Hospital-Related Cardiac Morbidity among Survivors of Breast Cancer: Long-Term Risks and Predictors

BACKGROUND: Myocardial fibrosis is an important therapeutic target given its contribution to chamber dilatation, progression of heart failure and lethal cardiac arrhythmias. Myocardial fibrosis is mediated by persistent activation of resident (myo)fibroblasts that secrete extracellular matrix (ECM). Tetrandrine (TTD) is a calcium channel blocker (CCB) with documented anti-fibrotic actions believed to be secondary to its effects on lowering blood pressure. In this study, for the first time, we identified that TTD can directly reverse in vitro human cardiac fibroblast activation and limits in vivo myocardial fibrosis independent of hemodynamic load. METHODS AND RESULTS: IN VITRO: Human atrial biopsies were obtained from cardiac surgery patients (N1⁄410). Cardiac fibroblasts were isolated, expanded in culture, and seeded in 3D collagen matrices. Cell-collagen constructs were exposed to TGFb1 (10 ng/mL), with or without TTD (1 or 5mM) for 48 hours. Extent of collagen gel contraction, myofibroblast activation (a-SMAexpression), expression of pro-fibroticmRNAs and rate of collagen protein synthesis was compared. TTD exposure significantly decreased TGF-b1 induced human cardiac myofibroblast mediated collagen gel contraction (79.66 1.28 control; 67.05 3.91TTD1mM;60.05 8.85TTD5mM;both P< 0.01). Cell viability was similar between groups (annexin positive cells: control 1.7%, TTD (1mM) 1.1% and TTD(5mM) 1.4%). TTD significantly attenuated myofibroblast activation as assessed by aSMA expression using flow cytometer. TTD decreased collagen synthesis ([3H]-proline incorporation 171 (control); 119 (TTD1uM); and 43 (TTD5mM) DPM/mg protein) and expression ofa-SMAand collagenmRNAs (ACTA2;Col1A1;Col1A2). TTD inhibited collagen gel contraction in the presence of T-type and L-type CCB (Mibefradil (5mM) or verapamil (20mM)), and a calcium chelator BAPTA-AM (15mM) suggesting that the observed effects are notmediated by calcium channel blockade. IN VIVO: Dahl salt-sensitive rat model of pressure-overload was employed. Animals were divided into untreated controls and compared to TTD treatment groups (7.5mg; 15mg; 30mg/kg by IP injection over 4weeks). Systemic blood pressure wasmonitored by tail cuff. Left ventricular compliance and myocardial fibrosis was assessed by passive pressure-volume analysis, echocardiography and PSR staining. Myocardial fibrosis was attenuated and LV compliance preserved after 4weeks of TTD30mg/kg treatment as compared to untreated controls (% collagen area: 4.21 0.56 versus 2.13 0.31, P<0.01). TTD-induced changes in fibrosis and chamber compliance independent of hemodynamic load. CONCLUSION: TTD reverses human cardiac myofibroblast activation and myocardial fibrosis independent of calcium channel blockade and hemodynamic load. The mechanisms that mediate the anti-fibrotic activity of TTD should be further explored and leveraged for its therapeutic potential. 134 HOSPITAL-RELATED CARDIAC MORBIDITY AMONG SURVIVORS OF BREAST CANCER: LONG-TERM RISKS AND PREDICTORS