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Older kidney transplant recipients demonstrate increased rates of infection, and lower rates of rejection, compared with younger kidney transplant recipients. However, the mechanism behind this observation remains unknown. To develop a multifaceted view of age-associated immune dysfunction, we determined the function and phenotype of T cells predisposing to vulnerability to infection on a molecular level. Overlapping peptide pools representing the dominant CMV antigens were used to stimulate PBMC collected from 51 kidney transplant recipients, using cytokine secretion to determine specificity and intensity of response. Staphylococcal endotoxin B (SEB) was analyzed in parallel. To define immune cell subsets, we used single cell RNA sequencing (scRNAseq) to evaluate cellular surface markers and gene expression. We found increased frequency of SEB- and CMV-specific T cells was associated with freedom from infection, especially in older patients. Spatialized t-SNE analysis revealed decreased frequency of naïve T cells, increased frequency of TEMRA cells, and decreased frequency of IFNγ secreting T cells in patients with infection. Application of scRNAseq analysis revealed increased frequency of terminally differentiated T cells expressing NK-associated receptors and inhibitory markers. These findings offer unique insight into the mechanism behind vulnerability to infection in the kidney transplant recipient, revealing a specific T cell subtype of impaired antigen response and terminal effector phenotype as markers of T cell senescence.

Noninvasive immunologic analysis of peripheral blood holds promise for explaining the mechanism of development of adverse clinical outcomes, and may also become a method for patient risk stratification before or after mechanical circulatory support device (MCSD) implantation. Dysregulation of the innate immune system is associated with increased patient age but has yet to be evaluated in the older patient with advanced heart failure undergoing MCSD surgery. Patients pre- and post-MCSD implantation had peripheral blood mononuclear cells (PBMC) and serum isolated. Multiparameter flow cytometry was used to analyze markers of innate cell function, including monocyte subtypes. Multiplex cytokine analysis was performed. MELD-XI and SOFA scores were utilized as surrogate markers of outcomes. Increased levels of pro-inflammatory cytokines including IL-15, TNF-α, and IL-10 were associated with increased MELD-XI and SOFA scores. IL-8, TNF- α, and IL-10 were associated with risk of death after MCSD implantation, even with correction for patient age. Increased frequency of 'classical' monocytes (CD14 + CD16-) were associated with increased MELD-XI and SOFA scores. This suggests that inflammation and innate immune system activation contribute to progression to multiorgan system failure and death after MCSD surgery. Development of noninvasive monitoring of peripheral blood holds promise for biomarker development for candidate selection and patient risk stratification.

Immunologic impairment may contribute to poor outcomes after implantation of mechanical circulatory support device (MCSD), with infection often as a terminal event. The study of immune dysfunction is of special relevance given the growing numbers of older patients with heart disease. The aim of the study was to define which immunologic characteristics are associated with development of adverse clinical outcomes after MCSD implantation. We isolated peripheral blood mononuclear cells (PBMC) from patients pre- and up to 20 days post-MCSD implantation and analyzed them by multiparameter flow cytometry for T cell dysfunction, including terminal differentiation, exhaustion, and senescence. We used MELD-XI and SOFA scores measured at each time point as surrogate markers of clinical outcome. Older patients demonstrated increased frequencies of terminally differentiated T cells as well as NKT cells. Increased frequency of terminally differentiated and immune senescent T cells were associated with worse clinical outcome as measured by MELD-XI and SOFA scores, and with progression to infection and death. In conclusion, our data suggest that T cell dysfunction, independently from age, is associated with poor outcomes after MCSD implantation, providing a potential immunologic mechanism behind patient vulnerability to multiorgan dysfunction and death. This noninvasive approach to PBMC evaluation holds promise for candidate evaluation and patient monitoring.

Aim A main cause of death in Advanced Heart Failure (AdHF) after Mechanical Circulatory Support Device (MCSD) is the Multiorgan Dysfunction Syndrome (MOD), mediated by an aberrant immune response. Since implantation of MCSD has immune modulating effects that may influence patient outcome, we hypothesized that T cell immunophenotyping would reflect assessment of patient risk before and after MCSD implant. Methods Peripheral blood was collected from 11 consented patients at days −1, 1, 3, 5 and 7 before and after MCSD surgery, and at 5 corresponding time points from 4 healthy controls. Multi-parameter flow cytometry was performed to measure CD4 and CD8 T cell maturation state as well as experienced and exhausted T cell subpopulations. Clinical risk was calculated using a modified Sequential Organ Failure Assessment (SOFA) score and the MELD-XI score. Results Multi-parameter immunophenotyping generated 34 CD4 and CD8 subpopulations. High scored MELD samples, obtained from higher risk patients, exhibited higher percentage of both effector and central memory CD8 cells (p = 0.0071 and p = 0.0001) and higher percentage of effector memory CD4 cells (p = 0.0074). There was a trend towards association with higher percentage of CD8+KLRG1+CD28 (p = 0.06) and lower %PD1+CD57+CD4+cells (p = 0.0785) compared with lower MELD scored patient samples. We further analyzed the data using stepwise and penalized regression methods. We reduced the number of predictors down to 3 T-cell subpopulations with immunophenotyping profile of chronically activated T cells: CD8+CD38+CD57+, CD4+CD38+CD57+ and CD4+CD38+KLRG1+. We used these subpopulations as the inputs to a linear mixed effect model to successfully predict the SOFA scores of patients’ samples. The overall model was found to be statistically significant (p = 0.006), and the regression coefficient for CD8+CD38+CD57 was statistically significant (p < 0.05). Conclussion Statistical modeling of T-cell subpopulation frequencies in peripheral blood of MOD patients can provide significant information predictive of their clinical phenotype, warranting further study for developing models to better predict patient outcomes. [ABSTRACT FROM AUTHOR]