scholarly journals Study of Stem Cells Influence on Cardiac Cells Cultured with a Cyanide-P-Trifluoromethoxyphenylhydrazone in Organ-on-a-Chip System

Biosensors ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 131
Author(s):  
Anna Kobuszewska ◽  
Dominik Kolodziejek ◽  
Michal Wojasinski ◽  
Tomasz Ciach ◽  
Zbigniew Brzozka ◽  
...  
Keyword(s):  
Stem Cells ◽  
Metabolic Activity ◽  
Microfluidic System ◽  
Cardiac Cells ◽  
Effective Solution ◽  
Mitochondrial Oxidative Phosphorylation ◽  
Human Cardiomyocytes ◽  
Bioluminescence Assay ◽  
Nanofiber Mats

Regenerative medicine and stem cells could prove to be an effective solution to the problem of treating heart failure caused by ischemic heart disease. However, further studies on the understanding of the processes which occur during the regeneration of damaged tissue are needed. Microfluidic systems, which provide conditions similar to in vivo, could be useful tools for the development of new therapies using stem cells. We investigated how mesenchymal stem cells (MSCs) affect the metabolic activity of cardiac cells (rat cardiomyoblasts and human cardiomyocytes) incubated with a potent uncoupler of mitochondrial oxidative phosphorylation under microfluidic conditions. A cyanide p-trifluoromethoxyphenylhydrazone (FCCP) was used to mimic disfunctions of mitochondria of cardiac cells. The study was performed in a microfluidic system integrated with nanofiber mats made of poly-l-lactid acid (PLLA) or polyurethane (PU). The microsystem geometry allows four different cell cultures to be conducted under different conditions (which we called: normal, abnormal—as both a mono- and co-culture). Metabolic activity of the cells, based on the bioluminescence assay, was assessed in the culture’s performed in the microsystem. It was proved that stem cells increased metabolic activity of cardiac cells maintained with FCCP.

scholarly journals “Empowering” Cardiac Cells via Stem Cell Derived Mitochondrial Transplantation- Does Age Matter?

2021 ◽  
Vol 22 (4) ◽  
pp. 1824
Author(s):  
Matthias Mietsch ◽  
Rabea Hinkel
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Treatment Strategies ◽  
Cardiac Cells ◽  
Aging Effects ◽  
Beneficial Effects ◽  
Micro Rnas ◽  
New Treatment

With cardiovascular diseases affecting millions of patients, new treatment strategies are urgently needed. The use of stem cell based approaches has been investigated during the last decades and promising effects have been achieved. However, the beneficial effect of stem cells has been found to being partly due to paracrine functions by alterations of their microenvironment and so an interesting field of research, the “stem- less” approaches has emerged over the last years using or altering the microenvironment, for example, via deletion of senescent cells, application of micro RNAs or by modifying the cellular energy metabolism via targeting mitochondria. Using autologous muscle-derived mitochondria for transplantations into the affected tissues has resulted in promising reports of improvements of cardiac functions in vitro and in vivo. However, since the targeted treatment group represents mainly elderly or otherwise sick patients, it is unclear whether and to what extent autologous mitochondria would exert their beneficial effects in these cases. Stem cells might represent better sources for mitochondria and could enhance the effect of mitochondrial transplantations. Therefore in this review we aim to provide an overview on aging effects of stem cells and mitochondria which might be important for mitochondrial transplantation and to give an overview on the current state in this field together with considerations worthwhile for further investigations.

scholarly journals Simulation of hypoxia of myocardial cells in microfluidic systems

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Anna Kobuszewska ◽  
Elżbieta Jastrzębska ◽  
Kamil Żukowski ◽  
Zbigniew Brzózka
Keyword(s):  
Fluorescent Dyes ◽  
Microfluidic System ◽  
Cardiac Cells ◽  
Human Cardiomyocytes ◽  
New Methods ◽  
Biochemical Simulation ◽  
Carbonyl Cyanide ◽  
Myocardial Hypoxia ◽  
Biochemical Approach ◽  
Methods Of Treatment

Abstract The paper presents a newly designed microfluidic system that allows simulation of myocardial hypoxia by biochemical method. The geometry of the microsystem was designed in such a way, that quantitative fluorescent measurements using a spectrofluorometric plate reader was possible. Biochemical simulation of hypoxia was carried out using potent mitochondrial oxidative phosphorylation uncoupler—Carbonyl cyanide-4-(trifluoromethoxy)phenylhydrazone (FCCP). Two cardiac cell lines were used in the study—rat cardiomyoblasts (H9C2) and human cardiomyocytes. The effectiveness of biochemical simulation of hypoxia was studied using two fluorescent dyes: carbocyanine iodide (JC-1) and Fluo-4. Changes in the mitochondrial membrane potential and concentration of intracellular calcium ions were tested. The major novelty of this research was the applying the microfluidic system to create hypoxia conditions for cardiac cells using the biochemical approach. In further studies, the presented hypoxia model could be used to develop new methods of treatment of ischemic heart disease for example in cell therapy based on stem cells.

scholarly journals Mcl-1/CDK9 Targeting By AZD5991/AZD4573 Overcomes Intrinsic and Acquired Venetoclax Resistance in Vitro and In Vivo in PDX Model of AML through Modulation of Cell Death and Metabolic Functions

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 768-768 ◽  
Author(s):  
Bing Z Carter ◽  
Po Yee Mak ◽  
Wenjing Tao ◽  
Mark Warmoes ◽  
Philip L Lorenzi ◽  
...  
Keyword(s):  
Stem Cells ◽  
Research Funding ◽  
Single Agent ◽  
O2 Consumption ◽  
Mitochondrial Oxidative Phosphorylation ◽  
Metabolic Functions ◽  
Pdx Model ◽  
Induced Apoptosis

Abstract Mcl-1 and Bcl-2 are two major anti-apoptotic Bcl-2 proteins frequently overexpressed in malignant cells. They cooperatively support cell survival and are associated with therapy resistance. ABT-199 (venetoclax), a highly selective Bcl-2 inhibitor, showed potent preclinical activity but limited clinical efficacy in AML as a single agent. Mcl-1 is induced by and a major resistance factor to ABT-199. Mcl-1 was recently found to also positively regulate mitochondrial oxidative phosphorylation that induces cancer stem cells and promotes chemoresistance. Mcl-1 is essential for the development of AML and for the survival of AML cells and stem cells. Increased mitochondrial oxidative phosphorylation has been demonstrated in these cells. First we found that Mcl-1 overexpressing (OE)/knockdown (KD) AML cells were markedly more resistant/sensitive to ABT-199 than were corresponding control cells, supporting the notion of Mcl-1 as a resistance factor to ABT-199. Inhibition of Mcl-1 by the selective Mcl-1 inhibitor AZD5991 or the CDK9 inhibitor AZD4573, which down-regulates short-lived proteins such as Mcl-1, induced apoptosis and showed strong synergy when combined with ABT-199 in AML cell lines, primary AML blasts, and stem/progenitor cells from patients. Importantly, combinations of AZD5991 or AZD4573 with ABT-199 synergistically induced apoptosis in OCI-AML3 and Mcl-1 OE cells intrinsically resistant to ABT-199 and in AML cell lines and primary patient cells with acquired resistance to ABT-199. Although OE/KD Mcl-1 in AML cells did not show obvious alterations in baseline cell viability, NSGS mice harboring Mcl-1 OE/KD OCI-AML3 cells survived significantly shorter/longer than those transplanted with control cells, supporting additional, non-apoptogenic functions of Mcl-1 in AML. We observed that genetic modulation of Mcl-1 alters cellular mitochondrial respiration and ROS levels. AML cells with Mcl-1 OE/KD increased/decreased O2 consumption and mitochondrial ATP and ROS generation. Consistent with this finding, inhibition of Mcl-1 by AZD5991 or AZD4573 decreased O2 consumption and ATP generation in AML cells and also in MV4-11 cells with acquired ABT-199 resistance. Mass spectrometry-based stable isotope tracing experiments using 1,2-13C-glucose showed that both genetic and pharmacological inhibition of Mcl-1 decreased flux of glucose carbon through glycolysis, the TCA cycle, and the pentose phosphate pathway, suggesting a role for Mcl-1 in cellular respiration and redox metabolism. To further assess the efficacy of combined Mcl-1 and Bcl-2 inhibition in primary AML cells resistant to ABT-199, we developed a PDX model using cells from an AML patient who initially responded to ABT-199/demethylating agent and then relapsed. NSGS mice engrafted with these PDX cells were treated with ABT-199 (50 mg/kg, oval gavage qd), AZD5991 (60 mg/kg, i.v. weekly), AZD4573 (15 mg/kg, i.p. bid with 2 h interval for two consequent days/week), ABT-199+AZD5991, or ABT-199+AZD4573 for 6 wks. Flow cytometric analysis of circulating human CD45+ cells on day 18 of therapy showed that each agent significantly decreased leukemia burden and that the combinations were significantly more effective (P<0.01) than each single agent. CyTOF analysis of BM cells (day 25) showed that both combinations markedly reduced (P<0.001) human CD45+ cells and, more importantly, human CD34+CD38+/CD38- and CD34+CD38+/CD38-CD123+ cells. Those combination treatments also decreased Mcl-1, Bcl-2, b-catenin, c-Myc, and FAK protein expression in CD34+CD38-CD123+ cells. Interestingly, AZD5991, AZD4573, or their combinations with ABT-199 greatly decreased CXCR4 in all cell populations. Ultimately, each single agent only marginally prolonged survival, whereas ABT-199+AZD4573 and even more so ABT-199+AZD5991 markedly improved survival in this highly ABT-199 resistant PDX model (Fig). Conclusion: we demonstrate that Mcl-1 has metabolic functions in AML and that inhibition of Mcl-1 enhances ABT-199 apoptogenic activity and overcomes intrinsic and acquired ABT-199 resistance in vitro and in vivo in a PDX murine model of AML, suggesting that inhibition of Mcl-1 improves the efficacy of ABT-199, and overcomes established resistance to Bcl-2 inhibition. Suppressing metabolic activity and CXCR4 inhibition may also contribute to the efficacy of this combination against AML stem cells in the BM microenvironment. Figure. Figure. Disclosures Carter: AstraZeneca: Research Funding; novartis: Research Funding. Lorenzi:Erytech Pharma: Consultancy; NIH: Patents & Royalties. Cidado:AstraZeneca: Employment, Equity Ownership. Drew:AstraZeneca: Employment. Andreeff:AstraZeneca: Research Funding.

scholarly journals From Human Pluripotent Stem Cells to 3D Cardiac Microtissues: Progress, Applications and Challenges

2020 ◽  
Vol 7 (3) ◽  
pp. 92
Author(s):  
Mariana A. Branco ◽  
Joaquim M.S. Cabral ◽  
Maria Margarida Diogo
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
High Throughput Screening ◽  
Cardiac Development ◽  
Human Pluripotent Stem Cells ◽  
Cardiac Cells ◽  
Cardiac Disorders ◽  
The Impact

The knowledge acquired throughout the years concerning the in vivo regulation of cardiac development has promoted the establishment of directed differentiation protocols to obtain cardiomyocytes (CMs) and other cardiac cells from human pluripotent stem cells (hPSCs), which play a crucial role in the function and homeostasis of the heart. Among other developments in the field, the transition from homogeneous cultures of CMs to more complex multicellular cardiac microtissues (MTs) has increased the potential of these models for studying cardiac disorders in vitro and for clinically relevant applications such as drug screening and cardiotoxicity tests. This review addresses the state of the art of the generation of different cardiac cells from hPSCs and the impact of transitioning CM differentiation from 2D culture to a 3D environment. Additionally, current methods that may be employed to generate 3D cardiac MTs are reviewed and, finally, the adoption of these models for in vitro applications and their adaptation to medium- to high-throughput screening settings are also highlighted.

Abstract 159: Empowering Cardiac Progenitor Cell-mediated Repair of Injured Myocardium by Overexpressing P2Y2 Nucleotide Receptor

2015 ◽  
Vol 117 (suppl_1) ◽  
Author(s):  
Farid El-Sayed ◽  
Mark Sussman
Keyword(s):  
Stem Cells ◽  
Preliminary Data ◽  
Purinergic Signaling ◽  
Extracellular Nucleotides ◽  
Hippo Signaling ◽  
Functional Responses ◽  
Human Cardiomyocytes ◽  
Injured Myocardium ◽  
Wide Range

Heart failure is a leading cause of death in the US due to the limited capability of adult mammalian heart to regenerate following injury. Autologous stem cell therapy holds promise for regeneration of injured myocardium after myocardial infarction. However, stem cells derived from diseased organs exhibit impaired proliferative and migratory capabilities and increased susceptibility to cell death. Empowering stem cells from diverse origins, including cardiac progenitor cells (CPCs), with pro-survival genes has been attempted. Despite the well-established roles of purinergic signaling mediated by extracellular nucleotides in regulating diverse cellular responses in cardiovascular diseases, it has not been well-defined in CPCs. Our preliminary data show, for the first time, that the majority of P2 purinergic receptors are expressed and exhibit functional responses to ATP and UTP in mouse and human CPCs. Since previous findings have shown that the G protein-coupled P2Y 2 receptor (P2Y 2 R) induces cardioprotective responses in animal models as well as human cardiomyocytes and regulates a wide range of signaling pathways that are crucial to tissue repair in various experimental models and in stem cells from diverse origins, we aim to determine whether P2Y 2 R plays similar roles in CPCs. Our preliminary data show that the P2Y 2 R agonists ATP and UTP enhance human CPC (hCPC) proliferation, migration and survival. Interestingly, hCPCs that exhibit relatively slower growth kinetics and higher levels of senescence markers show a dramatic decrease in P2Y 2 R expression as compared to fast-growing hCPCs consistent with our hypothesis that overexpressing P2Y 2 R participates in rejuvenating hCPCs and improving their growth capabilities. This hypothesis will be tested in vivo by determining whether P2Y 2 R overexpression in hCPCs improves their reparative potential for injured mouse myocardium. We also introduce the novel hypothesis that P2Y 2 R-induced regenerative responses in hCPCs involve the activation of Hippo signaling that is known to be regulated by different GPCRs, which links the extracellular nucleotides released during cardiac ischemia to extracellular matrix sensing and Hippo signaling that have been recently implicated in cardiac regeneration.

A Perfused Microfluidic System to Study the Differentiation of Neural Stem Cells in vitro

2018 ◽  
Vol 206 (3) ◽  
pp. 157-164 ◽  
Author(s):  
Qingxi Liu ◽  
Sijie Xing ◽  
Yupeng Liu ◽  
Zijiang Zhang ◽  
Lihui Lv ◽  
...  
Keyword(s):  
Stem Cells ◽  
Neural Stem Cells ◽  
Drug Evaluation ◽  
Microfluidic System ◽  
Quantitative Real Time Pcr ◽  
Culture Systems ◽  
Adult Neuron ◽  
Cellular Microenvironments

Introduction: Due to the ability to mimic in vivo cellular microenvironments, the development of multicell culture systems has received increasing interest for use as research models and serving as platforms for drug evaluation. Methods: In this study, we developed a perfused microfluidic system to resemble the in vivo intercellular environment and applied it to study the differentiation from neural stem cells into neurons. Results: As determined by immunofluorescence chemistry and quantitative real-time PCR, the neural stem cells grown in this microfluidic system showed an elevated differentiation rate toward the formation of neurons as determined by the increased level of βIII-tubulin production, which is 4 times higher than that of culturing neural stem cells only. Conclusion: These results revealed that some factors secreted into the intercellular microenvironment by adult neuron cells can stimulate the differentiation of neural stem cells, pointing to the importance of developing multicellular culture systems such as the perfused microfluidic system we report here to better resemble the in vivo situation.

scholarly journals Low frequency electromagnetic field decreases ischemia–reperfusion injury of human cardiomyocytes and supports their metabolic function

2018 ◽  
Vol 243 (10) ◽  
pp. 809-816 ◽  
Author(s):  
Dariusz Biały ◽  
Magdalena Wawrzyńska ◽  
Iwona Bil-Lula ◽  
Anna Krzywonos-Zawadzka ◽  
Agnieszka Sapa-Wojciechowska ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Electromagnetic Field ◽  
Reperfusion Injury ◽  
Cardiac Myocytes ◽  
Metabolic Activity ◽  
Ischemia Reperfusion Injury ◽  
Ischemia Reperfusion ◽  
Low Frequency ◽  
Cardiac Cells ◽  
Human Cardiomyocytes

Electromagnetic field at extremely low frequencies plays a significant role in the physiological function of human tissues and systems. It is shown that electromagnetic field inhibits production of reactive oxygen species which are involved in heart injury triggered by oxidative stress. We hypothesize that low frequency electromagnetic field protects function of cardiac cells from ischemia–reperfusion injury. Human cardiac myocytes, endothelial cells, and cardiac fibroblast underwent ischemia–reperfusion conditions in the presence or in the absence of low frequency electromagnetic field. LDH and MMP-2 activities (as markers of cell injury), and cell metabolic activity (by fluorescein diacetate staining) were measured to determine the protective role of low frequency electromagnetic field. Our data showed that short courses of low frequency electromagnetic field protect cardiac cells from cellular damage and preserve their metabolic activity during ischemia–reperfusion. This study demonstrates the possibility to use of low frequency electromagnetic field as strategy for the prevention or therapy of ischemia–reperfusion injury. Impact statement In our study, we showed that LF-EMF may be protective for heart during ischemia–reperfusion (I/R). Following is the short description of the main findings: (a) the response to the I/R injury was different for endothelial cells, fibroblasts, and cardiomyocytes; (b) I/R decreases MMP-2 activity in cardiac myocytes and fibroblasts; (c) I/R increases MMP-2 activity in endothelial cells; (d) LF-EMF reverses these changes; (e) LF-EMF protects cells from injury and preserves their metabolic activity.

scholarly journals A Micro-Optic Stalk (μOS) System to Model the Collective Migration of Retinal Neuroblasts

Micromachines ◽  
2020 ◽  
Vol 11 (4) ◽  
pp. 363
Author(s):  
Stephanie Zhang ◽  
Miles Markey ◽  
Caroline D. Pena ◽  
Tadmiri Venkatesh ◽  
Maribel Vazquez
Keyword(s):  
Stem Cells ◽  
Visual System ◽  
Neural Stem Cells ◽  
Fruit Fly ◽  
Microfluidic System ◽  
Collective Migration ◽  
Optic Stalk ◽  
Key Factor ◽  
Regenerative Therapies

Contemporary regenerative therapies have introduced stem-like cells to replace damaged neurons in the visual system by recapitulating critical processes of eye development. The collective migration of neural stem cells is fundamental to retinogenesis and has been exceptionally well-studied using the fruit fly model of Drosophila Melanogaster. However, the migratory behavior of its retinal neuroblasts (RNBs) has been surprisingly understudied, despite being critical to retinal development in this invertebrate model. The current project developed a new microfluidic system to examine the collective migration of RNBs extracted from the developing visual system of Drosophila as a model for the collective motile processes of replacement neural stem cells. The system scales with the microstructure of the Drosophila optic stalk, which is a pre-cursor to the optic nerve, to produce signaling fields spatially comparable to in vivo RNB stimuli. Experiments used the micro-optic stalk system, or μOS, to demonstrate the preferred sizing and directional migration of collective, motile RNB groups in response to changes in exogenous concentrations of fibroblast growth factor (FGF), which is a key factor in development. Our data highlight the importance of cell-to-cell contacts in enabling cell cohesion during collective RNB migration and point to the unexplored synergy of invertebrate cell study and microfluidic platforms to advance regenerative strategies.

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