assessing Treg potency.
• To establish a reliable surrogate for Treg functional potential, suitable for therapeutic evaluations.

Main Methods:

• Developed a 7-20 hour assay measuring Treg suppression of T cell activation marker expression (CD69, CD154).
• Utilized flow cytometry to analyze T cell activation following stimulation with CD3/CD28 beads.
• Sorted highly purified Treg (CD4+CD25+CD127(lo/⁻)CD45RA+) from healthy volunteers using BD FACSAria™ II.
• Assessed Treg function using autologous peripheral blood mononuclear cells (PBMC) as responder cells after in vitro expansion.

Main Results:

• The short-term assay successfully measured Treg suppression of T cell activation markers.
• Data indicate that this assay is a reliable surrogate for assessing Treg functional potential.
• The method allows for a significantly reduced timeframe compared to traditional proliferation assays.

Conclusions:

• A novel short-term functional assay provides a valuable tool for evaluating Treg potency.
• This assay could accelerate the assessment of Treg for therapeutic applications.
• The findings suggest this method is a reliable alternative to conventional Treg functional assays.