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Cardiovascular Research 2004 62(2):426-436; doi:10.1016/j.cardiores.2003.12.010
© 2004 by European Society of Cardiology
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Copyright © 2004, European Society of Cardiology

Connexin 43 expression and distribution in compensated and decompensated cardiac hypertrophy in patients with aortic stenosis

Sawa Kostin*,a, Sebastian Dammera, Stefan Heinb, Wolf P Klovekornb, Erwin P Bauerb and Jutta Schapera

aDepartment of Experimental Cardiology, Max-Planck-Institute, Benekestr. 2D-61231, Bad Nauheim, Germany
bDepartment of Cardiac Surgery, Kerckhoff Clinic, Bad Nauheim, Germany

* Corresponding author. Tel.: +49-6032-705402; fax: +49-6032-705-419. Email address: skostin{at}kerckhoff.mpg.de

Received 7 October 2003; revised 7 December 2003; accepted 10 December 2003


   Abstract
Top
 Abstract
1. Introduction
 2. Materials and methods
 3. Results
 4. Discussion
 5. Conclusions
 References
 
Objectives: Gap junctions (GJ) are important determinants of conduction. In advanced heart failure alterations of the major ventricular GJ protein, connexin 43 (Cx43) are found. However, changes in Cx43 expression during the progression from compensated cardiac hypertrophy to heart failure, especially in humans, have not been studied extensively. The aim of the present study was to investigate changes in Cx43 expression and distribution in compensated and decompensated left ventricular (LV) hypertrophy in pressure-overloaded human hearts with valvular aortic stenosis (AS). Methods: We measured Cx43 levels by Western blot and quantitative immunoconfocal microscopy of LV septum biopsies from three groups of patients with AS (group I (n=9): ejection fraction (EF)>50%; group II (n=12): EF 30–50%; group III (n=9): EF<30%). LV biopsies from six patients with mitral valve stenosis and two donor hearts served as controls. Results: Only the early phase of LV hypertrophy (AS-I) was characterized by extensive Cx43 lateral staining. As compared to controls, the AS-I group showed a 44.3% increase in Cx43 protein, which was reflected in an augmented number of GJs per 100 µm2 intercalated disc area (control: 62.5±6.4 vs. AS-I: 79.8±4, p<0.001) and an increased GJ surface density (control: 0.00547 vs. AS-I: 0.00724 µm2/µm3, p<0.01). Decompensated LV hypertrophy (AS-III) was specified by reduced percentage of the Cx43 signal per myocyte area (control: 1.74% vs. AS-III: 1.31%, p<0.01) or per intercalated disc (control: 18.3% vs. AS-III: 11.3%, p<0.005). Mean GJ area and GJ number per intercalated discs in the AS-III group were decreased significantly by, respectively, 42.5% and 36.4% as compared to control. In addition, decompensated LV myocardium showed a markedly heterogeneous spatial distribution of Cx43. Conclusion: The quantity and spatial distribution of Cx43 differs markedly between compensated and decompensated LV hypertrophy in human patients with AS. Upregulation of Cx43 in compensated hypertrophy may represent the immediate adaptive response to increased load, whereas diminished and heterogeneous Cx43 distribution in decompensated hypertrophy may play maladaptive roles culminating in heart failure and ventricular arrhythmias.

KEYWORDS Gap junctions; Connexins; Remodeling; Hypertrophy


   1. Introduction
Top
 Abstract
 1. Introduction
2. Materials and methods
 3. Results
 4. Discussion
 5. Conclusions
 References
 
Central to the highly organizational pattern of the direct cell-to-cell metabolic and electrical coupling are the gap junctions (GJ) and their constituent proteins, the connexins. Growing experimental and clinical evidence suggest that alterations in the amount and distribution of the major ventricular GJ protein connexin43 (Cx43) can lead to altered patterns of conduction, re-entrant arrhythmias and sudden death (reviewed in Refs. [1–6]). Reduced expression of Cx43 has consistently been observed as a common feature in a variety of chronic human heart diseases, such as idiopathic cardiomyopathies, ischemic injury or myocarditis [7–13], that are associated with an increased risk of cardiac arrhythmias and sudden death. In addition, disrupted patterns of GJ distribution and decreased expression of Cx43 are also common features of cardiac remodeling observed in a variety of animal heart failure models [14–17]. Thus, the concept that GJs in the failing myocardium undergo dramatic remodeling is now well documented.

In addition to studies in failing hearts on Cx43 expression and distribution, a number of experimental studies have investigated alterations in Cx43 and GJs in the hypertrophied myocardium [15,18–23]. In marked contrast to the bulk of experimental data, the knowledge of Cx43 expression and distribution in human patients with left ventricular (LV) hypertrophy is extremely limited. At present, the information on Cx43 expression in LV hypertrophy in humans is restricted to only 5 patients with pressure overloaded hearts [10]. In the present study, we have therefore studied LV samples obtained from 30 patients with LV hypertrophy due to aortic stenosis (AS) at different stages of compensation by employing multiple quantitative methods to determine Cx43 expression and distribution. By comparing with LV specimens obtained from eight control hearts, we show that compensated pressure-induced hypertrophy differs markedly from decompensated hypertrophy in both, quantity and spatial distribution of Cx43.


   2. Materials and methods
Top
 Abstract
 1. Introduction
 2. Materials and methods
3. Results
 4. Discussion
 5. Conclusions
 References
 
2.1. Patients
The institutional Ethical Committee approved the study and all patients gave written informed consent. Thirty patients with isolated AS were subdivided in three subgroups on the basis of ejection fraction (EF) determined by quantitative echocardiography at the time of hospitalization: group I (AS-I), EF>50%; group II (AS-II), EF 30%–50%; and group III (AS-III), EF<30%. All patients underwent surgical aortic valve replacement. Based on clinical and hemodynamic data (Table 1), the myocardium from the AS-I group was regarded as representing compensated LV hypertrophy, the AS-II group as the transitional stage and the AS-III group as representing the failing myocardium. LV myocardium from two donor hearts that for technical reasons were not used for transplantation and intraoperative myocardial samples from papillary muscles obtained from six patients with mitral valve stenosis with normal LV function served as control tissues. Previous ultrastructural and immunoconfocal studies showed that quantitative relationships of GJs and intercalated discs in the papillary muscles closely resemble those in areas of the LV free walls [24,25].


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  		Table 1 Clinical preoperative data

 
2.2. Tissue preparation and immunofluorescent labeling
During open heart surgery, myectomy samples were removed from the LV septum, immediately frozen in liquid nitrogen and stored at –80 °C. Before immunolabeling, tissue characterization and orientation was recorded by hematoxylin–eosin staining. Frozen sections were fixed for 15 min with 4% paraformaldehyde and then incubated with the primary antibodies: polyclonal anti-Cx43 (Zymed), monoclonal anti-N-cadherin (clone GC-4, Sigma) and monoclonal anti-{alpha}-actinin (clone EA-53, Sigma). Anti-mouse or anti-rabbit IgG-conjugated with Cy3 or Cy2 (Biotrend) served as detection systems. The nuclei were stained with TOTO-3 (Molecular Probes). F-actin was fluorescently stained using TRITC-conjugated phalloidin (Sigma).

2.3. Confocal microscopy
The samples were examined by confocal scanning laser microscopy (Leica TCSNT), equipped with argon/krypton and helium/neon lasers. Confocal images were obtained using different Leica objectives and three confocal detectors for reflected fluorescence. Each recorded image was taken using multi-channel scanning and consisted of 1024 x 1024 pixels. Series of confocal optical sections were taken through the depth of the tissue sample at 0.5–1-µm intervals. In order to improve image quality and to obtain a high signal/noise ratio each image from the series was signal-averaged. After data acquisition, the images were transferred to a Silicon Graphics Indy or Octane workstations (Silicon Graphics) for image restoration and reconstruction using Imaris®, the multichannel image processing software (Bitplane, Zürich, Switzerland). The principles of this method have been previously described [26,27].

2.4. Quantitative analysis
2.4.1. Quantification of Cx43 in longitudinal sections
Areas selected for quantitative analysis of Cx43 consisted of LV myocytes cut in a plane parallel to the long axis of the cells. To reduce variability between preparations, all sections were immunolabeled simultaneously using identical dilutions of primary and secondary antibodies. The standardized parameters of imaging, zoom, pinholes, objective and laser power were kept constant for recording of data in all measurements. From each optical field (size 200 x 200 µm), 10 confocal slides were obtained at a 1-µm interval. The values of myocyte area and myocyte volume were immediately determined using the Leica TCNT software and were based on clearly delineated cell borders by N-cadherin labeling. From the fifth of a total 10 sections forming a stack of confocal images, several histograms of Cx43 fluorescence intensity (FI) were obtained. The first histogram represented the total FI of Cx43 per myocyte. The following histograms represented Cx43 signal intensity that was not associated with transverse boundaries of the cell, i.e. with the intercalated discs. The histograms were converted into Macintosh Excel data for calculation of Cx43 signal as previously described [26–28]. The area of positive Cx43 labeling was defined as the number of pixels with Cx43 signal intensity exceeding the threshold of 50 on the 0–255 FI scale. Thereafter, the total number of positive pixels was converted into square micrometers. The quantity of Cx43 per myocyte area was expressed as the percent of myocyte surface area occupied by Cx43 positive label. The quantity of Cx43 per myocyte volume (GJ surface density) was calculated from measurements of the total GJ area per myocyte surface divided by the values of the myocyte volume and expressed as µm2/µm3. In total, Cx43 expression in longitudinal sections was quantified in 1513 myocytes or in 43.3±15.8 myocytes per each patient (range 15–83 cells).

2.4.2. Quantification of Cx43 in transverse sections
To measure the number and size of individual GJs we used en face viewed intercalated discs derived from transversely sectioned tissue. It has been reported that in this view the overlap of individual GJs is minimized [29–31]. Quantification of Cx43 in transverse sections was done using protocols reported by Severs et al. [8,29–31] with several modifications. In brief, for quantification of Cx43 in transversely sectioned myocytes we used double-labeled tissue sections for Cx43 and N-cadherin. All sections from all patients were immunolabeled simultaneously using identical dilutions of primary and secondary antibodies. Sections were scanned under identical parameters of imaging, zoom, pinholes, objective and laser power. A 63 x objective, 2.0 zoom factor with a detector pinhole of 90-µm and a 1024 x 1024-pixel image gave an optical section thickness of 0.5-µm. From each microscopic field, a series of confocal images through the depth of tissue containing the complete disc was recorded. During acquisition, each confocal slide was signal-averaged and the entire series was projected as a single composite image by superimposition using Imaris® software (Bitplane, Zürich, Switzerland). This final image was quantitatively analyzed using NIH image software. A threshold value of 50 on a 0–255 units FI scale was applied to reduce any background levels, and the image was then edited automatically to outline individual GJs. The following parameters were analyzed: intercalated disc area, the proportion of GJs per disc area, the mean GJ length and area, and the number of GJs per 100 µm2 disc area. These parameters were determined in a total of 1002 intercalated discs or in 29.4±6.6 intercalated discs per each patient (range 21–44 discs), containing 4261±1577 GJs (range 2249–7752 GJs).

2.5. Western blot
Ten micrograms of total protein per line was run on 12% SDS polyacrylamide separating gels and electrophoretically transferred onto nitrocellulose membrane (Invitrogen) overnight. The membrane was incubated overnight with anti-Cx43 antibody (Zymed) in 1:1000 dilution, washed and incubated with alkaline phosphatase-conjugated secondary antibody and SuperSignal WestFemto detection system. Quantification was done by scanning of the immunoblots on a STROM 860 (Amersham Pharmacia Biotech) using ImageQuant software. In order to exclude the influence of fibrosis on myocyte proteins, immunoblotting for sarcomeric {alpha}-actin was performed and the values of Cx43 were standardized to sarcomeric actin.

2.6. Statistical analysis
Results are reported as means±S.D. For multiple comparisons we used ANOVA, followed by analysis with the Bonferroni t-test. Differences between groups were considered significant at p<0.05.


   3. Results
Top
 Abstract
 1. Introduction
 2. Materials and methods
 3. Results
4. Discussion
 5. Conclusions
 References
 
3.1. Cx43 distribution
3.1.1. Patterns of Cx43 distribution in longitudinal sections
In longitudinally sectioned LV myocytes from normal hearts, Cx43 is located mainly at the intercalated discs, with little signal at the lateral sarcolemma (Fig. 1A). In the AS-I group, Cx43 labeling was distributed not only at the intercalated discs, but also at the lateral sarcolemma (Fig. 1B). Lateralization of Cx43 was less evident in the AS-II and AS-III groups (data not shown). Similar to Cx43 labeling, N-cadherin in normal hearts was exclusively restricted to the transverse boundaries of the myocytes (Fig. 1C). In contrast to Cx43 distribution, N-cadherin in the AS-I group was prominently concentrated at the myocyte termini (Fig. 1D). Similar patterns of N-cadherin labeling were found in the AS-II and AS-III groups (data not shown). Double labeling for Cx43 and N-cadherin clearly revealed that GJs in the AS-I group displayed a marked dispersion over the lateral myocyte surface distant from N-cadherin labeling (Fig. 1E). In contrast, in control and in the AS-II and AS-III groups, Cx43 co-localized mainly with N-cadherin at the intercalated discs (not shown).


Figure 1
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  		Fig. 1 Confocal images of Cx43 (green) and {alpha}-actinin (red) labeling in LV samples sectioned in longitudinal planes to the long myocyte axis in a control patient (A) and in a patient from group I (B). Arrows indicate Cx43 labeling confined to the intercalated discs; arrowheads point to the lateral Cx43 immunostaining. (C, D) Confocal images showing that N-cadherin signal (green) is exclusively confined to the transverse boundaries of the myocytes in control (C) and in AS-I (D). (E) Double labeling for Cx43 (red) and N-cadherin (green) in a patient with AS-I. The arrow denotes Cx43 labeling confined to the intercalated disc, arrowheads point to the lateral Cx43 immunostaining. In all micrographs, nuclei are stained blue with TOTO-3. In C and D, myofibrils are stained red with TRITC-phalloidin.

 
Quantitative analysis revealed that in control patients most of the Cx43 signal (91.7±5.3% from the total Cx43 per myocyte) is confined to the intercalated discs. In contrast, in the AS-I group, only 60.3±17% (p<0.01, compared to control) was restricted to the intercalated discs, whereas the remaining signal represented lateral Cx43 labeling. In the AS-II and AS-III groups, the proportion of Cx43 signal confined to the intercalated discs averaged, respectively, 82.5±10.8% and 87.2±4.7% and did not differ statistically from control values. These data indicate that only the compensated phase of LV hypertrophy is characterized by lateralization of Cx43.

Typical of normal ventricular myocardium is a homogeneous distribution of Cx43. Fig. 2A and B compare the uniform distribution of Cx43 with that of N-cadherin in a control patient. A consistent feature of the failing myocardium due to AS is the heterogeneous distribution of Cx43. Fig. 2D shows a homogeneous distribution of N-cadherin in a patient from the AS-III group which contrasts with heterogeneously distributed Cx43 in terms of diminished Cx43 signal (Fig. 2C). Cx43 heterogeneity was not observed in control or AS-I and AS-II groups, whereas in the AS-III group it affected approximately 10–15% of the myocardium.


Figure 2
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  		Fig. 2 Double labeling for Cx43 (red) and N-cadherin (green) in a control patient (panels A and B) and in a patient from group III (C and D). Note the homogeneous distribution of both N-cadherin and Cx43 in panels A and B. In panel D, the area of the diminished Cx43 signal is traced with a dashed line. In all micrographs, nuclei are stained blue with TOTO-3.

 
3.1.2. Patterns of Cx43 distribution in transverse sections
Fig. 3 compares Cx43 distribution in relation to N-cadherin-positive regions of the intercalated discs in transversely sectioned myocytes. Consistent with previous observations [8,10,19,22,29–32], the intercalated discs in this view contain numerous centrally located small GJs surrounded by large GJs at the disc periphery. In the AS-I group, the overall distribution of GJs at the intercalated discs was essentially similar to that in control (Fig. 3A and B); however, centrally located small GJs were noticeably increased in number (Fig. 3C and D). The intercalated discs in the failing hearts (AS-III group) showed a striking loss of the centrally located small GJs and an apparent loss of the peripherally situated large GJs exhibiting concomitantly a decreased plaque size (Fig. 3E and F).


Figure 3
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  		Fig. 3 Confocal images of Cx43 (red) and N-cadherin (green) in myocytes sectioned transversely from a control patient (A, B) and in patients with AS-I (C, D) and AS-III (E, F).

 
3.2. Quantification
3.2.1. Quantitative analysis of Cx43 in myocytes sectioned longitudinally
Fig. 4 shows that in adult human myocardium, the Cx43 area occupies 1.74% of the myocyte area resulting in a gap junction surface density of 0.00547 µm2/µm3 of myocyte volume. The percent of GJs per cell area had a trend to be increased in the compensated LV hypertrophy (AS-I), whereas in decompensated hearts (AS-III), the proportion of Cx43 per cell was markedly diminished and occupied only 1.3±0.11% of the myocyte area (p<0.01). The differences between groups were even more evident when the quantity of Cx43 was calculated per myocyte volume. Thus, in the AS-I group, the GJ surface density was significantly increased by 33.4±27%. In contrast, the failing heart (AS-III group) displayed a 34.5±5.6% (p<0.01) reduction of Cx43 containing GJs per myocyte volume. There were no differences in this parameter between controls and the AS-II group.


Figure 4
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  		Fig. 4 Quantitative data of Cx43 labeling in myocytes sectioned in a longitudinal plane in controls and AS patients.

 
3.2.2. Quantitative analysis of Cx43 in LV samples sectioned transversely
Using double labeling for N-cadherin and Cx43 we have measured the intercalated disc area, the number and size of GJs. Fig. 5 shows that in all AS groups, the disc area was significantly increased as compared to controls. The proportion of Cx43 at the intercalated disc had a trend to be increased in AS-I, but was significantly lower in the failing hearts (AS-III). In control tissue, 100 µm2 of the intercalated disc area encompasses 62.5±6.4 GJs. In compensated hypertrophy (AS-I), there was a remarkable increase in the number of GJs, up to 80 junctions per 100 µm2 of the disc area. In contrast, the failing hearts (AS-III) showed a nearly two-fold loss of the number of GJs (39.9±4.2) as compared with AS-I (p<0.001). The mean GJ area in the control tissue was 0.65 µm2. When expressed as relative value, the mean GJ area was significantly reduced compared to control by 17±8.4% (p<0.01) and 42.5±8.6% (p<0.001), respectively, in the AS-II and AS-III groups or had a trend to be reduced in compensated hypertrophy (AS-I).


Figure 5
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  		Fig. 5 Quantitative data of the intercalated discs and Cx43 labeling in myocytes sectioned transversely.

 
Histograms comparing frequency distributions of GJ size classes are shown in Fig. 6. As reported previously [8,16,27,29,31,32], more than 50% of GJs are smaller than 0.5 µm in length. In the AS-I group, there was a significant augmentation of small GJs, whereas such GJs in the AS-III group were markedly reduced. These data indicate that compensated hypertrophy is characterized by a preferential proliferation of small GJs, whereas decompensated hypertrophy is associated with a concomitant decrease in the number of small centrally located GJs, as well as large GJs (>3 µm in length) situated at the disc periphery.


Figure 6
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  		Fig. 6 Frequency distribution of GJ size classes in control and AS patients.

 
3.2.3. Western blot
Densitometric quantification of the immunoblots revealed that the amount of Cx43 in compensated hypertrophy (AS-1 group) was significantly higher by 44.3±17.7% than that in control, whereas decompensated hearts (AS-III group) showed a 41.1±13.8% reduction in Cx43 protein as compared to control (Fig. 7). There were no differences in the amount of Cx43 between control and the AS-II group.


Figure 7
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  		Fig. 7 Representative Western blots and the amount of Cx43 normalized to actin. Data are expressed as percent relative to control.

 

   4. Discussion
Top
 Abstract
 1. Introduction
 2. Materials and methods
 3. Results
 4. Discussion
5. Conclusions
 References
 
4.1. Major findings
The present study shows that GJ remodeling in pressure overload-induced LV hypertrophy in human patients is a dynamic and multifaceted process that occurs in different levels: within the intercalated disc (changes in GJ number, loss of small centrally located or large peripheral GJs, or both), within the myocyte (dispersion and lateralization of GJs) and within the myocardial tissue (heterogeneous Cx43 distribution). Furthermore, we show that these patterns of GJ remodeling are associated with changes in Cx43 expression levels in a hypertrophy stage-dependent manner. The early phase of LV hypertrophy was characterized by increased Cx43 expression which was reflected in an increased number of GJs per intercalated disc, an augmented Cx43 immunofluorescent signal per myocyte volume and extensive Cx43 lateral labeling. The decompensated stage of cardiac hypertrophy was specified by overall reduced Cx43 signal per myocyte area, volume or per intercalated disc, decreased GJ size and number, and a heterogeneous pattern of Cx43 distribution. These conclusions are based on rigorous quantitative immunoconfocal measurements of Cx43 in longitudinally and transversely sectioned myocytes and complemented by immunoblotting analysis.

4.2. Cx43 in compensated cardiac hypertrophy
Compensatory hypertrophic growth of cardiac myocytes in response to an increased load is certainly a dynamic and complex process involving progressive changes in gene expression leading to increased expression of contractile proteins, assembly of new sarcomeres and improved contractile function [33]. Our data indicate that the hypertrophic response in human patients is associated with increased expression of Cx43 leading to an increased number of GJs. Similar findings were observed in the pressure-overloaded guinea pig hearts and in the volume overloaded pig hearts due to creation of an experimental aorto-caval fistula [20,21]. In contrast, in a rabbit model of LV volume overload due to aortic regurgitation, Cx43 was found to be decreased in the early stage and increased as lesion duration progressed [34]. On the other hand, in rats with the pressure overloaded hearts induced by monocrotaline treatment or by aortic banding, the cellular content of Cx43 was found to be unchanged as compared with that in control hearts [19,22]. One possible explanation of such contradictory results may include the use of different species, sampling periods, rapidity and severity of the hypertrophic response, or may stem from the fundamental difference in the effects of pressure versus volume overload on myocardial cell biology.

The very first evidence for enhanced Cx43 expression and intercellular coupling during initial stages of the hypertrophic response came from experimental studies on cultured cardiomyocytes exposed to early mediators of heart hypertrophy, such as cAMP and angiotensin II [35,36]. More recent studies have shown that mechanical load produced by pulsatile stretch causes rapid and marked upregulation (within 1 h) of Cx43 and a significant increase in conduction velocity in cultured ventricular myocytes [37,38]. It is well established that mechanical stretch stimulates signal transduction pathways that induce key features of hypertrophic remodeling [39], including upregulation of Cx43 expression by cardiac myocytes [37,38]. Recently, it has been shown that angiotensin II, endothelin I, vascular endothelial growth factor and transforming growth factor-β are important mediators of stretch-induced upregulation of Cx43 expression [40–43]. Previously, we have demonstrated in similar patients with AS that angiotensin II and transforming growth factor-β already increased in the early stages of cardiac hypertrophy [44,45]. Taken together, these clinical and experimental data indicate that a number of biochemical stimulators and signal transduction pathways plausibly converge on upregulation of Cx43—a feature consistently found in our patients with compensated hypertrophy. Thus, upregulation of the Cx43 protein in compensated hypertrophy constitutes an important part of the immediate adaptive response of the heart to increased workload.

Apart from the upregulation of Cx43, the compensated stage of LV hypertrophy in our patients with AS was characterized by a marked lateralization of Cx43. Such patterns of GJ remodeling have previously been observed in the pressure-overloaded rat hearts, in hypertrophic cardiomyopathy, and in the border zone of human, canine and rat myocardial infarcts [9,13,17,19,22,46]. Although it is not yet convincingly proven whether laterally dispersed Cx43 may contribute to a pro-arrhythmogenic substrate, several experimental studies have shown that lateralization of GJs is associated with the alterations of anisotropic conduction properties of different cardiac tissues [47,48], including the ventricular myocardium [22]. Whether lateral Cx43 may influence the electrophysiological properties of the compensated hypertrophied myocardium in human patients awaits additional studies.

4.3. Cx43 in decompensated cardiac hypertrophy
In chronically diseased human hearts due to cardiomyopathies, ischemic injury or myocarditis, reduced expression of Cx43 has been commonly observed [7–10,12,13]. Our results in myocardium failing because of pressure overload provide further support to the concept that downregulation of Cx43 is a typical feature of myocardial remodeling in heart failure regardless of its origin. This conclusion is in good agreement with the data obtained in a variety of animal heart failure models, including pressure overload in guinea pigs or rats [19,23], ischemia in dogs or rats [16,17,46], in monocrotaline-induced pulmonary hypertension [22] and lipopolysaccharide treatment [15].

The present findings of reduced levels of Cx43 and heterogeneous distribution of GJs in the failing myocardium due to AS would be expected to increase arrhythmia susceptibility, likely because of slowed ventricular conduction and unidirectional conduction block. This prediction is in good agreement with the observations of Cooklin et al. [49], who have shown that conduction velocity significantly decreases with increasing severity of hypertrophy and, most importantly, that it is affected by increases in GJ-mediated intercellular resistances. That reduced levels of Cx43 can result in slowed conduction and ventricular arrhythmia has also been demonstrated in studies carried out in mice heterozygous for a null mutation of the Cx43 gene with {approx}50% reduction in Cx43 content [50], that is comparable to the degree of reduction of Cx43 observed in our patients with AS. Furthermore, in our patients with decompensated LV, significant reductions of Cx43 were frequently superimposed on the heterogeneity of Cx43 distribution that could potentially lead to a non-uniform pattern of wave-propagation. Heterogeneity of Cx43 distribution in ventricular myocardium has consistently been observed in the ventricular myocardium of human patients [7] and experimental animals with heart failure [51]. Importantly, Kitamura et al. [52] have shown that the heterogeneous loss of Cx43 is responsible for the development of malignant ventricular arrhythmia in patients with dilated cardiomyopathy. Furthermore, a recent study of a murine model of heterogeneous GJ channel expression provided a direct causal link between Cx43 heterogeneity and conduction defects, and the occurrence of sudden death [53]. Collectively, these observations add substantial evidence that diminished Cx43 expression and GJ remodeling are important substrates of ventricular arrhythmias in the hypertrophied and failing myocardium.


   5. Conclusions
Top
 Abstract
 1. Introduction
 2. Materials and methods
 3. Results
 4. Discussion
 5. Conclusions
References
 
Disease severity-related changes in myocardial Cx43 expression and distribution exist between compensated and decompensated pressure-overloaded human hearts suggesting both adaptive and maladaptive roles of GJs in the transition from a phase of compensated structural and functional adaptation to a maladaptive state culminating in heart failure and ventricular arrhythmias.


   Acknowledgements
 
This study was supported by grants from the Kerckhoff Clinic, Bad Nauheim (Forschungsprojekt PFOR-371 to S.K.).


   Notes
 
Time for primary review 24 days


   References
Top
 Abstract
 1. Introduction
 2. Materials and methods
 3. Results
 4. Discussion
 5. Conclusions
 References
 

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