Peripheral blood immune cell profiling of acute corneal transplant rejection

Abstract

Acute rejection (AR) of corneal transplants (CT) has a profound effect on subsequent graft survival but detailed immunological studies in human CT recipients are lacking. In this multi-site, cross-sectional study, clinical details and blood samples were collected from adults with clinically diagnosed AR of full-thickness (FT)-CT (n = 35) and posterior lamellar (PL)-CT (n = 21) along with Stable CT recipients (n = 177) and adults with non-transplanted corneal disease (n = 40). For those with AR, additional samples were collected 3 months later. Immune cell analysis was performed by whole-genome microarrays (whole blood) and high-dimensional multi-color flow cytometry (peripheral blood mononuclear cells). For both, no activation signature was identified within the B cell and T cell repertoire at the time of AR diagnosis. Nonetheless, in FT- but not PL-CT recipients, AR was associated with differences in B cell maturity and regulatory CD4⁺ T cell frequency compared to stable allografts. These data suggest that circulating B cell and T cell subpopulations may provide insights into the regulation of anti-donor immune response in human CT recipients with differing AR risk. Our results suggest that, in contrast to solid organ transplants, genetic or cellular assays of peripheral blood are unlikely to be clinically exploitable for prediction or diagnosis of AR.

Keywords: biomarker; corneal transplantation/ophthalmology; flow cytometry; rejection: acute; translational research/science.

FIGURE 1

Stability of the immune profiles of patients with CT with stable graft function. Immune profiling of main immune populations (B cell, CD4 T, CD8 T; A), B cell subsets (Naïve, Switched, Non‐switched, Transitional, other; B), CD4 and CD8 T cell subsets (NAÏVE, EM, CM, TEMRA, and CD4 T_REG; C and D) and markers associated with T cell differentiation and immune activation (E) were analyses in CT with a stable graft function and sampled less than 3 years (yellow; n = 95) or more than 3 years (blue; n = 82). Each point represents a single patient and the boxplot represent median, IQR_25–75, and IQR_10–90.

FIGURE 2

Immune signature of acute rejection of CT. Immune profiling of CT patients with acute rejection (yellow; n = 56) or stable graft (blue; n = 177) and patients with corneal diseases that have not been subjected to CT (gray; n = 40). The frequencies of the main immune populations (B cell, CD4 T, CD8 T; A), B cell subsets (Naïve, Switched, Non‐switched, Transitional, other; B), CD4 (C, D) and CD8 (E) T cell subsets (NAÏVE, EM, CM, TEMRA and CD4 T_REG) and markers associated with T cell differentiation and immune activation (F) are shown for each patient (point) and summarize using boxplot (median, IQR_25–75, and IQR_10–90).

FIGURE 3

Stability of the immune signature of acute rejection of CT after therapeutic adjustment. Immune signature of acute rejection was measured at the time of acute rejection diagnosis and 3‐month after in 28 FT‐CT patients. The frequency of each subset is shown for each patient (point) and gray box summarizes the value of stable CT patients (median, dot line; IQR_25–75, gray box).