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. 2024 Feb 22;9(7):e167457. doi: 10.1172/jci.insight.167457

Figure 7. Loss of suppressive capacity of migrated human Tregs is restored by rapamycin.

Figure 7

(A and B) Migrated HD-derived Tregs were treated with rapamycin (2 μM) for 4 hours, washed, and cocultured with Teff in a suppression assay. Showing representative plots of migrated Tregs with and without (vehicle) rapamycin treatment (B; 1:1). (C and D) HD-derived Tregs were treated with rapamycin (2 μM) for 4 hours and put on the Boyden chamber migration assay. After 24 hours, migrated Tregs were collected and cocultured with Teff. Showing representative plots of migrated Tregs with and without (vehicle) rapamycin treatment (D; 1:1). Percentage proliferation represents CellTrace dilution of Teff. Cell ratio is given as Teff/Treg. Relative proliferation is normalized to 1:0 condition (100%). Gating in Supplemental Figure 3. n = 3–6; 2-way ANOVA with Bonferroni’s multiple-comparison test. (E–G) Frozen PBMCs of HD and people with uRRMS were thawed, and Tregs were sorted by FACS. Immediately after isolation, Tregs were treated with rapamycin (2 μM) or vehicle for 4 hours, washed, and cocultured with Teff in a suppression assay (1:1). Showing representative plots of migrated Tregs with and without rapamycin treatment of HD (F) and persons with uRRMS (G). Horizontal bars represent group mean. n = 5 (HD), n = 5 (uRRMS); Wilcoxon test. **P < 0.01; ****P < 0.0001.