Adipose-derived stem cells combined with Neuregulin-1 delivery systems for heart tissue engineering

Myocardial infarction (MI) is the leading cause of death worldwide and extensive research has therefore been performed to find a cure. Neuregulin-1 (NRG) is a growth factor involved in cardiac repair after MI. We previously described how biocompatible and biodegradable microparticles, which are able to release NRG in a sustained manner, represent a valuable approach to avoid problems related to the short half-life after systemic administration of proteins. The effectiveness of this strategy could be improved by combining NRG with several cytokines involved in cardiac regeneration. The present study investigates the potential feasibility of using NRG-releasing particle scaffold combined with adipose derived stem cells (ADSC) as a multiple growth factor delivery-based tissue engineering strategy for implantation in the infarcted myocardium. NRG-releasing particle scaffolds with a suitable size for intramyocardial implantation were prepared by TROMS. Next, ADSC were adhered to particle scaffolds and their potential for heart administration was assessed in a MI rat model. NRG was successfully encapsulated reaching encapsulation efficiencies of 92.58 ± 3.84 %. NRG maintained its biological activity after the microencapsulation process. ADSC cells adhered efficiently to particle scaffolds within a few hours. The ADSC-cytokine delivery system developed proved to be compatible with intramyocardial administration in terms of injectability through a 23-gauge needle and tissue response. Interestingly, ADSC-scaffolds were present in the peri-infarted tissue two weeks after implantation. This proof of concept study provides important evidence required for future effectiveness studies and for the translation of this approach

Heart regeneration after miocardial infarction using synthetic biomaterials

Myocardial infarction causes almost 7.3 million deaths each year worldwide. However, current treatments are more palliative than curative. Presently, cell and protein therapies are considered the most promising alternative treatments. Clinical trials performed until now have demonstrated that these therapies are limited by protein short half‐life and by low transplanted cell survival rate, prompting the development of novel cell and protein delivery systems able to overcome such limitations. In this review we discuss the advances made in the last 10 years in the emerging field of cardiac repair using biomaterial‐based delivery systems with focus on the progress made on preclinical in vivo studies. Then, we focus in cardiac tissue engineering approaches, and how the incorporation of both cells and proteins together into biomaterials has opened new horizons in the myocardial infarction treatment. Finally, the ongoing challenges and the perspectives for future work in cardiac tissue engineering will also be discussed

Biodegradation and heart retention of polymeric microparticles in a rat model of myocardial ischemia

Poly-lactide-co-glycolide (PLGA) microparticles emerged as one of the most promising strategies to achieve site-specific drug delivery. Although these microparticles have been demonstrated to be effective in several wound healing models, their potential in cardiac regeneration has not yet been fully assessed. The present work sought to explore PLGA microparticles as cardiac drug delivery systems. PLGA microparticles were prepared by Total Recirculation One-Machine System (TROMS) after the formation of a multiple emulsion. Microparticles of different size were prepared and characterized to select the most suitable size for intramyocardial administration. Next, the potential of PLGA microparticles for administration in the heart was assessed in a MI rat model. Particle biodegradation over time and myocardial tissue reaction were studied by routine staining and confocal microscopy. Results showed that microparticles with a diameter of 5 μm were the most compatible with intramyocardial administration in terms of injectability through a 29-gauge needle and tissue response. Particles were present in the heart tissue for up to three months post-implantation and no particle migration towards other solid organs was observed, demonstrating good myocardial retention. CD68 immunolabeling revealed 31%, 47% and below 4% microparticle uptake by macrophages one week, one month and three months after injection, respectively (P<0.001). Taken together, these findings support the feasibility of the developed PLGA microparticles as vehicles for delivering growth factors in the infarcted myocardium

Controlled delivery of fibroblast growth factor-1 and neuregulin-1 from biodegradable microparticles promotes cardiac repair in a rat myocardial infarction model through activation of endogenous regeneration

Acidic fibroblast growth factor (FGF1) and neuregulin-1 (NRG1) are growth factors involved in cardiac development and regeneration. Microparticles (MPs) mediate cytokine sustained release, and can be utilized to overcome issues related to the limited therapeutic protein stability during systemic administration. We sought to examine whether the administration of microparticles (MPs) containing FGF1 and NRG1 could promote cardiac regeneration in a myocardial infarction (MI) rat model. We investigated the possible underlying mechanisms contributing to the beneficial effects of this therapy, especially those linked to endogenous regeneration. FGF1- and NRG1-loaded MPs were prepared using a multiple emulsion solvent evaporation technique. Seventy-three female Sprague-Dawley rats underwent permanent left anterior descending coronary artery occlusion, and MPs were intramyocardially injected in the peri-infarcted zone four days later. Cardiac function, heart tissue remodeling, revascularization, apoptosis, cardiomyocyte proliferation, and stem cell homing were evaluated one week and three months after treatment. MPs were shown to efficiently encapsulate FGF1 and NRG1, releasing the bioactive proteins in a sustained manner. Three months after treatment, a statistically significant improvement in cardiac function was detected in rats treated with growth factor-loaded MPs (FGF1, NRG1, or FGF1/NRG1). The therapy led to inhibition of cardiac remodeling with smaller infarct size, a lower fibrosis degree and induction of tissue revascularization. Cardiomyocyte proliferation and progenitor cell recruitment was detected. Our data support the therapeutic benefit of NRG1 and FGF1 when combined with protein delivery systems for cardiac regeneration. This approach could be scaled up for use in pre-clinical and clinical studies

Clonal chromosomal mosaicism and loss of chromosome Y in elderly men increase vulnerability for SARS-CoV-2

The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2, COVID-19) had an estimated overall case fatality ratio of 1.38% (pre-vaccination), being 53% higher in males and increasing exponentially with age. Among 9578 individuals diagnosed with COVID-19 in the SCOURGE study, we found 133 cases (1.42%) with detectable clonal mosaicism for chromosome alterations (mCA) and 226 males (5.08%) with acquired loss of chromosome Y (LOY). Individuals with clonal mosaic events (mCA and/or LOY) showed a 54% increase in the risk of COVID-19 lethality. LOY is associated with transcriptomic biomarkers of immune dysfunction, pro-coagulation activity and cardiovascular risk. Interferon-induced genes involved in the initial immune response to SARS-CoV-2 are also down-regulated in LOY. Thus, mCA and LOY underlie at least part of the sex-biased severity and mortality of COVID-19 in aging patients. Given its potential therapeutic and prognostic relevance, evaluation of clonal mosaicism should be implemented as biomarker of COVID-19 severity in elderly people. Among 9578 individuals diagnosed with COVID-19 in the SCOURGE study, individuals with clonal mosaic events (clonal mosaicism for chromosome alterations and/or loss of chromosome Y) showed an increased risk of COVID-19 lethality

Novel tissue engineering strategies based on the combination of the polymeric devices and adult stem cells for cardiac repair

Cardiovascular diseases (CVD) are the leading cause of death worldwide. Among them, myocardial infarction (MI) is the main CVDs, causing 7.4 million deaths each year. The current therapies for MI are palliative rather than regenerative. Since that, the tissue engineering (TE) strategy has grown investigators attention. With the aim of improving heart repair after MI, we have developed two TE strategies. The first strategy was the use of poly(lactic co-glycolic acid) (PLGA) microparticles (MPs) containing neuregulin (NRG), as support for attaching adipose-derived stem cells (ADSC): ADSC-NRG-MP. The second strategy was the combination of two different polymeric devices, MPs and hydrogels. The MPs loaded with NRG were embedded together with the ADSCs in hydrogels composed of different ratios of dextran (Dex) and hyaluronic acid (HA). Our hypothesis was that the developed systems would increase cell survival and would activate different pathways to favor heart regeneration. The ADSC-NRG-MP induced a more pronounced regeneration of the infarcted heart in a rat MI model (reduction in infarct size, higher left ventricle thickness and vasculogenesis). Also the employment of MPs as support for the ADSC, favored a long-term survival of the cells once in the tissue, being detectable three months after their administration. Moreover, among the hydrogels developed, the 50:50 Dex:HA hydrogel embedding 1 mg of NRG-MP and 500,000 ADSC showed to present the best characteristics (adequate stiffness for heart administration, prolonged gelation time to allow its injectability and degradation rate to allow cell survival). To conclude, the two strategies developed have shown to be promising candidates for heart repair after a MI

Novel tissue engineering strategies based on the combination of the polymeric devices and adult stem cells for cardiac repair

Cardiovascular diseases (CVD) are the leading cause of death worldwide. Among them, myocardial infarction (MI) is the main CVDs, causing 7.4 million deaths each year. The current therapies for MI are palliative rather than regenerative. Since that, the tissue engineering (TE) strategy has grown investigators attention. With the aim of improving heart repair after MI, we have developed two TE strategies. The first strategy was the use of poly(lactic co-glycolic acid) (PLGA) microparticles (MPs) containing neuregulin (NRG), as support for attaching adipose-derived stem cells (ADSC): ADSC-NRG-MP. The second strategy was the combination of two different polymeric devices, MPs and hydrogels. The MPs loaded with NRG were embedded together with the ADSCs in hydrogels composed of different ratios of dextran (Dex) and hyaluronic acid (HA). Our hypothesis was that the developed systems would increase cell survival and would activate different pathways to favor heart regeneration. The ADSC-NRG-MP induced a more pronounced regeneration of the infarcted heart in a rat MI model (reduction in infarct size, higher left ventricle thickness and vasculogenesis). Also the employment of MPs as support for the ADSC, favored a long-term survival of the cells once in the tissue, being detectable three months after their administration. Moreover, among the hydrogels developed, the 50:50 Dex:HA hydrogel embedding 1 mg of NRG-MP and 500,000 ADSC showed to present the best characteristics (adequate stiffness for heart administration, prolonged gelation time to allow its injectability and degradation rate to allow cell survival). To conclude, the two strategies developed have shown to be promising candidates for heart repair after a MI

New tools for genome preservation: Grafting germinal cells in brown trout (Salmo trutta)

Primordial germ cells (PGCs) are the only cells in developing embryos with the potential to transmit genetic information to the next generation, making them excellent candidates for gene banking and cryopreservation (Saito et al., 2008). PGCs originate from the germ cell line, producing spermatogonia and oogonia, which, in turn, preserve two main functions: self-renewal and production of gametes. The potential of primordial germ cells to resume gametogenesis after intra- or inter-specific grafting has been reported for a limited number of species during the last years (Takeuchi et al., 2004; Yoshizaki et al., 2010). These findings opened great expectations for the development of new biotechnologies in aquaculture, the possibility to generate germ-line quimeras for surrogate production being one of the most suggested option. Recent studies showed that this ability is maintained in more differentiated cells of the germinal line, the spermatogonia (Okutsu et al., 2006; Lacerda et al., 2010; Yazawa et al., 2010), which are easier to obtain from immature testes. The objective of the present study was, therefore, to evaluate the use of these tools in the conservation programmes of brown trout populations from the Northwest of Spain, which are threatened by extinction. In this work we wanted to check the effectiveness of the protocols for testes dissociation and spermatogonia enrichment of this species and evaluate the ability of cells to colonize the host gonads after intra-specific grafting.Research supported by the Spanish Ministry of Science and Innovation, PGCFISH (AGL2008-02172) project.Peer Reviewe

Stabilization of sperm membrane lipids during cryopreservation by the addition of AFPs

Trabajo presentado en el 3rd International Workshop on Biology of Fish Gametes, celebrado en Budapest del 7 al 9 de septiembre de 2011.Changes in the plasma membrane lipid composition during cryopreservation have been related to a decreasein sperm quality (Muller et al., 2008). In the present work we tested the addition of antifreeze proteins(AFPs) to the cryopreservation extender because of their ability to depress the freezing point and their potential interaction with membranesThis work was supported by the project Petri (PTR 95.1026.OP.01). J. Beirão was supported by a PhD grant from the Portuguese Ministry of Science and Technology (SFRH/BD/31990/2006) co-foundedby POPH - QREN (FEDER and MCTES).Peer Reviewe

Fatty acid composition of the head membrane and flagella affects Sparus aurata sperm quality

Special Issue: Proceedings of the Third International Workshop on the “Biology of Fish Gametes” : Budapest, Hungary, 7–9 September 2011.The successful fertilization achievement is, theoretically, affected by the fatty acids composition of the different sperm membrane domains, which has effects on plasma membrane fluidity, fusogenicity, signal transduction, and other processes. In the present study we analyzed the fatty acids composition of two different membrane domains (head membrane and flagella) of Sparus aurata sperm samples, and its relation to other quality parameters: motility and viability. Results showed that flagella had a higher rate of unsaturated (MUFA and PUFA) fatty acids compared with the head membrane. While flagellar fatty acid composition correlated with motility parameters, suggesting their relation with the need of fluidity for the flagellar beating, head membrane composition better correlated with preservation of sperm membrane integrity. © 2012 Blackwell Verlag GmbH.This work was supported by the project Petri (PTR 95.1026.OP.01). J. Beirão was supported by a PhD grant from the Portuguese Ministry of Science and Technology (SFRH/BD/31990/2006) co-founded by POPH – QREN (FEDER and MCTES).Peer Reviewe