TR3-56 and Treg Regulatory T Cell Subsets as Potential Indicators of Graft Tolerance Control in Kidney Transplant Recipients

Abstract

Identification of early signatures of immune rejection represents a key challenge in the clinical management of kidney transplant. To address such an issue, we enrolled 53 kidney transplant recipients without signs of graft rejection, no infectious episodes and no change in the immunosuppressive regimen in the last 6 months. An extensive immune profile revealed increased activation of the T cells, a decreased amount and growth ability of the Treg and a higher level of the T_R3-56 regulatory T cell subset, described by us as involved in the preferential control of cytotoxic T lymphocytes. In renal transplant recipients, the high level of the T_R3-56 cells associates with a reduction in both the amount and the growth ability of the Treg. Moreover, when the transplanted subjects were categorised according to their stable or unstable disease status, as defined by changes in serum creatinine ≥0.2 mg/dL in two consecutive detections, a higher T_R3-56 level and defective Treg growth ability were observed to characterise patients with unstable graft control. Further studies are required to substantiate the hypothesis that immune profiling, including T_R3-56 evaluation, might represent a valuable diagnostic tool to identify patients at risk of developing significant anti-donor allo-immune responses.

Keywords: TR3-56; Treg; immune profile; kidney transplant recipients; regulatory T cells.

Figure 1

Increased amount of the circulating CTL and of the T_R3-56 regulatory T cells, higher expression of the CD54 activation molecule on T cell effectors and decreased amount and growth ability of the Treg subset characterise a cohort of allograft kidney recipients showing no rejection episodes, no infections and no changes in immuno-suppression therapy in the previous six months. White and grey columns indicate data obtained in healthy controls and kidney transplanted subjects, respectively. (A,B) Indicate percentage and number of circulating T, CD4⁺ and CD8⁺ T lymphocytes, as indicated; (C) Indicates CD4/CD8 ratio; (D) Refers CD54 expression level in CD4⁺ and CD8⁺ T lymphocytes, as indicated; as detailed in the Section 4, CD54 expression on the T cell effectors has been expressed as ratio of the mean intensity fluorescence (MIF) value for CD4⁺ and CD8⁺ T cells and the control MIF value obtained after staining the same cell populations with the isotype control mAb, as described [24]. (E,F) Indicate percentage and number of the circulating T_R3-56 regulatory T cells, respectively; (G,H) Refer to percentage and number of the circulating Treg population; (I,J) Indicate the growth ability of the circulating T_R3-56 and Treg populations, as represented by their intracellular expression of the ki67 molecule; Statistical evaluation of data was performed by means of the Mann–Whitney test. Statistical significance values are indicated.

Figure 2

Allograft kidney recipients show association of highest level of circulating T_R3-56 regulatory T cells with significant decrease in the Treg growth ability and increasing CD54 expression by the CD4⁺ T cell population. White columns indicate healthy controls; light and dark grey columns indicate transplanted subjects showing circulating T_R3-56 levels <9.16% or ≥9.16% of the T cell population, respectively; the 9.16 cut-off value was obtained by increasing by three standard errors the median value observed in healthy controls (see patient and method section for details). (A,B) Indicate percentage and number of circulating T, CD4⁺ and CD8⁺ T lymphocytes; (C) Shows CD4/CD8 ratio; (D) Refers to CD54 expression level in CD4⁺ and CD8⁺ T lymphocytes; as detailed in Section 4, CD54 expression on the T cell effectors was expressed as a ratio of the mean intensity fluorescence (MIF) value for CD4⁺ and CD8⁺ T cells and the control MIF value obtained after staining the same cell populations with the isotype control mAb. (E,F) Indicate the percentage and number of the circulating Treg in the different cohorts; (G) Indicates the growth ability of the circulating Treg population, as represented by their intracellular expression of the ki67 molecule; Statistical evaluation of the data was performed by means of the Mann–Whitney test. Statistical significance values are indicated.

Figure 3

Increasing amount of circulating T_R3-56 lymphocytes and reduced growth ability of the Treg population characterise kidney transplant recipients showing unstable control of the graft. White columns indicate healthy controls; light and dark blue columns indicate transplanted subjects categorised, according to their clinical and laboratory profile, as belonging to the Stable or Unstable transplant recipient sub-groups, respectively. See Patient and Methods section for details. (A,B) Indicate percentage and number of circulating T, CD4⁺ and CD8⁺ T lymphocytes; (C) Indicates CD4/CD8 ratio; (D) Refers to CD54 expression level in CD4⁺ and CD8⁺ T lymphocytes; CD54 expression on the T cell effectors was expressed as a ratio of the mean intensity fluorescence (MIF) value for CD4⁺ and CD8⁺ T cells and the control MIF value obtained after staining the same cell populations with the isotype control mAb, as described [28]. (E,F) Show percentage and number of the circulating T_R3-56 lymphocytes; (G,H) Show percentage and number of circulating Treg; (I,J) Indicate the growth ability, as represented by their expression of the ki67 molecule, of the circulating T_R3-56 and Treg population, respectively; Statistical evaluation of data was performed by means of the Mann–Whitney test. Statistical significance values are indicated.