Mesenchymal stem cells (MSCs) have been reported to improve
survival of cardiomyocytes (CMCs) and overall regeneration of
cardiac tissue. Despite promising preclinical results, interactions
of MSCs and CMCs, both direct and indirect, remain unclear. In
this study, porcine bone marrow MSCs and freshly isolated
porcine primary adult CMCs were used for non-contact co-culture
experiments. Morphology, viability and functional parameters of
CMCs were measured over time and compared between CMCs
cultured alone and CMCs co-cultured with MSCs. In non-contact
co-culture, MSCs improved survival of CMCs. CMCs co-cultured
with MSCs maintained CMCs morphology and viability in
significantly higher percentage than CMCs cultured alone. In
viable CMCs, mitochondrial respiration was preserved in both
CMCs cultured alone and in CMCs co-cultured with MSCs.
Comparison of cellular contractility and calcium handling,
measured in single CMCs, revealed no significant differences
between viable CMCs from co-culture and CMCs cultured alone.
In conclusion, non-contact co-culture of porcine MSCs and CMCs
improved survival of CMCs with a sufficient preservation of
functional and mitochondrial parameters.
We previously found that Endothelin-11-31 (ET-11-31) exhibited a pro-arrhythmogenic effect in isolated rat hearts. In this study, we further investigated the effects of ET-11-31 on a cell viability and observed [Ca2+]i in cultured cardiomyocytes. Cultured neonatal rat cardiomyocytes were treated with 0.1, 1, and 10 nM ET-11-31 for 24h in the presence or absence of ETA receptor antagonist (BQ123) or phosphoramidon, a NEP/ECE inhibitor. Cell injury was evaluated by supernatant lactate dehydrogenase (LDH) assay, superoxide dismutase (SOD activity, and malondialdehyde (MDA) content. Cell viability was assessed by MTT assay. [Ca2+]i was measured with Fluo-3/AM under a laser confocal microscope. 1) ET-11-31 dose-dependently increased LDH release and decreased cell viability. 2) LDH and MDA levels were significantly elevated and SOD activity decreased after administration of 1 nM ET-11-31 for 24h, and these changes were markedly attenuated by 1 uM BQ123. 3) Exposure to 10 nM ET-11-31 caused a continuous increase in [Ca2+]i to cultured beating cardiomyocytes and termination of [Ca2+]i transient within 6 min, and this change was reversed by 1 uM BQ123 and attenuated by 0.5 mM phosphoramidon. These results suggest that ET-11-31 could cause cell injury, and that the effect of ET-11-31 on [Ca2+]i transients is mainly mediated by ETA receptor and partially attributed to the conversion of ET-11-31 to ET-11-21., A.-J. Ren, X. Yuan, L. Lin, Y.-X. Pan, Y.-W. Qing, W.-J. Yuan., and Obsahuje bibliografii a bibliografické odkazy
MicroRNAs are emerging as important regulators of cardiac function. This study investigated the role of microRNA-24 (miR-24) in ischemic cardiomyocytes, based on the observation that miR-24 expression was significantly enhanced in the ischemic myocardium of rats. Using primary cultured rat cardiomyocytes, cell injury was induced by ischemic conditions, and the cells were evaluated for changes in lactate dehydrogenase (LDH) release, cell viability, apoptosis and necrosis. The results showed that miR-24 was increased in myocytes exposed to ischemia. When miR-24 was further overexpressed in ischemic myocytes using the mimic RNA sequence, LDH release was reduced, cell viability was enhanced, and apoptosis and necrosis rates were both decreased. By contrast, a deficiency in miR-24 resulted in the largest LDH release, lowest cell viability and highest apoptosis and necrosis rates in normal and ischemic myocytes, with significant changes compared to that of non-transfected myocytes. Additionally, the mRNA and protein levels of the pro-apoptotic gene, BCL2L11, were down-regulated by miR-24 overexpression and up-regulated by miR-24 deficiency. The luciferase reporter assay confirmed BCL2L11 to be a target of miR-24. Overall, this study showed a protective role for miR-24 against myocardial ischemia by inhibiting BCL2L11, and may represent a potential novel treatment for ischemic heart disease., D.-F. Li ... [et al.]., and Obsahuje seznam literatury
Infection with Trypanosoma cruzi Chagas, 1909 is reported to increase the production of reactive oxygen species in patients with Chagas disease. Mitochondria dysfunction, host inflammatory response and inadequate antioxidant response are described as the main factors leading to oxidative stress during acute and chronic stages of the disease. The Seahorse XFe24 extracellular flux platform allows energy metabolism determination through mitochondrial respiration and glycolysis measurements. XFe24 platform can be used in in vitro models of T. cruzi-infected cells, which allow the assessment and even modulation of endogenous conditions of infected cells, generating readouts of real-time cellular bioenergetics changes. In this protocol, we standardised the use of XFe24 technology in T. cruzi infected AC16 cardiomyocytes and SGHPL-5 trophoblasts. In addition, we provide a list of optimised assay specifications, advantages and critical steps to be considered during the process. Cardiomyocytes and trophoblasts are attractive target cells to evaluate the metabolic environment in acute, chronic and congenital Chagas transmission scenarios.