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Determination of Absolute Counts of Circulating Regulatory T Cells in Cynomolgus Macaques Using an Optimized Flow Cytometric Method

By Stacie M. Clark, Padma K. Narayanan, Madeline and M. Fort


Regulatory T cells (Tregs) are a rare subset of lymphocytes that inhibit the activation and effector functions of T cells and are important regulators of immune responses. Although Tregs are well characterized in humans and rodents, little is known about their immunophenotyping (IP) profile in cynomolgus macaques (Macaca fascicularis), which is an important species for pharmacological and toxicological evaluation of potential immune modulators because of their similar physiologic, genetic, and metabolic response patterns to humans. The authors have developed an immunophe-notyping panel using a high-throughput 96-well microtiter plate–based assay to detect circulating Tregs (CD3þCD4þCD25hiFoxP3þ) and have deter-mined the normal range for the number of Tregs in naive healthy cynomolgus macaques to be 56.4 to 179.7 cells/mL (mean+ SEM 113.6+ 5.1 cells/mL; n 25). Furthermore, the authors compared the resulting FoxP3þ Treg profiles with a CD127lo cell-surface panel (CD3þCD4þCD25hi CD127lo) and found a close correlation between the absolute numbers of CD3þCD4þCD25hiFoxP3þ and CD3þCD4þCD25hiCD127lo cells (mean + SD 120+ 8.0 cells/mL). Quantification of circulating Tregs in cynomolgus macaques in this high-throughput assay may help to identify drug candidates that affect this rare, but critical, immunoregulatory cell population. Keywords: flow cytometry; regulatory T cells; immunophenotyping; FoxP3; CD127; cynomolgus macaques The impact on immune status and function in preclinical studies in animals is an important component of safety assess-ment of immunomodulatory agents. Although the benefits o

Year: 2016
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