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Title: Integrated molecular, biochemical, and physiological assessment unravels key extraction method mediated influences on rat neonatal cardiomyocytes
Authors: JENSEN, LeonardoNERI, ElidaBASSANEZE, ViniciusOLIVEIRA, Nathalia C. De AlmeidaDARIOLLI, RafaelTURACA, Lauro T.LEVY, DeboraVERONEZ, DouglasFERRAZ, Mariana S. A.ALENCAR, Adriano M.BYDLOWSKI, Sergio P.CESTARI, Idagene A.KRIEGER, Jose Eduardo
Citation: JOURNAL OF CELLULAR PHYSIOLOGY, v.233, n.7, p.5420-5430, 2018
Abstract: Neonatal cardiomyocytes are instrumental for disease modeling, but the effects of different cell extraction methods on basic cell biological processes remain poorly understood. We assessed the influence of two popular methods to extract rat neonatal cardiomyocytes, Pre-plating (PP), and Percoll (PC) on cell structure, metabolism, and function. Cardiomyocytes obtained from PP showed higher gene expression for troponins, titin, and potassium and sodium channels compared to PC. Also, PP cells displayed higher levels of troponin I protein. Cells obtained from PC displayed higher lactate dehydrogenase activity and lactate production than PP cells, indicating higher anaerobic metabolism after 8 days of culture. In contrast, reactive oxygen species levels were higher in PP cells as indicated by ethidium and hydroxyethidium production. Consistent with these data, protein nitration was higher in PP cells, as well as nitrite accumulation in cell medium. Moreover, PP cells showed higher global intracellular calcium under basal and 1mM isoprenaline conditions. In a calcium-transient assessment under electrical stimulation (0.5Hz), PP cells displayed higher calcium amplitude than cardiomyocytes obtained from PC and using a traction force microscope technique we observed that PP cardiomyocytes showed the highest relaxation. Collectively, we demonstrated that extraction methods influence parameters related to cell structure, metabolism, and function. Overall, PP derived cells are more active and mature than PC cells, displaying higher contractile function and generating more reactive oxygen species. On the other hand, PC derived cells display higher anaerobic metabolism, despite comparable high yields from both protocols.
Appears in Collections:

Artigos e Materiais de Revistas Científicas - FM/MCM
Departamento de Clínica Médica - FM/MCM

Artigos e Materiais de Revistas Científicas - FM/MCP
Departamento de Cardio-Pneumologia - FM/MCP

Artigos e Materiais de Revistas Científicas - HC/InCor
Instituto do Coração - HC/InCor

Artigos e Materiais de Revistas Científicas - LIM/13
LIM/13 - Laboratório de Genética e Cardiologia Molecular

Artigos e Materiais de Revistas Científicas - LIM/31
LIM/31 - Laboratório de Genética e Hematologia Molecular

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