Cardiac hypertrophy and cardiac fibrosis is a pathophysiological driving force for heart failure (129, 150, 265, 287-289). In response to various stress stimuli myocytes grow either in
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length and/or width as a result of increased cardiac afterload (287, 445). Since, upregulation of the HO system by hemin suppressed inflammatory/oxidative markers, macrophage infiltration and potentiated insulin signaling, I examined whether the cytoprotective role of hemin can be extended to cardiac tissue in the form of improvement in cardiac morphological lesions and function.
In Chapter 2 and Chapter 3, I explored the effects of hemin treatment on cardiac structure and function. In ZF and ZDF rats, hemin treatment significantly reduced left-ventricular hypertrophy, cardiac fibrosis and cardiomyocyte longitudinal muscle fiber thickness and collagen deposition. However, co-administration of hemin and SnMP nullified the cytoprotective effects of hemin. Histological and morphometric analysis revealed that obesity-induced interstitial and perivascular collagen deposition, scarring of cardiomyocytes and cardiac fibrosis as observed in ZF rats was significantly attenuated by the hemin treatment (Chapter 2). As a parallel project, another novel and important finding of our study was that hemin therapy successfully reduced pericardial adipocyte hypertrophy as observed in untreated ZF rats and restored it to a level comparable to their ZL counterparts (54). The present research and previous studies support the existence of inter-organ crosstalk between the heart and pericardial adipose tissue (75, 78, 79). Thus, the movement of inflammatory/oxidative mediators and other atherogenic factors from the pericardial fat to the myocardium or vice versa in a paracrine manner might be suggestive of a mechanism for the subsequent development of insulin resistance and related cardiac complications (72, 129, 150, 260, 261, 265, 287-289). Altogether, the data from the present study and recent publication (54), strongly suggests that hemin-mediated suppression of pericardial adiposity and the parallel reduction of pro-inflammatory cytokines and chemokines may attenuate the adverse effects of inflammatory mediators on the
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myocardium derived from pericardial adipose tissue through a paracrine mechanism. Although, further research is needed to establish this interconnection, the results obtained with hemin treatment in the present study have given important suggestions for future work in this direction.
Myocardial remodeling is an important pathophysiological process characterized by abnormal deposition of interstitial collagen and over expression of extracellular matrix proteins such as TGF-β, fibronectin and collagen that leads to increased myocardial stiffness and reduced myocardial contractility (129, 248, 265, 281, 283, 286, 292). This suggests that attenuation of cardiac fibrosis in turn might improve cardiac function. Another important observation of my thesis work is that hemin treatment improved cardiac function through a mechanism that involves the reduction of ECM/profibrotic proteins including TGF-β, fibronectin and collagen IV. Collagen IV plays an important role in cardiac hypertrophy and fibrosis (21, 446), In addition, TGF-β mobilizes ECM through the induction of collagen and fibronectin, and contributes to cardiac injury and fibrosis (21). Thus, reduction in TGF-β expression will in turn suppresses fibronectin and collagen protein expression. Interestingly, my work also showed that hemin treatment in ZF rats reduced the expression of collagen IV and TGF-β by 2.8 fold and 3.4 fold compared to 6.9 fold and 4.6 fold above basal expression of collagen IV and TGF-β in untreated ZF rats. In a similar manner the 7.5 fold increase in basal expression of fibronectin in ZF rats was significantly suppressed 4.5 fold in hemin treated ZF rats (Chapter 2).
To further confirm the cardioprotective effects of an upregulated HO system by hemin, I assessed the effect of hemin on markers of heart failure such as osteopontin and osteoprotegerin.
Osteopontin and osteoprotegerin have been implicated to play pathophysiological roles in various biological processes such as adipose tissue inflammation, fibrosis, tissue remodeling, hypertrophy and insulin resistance (294, 295, 297-309, 311, 447). Osteopontin and
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osteoprotegerin are highly activated in response to various inflammatory molecules such as LPS, TNF-α, IFN-γ, TGF-β, angiotensin-II, IL-1β (295, 297, 299, 309). It is suggested that the elevated expression of osteopontin and osteoprotegerin in the circulation and in cardiac tissue is a predictor for impaired cardiac function and subsequent heart failure (295, 297, 298, 303, 309, 311). In Chapter 2, I have shown that in ZF rats, basal expression of osteopontin and osteoprotegerin in cardiac tissue was significantly elevated by 4.6 and 7.1 fold, respectively, as compared to ZL controls. However, hemin therapy in ZF rats reduced the expressions of osteopontin and osteoprotegerin by 3.5 and 3.3 fold, respectively. Similarly, our recent publication showed that hemin treatment significantly reduced the expression of osteopontin in pericardial adipose tissue of ZF rats (54).
The novel and central observation of my thesis work is that hemin treatment improved cardiac structure and function as evidenced by improvement in various hemodynamic and echocardiographic parameters in ZF rats and ZDF rats (Chapter 2 and Chapter 3). It should be noted that cardiomyocyte hypertrophy and myocardial fibrosis are microscopic changes which are observed at initial stages in heart failure (286, 292). Later, macroscopic changes such as increased left-ventricular wall thickness, altered cardiac hemodynamics, impaired diastolic function and subsequent systolic dysfunction becomes more prominent (287, 445). My thesis data showed that hemin treatment suppressed cardiac hypertrophy, fibrosis and left-ventricular longitudinal muscle fiber thickness (Chapter 2 and Chapter 3). Therefore, another possible mechanism that might be responsible for improved cardiac function by an upregulated HO system include improved hemodynamic and echocardiographic parameters that eventually leads to a healthier heart accompanied by improved ventricular contractility (448). Interestingly, my thesis data showed that hemin treatment significantly reduced left-ventricular diastolic wall
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thickness and left-ventricular systolic wall thickness and decreased mean-arterial pressure, arterial-diastolic pressure and arterial-systolic pressure in ZF rats and ZDF rats (Chapter 2 and Chapter 3). Furthermore, reduction in left-ventricular developed pressure, +dp/dt, and total peripheral resistance along with enhanced cardiac output was observed in hemin treated ZF rats (Chapter 2). Reduction in the pressure gradient that is required to maintain adequate systemic blood supply will reduce the left-ventricular workload and oxygen consumption and decrease the risk for cardiovascular abnormalities (428). Altogether, from these results, it can be inferred that the improvement in hemodynamic and echocardiographic parameters by hemin decreased left-ventricular afterload and thereby, protects against the onset of left-ventricular dysfunction that would affect cardiac performance and lead to heart failure (80).
Overall, my study clearly showed the beneficial effects of upregulated HO by hemin against obesity-induced altered cardiac structure and function. Suppression of cardiac hypertrophy, fibrosis and ECM/pro-fibrotic proteins are some of the mechanisms by which the HO system improved cardiac function in obese ZF rats.