Exosomes play a crucial role in the progression of infectious diseases, as exosome release and biogenesis are affected by external factors, such as pathogenic infections. Pyrogens may aide in the progression of diseases by triggering inflammation, endothelial cell injury, and arterial plaque rupture, all of which can lead to acute coronary disease, resulting in cardiac tissue death and the onset of a cardiac event (CE). To better understand the effects of Gram-negative bacterial infections on exosome composition and biogenesis, we examined exosome characteristics after treatment of AC16 human cardiomyocytes with lipopolysaccharide (LPS), which served as a model system for Gram-negative bacterial infection. Using increasing doses (0, 0.1, 1, or 10 µg) of LPS, we showed that treatment with LPS substantially altered the composition of AC16-derived exosomes. Both the relative size and the quantity (particles/mL) of exosomes were decreased significantly at all tested concentrations of LPS treatment compared to the untreated group. In addition, LPS administration reduced the expression of exosomal proteins that are related to exosomal biogenesis. Conversely, we observed an increase in immunomodulators present after LPS administration. This evaluation of the impact of LPS on cardiac cell death and exosome composition will yield new insight into the importance of exosomes in a variety of physiological and pathological processes as it relates to disease progression, diagnosis, and treatment.
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Beta-Amyloid Instigates Dysfunction of Mitochondria in Cardiac Cells
Alzheimer’s disease (AD) includes the formation of extracellular deposits comprising aggregated β-amyloid (Aβ) fibers associated with oxidative stress, inflammation, mitochondrial abnormalities, and neuronal loss. There is an associative link between AD and cardiac diseases; however, the mechanisms underlying the potential role of AD, particularly Aβ in cardiac cells, remain unknown. Here, we investigated the role of mitochondria in mediating the effects of Aβ1-40 and Aβ1-42 in cultured cardiomyocytes and primary coronary endothelial cells. Our results demonstrated that Aβ1-40 and Aβ1-42 are differently accumulated in cardiomyocytes and coronary endothelial cells. Aβ1-42 had more adverse effects than Aβ1-40 on cell viability and mitochondrial function in both types of cells. Mitochondrial and cellular ROS were significantly increased, whereas mitochondrial membrane potential and calcium retention capacity decreased in both types of cells in response to Aβ1-42. Mitochondrial dysfunction induced by Aβ was associated with apoptosis of the cells. The effects of Aβ1-42 on mitochondria and cell death were more evident in coronary endothelial cells. In addition, Aβ1-40 and Aβ1-42 significantly increased Ca2+ -induced swelling in mitochondria isolated from the intact rat hearts. In conclusion, this study demonstrates the toxic effects of Aβ on cell survival and mitochondria function in cardiac cells.
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- Award ID(s):
- 2006477
- NSF-PAR ID:
- 10320684
- Date Published:
- Journal Name:
- Cells
- Volume:
- 11
- Issue:
- 3
- ISSN:
- 2073-4409
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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Cardiovascular disease has been associated with increased levels of reactive oxygen species (ROS). Recently, we have shown that a critical balance between cytosolic ROS and mitochondrial ROS is crucial in cardiovascular health and that modulation of mitochondrial ROS helps prevent detrimental effects of cytosolic ROS on endothelial cells (EC) in transgenic animals. Here, we report the development of a controlled delivery system for a mitochondria‐targeted antioxidant, JP4‐039, from an electrospun scaffold made of FDA‐approved biocompatible polymeric nanofibers. We demonstrate that the active antioxidant moiety was preserved in released JP4‐039 for over 72 h using electron paramagnetic resonance. We also show that both the initial burst release of the drug within the first 20 min and the ensuing slow and sustained release that occurred over the next 24 h improved tube formation in human coronary artery ECs (HCAEC)
in vitro . Taken together, these findings suggest that electrospinning methods can be used to upload mitochondrial antioxidant (JP4‐039) onto a biocompatible nanofibrous PLGA scaffold, and the uploaded drug (JP4‐039) retains nitroxide antioxidant properties upon release from the scaffold, which in turn can reduce mitochondrial ROS and improve EC functionin vitro . -
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Abstract Background and Aims Fibrotic tissue formed after myocardial infarction (MI) can be as detrimental as MI itself. However, current in vitro cardiac fibrosis models fail to recapitulate the complexities of post‐MI tissue. Moreover, although MI and subsequent fibrosis is most prominent in the aged population, the field suffers from inadequate aged tissue models. Herein, an aged human post‐MI tissue model, representing the native microenvironment weeks after initial infarction, is engineered using three‐dimensional bioprinting via creation of individual bioinks to specifically mimic three distinct regions: remote, border, and scar.
Methods The aged post‐MI tissue model is engineered through combination of gelatin methacryloyl, methacrylated hyaluronic acid, aged type I collagen, and photoinitiator at variable concentrations with different cell types, including aged human induced pluripotent stem cell‐derived cardiomyocytes, endothelial cells, cardiac fibroblasts, and cardiac myofibroblasts, by introducing a methodology which utilizes three printheads of the bioprinter to model aged myocardium. Then, using cell‐specific proteins, the cell types that comprised each region are confirmed using immunofluorescence. Next, the beating characteristics are analyzed. Finally, the engineered aged post‐MI tissue model is used as a benchtop platform to assess the therapeutic effects of stem cell‐derived extracellular vesicles on the scar region.
Results As a result, high viability (>74%) was observed in each region of the printed model. Constructs demonstrated functional behavior, exhibiting a beating velocity of 6.7 μm/s and a frequency of 0.3 Hz. Finally, the effectiveness of hiPSC‐EV and MSC‐EV treatment was assessed. While hiPSC‐EV treatment showed no significant changes, MSC‐EV treatment notably increased cardiomyocyte beating velocity, frequency, and confluency, suggesting a regenerative potential.
Conclusion In conclusion, we envision that our approach of modeling post‐MI aged myocardium utilizing three printheads of the bioprinter may be utilized for various applications in aged cardiac microenvironment modeling and testing novel therapeutics.
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Key points Association of plasma membrane BKCachannels with BK‐β subunits shapes their biophysical properties and physiological roles; however, functional modulation of the mitochondrial BKCachannel (mitoBKCa) by BK‐β subunits is not established.
MitoBKCa‐α and the regulatory BK‐β1 subunit associate in mouse cardiac mitochondria.
A large fraction of mitoBKCadisplay properties similar to that of plasma membrane BKCawhen associated with BK‐β1 (left‐shifted voltage dependence of activation,
V 1/2 = −55 mV, 12 µm matrix Ca2+).In BK‐β1 knockout mice, cardiac mitoBKCadisplayed a low
P oand a depolarizedV 1/2of activation (+47 mV at 12 µm matrix Ca2+)Co‐expression of BKCawith the BK‐β1 subunit in HeLa cells doubled the density of BKCain mitochondria.
The present study supports the view that the cardiac mitoBKCachannel is functionally modulated by the BK‐β1 subunit; proper targeting and activation of mitoBKCashapes mitochondrial Ca2+handling.
Abstract Association of the plasma membrane BKCachannel with auxiliary BK‐β1–4 subunits profoundly affects the regulatory mechanisms and physiological processes in which this channel participates. However, functional association of mitochondrial BK (mitoBKCa) with regulatory subunits is unknown. We report that mitoBKCafunctionally associates with its regulatory subunit BK‐β1 in adult rodent cardiomyocytes. Cardiac mitoBKCais a calcium‐ and voltage‐activated channel that is sensitive to paxilline with a large conductance for K+of 300 pS. Additionally, mitoBKCadisplays a high open probability (
P o) and voltage half‐activation (V 1/2 = −55 mV,n = 7) resembling that of plasma membrane BKCawhen associated with its regulatory BK‐β1 subunit. Immunochemistry assays demonstrated an interaction between mitochondrial BKCa‐α and its BK‐β1 subunit. Mitochondria from the BK‐β1 knockout (KO) mice showed sparse mitoBKCacurrents (five patches with mitoBKCaactivity out of 28 total patches fromn = 5 different hearts), displaying a depolarizedV 1/2of activation (+47 mV in 12 µm matrix Ca2+). The reduced activity of mitoBKCawas accompanied by a high expression of BKCatranscript in the BK‐β1 KO, suggesting a lower abundance of mitoBKCachannels in this genotype. Accordingly, BK‐β1subunit increased the localization of BKDEC (i.e. the splice variant of BKCathat specifically targets mitochondria) into mitochondria by two‐fold. Importantly, both paxilline‐treated and BK‐β1 KO mitochondria displayed a more rapid Ca2+overload, featuring an early opening of the mitochondrial transition pore. We provide strong evidence that mitoBKCaassociates with its regulatory BK‐β1 subunit in cardiac mitochondria, ensuring proper targeting and activation of the mitoBKCachannel that helps to maintain mitochondrial Ca2+homeostasis. -
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Abstract Background Cardiac pathological outcome of metabolic remodeling is difficult to model using cardiomyocytes derived from human-induced pluripotent stem cells (hiPSC-CMs) due to low metabolic maturation.
Methods hiPSC-CM spheres were treated with AMP-activated protein kinase (AMPK) activators and examined for hiPSC-CM maturation features, molecular changes and the response to pathological stimuli.
Results Treatment of hiPSC-CMs with AMPK activators increased ATP content, mitochondrial membrane potential and content, mitochondrial DNA, mitochondrial function and fatty acid uptake, indicating increased metabolic maturation. Conversely, the knockdown of AMPK inhibited mitochondrial maturation of hiPSC-CMs. In addition, AMPK activator-treated hiPSC-CMs had improved structural development and functional features—including enhanced Ca2+transient kinetics and increased contraction. Transcriptomic, proteomic and metabolomic profiling identified differential levels of expression of genes, proteins and metabolites associated with a molecular signature of mature cardiomyocytes in AMPK activator-treated hiPSC-CMs. In response to pathological stimuli, AMPK activator-treated hiPSC-CMs had increased glycolysis, and other pathological outcomes compared to untreated cells.
Conclusion AMPK activator-treated cardiac spheres could serve as a valuable model to gain novel insights into cardiac diseases.
- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.3390/cells11030373
