Memory CD8 T cells do not retain α4β7 expression regardless of anatomical location or immunization route. Naive Thy1.1⁺ P14 were transferred to naive mice. (A and B) The next day, mice were infected intranasally with 500 pfu of recombinant influenza virus that expresses gp33. 5 (A) or 86 (B) d later, lymphocytes were isolated from the indicated tissues and stained with α-Thy1.1, CD8, and α4β7 or CD44. Plots are gated on CD8⁺ lymphocytes. (C and D) Alternatively, 200 µg DNA that expresses the glycoprotein of LCMV under control of the CMVie promoter was administered intramuscularly into both anterior tibialis muscles. (C) 9 and 12 d later, Thy1.1⁺ cells were examined for expression of α4β7 (gated on CD8⁺ Thy1.1⁺ lymphocytes). (D) 12 d after immunization, lymphocytes were isolated from spleen and IEL and stained with α-CD8α, Thy1.1, and CD44 antibodies. Plots are gated on CD8α⁺ lymphocytes. (E) Naive Thy1.1⁺ P14 were transferred to naive C57BL/6J mice. Control mice received normal drinking water, whereas treated mice were exposed to 2 µg/ml FTY720 in the drinking water ad libitum for the duration of the experiment. The next day, both groups of mice were immunized with LCMV, and α4β7 expression among Thy1.1⁺ P14 in spleen was compared among control mice (black line), FTY720-treated mice (dashed line), and CD44^lo CD8 T cells isolated from spleens of naive mice (gray histogram). Plots are gated on Thy1.1⁺ CD8⁺ lymphocytes. All data are representative of two experiments with at least three mice per group in each experiment.