Research Article: 2-Cl-C.OXT-A stimulates contraction through the suppression of phosphodiesterase activity in human induced pluripotent stem cell-derived cardiac organoids

Date Published: July 11, 2019

Publisher: Public Library of Science

Author(s): Takahiro Kitsuka, Manabu Itoh, Sojiro Amamoto, Ken-ichi Arai, Junichi Oyama, Koichi Node, Shuji Toda, Shigeki Morita, Takahiro Nishida, Koichi Nakayama, Johnson Rajasingh.


2-Cl-C.OXT-A (COA-Cl) is a novel synthesized adenosine analog that activates Sphingosine-1-phosphate 1 receptor (S1P1R) and combines with the adenosine A1 receptor (A1R) in G proteins and was shown to enhance angiogenesis and improve the brain function in rat stroke models. However, the role of COA-Cl in hearts remains unclear. COA-Cl, which has a similar structure to xanthine derivatives, has the potential to suppress phosphodiesterase (PDE), which is an important factor involved in the beating of heart muscle.

Cardiac organoids with fibroblasts, human induced pluripotent stem cell-derived cardiac myocytes (hiPSC-CMs), and hiPSC-derived endothelial cells (hiPSC-ECs) were cultured until they started beating. The beating and contraction of organoids were observed before and after the application of COA-Cl. COA-Cl significantly increased the beating rate and fractional area change in organoids. To elucidate the mechanism underlying these effects of COA-Cl on cardiac myocytes, pure hiPSC-CM spheroids were evaluated in the presence/absence of Suramin (antagonist of A1R). The effects of COA-Cl, SEW2871 (direct stimulator of S1P1R), two positive inotropes (Isoproterenol [ISO] and Forskolin [FSK]), and negative inotrope (Propranolol [PRP]) on spheroids were assessed based on the beating rates and cAMP levels. COA-Cl stimulated the beating rates about 1.5-fold compared with ISO and FSK, while PRP suppressed the beating rate. However, no marked changes were observed with SEW2871. COA-Cl, ISO, and FSK increased the cAMP level. In contrast, the level of cAMP did not change with PRP or SEW2871 treatment. The results were the same in the presence of Suramin as absence. Furthermore, an enzyme analysis showed that COA-Cl suppressed the PDE activity by half.

COA-Cl, which has neovascularization effects, suppressed PDE and increased the contraction of cardiac organoids, independent of S1P1R and A1R. These findings suggest that COA-Cl may be useful as an inotropic agent for promoting angiogenesis in the future.

Partial Text

Positive inotropic agents are considered to enhance the hemodynamic profile in terms of elevating the cardiac output, decreasing the cardiac filling pressure, and improving the organ perfusion in patients with heart disease. In clinical trials in which inotropic agents were administered to promote the contraction of the dysfunctional heart, safety was a concern from the viewpoint of the long-term prognosis [1]. However, in the clinical setting, inotropic agents remain a major treatment for patients in the decompensated phase of severe heart failure. Positive inotropic drugs are useful for increasing the cardiac output in order to resolve various problems associated with heart failure, especially for patients with a low systolic blood pressure or low cardiac output [2].

In this study, we showed that COA-Cl suppresses PDE and increases the contraction of cardiac organoids, independent of S1P1R and A1R. COA-Cl can produce signaling molecules and tube formation in cultured human vascular endothelial cells through S1P1R-mediated extracellular stimulation [3]. COA-Cl has been shown to exert a neuroprotective effect against intracerebral hemorrhaging [6]. It has also been reported that COA-Cl may reduce oxidative stress, which may be one of the mechanisms underlying its neuroprotective effect [5]. In addition, in the perfused heart, pretreatment with S1P has been shown to significantly restore the heart function after ischemia. Interestingly, the protective effect of ischemic preconditioning on ischemia/reperfusion injury was promoted in the hearts of rats treated with S1P1R and S1P3R antagonists [18][19].

COA-Cl was shown to act as a partial PDE inhibitor of cardiac organoids. Cardiac organoids are useful for drug discovery, facilitating assay performance. The results of the present study suggest their possible application in cardiac transplantation for regenerative medicine. COA-Cl can be used as a cardiac inotropic agent, as a partial PDE inhibitor, and to promote angiogenesis in patients with ischemic heart disease and heart failure.




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