Characterization and Standardization of Cultured Cardiac Fibroblasts for Ex Vivo Models of Heart Fibrosis and Heart Ischemia
Abstract
A full understanding of cardiac fibroblast (cFB) biology is essential to study the adverse cardiac remodeling and recovery of myocardium infarction. However, compared to cardiac myocytes, cFBs are less well characterized. Important questions, including the variability introduced by cell age (neonatal vs. adult), culture conditions (passage, plate coating, and culture medium), and responses to stimuli (e.g., hypoxia and drug treatments), have not been well addressed and standardization of techniques is lacking. This variability invites inconsistency and the confounding of study conclusions. Thus, we here focus on characterizing cell responses and standardizing procedures for cFB isolation and culture conditions to provide reliable platforms to address important questions about cFB proliferation, activation, collagen matrix formation, and responses to relevant stimuli. Thirty litters of 1-3-day pups and 30 female (240-330 g) Sprague-Dawley rats were used to isolate neonatal and adult cFBs. We detail and validate procedures to isolate cFBs for the use of culture or direct analysis. We characterize the differences between neonatal and adult cFBs, define the changes of cFBs during serial passage, and identify the response of cFBs to...Continue Reading
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High-yield and high-purity isolation of hepatic stellate cells from normal and fibrotic mouse livers
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