Mast cells have a number of disparate functions throughout the body, however, until recently the role of these immune cells in the heart has been largely ignored. This changed with the advent of several studies reporting increases in cardiac mast cell density secondary to various myocardial pathologies [5
]. These cardiac mast cells have been shown to be of the connective tissue type, containing a variety of biologically active mediators including cytokines and proteases that have the ability to activate MMPs. Recently we identified such a relationship between mast cell activation and MMP activity in the heart [5
]. Treatment with the mast cell stabilizing drug, cromolyn, initiated prior to the imposition of volume overload induced by creation of an AV fistula, prevented the increases in cardiac mast cell density and MMP activation during the critical first 5 days of volume overload [5
]. In a subsequent study, chronic inhibition of mast cell degranulation over an 8 wk period post-fistula effectively prevented the adverse LV structural dilatation, increased compliance and decreased contractility present in the untreated fistula group [6
]. These studies used pharmacologic inhibitors of mast cell activation, however, a more direct assessment of mast cell mediated remodeling is possible in animals such as in the Ws/Ws rat that lack the normal complement of mast cells. Thus, the results presented herein, conclusively establish the direct role of cardiac mast cells in mediating the ventricular remodeling response induced secondary to chronic volume overload.
Activation of MMPs is a crucial step in producing the adverse ventricular remodeling post-fistula [12
]. In the current study, a marked increase in MMP-2 activation was found in the myocardium of WT rats. This was in stark contrast to the mast cell deficient Ws/Ws rats in which MMP-2 activation following 5 days of ventricular volume overload did not occur. We have previously documented that there are no basal differences in interstitial collagen density between normal Ws/Ws and WT hearts [19
]. Therefore, we would interpret the significant decrease in the concentration of fibrillar collagen in the WT fistula groups, concurrent with these differences in mast cell mediated MMP-2 activation, to be the result of collagen degradation.
Consistent with previous findings demonstrating the relationship between increased activation of MMP-2 and decreased CVF with ventricular dilatation [5
], we observed marked differences in the extent of ventricular dilatation between the Ws/Ws and WT groups. The relative increase in LV diameter in the WT group was substantially greater in the first 2 weeks and continued to further increase at all subsequent time points post-fistula. However, despite MMP-2 activation not being appreciable and collagen being preserved in the Ws/Ws group, a small increase in LV diameter did occur over the 8 wk period of volume overload. It is expected that this relatively small increase in ventricular diameter in the Ws/Ws could be due to either the significant increase in LV end diastolic pressure that typically occurs post-fistula [15
], or could reflect compensatory adaptation due to structural dilatation and/or increased chamber compliance induced by other mechanisms possibly related to cardiomyocyte remodeling [16
]. While neither of these possibilities can be discounted on the basis of an in vivo
measurement of diameter, the likelihood that this relatively small increase in LV diameter observed in the Ws/Ws hearts stems from this increase in LV filling pressure, and not pathological remodeling, would be consistent with the results of Brower et al. [6
] where there was no difference in the LV end diastolic pressure-volume relationship between sham-operated rats and fistula rats treated with a mast cell stabilizing drug for 8 wks of sustained ventricular volume overload. Under normal conditions, both the Ws/Ws and their WT are reported to have similar LV masses over a variety of ages. Further, given the similarity in LV weight post-fistula, there is no indication that mast cells influence the hypertrophic response. However, the smaller chamber dimension in the mast cell deficient Ws/Ws is comparable to the findings in previous studies using mast cell stabilizing compounds and ACE [20
] or MMP inhibitors [12
], with the corresponding increase in mass to volume ratio indicative of a compensated heart. Thus, mast cells appear essential to the pathological remodeling that occurs in response to volume overload of the heart, while alternative mechanisms are responsible for compensatory mechanisms.
Immuno-labeling of LV tissue from WT rats without an AV fistula demonstrated that concentrated TNF-α expression was localized in cardiac mast cells, in addition to being diffusely distributed throughout the myocardium. While mast cells are clearly not the only source of TNF-α in the heart under normal conditions, inhibition of the marked increase in myocardial TNF-α in the Ws/Ws rats post-fistula suggests that mast cells are essential to the increased levels of TNF-α observed following imposition of volume overload. This could be due to either direct synthesis and release by mast cells, via their stimulation of other cells to produce TNF-α, or possibly MMP mediated cleavage of pro-TNF-α present in the heart. To this end, leukocytes and viable cardiomyocytes are major producers of TNF-α following microembolism [21
]. Our finding is consistent with recent ischemia-reperfusion studies showing that the initial release of TNF-α was cardiac mast cell-dependent, while subsequent induction was derived from other cell sources [22
]. TNF-α has been documented to exhibit multiple roles in the heart, which can be either pathological or cardioprotective [21
]. While it is theoretically possible that the basal levels of TNF-α present in the Ws/Ws hearts reflect the attenuated LV dilatation, several lines of evidence suggest that TNF-α induces activation of MMPs in the heart. A continuous 15 day infusion of TNF-α in normal rats led to a significant progressive increase in LV diameter associated with significant reductions in interstitial collagen as assessed by CVF [26
]. Further characterization at the ultrastructural level demonstrated that the fibrillar collagen weave surrounding cardiac myocytes was significantly disrupted or completely absent [26
]. This was thought by the authors to be the result of TNF-α mediated MMP activation. Our laboratory subsequently tested this supposition and found myocardial MMP-2 activity to be significantly elevated in normal rats during the first week of a TNF-α infusionrelative to that in saline-infused rats [27
]. As a result of this TNF-α mediated MMP activation, LV CVF was reduced. These findings together with the findings of the current study strongly suggest that mast cell-mediated increases in TNF-α play a significant role in the early stages of myocardial remodeling. This suggests the possibility that the failure of recent clinical trials examining the efficacy of TNF-α inhibition may be an issue of timing. Although TNF-α does not appear to be critical to worsening end-stage heart failure [28
], our findings are suggestive of TNF-α being important in the earlier stages of myocardial remodeling prior to the transition to a decompensated state. Thus, future consideration should be given to experimentation or trials focused on intervention directed against TNF-αin the early stages of heart failure (or even diastolic dysfunction). It should be remembered, however, that several other mast cell products, including tryptase and chymase, are also capable of activating MMPs [29
]. Furthermore, while not yet determined in cardiac mast cells, non-cardiac mast cells have been shown to produce various MMPs including MMP-1 [32
], -2 [33
], -9 [33
], and -13 [35
], as well as ADAM-9, -10 and -17 [33
]. It is possible that cardiac mast cells may also produce some or all of these MMPs. Therefore, although TNF-α appears to exert a prominent role in cardiac mast cell-induced myocardial remodeling, it is likely that the release of other substances, including tryptase, chymase or MMPs, from cardiac mast cells also contribute to collagen degradation in response to myocardial stress.
In summary, these results obtained in Ws/Ws rats definitively establish the central role of cardiac mast cells in initiating MMP activation, collagen degradation and subsequent myocardial dilatation and are suggestive of a role for mast cell-derived TNF-α in mediating the remodeling. These findings continue to expand the functional role of mast cells in pathologic conditions and emphasize the need to explore mast cell stabilizing compounds as a potential therapy for heart failure.