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Fibroblast growth factor-2 protects neonatal rat cardiac myocytes from doxorubicin-induced damage via protein kinase C- dependent effects on efflux drug transporters

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dc.contributor.supervisor Cattini, Peter A. (Physiology) en_US
dc.contributor.author Wang, Jie
dc.date.accessioned 2013-04-02T17:05:24Z
dc.date.available 2013-04-02T17:05:24Z
dc.date.issued 2013-01-22 en_US
dc.identifier.citation Cardiovasc Res. 2013 Apr 1;98(1):56-63. en_US
dc.identifier.uri http://hdl.handle.net/1993/18318
dc.description.abstract Introduction: Therapeutic agents like doxorubicin, an anthracycline antibiotic drug, are widely used in cancer chemotherapy. The use of doxorubicin is limited however by an increased risk of cardiac damage as a side effect, and an increased cancer cell drug resistance mediated by efflux drug transporters. Strategies are needed to protect the heart and still allow the benefits of drug treatment. “Basic” fibroblast growth factor-2 (FGF-2) is a multi-functional protein. It is angiogenic and cardioprotective against ischemia-reperfusion injury. FGF-2 can also regulate cancer cell drug resistance or sensitivity, however, so far, there is no evidence that FGF-2 protects against doxorubicin-induced cardiac damage through effects on efflux drug transporter levels or function. Aims: To investigate whether: (1) FGF-2 can increase resistance to doxorubicin-induced neonatal rat cardiac myocyte damage; and if so whether (2) an effect on efflux drug transporters might contribute to this cardioprotection by FGF-2. Methods: Neonatal rat cardiac myocyte cultures were treated with doxorubicin in the absence or presence of pre-treatment with FGF-2. To assess cell damage: (i) culture medium was tested for lactate dehydrogenase (LDH) activity as an indication of plasma membrane disruption; (ii) cells were stained with fluorescent apoptosis and necrosis biomarkers as well as (iii) terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and acridine orange to assess DNA fragmentation or compaction. The role of FGF receptor (FGFR) or protein kinase C (PKC) was addressed through use of inhibitors including SU5402, or chelerythrine as well as bisindomaleimide. Multidrug resistance gene 1a and 1b (MDR1a, 1b), multidrug resistance gene 2 (MDR2) and multidrug resistance-related protein 1 (MRP1) gene expression, as well as the function of MDRs and MRPs protein products were assessed by real-time reverse transcriptase-polymerase chain reaction (qPCR), as well as retention/extrusion of (fluorescent) doxorubicin/calcein in cardiac myocytes, respectively. Efflux drug transporter inhibitors, including 20 µM cyclosporine A (CsA), 2 µM verapamil and 1 µM Tariquidar (XR9576) were used to asssess for a direct effect of FGF-2 on transporter function. Fluorescence-activated cell sorting (FACS) was used to measure fluorescent doxorubicin/calcein levels inside treated cardiac myocytes. Results: Doxorubicin increased the incidence of programmed cell death, DNA damage, and lysosome and LDH activity, while decreasing cell number at 24 hours. FGF-2 prevented the detrimental effects of doxorubicin. In turn, the protective effects of FGF-2 were blocked in the presence of FGFR or PKC inhibitors. FGF-2 treatment significantly increased MDR1a, MDR1b, MDR2, MRP1 RNA levels by qPCR, and protein levels as assessed by function, and specifically extrusion of doxorubicin/calcein, in the presence of doxorubicin when compared to doxorubicin treatment alone. Furthermore, inhibition of efflux drug transporters with CsA and Tariquidar (XR9576) significantly reduced the ability of FGF-2 to protect against doxorubicin-induced damage; the beneficial effect of FGF-2 was completely blocked by pretreatment with verapamil. Conclusion(s): These data indicate for the first time that exogenous FGF-2 can increase resistance to doxorubicin-induced neonatal rat cardiac myocyte damage, and implicate PKC and regulation of efflux transporter protein levels and/or function in the mechanism. en_US
dc.publisher Cardiovascular Research en_US
dc.subject FGF-2 en_US
dc.subject Doxorubicin en_US
dc.subject efflux drug transporters en_US
dc.subject PKC en_US
dc.subject neonatal rat cardiomyocytes en_US
dc.title Fibroblast growth factor-2 protects neonatal rat cardiac myocytes from doxorubicin-induced damage via protein kinase C- dependent effects on efflux drug transporters en_US
dc.degree.discipline Physiology en_US
dc.contributor.examiningcommittee Kardami, Elissavet (Physiology) Czubryt, Michael (Physiology) Nachtigal, Mark (Biochemistry and Medical Genetics) en_US
dc.degree.level Master of Science (M.Sc.) en_US
dc.description.note May 2013 en_US


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