The Ratio of CD226 and TIGIT Expression in Tfh and PD-1+ICOS+Tfh Cells Are Potential Biomarkers for Chronic Antibody-Mediated Rejection in Kidney Transplantation

Abstract

Kidney transplantation is the ideal treatment for end-stage renal disease (ESRD). Chronic antibody-mediated rejection (CAMR) is the main cause of graft failure. Tfh and B cells are key immune cells that play important roles in CAMR. In this study, the populations of different Tfh cell phenotypes and B cell subsets in CAMR were investigated in a total of 36 patients. Based on Banff-2019, 15 patients were diagnosed with CAMR (CAMR group), 11 recipients were diagnosed with recurrent or de novo IgA nephropathy (IgAN group), and 10 patients displayed stable renal function (stable group). The Tfh and B cell subsets were analyzed by flow cytometry. The percentage and absolute number of PD-1⁺ICOS⁺Tfh cells were significantly higher in CAMR (p < 0.05), as was the ratio of CD226⁺Tfh cells to TIGIT⁺Tfh cells (p < 0.05). Compared with stable recipients, CAMR patients had lower naïve B cells and higher unswitched memory B cells, which were also significantly related to renal function (p < 0.05). Using the logistic regression model, we concluded that the estimated glomerular filtration rate (eGFR), absolute number of PD-1⁺ICOS⁺Tfh cells, and ratio of CD226⁺Tfh cells to TIGIT⁺Tfh cells were independent risk factors for CAMR. The combination of eGFR, PD-1⁺ICOS⁺Tfh cells, and the ratio of CD226⁺Tfh cells to TIGIT⁺Tfh cells showed better diagnostic efficacy for CAMR than each single parameter. The collective findings show that monitoring different Tfh phenotypes and B cell subsets is beneficial to kidney transplant recipients and implicate the combination of eGFR, number of PD-1⁺ICOS⁺Tfh cells, and ratio of CD226⁺Tfh cells to TIGIT⁺Tfh cells as a biomarker for diagnosing CAMR. The findings may also inform new strategies to identify and treat CAMR.

Figure 1

Gating strategy for analysis using logic gates. Measurements were performed with fresh blood samples. (a) Tfh cells were defined as CD4⁺CXCR5⁺ T cells in the lymphocyte population. The Tfh cell phenotypes were PD-1⁺ICOS⁺CD4⁺CXCR5⁺ T cell (PD-1⁺ICOS⁺Tfh cells), CD226⁺CD4⁺CXCR5⁺ T cell (CD226⁺Tfh cells), and TIGIT⁺CD4⁺CXCR5⁺ T cell (TIGIT⁺Tfh cells). (b) B cells were further defined as CD19-expressing cells in the lymphocyte population. CD19⁺ B cells were analyzed for the expressions of IgD and CD27, CD24, and CD38. The B cell populations included CD27⁻IgD⁺ (naïve B cells), CD27⁺IgD⁺ (unswitched memory B cells), CD27⁺IgD⁻ (switched memory B cells), CD24⁺⁺CD38⁺⁺ (transitional B cells), CD27⁺CD38⁺ (plasmablasts), and CD38⁺ B cells (plasma cells).

Figure 2

The percentage and absolute number of different Tfh phenotypes in the CAMR, IgAN, and stable groups. Percentages of (a) Tfh cells, (b) PD-1⁺ICOS⁺Tfh cells, (c) CD226⁺Tfh cells, and (d) TIGIT⁺Tfh cells. Absolute number of (e) Tfh cells, (f) PD-1⁺ICOS⁺Tfh cells, (g) CD226⁺Tfh cells, and (h) TIGIT⁺Tfh cells. (i) The ratio of CD226⁺Tfh cells to TIGIT⁺Tfh cells.

Figure 3

B cell differentiation subsets in patients with CAMR, recurrent de novo IgAN, and stable renal function. Percentages of (a) naïve B cells, (b) transitional B cells, (c) unswitched memory B cells, (d) switched memory B cells, (e) plasma cells, and (f) plasmablasts. Absolute numbers of (g) naïve B cells, (h) transitional B cells, (i) unswitched memory B cells, (j) switched memory B cells, (k) plasma cells, and (l) plasmablasts.

Figure 4

Correlation between different Tfh phenotypes, B cell subsets, and eGFR.

Figure 5

ROC curves for diagnosis of CAMR. (a) ROC curve of PD-1⁺ICOS⁺Tfh cells. (b) ROC curve of CD226⁺Tfh cell/TIGIT⁺Tfh cell ratio. (c) ROC curve of eGFR. (d) Combination of CD226⁺Tfh cell/TIGIT⁺Tfh cell ratio and PD-1⁺ICOS⁺Tfh cells. (e) ROC curve of combination of CD226⁺Tfh cell/TIGIT⁺Tfh cell ratio, PD-1⁺ICOS⁺Tfh cells, and eGFR.