Thymic Atrophy and Immune Dysregulation in Infants with Complex Congenital Heart Disease.

Congenital heart disease (CHD) is the most common birth defect, and up to 50% of infants with CHD require cardiovascular surgery early in life. Current clinical practice often involves thymus resection during cardiac surgery, detrimentally affecting T-cell immunity. However, epidemiological data indicate that CHD patients face an elevated risk for infections and immune-mediated diseases, independent of thymectomy. Hence, we examined whether the cardiac defect impacts thymus function in individuals with CHD. We investigated thymocyte development in 58 infants categorized by CHD complexity. To assess the relationship between CHD complexity and thymic function, we analyzed T-cell development, thymic output, and biomarkers linked to cardiac defects, stress, or inflammation. Patients with highly complex CHD exhibit thymic atrophy, resulting in low frequencies of recent thymic emigrants in peripheral blood, even prior to thymectomy. Elevated plasma cortisol levels were detected in all CHD patients, while high NT-proBNP and IL-6 levels were associated with thymic atrophy. Our findings reveal an association between complex CHD and thymic atrophy, resulting in reduced thymic output. Consequently, thymus preservation during cardiovascular surgery could significantly enhance immune function and the long-term health of CHD patients.

A potential future Fontan modification: preliminary in vitro data of a pressure-generating tube from engineered heart tissue.

OBJECTIVES: Univentricular malformations are severe cardiac lesions with limited therapeutic options and a poor long-term outcome. The staged surgical palliation (Fontan principle) results in a circulation in which venous return is conducted to the pulmonary arteries via passive laminar flow. We aimed to generate a contractile subpulmonary neo-ventricle from engineered heart tissue (EHT) to drive pulmonary flow actively. METHODS: A three-dimensional tubular EHT (1.8-cm length, 6-mm inner diameter, ca. 1-mm wall thickness) was created by casting human-induced pluripotent stem cell-derived cardiomyocytes (0.9 ml, 18 mio/ml) embedded in a fibrin-based hydrogel around a silicone tube. EHTs were cultured under continuous, pulsatile flow through the silicone tube for 23 days. RESULTS: The constructs started to beat macroscopically at days 8-14 and remained stable in size and shape over the whole culture period. Tubular EHTs showed a coherent beating pattern after 23 days in culture, and isovolumetric pressure measurements demonstrated a coherent pulsatile wave formation with an average frequency of 77 ± 5 beats/min and an average pressure of 0.2 mmHg. Histological analysis revealed cardiomyocytes mainly localized along the inner and outer curvature of the tubular wall with mainly longitudinal alignment. Cell density in the center of the tubular wall was lower. CONCLUSIONS: A simple tube-shaped contractile EHT was generated from human-induced pluripotent stem cells and developed a synchronous beating pattern. Further steps need to focus on optimizing support materials, flow rates and geometry to obtain a construct that creates sufficient pressures to support a directed and pulsatile blood flow.

OMIP 073: Analysis of human thymocyte development with a 14-color flow cytometry panel.

This panel was designed for the identification and detailed characterization of the different developmental steps of human thymocytes. We optimized the panel for fresh tissue in order to provide an unbiased analysis of T cell development. Accurate selection of antibodies and precise gating allow us to phenotype 14 major stages of human thymocyte development and illustrate the trajectories of T cell development from early thymic progenitors (ETP) to mature T cells that are ready to populate the periphery. The panel identifies ETPs, T-lineage-committed cells (TC), CD34-positive immature single-positive CD4 cells (ISP4 CD34+), CD34-negative immature single-positive CD4 cells (ISP4 CD34-), CD45-low early double-positive cells (EDP CD45low), CD45-high early double-positive cells (EDP CD45high), late double-positive cells (LDP), single-positive CD4 cells (SP4), single-positive CD8 cells (SP8), ready-to-egress single-positive CD4 cells (rSP4), ready-to-egress single-positive CD8 cells (rSP8), T γδ cells (Tγδ), T regulatory cells (Treg), and ready-to-egress T regulatory cells (rTreg). To highlight important checkpoints during T cell development, we added antibodies relevant for specific developmental steps to the panel. These include CD1a to define TCs, CD28 as a marker for ß-selection and CD69 in combination with CD45RA to determine the maturation stage of thymocytes shortly before they become ready to egress the thymus and colonize the periphery. Moreover, Annexin V, as a marker for apoptosis, provides valuable extra information concerning the apoptotic death of thymocytes. Currently, we use this panel to identify aberrations in T cell development in health and disease.