Identification of β2-adrenoceptors on guinea pig alveolar macrophages using (-)-3-[125I]iodocyanopindolol

The β-adrenoceptor antagonist (-)-3-[125I]iodocyanopindolol ([125I]ICYP) binds with high affinity and in a saturable way to membranes of guinea pig alveolar macrophages. The equilibrium dissociation constant for [125I]ICYP is 24.3 ± 1.2 pM, and the number of binding sites is 166.3 ± 13.7 fmol/mg protein (N=4, ±SEM). Displacement studies with selective antagonists showed that [125I]ICYP labels β2-adrenoceptors on guinea pig alveolar macrophages

A comparative study between catalase gene therapy and the cardioprotector monohydroxyethylrutoside (MonoHER) in protecting against doxorubicin-induced cardiotoxicity in vitro

A comparative study between catalase gene therapy and the cardioprotector monohydroxyethylrutoside (MonoHER) in protecting against doxorubicin-induced cardiotoxicity in vitro. Abou-El-Hassan MA, Rabelink MJ, van der Vijgh WJ, Bast A, Hoeben RC. Department of Medical Oncology, Free University Medical Center, PO Box 7057, 1007 MB Amsterdam, The Netherlands. [email protected] Cardiotoxicity is the main dose-limiting side effect of doxorubicin in the clinic. Being a free radical producer, doxorubicin affects the heart specifically because of its low antioxidant capacity. Among those antioxidants, catalase is present in very low levels in the heart compared to other organs. Since catalase is an essential enzyme in detoxifying hydrogen peroxide, the aim of the present study was to investigate the protective effect of catalase as delivered by an adenovirus vector against doxorubicin-induced cardiotoxicity in cultured neonatal rat cardiac myocytes (NeRCaMs). 7-Monohydroxyethylrutoside (MonoHER), a potent cardioprotector currently under clinical investigations, was included in the study as a reference. Neonatal rat cardiac myocytes were infected with different multiplicity of infections (MOIs) of adenovirus encoding catalase (AdCat). A control infection with an adenovirus vector encoding a nonrelated protein was included. The activity and content of catalase in infected cells were determined during 3 days postinfection. One group of NeRCaMs was infected with AdCat before treatment with doxorubicin (0-50 microM). The second and third group were treated with doxorubicin (0-50 microM) with and without 1 mM monohydroxyethylrutoside (monoHER), respectively. The LDH release and viability of treated cells were measured 24 and 48 h after doxorubicin treatment. The beating rate was followed in three other groups of cells receiving the same treatments within 3 days after doxorubicin (0-100 microM) treatment. Catalase activity increased in AdCat-infected cells, with different MOIs, starting from the second day after infection as compared to the mock-infected cells (P50 microM doxorubicin. A 3.5-fold increase in the activity of catalase did not protect NeRCaMs against any of the cytotoxic effects of doxorubicin on NeRCaMs. In contrast, monoHER (1 mM) significantly protected NeRCaMs against the lethal effects of doxorubicin on the survival, LDH release and the beating rate of NeRCaMs (P72 h). The present study (1) illustrates that the cytotoxicity of high MOI of AdCat (>50) limited the possibility to increase catalase activity more than 3.5-fold, which was not enough to protect infected NeRCaMs against doxorubicin-induced cardiotoxicity and (2) confirms the efficacy of monoHER as a cardioprotector. Thus, the use of monoHER proves more suitable for the prevention of doxorubicin-induced cardiotoxicity than catalase gene transfer employing adenovirus vector

The protective effect of cardiac gene transfer of CuZn-sod in comparison with the cardioprotector monohydroxyethylrutoside against doxorubicin-induced cardiotoxicity in cultured cells

The protective effect of cardiac gene transfer of CuZn-sod in comparison with the cardioprotector monohydroxyethylrutoside against doxorubicin-induced cardiotoxicity in cultured cells. Abou El Hassan MA, Heijn M, Rabelink MJ, van der Vijgh WJ, Bast A, Hoeben RC. Department of Medical Oncology, Free University Medical Center, PO Box 7057, 1007 MB Amsterdam, The Netherlands. [email protected] Doxorubicin-induced cardiotoxicity is related to its production of free radicals that specifically affect heart tissue because of its low antioxidant status. Monohydroxyethylrutoside (monoHER), a potent antioxidant flavonoid, is under development as a protector against doxorubicin-induced cardiotoxicity. The overexpression of high levels of superoxide dismutase (sod) protects against free radical damage in transgenic mice. Seeking alternatives besides the few cardioprotectors that are presently under investigation, the aim of the present study was to investigate the protective effect of cardiac gene transfer of CuZn-sod compared with that of the presently most promising cardioprotector monoHER against doxorubicin-induced cardiotoxic effects on neonatal rat cardiac myocytes (NeRCaMs) in vitro. NeRCaMs were infected with different multiplicity of infections (MOIs) of adenovirus encoding CuZn-sod (AdCuZn-sod). A control infection with an adenovirus vector encoding a nonrelated protein was included. The overexpression of CuZn-sod was characterized within 3 days postinfection. For doxorubicin treatment, NeRCaMs were divided into three groups. The first group was infected with AdCuZn-sod before treatment with doxorubicin (0-50 microM). The second and third groups were treated with doxorubicin (0-50 microM) alone and with 1 mM monoHER, respectively. The LDH release and survival of treated cells were measured 24 and 48 hours after doxorubicin treatment. The beating rate was followed during the 3 days after doxorubicin (0-100 microM) treatment. At the third day after infection with an MOI of 25 plaque-forming unit (PFU) of AdCuZn-sod/cell, the activity of CuZn-sod significantly increased (five-fold, P=.029). Higher MOI produced cytopathic effects (CPEs). Doxorubicin alone produced significant concentration- and time-dependent reduction in NeRCaMs beating rate and survival (P or =50 microM)-treated cells ceased to beat after 24 hours. This cytotoxicity was associated with an increase in the LDH release from the treated cells (P 72 hours in the presence of monoHER.The present study showed the lack of adenoviral CuZn-sod gene-transfer to protect myocardiocytes against doxorubicin-induced toxicity and confirms the efficacy of monoHER cardioprotection. Thus, a gene-therapy strategy involving overexpression of CuZn-sod to protect against doxorubicin-induced cardiotoxicity is not feasible with the currently available adenovirus vectors