Do Human iPSC-Derived Cardiomyocytes Cultured on PLA Scaffolds Induce Expression of CD28/CTLA-4 by T Lymphocytes?

Abstract

Many research groups have developed various types of tissue-engineered cardiac constructs. However, the immunological properties of such artificial tissues are not yet fully understood. Previously, we developed microfiber scaffolds carrying human iPSC-derived cardiomyocytes (hiPSC-CM). In this work, we evaluated the ability of these tissue-engineered constructs to activate the expression of CD28 and CTLA-4 proteins on T lymphocytes, which are early markers of the immune response. For this purpose, electrospun PLA microfiber scaffolds were seeded with hiPSC-CM and cultured for 2 weeks. Allogeneic mononuclear cells were then co-cultured for 48 h with three groups of samples: bare scaffolds, pure cardiomyocyte culture and tissue-engineered constructs, followed by analysis of CD28/CTLA-4 expression on T lymphocytes using flow cytometry. PLA scaffolds and concanavalin A stimulation (positive control) statistically significantly increased CD28 expression on CD4+ T cells (up to 61.3% and 66.3%) CD8+ T cells (up to 17.8% and 21.7%). CD28/CTLA-4 expression was not increased when T lymphocytes were co-cultured with cardiac tissue-engineered constructs and iPSC-CM monolayers. Thus, iPSC-CM in monolayers and on PLA microfiber scaffolds did not induce T cell activation, which suggests that such cardiac constructs would not be a cause of rejection after implantation.

Functional Activity of Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes on a Mouse Renal Subcapsular Xenograft Model

“…Between days 14 and 18 of differentiation, the cells were dissociated using TrypLE Express (Thermo Fisher Scientific, Waltham, MA, USA) and transferred into Matrigel™-coated 6-well plates in RPMI-1640 medium supplemented with 20% embryonic bovine serum (Autogene Bioclear, Calne, UK) and 10 µM Y-27632 supplement (StemRD, Burlingame, CA, USA). Two days after the transfer and within one week, metabolic cell selection was conducted to purify the population of cardiomyocytes [ 37 ]. The metabolic selection medium consisted of RPMI-1640 without D-glucose (Thermo Fisher Scientific, Waltham, MA, USA), 213 μg/mL L-ascorbic acid 2-phosphate (Sigma-Aldrich, Burlington, MA, USA), 500 µg/mL recombinant human albumin expressed in Oryza sativa (Sigma-Aldrich, Burlington, MA, USA), and 5 mM DL-sodium lactate L4263 (Sigma-Aldrich, Burlington, MA, USA).…”

Functional Activity of Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes on a Mouse Renal Subcapsular Xenograft Model

“…On days 14-18 of differentiation, the cells were dissociated using TrypLE Express (Thermo Fisher Scientific, USA) and transferred into 6-well plates coated with Matrigel™ (Corning, USA) in RPMI-1640 medium supplemented with 20% embryonic bovine serum (Autogene Bioclear, Calne, UK) and 10 µM Y-27632 supplement (StemRD, USA). Two days after the transfer and within one week, metabolic cell selection was carried out to purify the population of cardiomyocytes [15]. Composition of the metabolic selection medium: RPMI-1640 without D-glucose (Thermo Fisher Scientific, USA), 213 µg/mL L-ascorbic acid 2-phosphate (Sigma-Aldrich, Burlington, MA, USA), 500 µg/mL recombinant human albumin expressed in Oryza sativa (Sigma-Aldrich, USA) and 5 mM DL-sodium lactate L4263 (Sigma-Aldrich, USA).…”