scholarly journals Cardiac aorta-derived extracellular matrix scaffold enhances critical mediators of angiogenesis in isoproterenol-induced myocardial infarction mice

Author(s):  
Mahara Hosseinabadi ◽  
Zohreh Abdolmaleki ◽  
Seyed Hamed Shirazi Beheshtiha

AbstractAn incapability to improve lost cardiac muscle caused by acute ischemic injury remains the most important deficiency of current treatments to prevent heart failure. We investigated whether cardiomyocytes culturing on cardiac aorta-derived extracellular matrix scaffold has advantageous effects on cardiomyocytes survival and angiogenesis biomarkers’ expression. Ten male NMRI mice were randomly divided into two groups: (1) control (healthy mice) and (2) myocardial infarction (MI)-induced model group (Isoproterenol/subcutaneously injection/single dose of 85 mg/kg). Two days after isoproterenol injection, all animals were sacrificed to isolate cardiomyocytes from myocardium tissues. The fresh thoracic aorta was obtained from male NMRI mice and decellularized using 4% sodium deoxycholate and 2000 kU DNase-I treatments. Control and MI-derived cardiomyocytes were seeded on decellularized cardiac aorta (DCA) considered three-dimensional (3D) cultures. To compare, the isolated cardiomyocytes from control and MI groups were also cultured as a two-dimensional (2D) culture system for 14 days. The cell viability was examined by MTT assay. The expression levels of Hif-1α and VEGF genes and VEGFR1 protein were tested by real-time PCR and western blotting, respectively. Moreover, the amount of VEGF protein was evaluated in the conditional media of the 2D and 3D systems. The oxidative stress was assessed via MDA assay. Hif-1α and VEGF genes were downregulated in MI groups compared to controls. However, the resulting data showed that decellularized cardiac aorta matrices positively affect the expression of Hif-1α and VEGF genes. The expression level of VEGFR1 protein was significantly (p ≤ 0.01) upregulated in both MI and healthy cell groups cultured on decellularized cardiac aorta matrices as a 3D system compared to the MI cell group cultured in the 2D systems. Furthermore, MDA concentration significantly decreased in 3D-cultured cells (MI and healthy cell groups) rather than the 2D-cultured MI group (p ≤ 0.015). The findings suggest that cardiac aorta-derived extracellular scaffold by preserving VEGF, improving the cell viability, and stimulating angiogenesis via upregulating Hif-1α, VEGF, and VEGFR1 in cardiomyocytes could be considered as a potential approach along with another therapeutic method to reduce the complications of myocardial infarction and control the progressive pathological conditions related to MI.

2020 ◽  
Vol 17 (1) ◽  
pp. 11-17 ◽  
Author(s):  
Xiancan Wang ◽  
Yuqiang Shang ◽  
Shilin Dai ◽  
Wei Wu ◽  
Fan Yi ◽  
...  

Purpose: Myocardial infarction is a common cardiovascular disease. MicroRNA-16-5p (miR-16-5p) was upregulated in heart and kidney hypoxia/reoxygenation (H/R) injury. However, the role of miR-16-5p in myocardial infarction injury is still unclear. Methods: Human adult ventricular cardiomyocytes (AC16) were treated with ischemia/reperfusion (H/R). The miR-16-5p level was evaluated through real-time PCR. The activity of lactate dehydrogenase (LDH) and creatine kinase-MB (CK-MB) was detected via LDH and CK-MB monitoring kits. Cell viability was examined with 3-(4,5-dimethylthiazol-2-yl)-2,5diphenyltetra-zolium bromide (MTT) assay. Western blotting was used to analyze the protein levels. The luci-ferase report assay confirmed the relative luciferase activity. Results: miR-16-5p was elevated in H/R-treated AC16 cells. miR-16-5p overexpression and knockdown were carried out. miR-16-5p knockdown repressed cell apoptosis, attenuated LDH and CK-MB activities, and enhanced cell viability in H/R-treated AC16 cells. Moreover, miR-16-5p knockdown promoted angiogenesis in human microvascular endothelial cells (HMVEC), causing elevation of vascular endothelial growth factor (VEGF), insulin receptor substrates 1 (IRS1), minichromosome maintenance complex component 2 (MCM2) and proliferating cell nuclear antigen (PCNA) protein levels. Moreover, miR-16-5p was testified to target IRS1. IRS1 silencing alleviated miR-16-5p knockdown-mediated inhibition of apoptosis in AC16 cells. Conclusion: miR-16-5p knockdown increased cell viability and angiogenesis, as well as inhibited cell apoptosis by increasing IRS1. These findings indicated that miR-16-5p knockdown may be a new therapeutic target for myocardial infarction.


2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Naresh Polisetti ◽  
Anke Schmid ◽  
Ursula Schlötzer-Schrehardt ◽  
Philip Maier ◽  
Stefan J. Lang ◽  
...  

AbstractAllogenic transplants of the cornea are prone to rejection, especially in repetitive transplantation and in scarred or highly vascularized recipient sites. Patients with these ailments would particularly benefit from the possibility to use non-immunogenic decellularized tissue scaffolds for transplantation, which may be repopulated by host cells in situ or in vitro. So, the aim of this study was to develop a fast and efficient decellularization method for creating a human corneal extracellular matrix scaffold suitable for repopulation with human cells from the corneal limbus. To decellularize human donor corneas, sodium deoxycholate, deoxyribonuclease I, and dextran were assessed to remove cells and nuclei and to control tissue swelling, respectively. We evaluated the decellularization effects on the ultrastructure, optical, mechanical, and biological properties of the human cornea. Scaffold recellularization was studied using primary human limbal epithelial cells, stromal cells, and melanocytes in vitro and a lamellar transplantation approach ex vivo. Our data strongly suggest that this approach allowed the effective removal of cellular and nuclear material in a very short period of time while preserving extracellular matrix proteins, glycosaminoglycans, tissue structure, and optical transmission properties. In vitro recellularization demonstrated good biocompatibility of the decellularized human cornea and ex vivo transplantation revealed complete epithelialization and stromal repopulation from the host tissue. Thus, the generated decellularized human corneal scaffold could be a promising biological material for anterior corneal reconstruction in the treatment of corneal defects.


2021 ◽  
Vol 22 (8) ◽  
pp. 3901
Author(s):  
Mohsen Setayeshmehr ◽  
Shahzad Hafeez ◽  
Clemens van Blitterswijk ◽  
Lorenzo Moroni ◽  
Carlos Mota ◽  
...  

Various hydrogel systems have been developed as biomaterial inks for bioprinting, including natural and synthetic polymers. However, the available biomaterial inks, which allow printability, cell viability, and user-defined customization, remains limited. Incorporation of biological extracellular matrix materials into tunable synthetic polymers can merge the benefits of both systems towards versatile materials for biofabrication. The aim of this study was to develop novel, cell compatible dual-component biomaterial inks and bioinks based on poly(vinyl alcohol) (PVA) and solubilized decellularized cartilage matrix (SDCM) hydrogels that can be utilized for cartilage bioprinting. In a first approach, PVA was modified with amine groups (PVA-A), and mixed with SDCM. The printability of the PVA-A/SDCM formulations cross-linked by genipin was evaluated. On the second approach, the PVA was functionalized with cis-5-norbornene-endo-2,3-dicarboxylic anhydride (PVA-Nb) to allow an ultrafast light-curing thiol-ene cross-linking. Comprehensive experiments were conducted to evaluate the influence of the SDCM ratio in mechanical properties, water uptake, swelling, cell viability, and printability of the PVA-based formulations. The studies performed with the PVA-A/SDCM formulations cross-linked by genipin showed printability, but poor shape retention due to slow cross-linking kinetics. On the other hand, the PVA-Nb/SDCM showed good printability. The results showed that incorporation of SDCM into PVA-Nb reduces the compression modulus, enhance cell viability, and bioprintability and modulate the swelling ratio of the resulted hydrogels. Results indicated that PVA-Nb hydrogels containing SDCM could be considered as versatile bioinks for cartilage bioprinting.


2010 ◽  
Vol 2010 ◽  
pp. 1-4 ◽  
Author(s):  
Jirapa Chetsawang ◽  
Piyarat Govitrapong ◽  
Banthit Chetsawang

It has been reported that overproduction of reactive oxygen species occurs after brain injury and mediates neuronal cells degeneration. In the present study, we examined the role of Ras signaling on hydrogen peroxide-induced neuronal cells degeneration in dopaminergic neuroblastoma SH-SY5Y cells. Hydrogen peroxide significantly reduced cell viability in SH-SY5Y cultured cells. An inhibitor of the enzyme that catalyzes the farnesylation of Ras proteins, FTI-277, and a competitive inhibitor of GTP-binding proteins, GDP-beta-S significantly decreased hydrogen peroxide-induced reduction in cell viability in SH-SY5Y cultured cells. The results of this study might indicate that a Ras-dependent signaling pathway plays a role in hydrogen peroxide-induced toxicity in neuronal cells.


2000 ◽  
Vol 1 (4) ◽  
pp. 369-378 ◽  
Author(s):  
Marwan E El-Sabban ◽  
Khaled A Hassan ◽  
Adel E Birbari ◽  
Khalil M Bitar ◽  
Anwar B Bikhazi

2015 ◽  
Vol 35 (suppl_1) ◽  
Author(s):  
Irene Cuadrado ◽  
Maria Jose Garcia Miguel ◽  
Irene Herruzo ◽  
Mari Carmen Turpin ◽  
Ana Martin ◽  
...  

Extracellular matrix metalloproteinase inducer EMMPRIN, is highly expressed in patients with acute myocardial infarction (AMI), and induces activation of several matrix metalloproteinases (MMPs), including MMP-9 and MMP-13. To prevent Extracellular matrix degradation and cardiac cell death we targeted EMMPRIN with paramagnetic/fluorescent micellar nanoparticles with an EMMPRIN binding peptide AP9 conjugated (NAP9), or an AP9 scramble peptide as a negative control (NAPSC). NAP9 binds to endogenous EMMPRIN as detected by confocal microscopy of cardiac myocytes and macrophages incubated with NAP and NAPSC in vitro, and in vivo in mouse hearts subjected to left anterior descending coronary artery occlusion (IV injection 50mγ/Kg NAP9 or NAP9SC). Administration of NAP9 at the same time or 1 hour after AMI reduced infarct size over a 20% respect to untreated and NAPSC injected mice, recovered left ventricle ejection fraction (LVEF) similar to healthy controls, and reduced EMMPRIN downstream MMP9 expression. In magnetic resonance scans of mouse hearts 2 days after AMI and injected with NAP9, we detected a significant gadolinium enhancement in the left ventricle respect to non-injected mice and to mice injected with NAPSC. Late gadolinium enhancement assays exhibited NAP9-mediated left ventricle signal enhancement as early as 30 minutes after nanoprobe injection, in which a close correlation between the MRI signal enhancement and left ventricle infarct size was detected. Taken together, these results point EMMPRIN targeted nanoprobes as a new tool for the treatment of AMI.


2015 ◽  
Vol 117 (suppl_1) ◽  
Author(s):  
Brisa Pena ◽  
Valentina Martinelli ◽  
Susanna Bosi ◽  
Carmen Sucharov ◽  
Mark Jeong ◽  
...  

Background: Advances in cell therapy and material science have made tissue engineering a promising strategy for heart regeneration. We developed an injectable biomimetic reverse thermal gel (RTG) that is liquid at room temperature but gel-like at body temperature, with the ultimate goal of being able to serve as a vehicle for cell-based delivery (liquid) to targeted tissue areas (gel-phase at 37°C). In this study we tested the suitability of this biomimetic RTG on cell viability. Methods and results: We tested different biomimetic RTG systems with and without the chemical incorporation of lysine. In vitro 3D culture experiments were performed with neonatal rat ventricular myocytes (NRVM) by mixing 3x104 cells with 50 μl of polymeric solution and allowing gel formation at 37°C. The cultured cells were incubated for 21 days. For controls we used NRVMs plated on 2D traditional gelatin coated dishes. We found that the 3D polymeric matrix induces rapid coordinated contraction with improved functionality when compared with standard 2D-cultured NRVM. By immunostaining for the morphology of the sarcomere (alpha-actinin) and DAPI, we also observed that the 3D polymeric matrix stimulates cells to spread and form 3D syncytia. Conclusion: These proof-of-concept results demonstrate long-term cell viability in this unique biomimetic system and therefore provide feasibility of a polymeric cell delivery system that permits reversible liquid-to-gel transition at body temperature. These results offer potential for a tissue engineering approach to cardiac regeneration.


2001 ◽  
Vol 114 (1) ◽  
pp. 187-197 ◽  
Author(s):  
C. Unsold ◽  
M. Hyytiainen ◽  
L. Bruckner-Tuderman ◽  
J. Keski-Oja

Latent TGF-beta binding proteins (LTBPs) are components of the extracellular matrix (ECM). They belong to the fibrillin/LTBP-superfamily, and are high molecular weight glycoproteins characterized by EGF-like repeats and 8-Cys repeats. Most LTBPs associate with the small latent forms of TGF-beta. Their roles include to facilitate the secretion of latent TGF-beta and to target it to the ECM. In order to identify new matrix-binding domains of LTBP-1 and to characterize their association with the extracellular matrix, we have produced (in a mammalian expression system) partly overlapping recombinant fragments of its shorter form, LTBP-1S, and analyzed the binding of the purified fusion proteins to extracellular matrices of cultured human dermal and lung fibroblasts. Recombinant fragments from three different regions of the N- and C-termini showed affinity to the matrix. These interacting regions contain either the first (hybrid), second or fourth 8-Cys domains of the LTBP-1S molecule. They bound independently to the matrix. Each of them had an ability to inhibit the association of native exogenous LTBP-1 with fibroblast extracellular matrix. The interactions of the LTBP-1 fragments with the extracellular matrix resisted treatment with sodium deoxycholate, suggesting strong, possibly covalent binding. The binding occurred in a time- and dose-dependent fashion. The N-terminal fragments bound more readily to the matrices. With all fragments the binding took place both with intact fibroblast matrices and with matrices isolated by sodium deoxycholate. When using CHO cell layers, which form sparse matrices, only the N-terminal fragment of LTBP-1 was efficiently incorporated. The association of the binding fragments with isolated matrices was enhanced by soluble, cell-derived factors. The current data suggest that LTBP-1 contains three different domains with an ability to associate with the extracellular matrix.


Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Marcin Dobaczewski ◽  
Marcin Bujak ◽  
Carlos Gonzalez ◽  
Na Li ◽  
Xiao-Fan Wang ◽  
...  

We have recently demonstrated that the Transforming Growth Factor (TGF)-β/Smad3 pathway is activated in healing infarcts and plays an essential role in the pathogenesis of cardiac remodeling. Smad3 −/− mice were protected from the development of ventricular dilation following infarction and exhibited markedly reduced fibrosis of the peri-infarct area and the remodeling non-infarcted heart. Accordingly, we hypothesized that Smad3 signaling plays an essential role in regulating cardiac fibroblast function and gene expression in myocardial infarction. Surprisingly, Smad3 −/− infarcts exhibited increased peak infiltration with myofibroblasts, associated with evidence of enhanced proliferative activity. Smad3 −/− mice had a higher density of Ki-67-positive proliferating myofibroblasts in the infarcted myocardium in comparison with wildtype (WT) animals (Smad3−/− 917±291 cells/mm 2 vs. WT 614±115 cells/mm 2 , p<0.05). In vitro experiments suggested that TGF-β inhibits murine cardiac fibroblast proliferation in a concentration-dependent manner and that the antiproliferative effects of TGF-β are abrogated in Smad3 −/− fibroblasts. On the other hand Smad3 signaling was essential for extracellular matrix protein synthesis by cardiac fibroblasts. TGF-β-mediated induction of procollagen type III and of the matricellular protein tenascin-C in cardiac fibroblasts was dependent on Smad3. In addition, TGF-β-induced Tissue Inhibitor of Metalloproteinases (TIMP)-1 and -2 upregulation was also abrogated in Smad3 −/− fibroblasts, suggesting that Smad3 signaling regulates matrix metabolism. In vivo, Smad3 −/− infarcts exhibited attenuated tenascin-C and collagen deposition in the infarct and in the remodeling non-infarcted heart. Our findings suggest that the Smad3 pathway critically regulates fibroblast function in healing myocardial infarction. In Smad3 −/− mice, the healing infarct contains abundant myofibroblasts that exhibit enhanced proliferative activity, but have markedly decreased ability to synthesize extracellular matrix proteins and to produce TIMPs. In the absence of Smad3, attenuated matrix deposition in the remodeling non-infarcted heart results in decreased dilation and ameliorated diastolic dysfunction. This research has received full or partial funding support from the American Heart Association, AHA South Central Affiliate (Arkansas, New Mexico, Oklahoma & Texas).


2006 ◽  
Vol 96 (6) ◽  
pp. 3257-3265 ◽  
Author(s):  
Ekaterina Likhtik ◽  
Joe Guillaume Pelletier ◽  
Andrei T. Popescu ◽  
Denis Paré

This study tested whether firing rate and spike shape could be used to distinguish projection cells from interneurons in extracellular recordings of basolateral amygdala (BLA) neurons. To this end, we recorded BLA neurons in isoflurane-anesthetized animals with tungsten microelectrodes. Projection cells were identified by antidromic activation from cortical projection sites of the BLA. Although most projection cells fired spontaneously at low rates (<1 Hz), an important subset fired at higher rates (up to 6.8 Hz). In fact, the distribution of firing rates in projection cells and unidentified BLA neurons overlapped extensively, even though the latter cell group presumably contains a higher proportion of interneurons. The only difference between the two distributions was a small subset (5.1%) of unidentified neurons with unusually high firing rates (9–16 Hz). Similarly, distributions of spike durations in both cell groups were indistinguishable, although most of the fast-firing neurons had spike durations at the low end of the distribution. However, we observed that spike durations depended on the exact position of the electrode with respect to the recorded cell, varying by as much as 0.7 ms. Thus neither firing rate nor spike waveform allowed for unequivocal separation of projection cells from interneurons. Nevertheless, we propose the use of two firing rate cutoffs to obtain relatively pure samples of projection cells and interneurons: ≤1 Hz for projection cells and ≥7 Hz for fast-spiking interneurons. Supplemented with spike-duration cutoffs of ≥0.7 ms for projection cells and ≤0.5 ms for interneurons, this approach should keep instances of misclassifications to a minimum.


Sign in / Sign up

Export Citation Format

Share Document