Background: For clinical application of cardiomyocytes derived from induced pluripotent stem cells (iPSC-CM), immunological response (IR) to transplant should be great concern for effectiveness in treatment for heart failure (HF). However, IR including innate immunity after in vivo transplantation is largely unknown. We hypothesized that natural killer cells (NKCs) may play an important role in innate immune rejection to transplanted iPSC-CMs.
Method: Cardiomyogenic differentiation was performed using C57BL/6 mouse derived iPSCs with expression in Luciferase. The iPSC-CMs were transplanted into subcutaneous tissue of C57BL/6 mouse, of which NKCs were depleted (none-NK, n=5) or were not (control, n=5). To evaluate the quantity of transplanted iPSC-CMs, the luminescence intensity (LI) of iPSC-CMs was measured using bioluminescence imaging system (BLI). iPSC-CM and lymphocyte (Lym) extracted from spleen were used for flow cytometric analysis (FCM) to evaluate the expression of NKC related-markers.
Result: FCM showed 64.8% of iPSC-CMs expressed cardiac troponin T with the significantly lower expressions of major histocompatibility complex class1, which is inhibitory NKC ligand, compared with Lyms (Lyms vs iPSC-CMs; H2Db; 99.6% vs 2.5%, H2Kb; 99.9% vs 0.9% and Q2; 78.6% vs 0.4%; p<0.05), with the expression in NKC activating ligands in iPSC-CMs (CD112; 27.7%, CD155; 24.3%). BLI demonstrated that LI in none-NK at post-transplantation day 7 was significantly higher than that in control (85±26% vs 31±8%, p<0.05, respectively). Immunostaining analysis revealed that CD335-positive NKCs, which also expressed NKC activation marker CD107a, infiltrated into transplant containing iPSC-CMs in control and the quantity of Annexin V-positive iPSC-CMs in control was significantly larger than those in none-NK (Apotosis Index; 22.2±2.9% vs 8.1±0.4%, p<0.05, respectively). The percentage of CD107a-positive NKCs in control was significantly larger than that in mouse without cell transplantation. (37.4±3.5% vs 7.2±2.1%, p<0.05).
Conclusion: NKCs have a crucial role in the rejection of transplanted syngeneic iPSC-CMs, suggesting that intensive care for innate immune system after transplantation may promise adequate effectiveness in iPSCs therapy for HF.