Characterization of peripheral immune cells in kidney transplantation recipients under different immunosuppressive treatments

Abstract

Background: A comprehensive peripheral immune cell characterization including novel immunosuppressive subsets myeloid-derived suppressive cells (MDSCs) in kidney transplant recipients (KTRs) under different immunosuppressive treatments can help: 1) Immunosuppression situation and allograft acceptance assessment; 2) Infection and rejection emergence indication; 3) Beneficial immunosuppressive regimens' selection.

Methods: 26 KTRs with an average transplant duration of 360 days and 13 healthy controls were enrolled in this study. 11KTRs were included in the SRL-based therapy group and the other 15 in the TAC-based therapy group. Flow cytometry was used to detect the percentages and absolute numbers of MDSCs, T cell populations, HLA-DR⁺ monocytes, neutrophil CD64 index, and cytokines in peripheral blood.

Results: In KTRs, the expression of G-MDSCs and M-MDSCs was significantly higher than the HCs, while the expression of HLA-DR⁺ monocytes, CD38⁺/CD28⁺ activated T cells, CD4⁺ naïve T cells, CD4⁺ effector memory T cells, and central memory T cells were significantly lower. The use of mTOR inhibitors in KTRs induced changes in the distribution of activated and naïve-memory T cell subsets and decreased proinflammatory cytokines.

Discussion: In KTRs, G-MDSCs and M-MDSCs accumulated while functionally activated, naïve-memory T cell populations and HLA-DR⁺ monocytes markedly decreased one year after transplantation. Additionally, the number of MDSCs and T cell subsets following transplantation is likely regulated by mTOR inhibitors.

Keywords: flow cytometry; immune cell biomarker; kidney transplantation; mTOR inhibitors; myeloid-derived suppressor cells.

Figure 1

The research flowchart for the sufficient monitoring of immune cell phenotypes in KTRs with different immunosuppressants and HCs. SRL, Sirolimus; TAC, Tacrolimus; MMF, Mycophenolate mofetil; Pred, Prednisolone.

Figure 2

Characterization of circulating MDSC subsets in KTRs applying different immunosuppressants administration and HCs. (A) Absolute numbers (left panel) and percentage (right panel) of G-MDSCs in HCs (squares), KTRs (circles), TAC-based therapy group (inverted triangles) and SRL-based therapy group (triangles). (B) Absolute numbers (left panel) and percentage (right panel) of M-MDSCs in HCs (squares), KTRs (circles), TAC-based therapy group (inverted triangles) and SRL-based therapy group (triangles). Lines indicate median values including interquartile range (IQR). *p < 0.05, **p < 0.01, ***p < 0.001 calculated with Mann-Whitney U test.

Figure 3

Characterization of nCD64 index and HLA-DR expression on CD14⁺ monocytes in KTRs applying different immunosuppressants administration and HCs. (A)nCD64 index in the HCs (squares), KTRs (circles), TAC-based therapy group (inverted triangles) and SRL-based therapy group (triangles). (B) Percentage of HLA-DR⁺ monocytes and (C) HLA-DR MFI on monocytes in the HCs (squares), KTRs (circles), TAC-based therapy group (inverted triangles) and SRL-based therapy group (triangles). Lines indicate median values including interquartile range (IQR). *p < 0.05, **p < 0.01, ***p< 0.001 calculated with Mann-Whitney U test.

Figure 4

Characterization of circulating regulatory and activated T cell subsets in KTRs applying different immunosuppressants administration and HC. (A) Absolute numbers of Tregs in KTRs (circles), HCs (squares), SRL-based therapy group (triangles) and TAC-based therapy group (inverted triangles). (B) Absolute numbers of CD38⁺ CD4⁺ T cells, (C) CD38⁺ CD8⁺ T cells, (D) CD28⁺ CD4⁺ T cells and (E) CD28⁺ CD8⁺ T cells, (F) HLA-DR⁺ CD4⁺ T cells and (G) HLA-DR⁺ CD8⁺ T cells in the HCs (squares), KTRs (circles), TAC-based therapy group (inverted triangles) and SRL-based therapy group (triangles). Lines indicate median values including interquartile range (IQR). *p < 0.05, **p < 0.01, ***p < 0.001 calculated with Mann-Whitney U test.

Figure 5

Characterization of circulating CD4⁺ naïve-memory T subsets in KTRs and HCs. (A) Absolute numbers (left panel) and percentage (right panel) of CD27⁺CD45RA⁺CD4⁺ naïve T cells, (B) CD27⁺CD45RA^-CD4⁺ central memory T cells, (C) CD27^-CD45RA^-CD4⁺ effector memory T cells and (D) CD27^-CD45RA⁺CD4⁺ effector T cells in the HCs (squares), KTRs (circles), TAC-based therapy group (inverted triangles) and SRL-based therapy group (triangles). Lines indicate median values including interquartile range (IQR). *p < 0.05, **p < 0.01, ***p < 0.001 calculated with Mann-Whitney U test.

Figure 6

Characterization of circulating CD8⁺ naïve-memory T subsets in KTRs and HCs. (A) Absolute numbers (left panel) and percentage (right panel) of CD27⁺CD45RA⁺CD8⁺ naïve T cells, (B) CD27⁺CD45RA^-CD8⁺ central memory T cells, (C) CD27^-CD45RA^-CD8⁺ effector memory T cells and (D) CD27^-CD45RA⁺CD8⁺ effector T cells in the HCs (squares), KTRs (circles), TAC-based therapy group (inverted triangles) and SRL-based therapy group (triangles). Lines indicate median values including interquartile range (IQR). *p < 0.05, **p < 0.01, ***p < 0.001 calculated with Mann-Whitney U test.

Figure 7

Characterization of circulating total B cells in KTRs under different immunosuppressive treatments. (A) Absolute numbers and (B) percentage of CD19⁺ B cells in the TAC-based therapy group (squares) and SRL-based therapy group(circles). Lines indicate median values including interquartile range (IQR). p was calculated with Mann-Whitney U test.

Figure 8

Concentration of circulating adaptive immune cell subsets between patients applying different immunosuppressants administration. (A) Concentration of IL-2, IL-4, IL-10, TNF-α, IFN-γ, IL-17A, IL-1b, IL-5, IL-12, IFN-α, IL-8 in KTRs (red histograms) and HCs (blue histograms). (B) Concentration of IL-2, IL-4, TNF-α, IL-5, IL-12 in SRL-based therapy group (red histograms) and TAC-based therapy group (blue histograms). Black lines indicate median values including interquartile range (IQR). *p < 0.05, **p < 0.01, ***p < 0.001 calculated with Mann-Whitney U test.

Figure 9

Heatmap of peripheral immune cell subset changes across study groups. The heatmap shows Z-score standardized values of selected immune cell subsets in HCs, KTRs, and immunosuppressive subgroups (TAC, SRL). Immune cell types are grouped into myeloid and T cell lineages. Red indicates relative upregulation; blue indicates relative downregulation.