C-type natriuretic peptide induces inotropic and lusitropic effects in human 3D-engineered cardiac tissue: Implications for the regulation of cardiac function in humans

Abstract

The role of C-type natriuretic peptide (CNP) in the regulation of cardiac function in humans remains to be established as previous investigations have been confined to animal model systems. Here, we used well-characterized engineered cardiac tissues (ECTs) generated from human stem cell-derived cardiomyocytes and fibroblasts to study the acute effects of CNP on contractility. Application of CNP elicited a positive inotropic response as evidenced by increases in maximum twitch amplitude, maximum contraction slope and maximum calcium amplitude. This inotropic response was accompanied by a positive lusitropic response as demonstrated by reductions in time from peak contraction to 90% of relaxation and time from peak calcium transient to 90% of decay that paralleled increases in maximum contraction decay slope and maximum calcium decay slope. To establish translatability, CNP-induced changes in contractility were also assessed in rat ex vivo (isolated heart) and in vivo models. Here, the effects on force kinetics observed in ECTs mirrored those observed in both the ex vivo and in vivo model systems, whereas the increase in maximal force generation with CNP application was only detected in ECTs. In conclusion, CNP induces a positive inotropic and lusitropic response in ECTs, thus supporting an important role for CNP in the regulation of human cardiac function. The high degree of translatability between ECTs, ex vivo and in vivo models further supports a regulatory role for CNP and expands the current understanding of the translational value of human ECTs. NEW FINDINGS: What is the central question of this study? What are the acute responses to C-type natriuretic peptide (CNP) in human-engineered cardiac tissues (ECTs) on cardiac function and how well do they translate to matched concentrations in animal ex vivo and in vivo models? What is the main finding and its importance? Acute stimulation of ECTs with CNP induced positive lusitropic and inotropic effects on cardiac contractility, which closely reflected the changes observed in rat ex vivo and in vivo cardiac models. These findings support an important role for CNP in the regulation of human cardiac function and highlight the translational value of ECTs.

Keywords: Langendorff isolated heart; contractility; human stem cell-derived cardiomyocytes; relaxation; translation.

FIGURE 1

Illustration of the engineered cardiac tissue model and measured variables. (a) A representative image of an ECT in the Biowire platform attached to the polymer wires. Twitch force is measured by recording a video of the contracting tissues under field stimulation and converting pixel displacement of the polymer wires into force measurements. (b) Measured parameters for contractility and calcium transients. CM, cardiomyocyte; ECT, engineered cardiac tissue.

FIGURE 2

Isoproterenol and dobutamine treatment induce positive inotropic and lusitropic contractility responses in 3D‐engineered cardiac tissues. All data are expressed as a percentage of baseline. Values are means ± SD and represent n = 6, except (c) where n = 3. Significant difference between baseline and dobutamine/isoproterenol: ***P < 0.001, ****P < 0.0001.

FIGURE 3

Isoproterenol and dobutamine treatment induce positive inotropic and lusitropic calcium responses in 3D‐engineered cardiac tissues. All data are expressed as a percentage of baseline. Values are means ± SD and represent n = 6. Significant difference between baseline and CNP: ****P < 0.0001. CNP, C‐type natriuretic peptide.

FIGURE 4

CNP treatment induces positive inotropic and lusitropic contractility responses in 3D‐engineered cardiac tissues. All data are expressed as a percentage of their own baseline. Values are means ± SD and represent n = 8 (vehicle) and n = 6 (CNP). Significant difference between vehicle and CNP: *P < 0.05, ***P < 0.001. CNP, C‐type natriuretic peptide; ISO, isoproterenol.

FIGURE 5

CNP treatment induces positive inotropic and lusitropic calcium responses in 3D‐engineered cardiac tissues. All data are expressed as a percentage of their own baseline. Values are means ± SD and represent n = 8 (vehicle) and n = 6 (CNP). Significant difference between vehicle and CNP: *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. CNP, C‐type natriuretic peptide; ISO, isoproterenol.

FIGURE 6

CNP treatment increases systolic and diastolic parameters in isolated hearts. Values are means ± SD and represent n = 14 (vehicle) and n = 15 (CNP). CNP was dissolved in Krebs–Henseleit buffer to a final concentration of 30 nM. Coronary perfusion was before and after 17.5 min of global ischaemia. Significant difference between vehicle and CNP: *P < 0.05, **P < 0.01, ***P < 0.001. CNP, C‐type natriuretic peptide; dP/dt _max and dP/dt _min, maximal slope of the systolic pressure increment, and the diastolic pressure decrement; LVDP, left ventricular developed pressure.

FIGURE 7

CNP infusion increases indices of cardiac performance in vivo without altering systemic hemodynamics. All data are expressed as a percentage of baseline (t = 0). Values are means ± SD and represent n = 7 (vehicle) and n = 6 (CNP). CNP was infused at a constant rate of 200 pmol/min/kg (plasma concentration of ∼10 nmol/l at steady state) via the tail vein. Significant difference between vehicle and CNP: ***P < 0.001, ****P < 0.0001. CNP, C‐type natriuretic peptide; DBP, diastolic blood pressure; dP/dt _max and dP/dt _min, maximal slope of the systolic pressure increment, and the diastolic pressure decrement; HR, heart rate; LVDP, left ventricular developed pressure; LVEDP, left ventricular end‐diastolic pressure; LVESP, left ventricular end‐systolic pressure; MABP, mean arterial blood pressure; RPP, rate‐pressure product; SBP, systolic blood pressure.