Regulatory T cell expression of CLA or α(4)β(7) and skin or gut acute GVHD outcomes

Abstract

Regulatory T cells (Tregs) are a suppressive subset of CD4(+) T lymphocytes implicated in the prevention of acute GVHD (aGVHD) after allo-SCT (ASCT). To determine whether increased frequency of Tregs with a skin-homing (cutaneous lymphocyte Ag, CLA(+)) or a gut-homing (α(4)β(7)(+)) phenotype is associated with reduced risk of skin or gut aGVHD, respectively, we quantified circulating CLA(+) or α(4)β(7)(+) on Tregs at the time of neutrophil engraftment in 43 patients undergoing ASCT. Increased CLA(+) Tregs at engraftment was associated with the prevention of skin aGVHD (2.6 vs 1.7%; P=0.038 (no skin aGVHD vs skin aGVHD)), and increased frequencies of CLA(+) and α(4)β(7)(+) Tregs were negatively correlated with severity of skin aGVHD (odds ratio (OR), 0.67; 95% confidence interval (CI), 0.46-0.98; P=0.041) or gut aGVHD (OR, 0.93; 95% CI, 0.88-0.99; P=0.031), respectively. This initial report suggests that Treg tissue-homing subsets help to regulate organ-specific risk and severity of aGVHD after human ASCT. These results need to be validated in a larger, multicenter cohort.

Conflict of interest statement

Conflict of interest

The authors declare no conflict of interest.

Figures

Figure 1

Gating hierarchy to identify Tregs and their tissue-homing subsets using 10-color multiparametric flow cytometry. (a) CD4 gating. Viable T lymphocytes were identified by gating on cells that expressed CD3 while excluding cells expressing CD14 or marking with the amine viability dye (data not shown). CD4+ T cells were then selected by gating on CD4+CD8− cells. (b) Setting gates for additional parameters. Gates for Foxp3, CD25, CD127, CD45R0, α4β7 and CLA were determined in two-parameter comparisons before use in subset analysis using the total CD4+ cell population (from panel a), to insure adequate cell numbers for accurate gating. (c) Data acquisition. Data on Treg subset frequency were acquired by sequentially applying gates established in panel b to the CD25+Foxp3+ Treg population derived from the parent CD4+ cell population (from panel a).

Figure 2

Association between Tregs, CLA+ Tregs and α4β7+ Tregs with aGVHD outcomes. During neutrophil engraftment, the frequency of Tregs, CLA+ Tregs and α4β7+ Tregs was measured by multiparametric flow cytometry. Box plots define the values for median, range, 25th and 75th percentiles. Two-tailed P-values were calculated using logistic regression to assess for differences in the median percentage of Tregs, CLA+ Tregs and α4β7+ Tregs for the following comparisons: (a) grade 0–I aGVHD vs grade II–IV aGVHD; (b) no skin aGVHD vs skin aGVHD; (c) no gut aGVHD vs gut aGVHD; (g) grade 0–I recurrent aGVHD vs grade II–IV recurrent aGVHD; (h) no recurrent skin aGVHD vs recurrent skin aGVHD; (i) no recurrent gut aGVHD vs recurrent gut aGVHD. Scatter plots indicate the percentage of Tregs, CLA+ Tregs and α4β7+ Tregs for patients with varying degrees of aGVHD severity. Line represents median values. The proportional odds model was used to calculate the odds of developing increasing grade of aGVHD (d), increasing stage of skin aGVHD (e) and increasing stage of gut aGVHD (f) for increasing frequencies of Tregs, CLA+ Tregs and α4β7+ Tregs, respectively.