Semiquantitative histomorphometric analysis of myocardium following partial left ventriculectomy: 1-year follow-up
Abstract
Background:
Although partial left ventriculectomy (PLV) may have beneficial clinical effects in patients with dilated cardiomyopathy (DCM), there are no reports on effects of PLV on myocardial histology. The objective of this study was to assess histological properties of the LV myocardium 1 year following PLV as compared to histology at the time of the operation.
Methods:
The study group consisted of 15 consecutive PLV survivors, predominantly male (13/15), aged 45±12 years. Surgical specimens and endomyocardial biopsies, taken 12 months postoperatively, were processed routinely and stained with Masson-trichrome. The following morphometric parameters were assessed semiquantitavely: (1) degree of hypertrophy and attenuation; (2) nuclear evidence of hypertrophy; (3) myofibrillar volume fraction; (4) degree of degenerative vacuolar changes; and (5) fibrosis volume fraction.
Results
Both New York Heart Association (NYHA) functional class and ejection fraction (EF) improved 12 months following surgery as compared to preoperative values (2.40±0.69 vs. 3.33±0.49, p<0.001, and 33.21±12.05% vs. 20.21±9.07%, p<0.001, respectively). Morphometric analysis demonstrated postoperative decrease in the degree of attenuation as compared to preoperative values (1.40±0.51 vs. 2.47±0.64, p<0.01), as well as a decrease in fibrosis volume fraction (2.07±0.80 vs. 2.67±0.49, p<0.001) and nuclear hypertrophy (1.27±0.46 vs. 1.67±0.62, p<0.05). On the other hand, postoperative increase in myofibrillar volume fraction (1.87±0.61 vs. 1.40±0.61, p<0.01) was noted.
Conclusion
One year postoperatively, PLV has favourable effects on myocardial morphology that parallels improvement in the patient's functional status and LV systolic function.
1. Introduction
Morphological and morphometric features of the myocardium in patients with advanced stage dilated cardiomyopathy (DCM) have been critically analyzed 1,2. The most frequently used method of morphometric assessment is measurement of volume fractions of myocardial fibers and interstitial fibrosis, as well as diameters of different cardiomyocyte structures (fibers, nucleus, vacuoles, etc.). These morphometric parameters are the major histological indicator of myocyte condition, and can be used for prognostic purposes 3,4. Commonly used methods for myocardial morphometry are: semiquantitative scoring, point-counting method, and measurement by computer-based software 5,6. We have demonstrated good correlation between the three methods of morphometry in myocardial tissue 7.
Partial left ventriculectomy (PLV) was introduced for treatment of patients with end-stage idiopathic DCM 8,9. Previous studies have demonstrated the beneficial hemodynamic effects of PLV on ventricular function 10,11. However, to our knowledge, no study has directly compared different myocardial histomorphometric parameters before and after PLV. In order to determine possible morphometric changes in the myocardium after PLV, we made semiquantitative assessment of major morphometric parameters in surgically obtained LV specimens during the operation and compared them with measurements performed on endomyocardial biopsy (EMB) samples obtained 1 year later.
2. Patients and methods
2.1. Patients
The study group included 15 consecutive patients with dilated cardiomyopathy in whom PLV was performed due to end-stage heart failure, and who survived 1 year following operation. The investigation conforms with the principles outlined in the Declaration of Helsinki. The study design was approved by Dedinje Cardiovascular Institute Ethics Committee as a part of extensive evaluation of the effects of PLV. Informed consent was obtained from all patients.
2.2. Clinical and hemodynamic data
New York Heart Association (NYHA) functional class was determined, by experienced cardiologist, preoperatively and 1 year postoperatively. Left ventricular ejection fraction (EF) was measured by 2D echocardiography using Simpson's biplane method. Cardiac index and pulmonary capillary wedge pressure (PCWP) were determined by Swan–Ganz catheter. Thermodilution method was used for calculation of cardiac index.
2.3. Surgical procedure
Surgical technique was described in detail elsewhere 9. A switch from beating heart technique described by Batista to cardioplegic arrest was undertaken to provide still operative field and to enable better approach to the mitral valve. Mitral valve was repaired in eight patients, while three patients in whom repair was not feasible underwent mitral valve replacement with Medtronic–Hall monoleaflet artificial valve (size 27 or 29). Tricuspid valve repair was performed on the beating heart in nine patients after ventricular closure.
2.4. Myocardial specimens
Myocardial specimens were obtained from patients who underwent PLV, as surgical biopsies from the excised part of the LV. One year after the operation, the same patients underwent EMB for diagnostic and morphometric purposes. In order to avoid sampling variability and to standardize source of histomorphometric information, EMB specimens in all patients were taken from the inferobasal and/or inferolateral segments of the left ventricle.
The EMB and surgical specimens were embedded in paraffin, routinely processed and stained with Masson-trichrome. The representative regions in each specimen were chosen for morphometric analysis, and particular attention was paid to exclude subendocardial or perivascular areas with high amount of fibrosis. Three to five surgical specimens and EMB samples were analyzed per patient, using high power enlargement (400×).
2.5. Semiquantitative analysis
The measurements in chosen representative fields were graded using a usual semiquantitative scoring system: 0=no changes (minimal), 1=mild, 2=moderate, and 3=severe changes. We measured: (1a) myofibrillar hypertrophy (cross-sectional diameter at nuclear level), (1b) myofibrillar attenuation, (2) nuclear evidence of hypertrophy (diameter), (3) myofibrillar volume fraction, (4) vacuolar degeneration, and (5) fibrosis volume fraction. Degrees of myofibrillar hypertrophy and attenuation were mutually exclusive (these findings could not be judged to be both predominantly severe at the same time, i.e., when hypertrophy was judged to be severe, attenuation had to be mild and vice versa). Semiquantitative measurements were done in blinded manner by cardiovascular pathologists (JVD and ZVP) with extensive experience in morphometric analysis 11,12.
2.6. Statistical analysis
Data are expressed as mean±S.D. Clinical, hemodynamic, and morphometric data were compared using t-test for paired samples. p value at the level <0.05 was considered significant. Logistic regression model, that included operative histomorphometric variables, was constructed in order to determine whether histologic findings at the time of operation can be predictive of >5% increase in ejection fraction at 1-year follow-up. For this analysis alone, a p value of <0.10 was considered significant.
3. Results
Basic demographic data of patients enrolled in the study are shown in Table 1. Briefly, patients were predominantly male with an average age of 45 years, ranging from 20 to 64 years. Viral etiology of otherwise idiopathic dilated cardiomyopathy was suspected in two patients and postpartum in one, but the causative factor in all patients remained unknown. Patients had a history of symptomatic heart failure of more than 3 years on average, with frequent hospitalizations due to congestive heart failure.
| Sex (male) | 13/15 |
| Age (years) | 45±12 |
| Suspected etiology of DCM | |
| Unknown | 12/15 |
| Viral | 2/15 |
| Postpartum | 1/15 |
| Duration of symptoms (months) | 38±36 |
| Number of hospital admissions for CHF | 4.1±1.5 |
- a Abbreviations: CHF, congestive heart failure; DCM, dilated cardiomyopathy.
At the end of 1-year follow-up, NYHA functional class significantly improved as compared to preoperative values (3.33±0.49 vs. 2.40±0.69, p<0.001). Similarly, left ventricular contractility, expressed by ejection fraction, also improved during the follow-up (20.21±9.07% vs. 31.21±12.05%, p<0.001). On the other hand, cardiac index and pulmonary capillary wedge pressure at the end of the follow-up were similar to preoperative values (Table 2).
| Preoperatively | At 1 year | |
|---|---|---|
| NYHA class | 3.33±0.49 | 2.40±0.69‡ |
| EF (%) | 20.21±9.07 | 31.21±12.05‡ |
| CI (L/min/m2) | 2.23±1.01 | 2.72±0.83 |
| PCWP (mm Hg) | 20.91±10.01 | 19.82±10.23 |
- a Abbreviations: NYHA, New York Heart Association; EF, ejection fraction, CI, cardiac index; PCWP, pulmonary capillary wedge pressure.
- ‡ p<0.001.
When the samples obtained 1 year postoperatively were compared to their operative counterparts, it became evident that there was substantial improvement in myocardial morphology (Table 3). Namely, decrease in fibrosis volume fraction (2.07±0.80 vs. 2.67±0.49, p<0.001), myofibrillar attenuation (1.40±0.51 vs. 2.47±0.64, p<0.001), and nuclear hypertrophy (1.27±0.46 vs. 1.67±0.62, p<0.05) were noted, as well as an increase in myofibrillar volume fraction (1.87±0.64 vs. 1.40±0.61, p<0.01). Although there was a trend toward decrease of myofibrillar hypertrophy and vacuolar degeneration, it did not reach statistical significance. A logistic regression model constructed for identification of preoperative histologic variables predictive of >5% increase in ejection fraction at 1-year follow-up failed to identify any of these variables as independent predictors of significant improvement in LV systolic function (Table 4).
| Preoperative | At 1 year | |
|---|---|---|
| Myofibrillar hypertrophy | 1.33±0.62 | 1.13±0.35 |
| Myofibrillar attenuation | 2.47±0.64 | 1.40±0.51‡ |
| Nuclear hypertrophy | 1.67±0.62 | 1.27±0.46* |
| Myofibrillar volume fraction | 1.40±0.61 | 1.87±0.64† |
| Vacuolar degeneration | 1.87±0.74 | 1.47±0.52 |
| Fibrosis volume fraction | 2.67±0.49 | 2.07±0.80‡ |
- ‡ p<0.001.
- * p<0.05.
- † p<0.01.
| p value | |
|---|---|
| Myofibrillar hypertrophy | 0.58 |
| Myofibrillar attenuation | 0.34 |
| Nuclear hypertrophy | 0.39 |
| Myofibrillar volume fraction | 0.60 |
| Vacuolar degeneration | 0.65 |
| Fibrosis volume fraction | 0.85 |
4. Discussion
Partial left ventriculectomy, as a surgical procedure for LV volume reduction, has been proposed as an alternative treatment approach for patients with end-stage dilated cardiomyopathy, mainly in countries with an inadequate heart transplant program 13,14. The main concept of the procedure is that the reduction of the LV diameters will result in mechanical improvement in LV emptying that will result in partial restoration of myocardial contractility. It has been suggested that LV ejection fraction improves mainly through a decrease in end-systolic stress 11. It is tempting to speculate that the mechanical improvement in ventricular performance will translate into histological remodelling of myocardial components. In addition to slowing the progression of the underlying disease, such structural changes could bring further improvement of LV function and longer survival expectancy in such patients.
Previous studies have demonstrated the beneficial hemodynamic effects of PLV on ventricular function, with improvement of left ventricular function in survivors 15. However, its wider clinical acceptance is limited by high early postoperative mortality and lack of definitive evidence of sustained improvement.
After the introduction of EMB, several authors noted the correlation between structural changes in the myocardium and its functional state 16,17. The assessment of heart muscle diseases by various methods has been the subject of numerous histological studies 18,19. It appears that the percent of fibrosis, degree of hypertrophy, attenuation and degeneration, as well as specific clinical parameters, may have important prognostic value in patients with end-stage heart muscle disease 16,17.
Our study demonstrated a decrease in the degree of myofibrillar attenuation, nuclear hypertrophy, and fibrosis volume fraction 1 year following PLV. Although the decrease of attenuation of myocardial fibers and diminution of nuclear signs of hypertrophy may be attributed to the reduction in heart dilatation, decrease of fibrosis volume fraction is a more relative estimate. It appears that consequent increase in myocardial volume fraction is also relative, due to lesser degree of interstitial edema and attenuation.
On the other hand, lack of pre- and postoperative myocardial hypertrophy and degenerative vacuolar changes in myocardial fibers indicates that substantial histomorphometric “rearrangement” did not occur. From the morphometric point of view, more extensive changes in fiber diameter and the amount of degeneration would be preferred over the changes noted in relative myocardial and fibrosis volume fractions. These findings could be attributed to the relatively short follow-up period, need for more accurate morphometric measurements, and/or irreversibility of the major morphological changes in end-stage dilated cardiomyopathy.
Interestingly, McCarthy et al. 20 demonstrated that insertion of left ventricular assist device (mean duration 76±34 days) was associated with the improvement of markers of acute myocyte damage (number of wavy fibers and contraction band necrosis), but also with an increase in myocardial fibrosis.
Recent studies have shown that among factors influencing outcome following PLV, only myocardial cell diameter and fibrosis severity significantly affected survival 8,21,22. However, logistic regression model in our study failed to identify any preoperative semiquantitative morphometric variable as an independent predictor of significant increase in LV systolic function. It should be noted that these studies did not compare morphometric parameters before and after surgery.
A major limitation of this study is that we excluded, by study design, patients who did not survive for 1 year following surgery and who presumably had worse histology than survivors. Consequently, it is not clear what effects PLV would have had on myocardial morphology in these patients. Another potential limitation is that endomyocardial biopsy, by its definition, gives regional information, but it is commonly used to predict overall histology, especially in subjects with idiopathic dilated cardiomyopathy.
In conclusion, our histomorphometric findings 1 year after PLV showed favourable effects on myocardial morphology that parallel the patient's functional status and LV systolic function. Although there is histological evidence of improvement, some of the functional and hemodynamic enhancements are difficult to explain with histomorphometric parameters only. Therefore, it appears that morphological findings and functional status are two close, but still separate categories.
