Cardiomyocyte cell cycle activation improves cardiac function after myocardial infarction

Abstract

Aims: Cardiomyocyte loss is a major contributor to the decreased cardiac function observed in diseased hearts. Previous studies have shown that cardiomyocyte-restricted cyclin D2 expression resulted in sustained cell cycle activity following myocardial injury in transgenic (MHC-cycD2) mice. Here, we investigated the effects of this cell cycle activation on cardiac function following myocardial infarction (MI).

Methods and results: MI was induced in transgenic and non-transgenic mice by left coronary artery occlusion. At 7, 60, and 180 days after MI, left ventricular pressure-volume measurements were recorded and histological analysis was performed. MI had a similar adverse effect on cardiac function in transgenic and non-transgenic mice at 7 days post-injury. No improvement in cardiac function was observed in non-transgenic mice at 60 and 180 days post-MI. In contrast, the transgenic animals exhibited a progressive and marked increase in cardiac function at subsequent time points. Improved cardiac function in the transgenic mice at 60 and 180 days post-MI correlated positively with the presence of newly formed myocardial tissue which was not apparent at 7 days post-MI. Intracellular calcium transient imaging indicated that cardiomyocytes present in the newly formed myocardium participated in a functional syncytium with the remote myocardium.

Conclusion: These findings indicate that cardiomyocyte cell cycle activation leads to improvement of cardiac function and morphology following MI and may represent an important clinical strategy to promote myocardial regeneration.

Figure 1

Characterization of myocardial infarcts in non-transgenic and MHC-cycD2 mice. (a) Representative Azan staining of coronal sections from non-transgenic and MHC-cycD2 hearts at 7 and 180 days post-MI. The sections were sampled at 2, 3 and 4 mm from the apex of the heart, as indicated. Arrows indicate regions of newly formed myocardium in the MHC-cycD2 heart at 180 days post-MI which were not present in the corresponding anatomical location at 7 days post-MI, nor in the non-transgenic mice. Arrowheads indicate where the infarct scar has been largely resolved near the base of the heart by 180 days post-MI in the MHC-cycD2. (b) Consecutive sections of the apical scar/myocardium border of an MHCcycD2 mouse at 180 days post-MI stained with Azan (left panel) and connexin-43 (right panel; HRP-conjugated secondary antibody). Arrow indicates the junctional complex between two cardiomyocytes within the newly formed myocardium; SC, scar; MY, myocardium.

Figure 2

Characterization of [Ca²⁺]_i transients in the newly formed myocardium. (a) Full-frame TPME image of cells at the apical scar/myocardium border of an MHC-cycD2 heart at 180 days post-MI (i.e., the newly formed myocardium). The white bar demarks the position of line-scan mode data acquisition and numbers indicate the position of individual cardiomyocytes; SC, scar; MY, myocardium. (b) Stacked line-scan image acquired during automatic depolarization from the heart depicted in panel (a). (c) Spatially integrated traces of the changes in rhod-2 (red) fluorescence during spontaneous depolarizations from the heart depicted in panel (a). (d) Superimposed tracings of spontaneous changes in rhod-2 fluorescence as a function of time from cardiomyocytes at the scar/myocardium border (upper traces) and from remotely-located cardiomyocytes (lower traces). For each cell, spatially averaged changes in rhod-2 fluorescence were obtained and subsequently normalized such that 0 represents the fluorescence intensity before the [Ca²⁺]_i transient and 1 represents the peak fluorescence intensity. Similar results were obtained when the preparations were paced via point stimulation at remote sites of the myocardium.

Figure 3

Functional parameters in infarcted non-transgenic (open bars) and MHC-cycD2 (black bars) mice, normalized to their respective sham-operated animals. Y-axis: % in sham operation groups. P<0,05 between txg (transgenic mice) and non-txg (non-transgenic mice) at 60 and 180 days post myocardial infarction.