Differential homing receptor expression by alloreactive T cells in anatomic distinct priming sites. CFSE-labeled allogeneic FVB/N-L2G85 (H-2^q, Thy1.1) splenocytes were transplanted into myeloablative conditioned Balb/c recipients (H-2^d, Thy1.2). On day 3 after aHCT, immediately before alloreactive T cells enter aGVHD target organs, cells were isolated from different SLOs for phenotypic comparison by flow cytometry. (A) Representative staining of α4β7 integrin and CCR9 on in vivo primed Thy1.1⁺ donor CD4⁺ T cells (top) and CD8⁺ T cells (bottom). Alloreactive T cells had undergone several cell divisions (color encoded) and markedly up-regulated α4β7 integrin and CCR9 after 5 cell divisions predominantly in mLN and spleen (1 of 4 representative experiments is shown). (B) Bar graphs depict percentage of dividing donor T cells expressing peripheral homing receptors. Of note, intestinal-associated lymphoid organs (PP, mLN) up-regulated mucosal-associated homing receptors, whereas pLNs (cLN and iLN) displayed more pronounced skin homing receptor expression (E-lig, P-lig). Most of the dividing cells in all SLO are CD44⁺. Cells were pooled from 5 mice, 3 independent experiments were performed. Bars depict means plus or minus SD.