Compromised Immune Status of Patients with Post-Liver Transplant Biliary Complications
Liver transplantation (LTx) is a life-saving procedure for patients with end-stage liver disease. However, post-transplant complications, particularly biliary complications (BC), remain a significant challenge. BC, including biliary strictures with cholangitis, are the most common complications after LTx. Immunosuppression (IS) is essential for LTx patients to prevent alloimmune responses mediated primarily by T cells. However, excessive IS can compromise T cell homeostasis, increasing the risk of opportunistic infections and malignancies. This study investigates the immune status, particularly T cell-mediated immunity, in LTx patients with BC, providing insights into the immunological characteristics and potential therapeutic implications.
T Cell Heterogeneity and Function in LTx Patients
Human T cells are heterogeneous, consisting of various subsets with distinct functions based on the expression of CD45RA and CD62L. These subsets include naïve T cells (Tn, CD45RA+ CD62L+), stem cell memory T cells (Tscm, CD45RA+ CD62L+ CD95+), central-memory T cells (Tcm, CD45RA- CD62L+), effector-memory T cells (Tem, CD45RA- CD62L-), and terminally differentiated effector subsets (CD45RA+ CD62L-). Memory T cells are more efficient in controlling pathogens compared to naïve T cells. Tscm, a specialized subset of memory T cells, differentiate directly from naïve precursors and can reconstitute the full diversity of memory T cells upon antigen priming while maintaining their pool size through self-renewal.
The differentiation of human Tscm to Tcm or Tem and their function in LTx patients remain unclear. This study aimed to elucidate these aspects by analyzing T cell subsets and their functional characteristics in LTx patients with and without BC.
Study Design and Patient Characteristics
The study included 42 patients who underwent orthotopic LTx within one year at the First Affiliated Hospital of Xi’an Jiaotong University. Patients with post-LTx BC, primarily biliary strictures confirmed by magnetic resonance cholangiopancreatography (MRCP) within one year after LTx, were included. Blood samples were collected the day after the confirmation of biliary stricture by MRCP. Age-matched patients with stable liver function and no BC within one year after LTx were categorized as the transplant group without BC. One patient with rejection was excluded from the final analysis. Eighteen age-matched healthy volunteers served as healthy controls (HCs).
Patient characteristics, including age, gender, primary liver disease, liver enzyme levels, concentration of immunosuppressants, and immune cell counts on the day of T cell analysis, were documented. The BC group had significantly higher levels of aspartate aminotransferase and total bilirubin compared to the group without BC. Complete blood count data revealed a decreased absolute number of white blood cells and neutrophils in patients with post-LTx BC, with a slightly increased proportion of lymphocytes, although the absolute count of lymphocytes was similar. Three patients with infections in the BC group had C-reactive protein levels greater than 10 mg/L on the day of T cell analysis.
T Cell Subsets and Their Correlation
Peripheral blood mononuclear cells (PBMCs) were isolated using Ficoll gradient centrifugation. PBMCs were either stained directly or stimulated with phorbol 12-myristate 13-acetate and ionomycin in the presence of brefeldin A for cytokine detection by flow cytometry. The flow cytometry gating scheme for T cell subsets was used to identify various T cell populations.
Patients with BC showed decreased numbers of CD4+ T cells and increased numbers of CD8+ T cells compared to patients without BC. Significantly decreased numbers of Tn and Tscm, and increased numbers of Tem were observed in CD8+ T cells, but not in CD4+ effector T cells (Teff) or regulatory T cells (Tregs).
A significant negative correlation between Tscm and Tem, rather than with Tcm, was found in both CD8+ and CD4+ T cells in HCs. This correlation suggests direct differentiation of Tscm to Tem. However, in LTx patients, a negative correlation between Tscm and Tem was observed in CD8+ T cells, but not in CD4+ T cells. In LTx patients without BC, the correlation coefficient between CD8+ Tscm and CD8+ Tem was similar to that in HCs, while the value in LTx patients with BC was smaller, indicating decelerated differentiation of Tscm to Tem.
Functional Analysis of T Cells
To further investigate the function of CD4+ Teff, CD8+ T cells, and Tregs in LTx patients with BC, the expression of co-inhibitory receptors programmed cell death protein 1 (PD-1) and T cell immunoglobulin and mucin domain-3 (Tim-3), along with CD39 and cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), was evaluated. Higher expression of PD-1 was observed in LTx patients with BC on CD4+ Teff, indicating Teff exhaustion. CD4+ Teff and CD8+ T cells from LTx patients with BC also showed decreased interferon-gamma or interleukin-2 production. However, no significant differences were found in CD39 and CTLA-4 expression on Tregs.
Discussion
The findings of this study highlight the compromised immune status of LTx patients with BC, particularly in T cell-mediated immunity. The decreased numbers of CD4+ T cells and increased numbers of CD8+ T cells in patients with BC suggest an altered immune environment. The significant decrease in Tn and Tscm, along with the increase in Tem in CD8+ T cells, indicates a shift towards a more differentiated and potentially exhausted T cell phenotype.
The negative correlation between Tscm and Tem in HCs supports the direct differentiation of Tscm to Tem. However, the decelerated differentiation of Tscm to Tem in LTx patients with BC, as evidenced by the smaller correlation coefficient, suggests impaired T cell differentiation in these patients. This impairment is more pronounced in CD4+ T cells, where no negative correlation between Tscm and Tem was observed.
The higher expression of PD-1 on CD4+ Teff and the weakened cytokine secretion capacity in CD4+ T cells further support T cell exhaustion in LTx patients with BC. The differences in immunosuppressive regimens, with 87.5% of patients with BC under FK506 suppression and 88% of patients without BC under cyclosporin A IS, may also play a role in the observed immune alterations and warrant further investigation.
Conclusion
In summary, this study demonstrates the compromised immune status of LTx patients with BC, characterized by altered T cell subsets, impaired T cell differentiation, and T cell exhaustion. These findings underscore the need for tailored IS therapy with precise immune monitoring of T cell differentiation in LTx patients with BC. Further studies with larger patient cohorts are necessary to validate these findings and explore the impact of different immunosuppressive regimens on T cell-mediated immunity in LTx patients.
doi.org/10.1097/CM9.0000000000001088
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