Cardiac remodeling and MMPs on the model of chronic daunorubicin-induced cardiomyopathy in rabbits.

The matrix metalloproteinases (MMPs) play a key role during cardiac remodeling. The aim of the study was to investigate the changes in collagenous proteins and MMPs in the model of non-ischemic, anthracycline-induced chronic cardiomyopathy in rabbits using both biochemical and histological approaches. The study was carried out in three groups of Chinchilla male rabbits: 1) daunorubicin (3 mg/kg, once weekly for 10 weeks), 2) control (saline in the same schedule), 3) daunorubicin with the cardioprotectant dexrazoxane (60 mg/kg, before each daunorubicin). Morphological changes in the myocardium of daunorubicin-treated animals were characterized by focal myocardial interstitial fibrosis of different intensity. The subsequent proliferation of the fibrotic tissue was marked by an increased content of both collagen types I and III, which resulted in their typical coexpression in the majority of bundles of fibers forming either smaller or larger scars. Biochemical analysis showed a significantly increased concentration of hydroxyproline, mainly in the pepsin-insoluble fraction of collagenous proteins, in the daunorubicin-treated group (1.42+/-0.12 mg/g) as compared with the control (1.03+/-0.04 mg/g) and dexrazoxane (1.07+/-0.07 mg/g) groups. Dexrazoxane co-administration remarkably reduced the cardiotoxic effects of daunorubicin to the extent comparable with the controls in all evaluated parameters. Using zymography, it was possible to detect only a gelatinolytic band corresponding to MMP-2 (MMP-9 activity was not detectable). However, no significant changes in MMP-2 activity were determined between individual groups. Immunohistochemical analysis revealed increased MMP-2 expression in both cardiomyocytes and fibroblasts. Thus, this study has revealed specific alterations in the collagen network in chronic anthracycline cardiotoxicity in relationship to the expression and activity of major MMPs.

Dexrazoxane-afforded protection against chronic anthracycline cardiotoxicity in vivo: effective rescue of cardiomyocytes from apoptotic cell death.

BACKGROUND: Dexrazoxane (DEX, ICRF-187) is the only clinically approved cardioprotectant against anthracycline cardiotoxicity. It has been traditionally postulated to undergo hydrolysis to iron-chelating agent ADR-925 and to prevent anthracycline-induced oxidative stress, progressive cardiomyocyte degeneration and subsequent non-programmed cell death. However, the additional capability of DEX to protect cardiomyocytes from apoptosis has remained unsubstantiated under clinically relevant in vivo conditions. METHODS: Chronic anthracycline cardiotoxicity was induced in rabbits by repeated daunorubicin (DAU) administrations (3 mg kg(-1) weekly for 10 weeks). Cardiomyocyte apoptosis was evaluated using TUNEL (terminal deoxynucleotidyl transferase biotin-dUTP nick end labelling) assay and activities of caspases 3/7, 8, 9 and 12. Lipoperoxidation was assayed using HPLC determination of myocardial malondialdehyde and 4-hydroxynonenal immunodetection. RESULTS: Dexrazoxane (60 mg kg(-1)) co-treatment was capable of overcoming DAU-induced mortality, left ventricular dysfunction, profound structural damage of the myocardium and release of cardiac troponin T and I to circulation. Moreover, for the first time, it has been shown that DEX affords significant and nearly complete cardioprotection against anthracycline-induced apoptosis in vivo and effectively suppresses the complex apoptotic signalling triggered by DAU. In individual animals, the severity of apoptotic parameters significantly correlated with cardiac function. However, this effective cardioprotection occurred without a significant decrease in anthracycline-induced lipoperoxidation. CONCLUSION: This study identifies inhibition of apoptosis as an important target for effective cardioprotection against chronic anthracycline cardiotoxicity and suggests that lipoperoxidation-independent mechanisms are involved in the cardioprotective action of DEX.

Anthracycline toxicity to cardiomyocytes or cancer cells is differently affected by iron chelation with salicylaldehyde isonicotinoyl hydrazone.

BACKGROUND AND PURPOSE: The clinical utility of anthracycline antineoplastic drugs is limited by the risk of cardiotoxicity, which has been traditionally attributed to iron-mediated production of reactive oxygen species (ROS). EXPERIMENTAL APPROACH: The aims of this study were to examine the strongly lipophilic iron chelator, salicylaldehyde isonicotinoyl hydrazone (SIH), for its ability to protect rat isolated cardiomyocytes against the toxicity of daunorubicin (DAU) and to investigate the effects of SIH on DAU-induced inhibition of proliferation in a leukaemic cell line. Cell toxicity was measured by release of lactate dehydrogenase and staining with Hoechst 33342 or propidium iodide and lipid peroxidation by malonaldehyde formation. KEY RESULTS: SIH fully protected cardiomyocytes against model oxidative injury induced by hydrogen peroxide exposure. SIH also significantly but only partially and with no apparent dose-dependency, reduced DAU-induced cardiomyocyte death. However, the observed protection was not accompanied by decreased lipid peroxidation. In the HL-60 acute promyelocytic leukaemia cell line, SIH did not blunt the antiproliferative efficacy of DAU. Instead, at concentrations that reduced DAU toxicity to cardiomyocytes, SIH enhanced the tumoricidal action of DAU. CONCLUSIONS AND IMPLICATIONS: This study demonstrates that iron is most likely involved in anthracycline cardiotoxicity and that iron chelation has protective potential, but apparently through mechanism(s) other than by inhibition of ROS-induced injury. In addition to cardioprotection, iron chelation may have considerable potential to improve the therapeutic action of anthracyclines by enhancing their anticancer efficiency and this potential warrants further investigation.

Evaluation of ECG time intervals in a rabbit model of anthracycline-induced cardiomyopathy: a useful tool for assessment of cardioprotective agents.

The aim of this study was to analyze the ECG time intervals in the course of the development of chronic anthracycline cardiomyopathy in rabbits. Furthermore, this approach was employed to study the effects of a model cardioprotective drug (dexrazoxane) and two novel iron chelating compounds--salicylaldehyde isonicotinoyl hydrazone (SIH) and pyridoxal 2-chlorobenzoyl hydrazone (o-108). Repeated daunorubicin administration induced a significant and progressive prolongation of the QRS complex commencing with the eighth week of administration. At the end of the study, we identified a significant correlation between QRS duration and the contractility index dP/dt(max) (r = -0.81; P<0.001) as well as with the plasma concentrations of cardiac troponin T (r = 0.78; P<0.001). In contrast, no alterations in ECG time intervals were revealed in the groups co-treated with either dexrazoxane or both novel cardioprotective drugs (SIH, o-108). Hence, in this study, the QRS duration is for the first time shown as a parameter suitable for the non-invasive evaluation of the anthracycline cardiotoxicity and cardioprotective effects of both well established and investigated drugs. Moreover, our results strongly suggest that novel iron chelators (SIH and o-108) merit further study as promising cardioprotective drugs against anthracycline cardiotoxicity.

Early detection of anthracycline cardiotoxicity in a rabbit model: left ventricle filling pattern versus troponin T determination.

Anthracycline cardiotoxicity represents a serious risk of anticancer chemotherapy. The aim of the present pilot study was to compare the potential of both the left ventricular (LV) filling pattern evaluation and cardiac troponin T (cTnT) plasma levels determination for the early detection of daunorubicin-induced cardiotoxicity in rabbits. The echocardiographic measurements of transmitral LV inflow as well as cTnT determinations were performed weekly for 10 weeks in daunorubicin (3 mg/kg weekly) and control groups (n=5, each). Surprisingly, no significant changes in LV-filling pattern were observed through the study, most likely due to the xylazine-containing anesthesia, necessary for appropriate resolving of the E and A waves. In contrast to the echographic measurement, the dP/dt(min) index obtained invasively at the end of the study revealed a significant impairment in LV relaxation, which was further supported by observed disturbances in myocardial collagen content and calcium homeostasis. However, at the same time cTnT plasma levels were progressively rising in the daunorubicin-treated animals from the fifth week (0.024+/-0.008 microg/l) until the end of the experiment (0.186+/-0.055 microg/l). Therefore, in contrast to complicated non-invasive evaluation of diastolic function, cTnT is shown to be an early and sensitive marker of anthracycline-induced cardiotoxicity in the rabbit model.

Safety and tolerability of repeated administration of pyridoxal 2-chlorobenzoyl hydrazone in rabbits.

Recently, pyridoxal 2-chlorobenzoyl hydrazone (o-108) has been identified as an effective iron chelator [Link et al., Blood 2003; 101: 4172-79]. Since chronic treatment would be necessary in its potential indications, in the present study, the safety and tolerability of this agent after repeated administration was determined. Three doses of o-108 (25, 50, 100 mg/kg, in 10% Cremophor EL) were administered intraperitoneally, once weekly, for 10 weeks to three groups (n=5 each) of Chinchilla male rabbits. The effects on biochemical, haematological and cardiovascular parameters were examined during the experiment; histopathological examination was performed at the end of the experiment. Results were compared with control (saline 2 mL/kg, n=11) and vehicle groups (10% Cremophor EL, 2 mL/kg, n=12). No premature deaths occurred; the well-being of animals was evidenced by their body weight gain, although lower gain was observed with the highest dose (100 mg/kg). Significant elevations of cardiac troponin T plasma concentrations were observed with the highest dose of o-108, but no abnormalities were found in the cardiovascular function and only minor and inconsistent changes in haematological and biochemical parameters were observed. Histopathological examinations of selected organs revealed only weak and reversible changes through all studied groups. Thus, the data from this study suggest that o-108 remains a promising drug from the standpoint of the possibility of its repeated administration and warrants further investigation.