Recipient-Derived Immune Cell Phenotyping of Bronchoalveolar Lavage Identifies Patients with Biopsy Proven Early Acute Cellular Rejection after Lung Transplantation: A Pilot Study

Abstract

Acute cellular rejection (ACR) is associated with the ultimate development of chronic lung allograft dysfunction. Transbronchial biopsy, the gold standard for the diagnosis of ACR, is associated with potential morbidity. We set out to define the correlation between the recipient-derived immune cell content in bronchoalveolar lavage (BAL) and biopsy proven early ACR (<4 months). We analyzed BAL samples from 8 lung transplant recipients with early ACR and 5 without ACR. Immunophenotyping was performed using spectral flow cytometry; cell origin (donor v recipient) was determined by staining for human leukocyte antigen. T test was used to identify differences in immune cell populations between those with and without ACR. Live CD45+ cells were defined as CD4+ and CD8+ T cells, B cells (CD19+, CD20+), CD56+ lymphocytes, and monocytes (CD14+CD206+/CD64+). BAL was obtained at 1.5 months (range 0.5-3.5) from transplant at time of ACR and at 1.6 months (range 0.5 - 2.8) for non-ACR. The presence of ACR was associated with a statistically significant decrease in the proportion of donor monocytes (Figure 1: Mean 85%, SD 15% v 34%, SD 16%; p <0.001) and donor-derived regulatory T cells (Tregs), defined as CD4+CD25+FOXP3+ (Mean 3%, SD 3% v Mean 9%, SD 3%; p = 0.004). When comparing absolute number of both donor and recipient immune cells found in the BAL, patients with ACR had a statistically significant increase in the number of recipient-derived CD4+ T cells (Mean 7%, SD 5% v 1%, SD 0.05%; p value = 0.02), and a trend towards increased recipient-derived CD8+ T cells (Mean 18%, SD 27% v <1%, SD 1%; p value = 0.17). Biopsy proven, early ACR is associated with a relative decline in donor monocytes and donor Tregs, and increased proportion of recipient-derived CD4+ T cells. These results support further investigation into the role of BAL immune cell phenotyping as a potential less-invasive biomarker for ACR.