LEA29Y expression affects myocardial scar formation after ischemia/reperfusion in pigs
Heinke Heymer1,2, Nadja Hornaschewitz1,2, Mara Corsten1,2, Li Xianghai1, Eckhard Wolf3, Andreas Parzefall4, Nikolai Klymiuk1,2, Christian Kupatt1, Andrea Bähr1,2.
1Department of Internal Medicine I, Klinikum rechts der Isar, TU Munich, Munich, Germany; 2Center for Innovative Medical Models, LMU Munich, Munich, Germany; 3Chair for Molecular Animal Breeding and Biotechnology, LMU Munich, Munich, Germany; 4Institute for Veterinary Pathology, LMU Munich, Munich, Germany
Background: The processes of scar formation during myocardial remodeling after ischemia/reperfusion (I/R) are intricate and tightly regulated by the immune system and its effector cells. The fusion protein LEA29Y is a widely used immunosuppressive medication that inhibits T-cell co-stimulation, thereby limiting immune cell activation. A Pig model with systemic LEA29Y-Expression has been established as a model for general immunosuppression, enabling investigation of I/R injury and remodeling under immunocompromised conditions. Investigations include an emphasis on collagen content as a representation of scar formation, as well as the quantification of immune cells.
Methods: Using a catheter-based Intervention, 60 minutes of Ischemia in the left anterior descending artery (LAD) was induced, followed by a permanent reperfusion, leading to an infarction in the Left Ventricle (LV). Animals were sampled after 3, 9, 14 and 26 days and compared to controls. Ejection fraction was calculated, ECG was recorded and complemented by histological stainings with HE, Sirius Red, IBA1, CD3-T-Cells and neutrophils. Histological Analysis was performed using “QuPath”, an open source bioimage analysis program that allows for the analysis of wholly scanned slides, thus enabling a high-resolution global measurement of stained tissue in per-cent and micrometers square.
Results: LEA29Y and WT animals displayed similar infarct sizes. LV angiography showed a significantly higher loss of ejection fraction in the transgenic pigs after 3 days (-15.48±1.069 vs. -7.554±1.873, p=0.011), but not after 9 days or later. QuPath analysis of stained collagen areas detects scar formation over the course of the investigation period. It showed a peak of collagen at d14, with less height and slower decline at d26 in LEA29Y Animals compared to WT (day 3: 13,33% of LEA29Y LV vs. 12,6% of WT LV; day 9: 18,36% of LEA29Y LV vs. 16,57% of WT LV; day 14: 18,4% of LEA29Y LV vs. 20,75% of WT LV; day 26: 16,49% of LEA29Y LV vs. 15,04% of WT LV).
Conclusion: Blocking of T-cell co-stimulation initially leads to a higher loss of pumping function of the LV and a delayed scar contraction. To further investigate the loss of T-cells and the connections between them and other immune cells such as macrophages and neutrophils, immunohistochemistry stainings are conducted.