Previous studies described the M-band as a dynamic element of the sarcomeric cytoskeleton which adapts to the contractile regime of a particular muscle by varying the type and number of different myomesin molecules cross-linking the thick filaments [3
]. Here, we characterize the M-band protein composition in mouse and human heart, in both healthy and pathological situations. We show that expression patterns of myomesins correlate with the physiological parameters of the heart muscle, and vary depending on species, developmental stage or type of cardiomyopathy (Tables , ).
Summary of expression data of myomesin proteins in the heart of mouse and human
Summary of alterations in expression of myomesins in DCM in mouse and human
The M-band protein composition is identical in embryonic heart of mouse and human and is restricted to the EH-myomesin isoform. However, this isoform is down-regulated around birth and the adult hearts show distinct expression patterns of myomesins (Table ). Human myocardium accumulates much more myomesin-3 compared to mouse heart. Thus, the M-band composition of mouse heart resembles fast skeletal fibers, while that in human heart is more close to intermediate speed fibers [36
]. This can be explained by different physiological demands to the M-bands. The mouse heart beats much faster than the human one, and the mouse heart myosin produces much higher contraction speed [7
]. Both parameters probably necessitate the presence of a “faster” M-band in mouse myocardium. Our observations showed the limitations in direct applicability of results obtained from mouse models for the human heart. The differences in physiological characteristics and in the cytoskeletal design between mouse and human hearts should always be taken into account by any prediction.
Our experiments showed an accumulation of EH-myomesin in both mouse models of DCM (MLP-KO and β-catenin c
ex3), which correlated well with the decrease of ejection fraction at the early stage of disease. This suggests EH-myomesin as an early marker of DCM. In addition to EH-myomesin up-regulation, we found that some DCM mouse models (e.g. MLP-KO mice) up-regulated myomesin-3 expression, which might represent a fine-tuning of the M-band to specific biomechanical alterations. As discussed above, the adult human heart has a different M-band composition compared to mouse. However, the up-regulation of EH-myomesin was a constant characteristic also of DCM in human heart, irrespective of the cause of the disease. We found that expression of EH-myomesin is dramatically up-regulated (41-fold, P
< 0.001) in the DCM group compared to the control and HCM groups. The up-regulation of EH-myomesin was much lower in DCM hearts supported by a LVAD.
The up-regulation of EH-myomesin in DCM is consistent with the reported re-expression of fetal isoforms of sarcomeric proteins in the dilated heart [10
]. Accordingly, the appearance of the myomesin isoform containing the additional elastic EH-fragment matches the up-regulation of compliant titin isoforms in human DCM [26
], suggestive of a correlative adaptation of both cytoskeletal structures in this pathological situation.
What might be the reason for such a remodeling of the sarcomere cytoskeleton? We propose that it is a general adaptation to the altered contractile mechanics of the sarcomere in the dilated heart. Indeed, our observations show that up-regulation of EH-myomesin is universal for all tested DCM hearts, irrespectively of species and regardless to the cause of the disease. Also, the increase of the EH-myomesin expression correlates well with the decrease of the heart pumping function and correspondingly to the increase of volume overload in the dilated heart. Previously, EH-myomesin expression was reported in fibers working in the eccentric contraction regime in mouse skeletal [4
] and rat extraocular muscles [39
]. The sarcomere in the dilated heart has to contract in overstretched conditions as well, requiring the switch to a “fuzzy sarcomere” design, which is more stable in eccentric conditions [2
]. The application of LVAD leads to a decreased load of the contracting sarcomere, which correlates with much lower levels of EH-myomesin in the LVAD supported hearts.
The remodeling to a more embryonic type of cytoskeleton might also be an adaptive response to calcium signaling defects in the failing heart. Indeed, the failing human heart shows slower relaxation kinetics caused by delayed Ca2+
] due to reduced SERCA function, enhanced NCX function, and greater Ca2+
leak from the sarcoplasmic reticulum [6
]. In the embryonic heart, the Ca2+
re-uptake is compromised by immature excitation–contraction coupling [11
]. The LVAD support improves the contractility of the failing heart and shortens the rate of the Ca2+
decay to the level of non-failing heart [8
]. This correlates with our finding that EH-myomesin expression in LVAD supported hearts does not differ significantly from the non-dilated control hearts.
We have found that the M-band composition changes in a cell-autonomous fashion in the failing heart. In the heart of β-catenin c
ex3 mice, some cardiomyocytes switch completely to the embryonic M-band phenotype by up-regulating EH-myomesin and down-regulating M-protein, whereas others keep the phenotype of adult cardiomyocytes with high levels of M-protein and very low levels of EH-myomesin. The down-regulation of M-protein is not detectable in the human DCM hearts, but EH-myomesin is also up-regulated in a cell-specific manner. We intend that increased myocardial strain in dilated heart might provoke the up-regulation of EH-myomesin and “patchy”, cell-specific expression pattern might be caused by a different mechanical burden on certain myocytes. This suggestion is supported by the observation that a strong up-regulation of EH-myomesin is observed in the scar region of the heart remodeled after myocardial infarction, as the ventricular wall is very thin in the scar region and the slim layer of remaining myocytes is subjected to huge contractile stress. This is a very important observation, because all molecular alterations found by previous studies in the diseased myocardium were averaged values for the whole heart and did not consider the divergence between individual cardiomyocytes. We, therefore, hypothesize that M-band alterations might be part of a general adaptation of the sarcomeric cytoskeleton to unfavorable working conditions in the failing heart, and might also affect the mechanical properties of the cardiomyocytes. The progressive cell-to-cell heterogeneity through the myocardial wall would impair the force transmission and might have important implications for the mechanism of cardiac failure. Computer models showed that occurrence of mechanical non-uniformities in myocardial muscle might lead to spontaneous Ca2+
release from sarcoplasmic reticulum and initiate arrhythmogenic waves [37
]. Thus, the cytoskeletal alterations by DCM, initially driven by a compensatory mechanism, may eventually become maladaptive with disease progression. However, further studies are needed to clarify the role of cytoskeletal non-uniformities in the development of arrhythmias in the failing heart.
Here, we present for the first time that EH-myomesin, the dominant myomesin isoform in the embryonic heart, is up-regulated in DCM. Since we have previously shown that this myomesin isoform is associated with a higher degree of M-band elasticity [35
], this finding might give new insights into the understanding of progressive ventricular dilation on a structural level. Furthermore, the up-regulation of EH-myomesin at early stages of the disease and its correlation to the impairment of ventricular function may be used for a better characterization of DCM in human patients.