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1.
Life Sci ; 283: 119849, 2021 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-34343539

RESUMO

AIMS: Cardiotoxicity of doxorubicin frequently complicates treatment outcome. Aberrantly activated calcium/calmodulin pathway can eventually trigger signaling cascades that mediate cardiotoxicity. Therefore, we tested the hypothesis that trifluoperazine, a strong calmodulin antagonist, may alleviate this morbidity. MATERIALS AND METHODS: Heart failure and cardiotoxicity were assessed via echocardiography, PCR, immunohistochemistry, histopathology, Masson's trichrome staining and transmission electron microscopy. Whereas liver and kidney structural and functional alterations were evaluated histopathologically and biochemically. KEY FINDINGS: Results revealed that combination treatment with trifluoperazine could overcome doxorubicin-induced heart failure with reduced ejection fraction. Moreover, heart weight/body weight ratio and histopathological examination showed that trifluoperazine mitigated doxorubicin-induced cardiac atrophy, inflammation and myofibril degeneration. Transmission electron microscopy further confirmed the marked restoration of the left ventricular ultrastructures by trifluoperazine pretreatment. In addition, Masson's trichrome staining revealed that trifluoperazine could significantly inhibit doxorubicin-induced left ventricular remodeling by fibrosis. Of note, doxorubicin induced the expression of myocardial nuclear NF-κB-p65 and caspase-3 which were markedly inhibited by trifluoperazine, suggesting that cardioprotection conferred by trifluoperazine involved, at least in part, suppression of NF-κB and apoptosis. Furthermore, biochemical and histopathological examinations showed that trifluoperazine improved doxorubicin-induced renal and hepatic impairments both functionally and structurally. SIGNIFICANCE: In conclusion, the present in vivo study is the first to provide evidences underscoring the protective effects of trifluoperazine that may pave the way for repurposing this calmodulin antagonist in ameliorating organ toxicity by doxorubicin.


Assuntos
Apoptose/efeitos dos fármacos , Cardiotoxicidade , Cardiotoxinas/efeitos adversos , Doxorrubicina/efeitos adversos , Miocárdio/metabolismo , Fator de Transcrição RelA/metabolismo , Trifluoperazina/farmacologia , Animais , Cardiotoxicidade/tratamento farmacológico , Cardiotoxicidade/metabolismo , Cardiotoxicidade/patologia , Cardiotoxinas/farmacologia , Caspase 3/metabolismo , Doxorrubicina/farmacologia , Masculino , Camundongos , Miocárdio/patologia
2.
Rev. urug. cardiol ; 36(1): e36107, abr. 2021. ilus, tab
Artigo em Espanhol | LILACS, BNUY, UY-BNMED | ID: biblio-1252372

RESUMO

Las nuevas terapias oncológicas han logrado aumentar la sobrevida del paciente con cáncer, observando, sin embargo, un incremento de la morbilidad y mortalidad vinculadas a sus efectos secundarios. El desarrollo de eventos cardiovasculares adversos impacta negativamente en el pronóstico durante el tratamiento del cáncer, pero también en los supervivientes al cáncer, donde las enfermedades cardiovasculares (ECV) y las segundas neoplasias son la principal causa de muerte1-5. La cardiotoxicidad inducida por el tratamiento del cáncer se define como el conjunto de ECV derivadas de los tratamientos oncológicos. Su manifestación es variada e incluye el desarrollo de disfunción ventricular, insuficiencia cardíaca (IC), isquemia miocárdica, hipertensión arterial y arritmias, entre otras. Puede ser consecuencia tanto del efecto directo del tratamiento sobre la estructura y función cardíacas, como del desarrollo acelerado de ECV6-9. Frecuentemente se utiliza el término cardiotoxicidad como sinónimo de disfunción ventricular por quimioterapia (DV-QT). Dado que la cardiotoxicidad abarca un espectro más amplio de afectación cardiovascular, creemos conveniente hablar de DV-QT para referirnos a la afectación de la función sistólica del ventrículo izquierdo. La DV-QT y el desarrollo de IC representan una de las complicaciones más temidas por su impacto pronóstico en la esfera cardiovascular y oncológica, dado que limitan el arsenal terapéutico para el tratamiento del cáncer5,10. Han sido creadas diversas sociedades de cardio-onco-hematología con el fin de generar recomendaciones de práctica clínica y formar profesionales capacitados para el manejo de las complicaciones cardiovasculares del tratamiento del cáncer11. La cardio-oncología es una disciplina en creciente y continuo desarrollo. Creemos que es fundamental realizar tareas de formación médica continua, así como también estimular el trabajo conjunto de diversas especialidades para brindar una mejor asistencia. Este texto es el resultado del trabajo de un equipo multidisciplinario que incluye cardiólogos, hematólogos y oncólogos, y pretende brindar información a los integrantes del equipo de salud involucrados en la asistencia de pacientes oncológicos. Debido a su extensión, hemos decidido fraccionar el contenido en tres partes para facilitar su publicación.


New oncological therapies have been successful in increasing cancer patient survival, but they have also led to an increase in morbidity and mortality linked to their side effects. During cancer treatment, the development of cardiovascular side effects has a negative impact in prognosis, but also in cancer survivors, in whom cardiovascular diseases and secondary malignancies are the main cause of death. Cancer related cardiotoxicity is defined as the development of cardiovascular diseases related to cancer treatment. Clinical presentation is broad involving ventricular dysfunction, heart failure, myocardial ischemia, arterial hypertension and arrhythmias among others. This may result from the direct cardiovascular effect of a cancer treatment or accelerated development of cardiovascular diseases. Frequently, in the literature cardiotoxicity and chemotherapy related ventricular dysfunction are used as synonyms. However, cardiotoxicity includes a broad spectrum of cardiovascular manifestations, thus in this text we refer to chemotherapy related ventricular dysfunction as the presence of left ventricular systolic impairment. Chemotherapy related ventricular dysfunction and heart failure are two of the most feared complications of cancer treatment due to its impact on cardiovascular and oncological prognosis, affecting treatment options. Numerous worldwide cardio-onco-hematology societies have emerged to generate clinical practice guidelines and improve the diagnosis and evaluation of cardiovascular cancer treatment side effects. Cardio-Oncology is a discipline in continuous growth and development. We strongly believe that continuum medical education and a multidisciplinary approach is necessary to provide a quality health care. This text is the result of a multidisciplinary work involving cardiologists, hematologists and oncologists. It is our goal to provide information to the health care team involved in the assistance of cancer patients. Due to its extension, it will be published in three parts.


O desenvolvimento de novas terapias oncológicas levou a um aumento na sobrevida dos pacientes, mas ao mesmo tempo traz consigo morbidades relacionadas aos tratamentos. O desenvolvimento de efeitos cardiovasculares adversos tem um impacto negativo no prognóstico dos pacientes em tratamento, bem como nos pacientes considerados curados, nos quais doença cardiovascular e malignidades secundárias são as principais causas de morte. Cardiotoxicidade relacionada ao câncer é definida como o desenvolvimento de doença cardiovascular secundária ao tratamento. A gama de apresentações clínicas é ampla, podendo se manifestar como disfunção ventricular, insuficiência cardíaca, isquemia miocárdica, hipertensão arterial, arritmias, entre outras. Isto pode ser resultante de desenvolvimento e progressão acelerados de doença cardiovascular ou por efeito direto das terapias. Frequentemente é dito na literatura que cardiotoxicidade e disfunção ventricular relacionada à quimioterapia são sinônimos. Entretanto, cardiotoxicidade engloba um amplo espectro de manifestações cardiovasculares. Neste texto, portanto, nos referimos à disfunção ventricular causada por quimioterápicos exclusivamente como a presença de disfunção sistólica ventricular esquerda. Disfunção ventricular relacionada à quimioterapia e insuficiência cardíaca são duas das mais temidas complicações do tratamento oncológico devido ao seu impacto no prognóstico cardiovascular e oncológico, podendo afetar ainda a escolha e manutenção das opções terapêuticas. Diversas sociedades cardio-onco-hematológicas surgiram ao redor do mundo com o objetivo de gerar diretriz clínicas práticas e melhorar o diagnóstico e tratamento das complicações cardiovasculares resultantes das terapias oncológicas. A cardio-oncologia é uma disciplina em contínuo crescimento e desenvolvimento. Nós acreditamos fortemente que educação médica continuada e uma abordagem multidisciplinar são necessárias para um cuidado médico de qualidade. Este texto é o resultado de um trabalho multidisciplinar envolvendo cardiologistas, hematologistas e oncologistas. Nosso objetivo é de oferecer informação à equipe de cuidados em saúde envolvido na assistência destes pacientes. Devido à sua extensão, este texto será publicado em três partes.


Assuntos
Humanos , Disfunção Ventricular/induzido quimicamente , Disfunção Ventricular/prevenção & controle , Disfunção Ventricular/diagnóstico por imagem , Cardiotoxinas/efeitos adversos , Cardiotoxinas/farmacologia , Antineoplásicos/efeitos adversos , Biomarcadores , Medição de Risco , Assistência ao Paciente/normas , Insuficiência Cardíaca/induzido quimicamente
3.
J Vis Exp ; (167)2021 01 23.
Artigo em Inglês | MEDLINE | ID: mdl-33554972

RESUMO

Despite several advances in cardiac tissue engineering, one of the major challenges to overcome remains the generation of a fully functional vascular network comprising several levels of complexity to provide oxygen and nutrients within bioengineered heart tissues. Our laboratory has developed a three-dimensional in vitro model of the human heart, known as the "cardiac spheroid" or "CS". This presents biochemical, physiological, and pharmacological features typical of the human heart and is generated by co-culturing its three major cell types, such as cardiac myocytes, endothelial cells, and fibroblasts. Human induced pluripotent stem cells-derived cardiomyocytes (hiPSC-CMs or iCMs) are co-cultured at ratios approximating the ones found in vivo with human cardiac fibroblasts (HCFs) and human coronary artery endothelial cells (HCAECs) in hanging drop culture plates for three to four days. The confocal analysis of CSs stained with antibodies against cardiac Troponin T, CD31 and vimentin (markers for cardiac myocytes, endothelial cells and fibroblasts, respectively) shows that CSs present a complex endothelial cell network, resembling the native one found in the human heart. This is confirmed by the 3D rendering analysis of these confocal images. CSs also present extracellular matrix (ECM) proteins typical of the human heart, such as collagen type IV, laminin and fibronectin. Finally, CSs present a contractile activity measured as syncytial contractility closer to the one typical of the human heart compared to CSs that contain iCMs only. When treated with a cardiotoxic anti-cancer agent, such as doxorubicin (DOX, used to treat leukemia, lymphoma and breast cancer), the viability of DOX-treated CSs is significantly reduced at 10 µM genetic and chemical inhibition of endothelial nitric oxide synthase, a downstream target of DOX in HCFs and HCAECs, reduced its toxicity in CSs. Given these unique features, CSs are currently used as in vitro models to study heart biochemistry, pathophysiology, and pharmacology.


Assuntos
Bioengenharia/métodos , Coração/fisiopatologia , Esferoides Celulares/citologia , Animais , Cardiotoxinas/farmacologia , Contagem de Células , Separação Celular , Sobrevivência Celular/efeitos dos fármacos , Técnicas de Cocultura , Colágeno/farmacologia , Doxorrubicina/farmacologia , Doxorrubicina/toxicidade , Células Endoteliais/citologia , Células Endoteliais/efeitos dos fármacos , Fibroblastos/citologia , Géis , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Miócitos Cardíacos/citologia , Miócitos Cardíacos/efeitos dos fármacos , Ratos , Fixação de Tecidos
4.
Theranostics ; 11(6): 2876-2891, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33456578

RESUMO

Rationale: Structural remodeling or damage as a result of disease or injury is often not evenly distributed throughout a tissue but strongly depends on localization and extent of damaging stimuli. Skeletal muscle as a mechanically active organ can express signs of local or even systemic myopathic damage, necrosis, or repair. Conventionally, muscle biopsies (patients) or whole muscles (animal models) are mechanically sliced and stained to assess structural alterations histologically. Three-dimensional tissue information can be obtained by applying deep imaging modalities, e.g. multiphoton or light-sheet microscopy. Chemical clearing approaches reduce scattering, e.g. through matching refractive tissue indices, to overcome optical penetration depth limits in thick tissues. Methods: Here, we optimized a range of different clearing protocols. We find aqueous solution-based protocols employing (20-80%) 2,2'-thiodiethanol (TDE) to be advantageous over organic solvents (dibenzyl ether, cinnamate) regarding the preservation of muscle morphology, ease-of-use, hazard level, and costs. Results: Applying TDE clearing to a mouse model of local cardiotoxin (CTX)-induced muscle necrosis, a complete loss of myosin-II signals was observed in necrotic areas with little change in fibrous collagen or autofluorescence (AF) signals. The 3D aspect of myofiber integrity could be assessed, and muscle necrosis in whole muscle was quantified locally via the ratios of detected AF, forward- and backward-scattered Second Harmonic Generation (fSHG, bSHG) signals. Conclusion: TDE optical clearing is a versatile tool to study muscle architecture in conjunction with label-free multiphoton imaging in 3D in injury/myopathy models and might also be useful in studying larger biofabricated constructs in regenerative medicine.


Assuntos
Microscopia Confocal/métodos , Microscopia de Fluorescência por Excitação Multifotônica/métodos , Músculo Esquelético/metabolismo , Necrose/diagnóstico , Animais , Cardiotoxinas/farmacologia , Colágeno/metabolismo , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/metabolismo , Imageamento Tridimensional/métodos , Camundongos , Camundongos Endogâmicos C57BL , Músculo Esquelético/efeitos dos fármacos , Miofibrilas/metabolismo , Miosina Tipo II/metabolismo , Necrose/induzido quimicamente , Necrose/metabolismo , Compostos de Sulfidrila/farmacologia
5.
Aging Cell ; 20(2): e13312, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33511781

RESUMO

Recruited immune cells play a critical role in muscle repair, in part by interacting with local stem cell populations to regulate muscle regeneration. How aging affects their communication during myogenesis is unclear. Here, we investigate how aging impacts the cellular function of these two cell types after muscle injury during normal aging or after immune rejuvenation using a young to old (Y-O) or old to old (O-O) bone marrow (BM) transplant model. We found that skeletal muscle from old mice (20 months) exhibited elevated basal inflammation and possessed fewer satellite cells compared with young mice (3 months). After cardiotoxin muscle injury (CTX), old mice exhibited a blunted inflammatory response compared with young mice and enhanced M2 macrophage recruitment and IL-10 expression. Temporal immune and cytokine responses of old mice were partially restored to a young phenotype following reconstitution with young cells (Y-O chimeras). Improved immune responses in Y-O chimeras were associated with greater satellite cell proliferation compared with O-O chimeras. To identify how immune cell aging affects myoblast function, conditioned media (CM) from activated young or old macrophages was applied to cultured C2C12 myoblasts. CM from young macrophages inhibited myogenesis while CM from old macrophages reduced proliferation. These functional differences coincided with age-related differences in macrophage cytokine expression. Together, this study examines the infiltration and proliferation of immune cells and satellite cells after injury in the context of aging and, using BM chimeras, demonstrates that young immune cells retain cell autonomy in an old host to increase satellite cell proliferation.


Assuntos
Senescência Celular/imunologia , Desenvolvimento Muscular/imunologia , Células Satélites de Músculo Esquelético/imunologia , Animais , Cardiotoxinas/farmacologia , Senescência Celular/efeitos dos fármacos , Camundongos , Desenvolvimento Muscular/efeitos dos fármacos , Células Satélites de Músculo Esquelético/efeitos dos fármacos
6.
Oxid Med Cell Longev ; 2020: 8819771, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33274007

RESUMO

The toxicity of doxorubicin (DOX) limits its clinical application. Nevertheless, at present, there is no effective drug to prevent DOX-induced cardiac injury. miR-204 is a newly discovered miRNA with many protective effects on cardiovascular diseases. However, little research has been done on the effects of miR-204 on DOX-induced cardiac injury. Our study is aimed at investigating the effect of miR-204 on DOX-induced myocardial injury. An adenoassociated virus system was used to achieve cardiac-specific overexpression of miR-204. Two weeks later, the mice were intraperitoneally injected with DOX (15 mg/kg) to induce cardiac injury. H9c2 myocardial cells were used to validate the role of miR-204 in vitro. Our study showed that miR-204 expression was decreased in DOX-treated hearts. miR-204 overexpression improved cardiac function and alleviated cardiac inflammation, apoptosis, and autophagy induced by DOX. In addition, our results showed that miR-204 prevented DOX-induced injury in cardiomyocytes by directly decreasing HMGB1 expression. Moreover, the overexpression of HMGB1 could offset the protective effects of miR-204 against DOX-induced cardiac injury. In summary, our study showed that miR-204 protected against DOX-induced cardiac injury via the inhibition of HMGB1, and increasing miR-204 expression may be a new treatment option for patients with DOX-induced cardiac injury.


Assuntos
Apoptose/efeitos dos fármacos , Cardiotoxinas/efeitos adversos , Doxorrubicina/efeitos adversos , Proteína HMGB1/metabolismo , MicroRNAs/metabolismo , Miócitos Cardíacos/metabolismo , Animais , Apoptose/genética , Cardiotoxinas/farmacologia , Doxorrubicina/farmacologia , Proteína HMGB1/genética , Inflamação/induzido quimicamente , Inflamação/genética , Inflamação/metabolismo , Inflamação/prevenção & controle , Masculino , Camundongos , MicroRNAs/genética , Miócitos Cardíacos/patologia
7.
Toxins (Basel) ; 12(4)2020 04 18.
Artigo em Inglês | MEDLINE | ID: mdl-32325789

RESUMO

It is widely accepted that snake venom cardiotoxins (CTXs) target the plasma membranes of cells. In the present study, we investigated the role of Asp residues in the interaction of Naja atra cardiotoxin 1 (CTX1) and cardiotoxin 3 (CTX3) with phospholipid bilayers using chemical modification. CTX1 contains three Asp residues at positions 29, 40, and 57; CTX3 contains two Asp residues at positions 40 and 57. Compared to Asp29 and Asp40, Asp57 was sparingly modified with semi-carbazide, as revealed by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass and mass/mass analyses. Thus, semi-carbazide-modified CTX1 (SEM-CTX1) mainly contained modified Asp29 and Asp40, while SEM-CTX3 contained modified Asp40. Compared to that of native toxins, trifluoroethanol easily induced structural transition of SEM-CTX1 and SEM-CTX3, suggesting that the structural flexibility of CTXs was constrained by Asp40. Modification of Asp29 and Asp40 markedly promoted the ability of CTX1 to induce permeability of cell membranes and lipid vesicles; CTX3 and SEM-CTX3 showed similar membrane-damaging activity. Modification of Asp residues did not affect the membrane-binding capability of CTXs. Circular dichroism spectra of SEM-CTX3 and CTX3 were similar, while the gross conformation of SEM-CTX1 was distinct from that of CTX1. The interaction of CTX1 with membrane was distinctly changed by Asp modification. Collectively, our data suggest that Asp29 of CTX1 suppresses the optimization of membrane-bound conformation to a fully active state and that the function of Asp40 in the structural constraints of CTX1 and CTX3 is not important for the manifestation of membrane-perturbing activity.


Assuntos
Ácido Aspártico/química , Cardiotoxinas , Proteínas Cardiotóxicas de Elapídeos , Bicamadas Lipídicas/metabolismo , Naja naja , Sequência de Aminoácidos , Animais , Cardiotoxinas/química , Cardiotoxinas/farmacologia , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Proteínas Cardiotóxicas de Elapídeos/química , Proteínas Cardiotóxicas de Elapídeos/farmacologia , Humanos , Células K562 , Permeabilidade/efeitos dos fármacos
8.
J Am Heart Assoc ; 8(21): e013018, 2019 11 05.
Artigo em Inglês | MEDLINE | ID: mdl-31617439

RESUMO

Background Small molecule kinase inhibitors (KIs) are a class of agents currently used for treatment of various cancers. Unfortunately, treatment of cancer patients with some of the KIs is associated with cardiotoxicity, and there is an unmet need for methods to predict their cardiotoxicity. Here, we utilized a novel computational method to identify protein kinases crucial for cardiomyocyte viability. Methods and Results One hundred forty KIs were screened for their toxicity in cultured neonatal cardiomyocytes. The kinase targets of KIs were determined based on integrated data from binding assays. The key kinases mediating the toxicity of KIs to cardiomyocytes were identified by using a novel machine learning method for target deconvolution that combines the information from the toxicity screen and from the kinase profiling assays. The top kinases identified by the model were phosphoinositide 3-kinase catalytic subunit alpha, mammalian target of rapamycin, and insulin-like growth factor 1 receptor. Knockdown of the individual kinases in cardiomyocytes confirmed their role in regulating cardiomyocyte viability. Conclusions Combining the data from analysis of KI toxicity on cardiomyocytes and KI target profiling provides a novel method to predict cardiomyocyte toxicity of KIs.


Assuntos
Sobrevivência Celular , Aprendizado de Máquina , Miócitos Cardíacos/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Receptor IGF Tipo 1/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Cardiotoxinas/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Miócitos Cardíacos/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Ratos
9.
J Vis Exp ; (151)2019 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-31609335

RESUMO

Skeletal muscle possesses an enormous capacity to regenerate after injury. This process is mainly driven by muscle stem cells, also termed satellite cells. Satellite cells are characterized by the expression of the transcription factor Pax7 and their location underneath the basal lamina in the resting skeletal muscle. Upon injury, satellite cells get activated, undergo self-renewal or differentiation to either form new myofibers or to fuse with damaged ones. The functionality of satellite cells in vivo can be investigated using a cardiotoxin based injury model of skeletal muscle. To study the function of one gene during the regeneration of skeletal muscle, transgenic mouse models are mostly used. Here, we present an alternative method to transgenic mice, to investigate the gene function in satellite cells during regeneration, e.g., in cases where transgenic mice are not available. We combine the cardiotoxin mediated injury of a specific skeletal muscle with the injection of a self-delivering siRNA into the regenerating muscle which is then taken up by satellite cells among other cells. Thereby, we provide a method to analyze gene function in satellite cells during regeneration under physiological conditions without the need for transgenic mice.


Assuntos
Cardiotoxinas/farmacologia , Músculo Esquelético/fisiologia , RNA Interferente Pequeno , Regeneração/fisiologia , Cicatrização , Animais , Diferenciação Celular , Separação Celular , Modelos Animais de Doenças , Feminino , Masculino , Camundongos , Camundongos Transgênicos , Músculo Esquelético/efeitos dos fármacos , Fator de Transcrição PAX7 , RNA Interferente Pequeno/metabolismo , Células Satélites de Músculo Esquelético/fisiologia , Venenos de Serpentes/farmacologia
10.
ACS Sens ; 4(10): 2623-2630, 2019 10 25.
Artigo em Inglês | MEDLINE | ID: mdl-31535848

RESUMO

Detection of adverse effects of cardiac toxicity at an early stage by in vitro methods is crucial for the preclinical drug screening. Over the years, several kinds of biosensing platforms have been proposed by the scientific society for the detection of cardiac toxicity. However, the proposed tissue platforms have been optimized to measure either mechanophysiology or electrophysiology of the cardiomyocytes but not both. Herein, we demonstrate in detail our successful attempt toward developing a novel "multifunctional microphysiological system" also known as "organs-on-chips" to measure simultaneously the mechanical and electrical characteristics of cardiomyocytes in vitro. The proposed device can rapidly recognize drug-induced cardiovascular toxicity in real time, which is one of the most significant factors for drug discovery and postmarketing surveillance. We confirm that the proposed sensor delivers the direct relationship between the contraction force and cell impedance of cardiomyocytes under the influence of different cardiovascular drugs such as verapamil, astemizole, and lidocaine. The obtained assay results provide a great potential for a deep understanding of the drug effects on the cardiomyocytes in vitro.


Assuntos
Técnicas Biossensoriais , Cardiotoxinas/farmacologia , Avaliação Pré-Clínica de Medicamentos/métodos , Miócitos Cardíacos/efeitos dos fármacos , Animais , Astemizol/farmacologia , Cardiotoxicidade , Células Cultivadas , Impedância Elétrica , Fenômenos Eletrofisiológicos , Lidocaína/farmacologia , Microeletrodos , Miócitos Cardíacos/fisiologia , Ratos , Verapamil/farmacologia
11.
PLoS One ; 14(8): e0216167, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31408461

RESUMO

Regulators of G Protein Signaling (RGS proteins) inhibit G protein-coupled receptor (GPCR) signaling by accelerating the GTP hydrolysis rate of activated Gα subunits. Some RGS proteins exert additional signal modulatory functions, and RGS12 is one such protein, with five additional, functional domains: a PDZ domain, a phosphotyrosine-binding domain, two Ras-binding domains, and a Gα·GDP-binding GoLoco motif. RGS12 expression is temporospatially regulated in developing mouse embryos, with notable expression in somites and developing skeletal muscle. We therefore examined whether RGS12 is involved in the skeletal muscle myogenic program. In the adult mouse, RGS12 is expressed in the tibialis anterior (TA) muscle, and its expression is increased early after cardiotoxin-induced injury, suggesting a role in muscle regeneration. Consistent with a potential role in coordinating myogenic signals, RGS12 is also expressed in primary myoblasts; as these cells undergo differentiation and fusion into myotubes, RGS12 protein abundance is reduced. Myoblasts isolated from mice lacking Rgs12 expression have an impaired ability to differentiate into myotubes ex vivo, suggesting that RGS12 may play a role as a modulator/switch for differentiation. We also assessed the muscle regenerative capacity of mice conditionally deficient in skeletal muscle Rgs12 expression (via Pax7-driven Cre recombinase expression), following cardiotoxin-induced damage to the TA muscle. Eight days post-damage, mice lacking RGS12 in skeletal muscle had attenuated repair of muscle fibers. However, when mice lacking skeletal muscle expression of Rgs12 were cross-bred with mdx mice (a model of human Duchenne muscular dystrophy), no increase in muscle degeneration was observed over time. These data support the hypothesis that RGS12 plays a role in coordinating signals during the myogenic program in select circumstances, but loss of the protein may be compensated for within model syndromes of prolonged bouts of muscle damage and repair.


Assuntos
Diferenciação Celular , Proliferação de Células , Músculo Esquelético/citologia , Distrofia Muscular Animal/patologia , Mioblastos/citologia , Proteínas RGS/fisiologia , Animais , Cardiotoxinas/farmacologia , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos mdx , Camundongos Knockout , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/lesões , Músculo Esquelético/metabolismo , Distrofia Muscular Animal/metabolismo , Mioblastos/efeitos dos fármacos , Mioblastos/metabolismo , Transdução de Sinais
12.
Int J Biol Macromol ; 136: 512-520, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31199971

RESUMO

Snake venom cardiotoxins (CTXs) present diverse pharmacological functions. Previous studies have reported that CTXs affect the activity of some serine proteases, namely, chymotrypsin, subtilisin, trypsin, and acetylcholinesterase. To elucidate the mode of action of CTXs, the interaction of CTXs with chymotrypsin was thus investigated. It was found that Naja atra CTX isotoxins concentration-dependently enhanced chymotrypsin activity. The capability of CTX1 and CTX5 in increasing chymotrypsin activity was higher than that of CTX2, CTX3, and CTX4. Removal of the molecular beacon-bound CTXs by chymotrypsin, circular dichroism measurement, and acrylamide quenching of Trp fluorescence indicated that CTXs bound to chymotrypsin. Chemical modification of Lys, Arg, or Met residues of CTX1 attenuated its capability to enhance chymotrypsin activity without impairing their bond with chymotrypsin. Catalytically inactive chymotrypsin retained the binding affinity for native and modified CTX1. CTX1 and chemically modified CTX1 differently altered the global conformation of chymotrypsin and inactivated chymotrypsin. Moreover, CTX1 did not reduce the interaction of 2-(p-toluidino)-naphthalene-6-sulfonate (TNS) with chymotrypsin and inactivated chymotrypsin. Together with previous results revealing that TNS can bind at the hydrophobic region of active site in chymotrypsin, our data suggest that CTXs can enhance chymotrypsin activity by binding to the region outside the enzyme's active site.


Assuntos
Cardiotoxinas/farmacologia , Quimotripsina/metabolismo , Naja naja , Sequência de Aminoácidos , Animais , Sítios de Ligação , Cardiotoxinas/química , Cardiotoxinas/metabolismo , Quimotripsina/química , Simulação de Acoplamento Molecular , Conformação Proteica/efeitos dos fármacos
13.
Zebrafish ; 16(4): 379-387, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31145051

RESUMO

Envenomation by the Venezuelan bushmaster snake (Lachesis muta muta) (Serpentes: Viperidae) is characterized by local and cardiac alterations. This study investigates the in vivo cardiac dysfunction, tissue destruction, and cellular processes triggered by Lachesis muta muta snake crude venom and a C-type lectin (CTL)-like toxin named Mutacytin-1 (MC-1). The 28 kDa MC-1 was obtained by molecular exclusion, ion exchange, and C-18 (checking pureness) reverse-phase chromatographies. N-terminal sequencing of the first eight amino acids (NNCPQ LLM) revealed 100% identity with Mutina (CTL-like) isolated from Lachesis stenophrys, which is a Ca2+-dependent-type galactoside-binding lectin from Bothrops jararaca and CTL BpLec from Bothrops pauloensis. The cardiotoxicity in zebrafish of MC-1 was evaluated by means of specific phenotypic expressions and larvae behavior at 5, 15, 30, 40 and 60 min post-treatment. The L. muta muta venom and MC-1 also produced heart rate/rhythm alterations, circulation modifications, and the presence of thrombus and apoptotic phenomenon with pericardial damages. Acridine orange (100 µg/mL) was used to visualize apoptosis cellular process in control and treated whole embryos. The cardiotoxic alterations happened in more than 90% of all larvae under the action of L. muta muta venom and MC-1. The findings have demonstrated the potential cardiotoxicity by L. muta muta venom, suggesting the possibility of cardiovascular damages to patients after bushmaster envenoming.


Assuntos
Cardiotoxicidade/embriologia , Cardiotoxinas/farmacologia , Crotalinae , Lectinas Tipo C , Proteínas de Répteis/química , Venenos de Serpentes/química , Peixe-Zebra/embriologia , Animais , Cardiotoxinas/química , Crotalinae/embriologia , Embrião não Mamífero/efeitos dos fármacos , Lectinas Tipo C/química , Proteínas de Répteis/farmacologia
14.
Biosci Rep ; 39(6)2019 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-31138757

RESUMO

Cancer is the leading cause of deaths around the world, especially in low- and middle- income countries. Pirarubicin (THP) is an effective drug for treatment of cancer, however, there still exists cardiotoxic effects of THP. Rutin is a kind of antioxidative compound extracted from plants, and might be a protective compound for cardiomyocytes. Phosphatidylinositol 3-hydroxy kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signaling pathway is critical for cellular survival, proliferation and metabolism, and thus we speculated rutin might perform a protective role in cardiomyocytes via PI3K/AKT/mTOR signaling pathway. And in this experiment, we first established a cardiotoxicity model of THP in mice model and cell models, and then found that rutin treatment could increase the proliferation of cells at low concentration. Then we explored the possible mechanism of the protective effect of rutin using Western blotting, quantitative polymerase chain reaction (qPCR) and ELISA methods, and found that the activation of PI3K/AKT/mTOR/nuclear factor-κB (NF-κB) signaling pathway was increased, and expression of downstream molecules involved in antioxidative stress were also increased. We further noticed that concentration of angiogenesis promoting factors were also increased in medium of cultured cells. Thus, we speculated that rutin could increase the activation of PI3K/AKT/mTOR signaling pathway, further decrease the oxidative stress level via increasing the expression of antioxidative stress enzymes with the increasing concentration of angiogenesis promoting factors, resulting in the protective role in cardiomyocytes and cardiac function.


Assuntos
Cardiotoxicidade/tratamento farmacológico , Cardiotoxinas/efeitos adversos , Doxorrubicina/análogos & derivados , Miócitos Cardíacos/metabolismo , Rutina/farmacologia , Transdução de Sinais/efeitos dos fármacos , Animais , Cardiotoxicidade/metabolismo , Cardiotoxicidade/patologia , Cardiotoxinas/farmacologia , Doxorrubicina/efeitos adversos , Doxorrubicina/farmacologia , Masculino , Camundongos , Miócitos Cardíacos/patologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Serina-Treonina Quinases TOR/metabolismo
15.
Sci Rep ; 9(1): 2514, 2019 02 21.
Artigo em Inglês | MEDLINE | ID: mdl-30792528

RESUMO

Doxorubicin (DOX) is a potent chemotherapeutic with distinct cardiotoxic properties. Understanding the underlying cardiotoxic mechanisms on a molecular level would enable the early detection of cardiotoxicity and implementation of prophylactic treatment. Our goal was to map the patterns of different radiopharmaceuticals as surrogate markers of specific metabolic pathways induced by chemotherapy. Therefore, cardiac distribution of 99mTc-sestamibi, 99mTc-Annexin V, 99mTc-glucaric acid and [18F]FDG and cardiac expression of Bcl-2, caspase-3 and -8, TUNEL, HIF-1α, and p53 were assessed in response to DOX exposure in mice. A total of 80 mice (64 treated, 16 controls) were evaluated. All radiopharmaceuticals showed significantly increased uptake compared to controls, with peak cardiac uptake after one (99mTc-Annexin V), two (99mTc-sestamibi), three ([18F]FDG), or four (99mTc-glucaric acid) cycles of DOX. Strong correlations (p < 0.01) were observed between 99mTc-Annexin V, caspase 3 and 8, and TUNEL, and between [18F]FDG and HIF-1α. This suggests that the cardiac DOX response starts with apoptosis at low exposure levels, as indicated by 99mTc-Annexin V and histological apoptosis markers. Late process membrane disintegration can possibly be detected by 99mTc-sestamibi and 99mTc-glucaric acid. [18F]FDG signifies an early adaptive response to DOX, which can be further exploited clinically in the near future.


Assuntos
Cardiotoxinas/farmacologia , Doxorrubicina/farmacologia , Coração/efeitos dos fármacos , Neoplasias/tratamento farmacológico , Animais , Anexina A5/genética , Apoptose/efeitos dos fármacos , Cardiotoxinas/efeitos adversos , Caspase 3/genética , Modelos Animais de Doenças , Doxorrubicina/efeitos adversos , Fluordesoxiglucose F18/farmacologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Coração/fisiopatologia , Humanos , Camundongos , Neoplasias/complicações , Neoplasias/patologia , Compostos de Organotecnécio/efeitos adversos , Compostos de Organotecnécio/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/genética , Compostos Radiofarmacêuticos/farmacologia
16.
PLoS One ; 13(11): e0204913, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30403670

RESUMO

Aspartame (ASP) has been used as an alternative to sucrose for diabetics and obese people worldwide. Co-administration of L-carnitine (LC) with ASP has a protective effect against the liver and kidney toxicity induced of ASP. The goal of the investigation was to assess the enhancement of LC effect on the cardiac toxicity caused of ASP. The rats were divided into 6 groups: control with saline, LC (10 mg/kg), ASP (75 mg/kg), ASP (150 mg/kg), LC with 75 mg/kg of ASP, and LC with 150 mg/kg ASP. The antioxidants were determined by measuring the activities of myeloperoxidase, xanthine oxidase, superoxide dismutase, catalase, and glutathione peroxidase, and by assessing the levels of lipid peroxidation, total thiols, and glutathione. There was a significant elevation in LPO, in conjunction with a significant decline in the enzymatic antioxidants superoxide dismutase, catalase, and glutathione peroxidase and the non-enzymatic antioxidants glutathione and thiols. The cardiac myofibrils were found in a disarrayed pattern in ASP treated-animals as compared to the control rats. The animals treated with ASP-HD showed more than one apoptotic cell with a large tail and a small head, and the relaxed loops of the damaged DNA were extended to form a comet-shaped structure. These effects may be due to the excessive generation of reactive oxygen species by ASP, which reduces cardiac function. Co-administration of LC with ASP improved all of the above-mentioned parameters that were disrupted of ASP alone. This study evidences a sufficient originality in showing how LC plays a positive role against cardiac toxicity of ASP.


Assuntos
Apoptose/efeitos dos fármacos , Aspartame/efeitos adversos , Cardiotoxinas/efeitos adversos , Carnitina/farmacologia , Miocárdio/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Animais , Antioxidantes/metabolismo , Aspartame/farmacologia , Cardiotoxinas/farmacologia , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Miocárdio/patologia , Oxirredutases/metabolismo , Ratos , Ratos Wistar , Espécies Reativas de Oxigênio/metabolismo
17.
Expert Rev Proteomics ; 15(11): 873-886, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30328726

RESUMO

INTRODUCTION: Being important representatives of various proteomes, membrane-active cationic peptides (CPs) are attractive objects as lead compounds in the design of new antibacterial, anticancer, antifungal, and antiviral molecules. Numerous CPs are found in insect and snake venoms, where many of them reveal cytolytic properties. Due to advances in omics technologies, the number of such peptides is growing dramatically. Areas covered: To understand structure-function relationships for CPs in a living cell, detailed analysis of their hydrophobic/hydrophilic properties is indispensable. We consider two structural classes of membrane-active CPs: latarcins (Ltc) from spider and cardiotoxins (CTXs) from snake venoms. While the former are void off disulfide bonds and conformationally flexible, the latter are structurally rigid and cross-linked with disulfide bonds. In order to elucidate structure-activity relationships behind their antibacterial, anticancer, and hemolytic effects, the properties of these polypeptides are considered on a side-by-side basis. Expert commentary: An ever-increasing number of venom-derived membrane-active polypeptides require new methods for identification of their functional propensities and sequence-based design of novel pharmacological substances. We address these issues considering a number of the designed peptides, based either on Ltc or CTX sequences. Experimental and computer modeling techniques required for these purposes are delineated.


Assuntos
Peptídeos Catiônicos Antimicrobianos/farmacologia , Cardiotoxinas/farmacologia , Peptídeos/química , Peptídeos/farmacologia , Animais , Antibacterianos/química , Antibacterianos/farmacologia , Antifúngicos/química , Antifúngicos/farmacologia , Peptídeos Catiônicos Antimicrobianos/química , Antineoplásicos/química , Antineoplásicos/farmacologia , Cardiotoxinas/química , Dissulfetos/química , Desenho de Fármacos , Hemolíticos/química , Hemolíticos/farmacologia , Humanos , Interações Hidrofóbicas e Hidrofílicas , Peptídeos/metabolismo , Venenos de Aranha/química , Relação Estrutura-Atividade
18.
F1000Res ; 72018.
Artigo em Inglês | MEDLINE | ID: mdl-30345014

RESUMO

Recent advances in cancer prevention and management have led to an exponential increase of cancer survivors worldwide. Regrettably, cardiovascular disease has risen in the aftermath as one of the most devastating consequences of cancer therapies. In this work, we define cancer therapeutics-induced cardiotoxicity as the direct or indirect cardiovascular injury or injurious effect caused by cancer therapies. We describe four progressive stages of this condition and four corresponding levels of prevention, each having a specific goal, focus, and means of action. We subsequently unfold this didactic framework, surveying mechanisms of cardiotoxicity, risk factors, cardioprotectants, biomarkers, and diagnostic imaging modalities. Finally, we outline the most current evidence-based recommendations in this area according to multidisciplinary expert consensus guidelines.


Assuntos
Antineoplásicos/efeitos adversos , Cardiotônicos/uso terapêutico , Cardiotoxicidade/etiologia , Neoplasias/complicações , Animais , Cardiotoxinas/farmacologia , Doenças Cardiovasculares/induzido quimicamente , Humanos , Neoplasias/tratamento farmacológico
19.
BMC Res Notes ; 10(1): 406, 2017 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-28807058

RESUMO

BACKGROUND: The dose-limiting toxic effect of cyclophosphamide (CY) is cardiotoxicity. The pathogenesis of myocardial damage is poorly understood, and there is no established means of prevention. In previous studies, we suggested that for CY-induced cardiotoxicity, whereas acrolein is the key toxic metabolite, carboxyethylphosphoramide mustard (CEPM) is protective. We sought to verify that acrolein is the main cause of cardiotoxicity and to investigate whether aldehyde dehydrogenase (ALDH), which is associated with greater CEPM production, is involved in the protective effect for cardiotoxicity. We also evaluated the protective effect of N-acetylcysteine (NAC), an amino acid with antioxidant activity and a known acrolein scavenger. METHODS: H9c2 cells were exposed to CY metabolites HCY (4-hydroxy-cyclophosphamide), acrolein or CEPM. The degree of cytotoxicity was evaluated by MTT assay, lactate dehydrogenase (LDH) release, and the production of reactive oxygen species (ROS). We also investigated how the myocardial cellular protective effects of CY metabolites were modified by NAC. To quantify acrolein levels, we measured the culture supernatants using high performance liquid chromatography. We measured ALDH activity after exposure to HCY or acrolein and the same with pre-treatment with NAC. RESULTS: Exposure of H9c2 cells to CEPM did not cause cytotoxicity. Increased ROS levels and myocardial cytotoxicity, however, were induced by HCY and acrolein. In cell cultures, HCY was metabolized to acrolein. Less ALDH activity was observed after exposure to HCY or acrolein. Treatment with NAC reduced acrolein concentrations. CONCLUSIONS: Increased ROS generation and decreased ALDH activity confirmed that CY metabolites HCY and acrolein are strongly implicated in cardiotoxicity. By inhibiting ROS generation, increasing ALDH activity and decreasing the presence of acrolein, NAC has the potential to prevent CY-induced cardiotoxicity.


Assuntos
Apoptose/efeitos dos fármacos , Cardiotoxinas/farmacologia , Ciclofosfamida/farmacologia , Miócitos Cardíacos/efeitos dos fármacos , Acetilcisteína/farmacologia , Acroleína/metabolismo , Acroleína/farmacologia , Acroleína/toxicidade , Aldeído Desidrogenase/metabolismo , Animais , Cardiotoxicidade/metabolismo , Cardiotoxicidade/prevenção & controle , Cardiotoxinas/toxicidade , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Ciclofosfamida/análogos & derivados , Ciclofosfamida/metabolismo , Ciclofosfamida/toxicidade , Sequestradores de Radicais Livres/farmacologia , Imunossupressores/metabolismo , Imunossupressores/farmacologia , Imunossupressores/toxicidade , Mostardas de Fosforamida/metabolismo , Mostardas de Fosforamida/farmacologia , Mostardas de Fosforamida/toxicidade , Ratos , Espécies Reativas de Oxigênio/metabolismo
20.
J Pharmacol Toxicol Methods ; 87: 48-52, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-28549786

RESUMO

Drug-induced cardiotoxicity poses a negative impact on public health and drug development. Cardiac safety pharmacology issues urged for the preclinical assessment of drug-induced ventricular arrhythmia leading to the design of several in vitro electrophysiological screening assays. In general, patch clamp systems allow for intracellular recordings, while multi-electrode array (MEA) technology detect extracellular activity. Here, we demonstrate a complementary metal oxide semiconductor (CMOS)-based MEA system as a reliable platform for non-invasive, long-term intracellular recording of cardiac action potentials at high resolution. Quinidine (8 concentrations from 10-7 to 2.10-5M) and verapamil (7 concentrations from 10-11 to 10-5M) were tested for dose-dependent responses in a network of cardiomyocytes. Electrophysiological parameters, such as the action potential duration (APD), rates of depolarization and repolarization and beating frequency were assessed. In hiPSC, quinidine prolonged APD with EC50 of 2.2·10-6M. Further analysis indicated a multifactorial action potential prolongation by quinidine: (1) decreasing fast repolarization with IC50 of 1.1·10-6M; (2) reducing maximum upstroke velocity with IC50 of 2.6·10-6M; and (3) suppressing spontaneous activity with EC50 of 3.8·10-6M. In rat neonatal cardiomyocytes, verapamil blocked spontaneous activity with EC50 of 5.3·10-8M and prolonged the APD with EC50 of 2.5·10-8M. Verapamil reduced rates of fast depolarization and repolarization with IC50s of 1.8 and 2.2·10-7M, respectively. In conclusion, the proposed action potential-based MEA platform offers high quality and stable long-term recordings with high information content allowing to characterize multi-ion channel blocking drugs. We anticipate application of the system as a screening platform to efficiently and cost-effectively test drugs for cardiac safety.


Assuntos
Potenciais de Ação/fisiologia , Antiarrítmicos/farmacologia , Cardiotoxinas/farmacologia , Células-Tronco Pluripotentes Induzidas/fisiologia , Miócitos Cardíacos/fisiologia , Semicondutores , Potenciais de Ação/efeitos dos fármacos , Animais , Animais Recém-Nascidos , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos/métodos , Humanos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Microeletrodos , Miócitos Cardíacos/efeitos dos fármacos , Quinidina/farmacologia , Ratos , Ratos Wistar
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