Early cell loss associated with mesenchymal stem cell cardiomyoplasty

Background: Human mesenchymal stem cells (hMSCs) show potential for therapeutic cellular cardiomyoplasty. However, a range of delivery methods, including direct intramyocardial injection, have resulted in poor engraftment in vivo. We used the in vivo rat heart model to study hMSC engraftment and retention in a normal beating heart. Materials and Methods: HMSCs transfected with green fluorescent protein were injected into the left ventricle (LV) of immunocompetent rats. Hearts were cryopreserved 30 minutes (Group A), 24 hours (Group B), and 5 days (Group C) post hMSC delivery. HMSC retention was estimated using confocal fluorescence microscopy and immunohistochemistry. Measured values were compared to projected GFP-positive cellular volumes. Immunohistochemical analyses probed for the presence of human cells with human prolyl hydroxylase beta (p4hβ) and an immune response with murine monocyte/macrophage antigen (CD68). Results: HMSC retention decreased significantly from 30 minutes to 5 days (p<0.05). In Group A the projected GFP positive cellular volume of 31% correlated with measured values and was significantly greater than the 1% predicted cellular volume in Group C. Moreover, human p4hβ was detected in Groups A and B, and not in Group C. Conversely, CD68 was detected in Groups B and C and not in Group A. Conclusions: In immunocompetent rats, engraftment and retention of hMSCs delivered intramyocardially significantly declines over a five day period. The influx of monocytes/macrophages suggests an unfavorable micro-environment for exogenous stem cell survival, confirmed by the absence of human cells detected five days post injection

Choice of Cell-Delivery Route for Skeletal Myoblast Transplantation for Treating Post-Infarction Chronic Heart Failure in Rat

Intramyocardial injection of skeletal myoblasts (SMB) has been shown to be a promising strategy for treating post-infarction chronic heart failure. However, insufficient therapeutic benefit and occurrence of ventricular arrhythmias are concerns. We hypothesised that the use of a retrograde intracoronary route for SMB-delivery might favourably alter the behaviour of the grafted SMB, consequently modulating the therapeutic effects and arrhythmogenicity.Three weeks after coronary artery ligation in female wild-type rats, 5x10(6) GFP-expressing SMB or PBS only (control) were injected via either the intramyocardial or retrograde intracoronary routes. Injection of SMB via either route similarly improved cardiac performance and physical activity, associated with reduced cardiomyocyte-hypertrophy and fibrosis. Grafted SMB via either route were only present in low numbers in the myocardium, analysed by real-time PCR for the Y-chromosome specific gene, Sry. Cardiomyogenic differentiation of grafted SMB was extremely rare. Continuous ECG monitoring by telemetry revealed that only intramyocardial injection of SMB produced spontaneous ventricular tachycardia up to 14 days, associated with local myocardial heterogeneity generated by clusters of injected SMB and accumulated inflammatory cells. A small number of ventricular premature contractions with latent ventricular tachycardia were detected in the late-phase of SMB injection regardless of the injection-route.Retrograde intracoronary injection of SMB provided significant therapeutic benefits with attenuated early-phase arrhythmogenicity in treating ischaemic cardiomyopathy, indicating the promising utility of this route for SMB-delivery. Late-phase arrhythmogenicity remains a concern, regardless of the delivery route

Improved pharmacokinetic and lymphatic uptake of Rose Bengal after transfersome intradermal deposition using hollow microneedles

The lymphatic system is active in several processes that regulate human diseases, among which cancer progression stands out. Thus, various drug delivery systems have been investigated to promote lymphatic drug targeting for cancer therapy; mainly, nanosized particles in the 10–150 nm range quickly achieve lymphatic vessels after an interstitial administration. Herein, a strategy to boost the lymphotropic delivery of Rose Bengal (RB), a hydrosoluble chemotherapeutic, is proposed, and it is based on the loading into Transfersomes (RBTF) and their intradermal deposition in vivo by microneedles. RBTF of 96.27 ± 13.96 nm (PDI = 0.29 ± 0.02) were prepared by a green reverse-phase evaporation technique, and they showed an RB encapsulation efficiency of 98.54 ± 0.09%. In vitro, RBTF remained physically stable under physiological conditions and avoided the release of RB. In vivo, intravenous injection of RBTF prolonged RB half-life of 50 min in healthy rats compared to RB intravenous injection; the RB half-life in rat body was further increased after intradermal injection reaching 24 h, regardless of the formulation used. Regarding lymphatic targeting, RBTF administered intravenously provided an RB accumulation in the lymph nodes of 12.3 ± 0.14 ng/mL after 2 h, whereas no RB accumulation was observed after RB intravenous injection. Intradermally administered RBTF resulted in the highest RB amount detected in lymph nodes after 2 h from the injection (84.2 ± 25.10 ng/mL), which was even visible to the naked eye based on the pink colouration of the drug. In the case of intradermally administered RB, RB in lymph node was detected only at 24 h (13.3 ± 1.41 ng/mL). In conclusion, RBTF proved an efficient carrier for RB delivery, enhancing its pharmacokinetics and promoting lymph-targeted delivery. Thus, RBTF represents a promising nanomedicine product for potentially facing the medical need for novel strategies for cancer therapy.<br/

Dissolving and Swelling Hydrogel-Based Microneedles: An Overview of Their Materials, Fabrication, Characterization Methods, and Challenges

YesPolymeric hydrogels are a complex class of materials with one common feature—the ability to form three-dimensional networks capable of imbibing large amounts of water or biological fluids without being dissolved, acting as self-sustained containers for various purposes, including pharmaceutical and biomedical applications. Transdermal pharmaceutical microneedles are a pain-free drug delivery system that continues on the path to widespread adoption—regulatory guidelines are on the horizon, and investments in the field continue to grow annually. Recently, hydrogels have generated interest in the field of transdermal microneedles due to their tunable properties, allowing them to be exploited as delivery systems and extraction tools. As hydrogel microneedles are a new emerging technology, their fabrication faces various challenges that must be resolved for them to redeem themselves as a viable pharmaceutical option. This article discusses hydrogel microneedles from a material perspective, regardless of their mechanism of action. It cites the recent advances in their formulation, presents relevant fabrication and characterization methods, and discusses manufacturing and regulatory challenges facing these emerging technologies before their approval

DLRmagazine 170 - Compute before flight

Modern engines should be as quiet and emission-free as possible without compromising on power. For decades, this has been the focus of DLR's aeronautics research institutes, who have been working on developing better engines and testing them using various test benches. But for some time now, there have been new components to this research – simulations and digital models. These innovations allow researchers to digitally test new concepts in advance without the need for costly prototypes, speeding up the development of new engine designs. In this issue, DLR researchers Stanislaus Reitenbach and Kai Becker talk about the role that the virtual engine could play in the future

Identification en thérapie cellulaire des patrons d’expression transcriptomique de cellules souches utilisées pour traiter la défaillance cardiaque afin d’en améliorer le potentiel thérapeutique

La cardiopathie ischémique incluant l’insuffisance cardiaque est la deuxième cause de mortalité annuelle au Canada. Bien que de nombreuses stratégies préventives et des thérapies pharmacologiques retardent la progression de la maladie, il n’existe aucune solution qui module directement aux les remaniements pathologiques et la perte de cardiomyocytes. Au cours des 25 dernières années, de multiples progrès dans les domaines de la médecine régénérative et de la thérapie cellulaire ont annoncé des résultats prometteurs, mais les résultats d’études cliniques contemporaines demeurent plutôt mitigés. COMPARE-AMI, une étude randomisée-contrôlée de phase II, a évalué l’effet d’injections intracoronariennes de cellules souches hématopoïétiques CD133+ chez des patients souffrant d’infarctus aigu. IMPACT-CABG, une ÉRC de phase II a également évalué l’effet d’injections intramyocardiques de cellules CD133+ chez les patients souffrant de cardiomyopathie ischémique chronique nécessitant une revascularisation chirurgicale. Nous avons émis l’hypothèse que les cellules CD133+ utilisées dans des études cliniques de cardiomyopathies ischémiques aiguës et chroniques des patients répondant à la thérapie cellulaire exhibent des signatures transcriptomiques communes responsables de leur effet thérapeutique. En classant les patients en tant que répondants et non-répondants selon leur fonction cardiaque, nous avons évalué, a posteriori, ces patrons d’expression. Les cellules CD133+ autologues de patients jugés répondants expriment des signatures qui sont hautement conservées entre elles (incluant l’angiogénèse, la régulation de la réponse au stress et la survie cellulaire) et uniques d’un modèle à l’autre et qui pourraient, en partie, exprimer les issus cliniques des patients. Afin de maximiser les effets de la thérapie cellulaire aux cellules souches, nous avons par la suite tenté de reproduire ces phénotypes par stimulation pharmacologique avec des inhibiteurs d’HSP90 pour leurs effets qui semblent reproduire ces signatures. Ainsi, nous avons démontré qu’une stimulation de cellules souches mésenchymateuses humaines (CSMh) au Célastrol (inhibiteur HSP90) pouvait répliquer certains de ces phénotypes. Notamment, des CSMh conditionnées activent des voies de signalisation de type ‘RISK’ et augmentent leur sécrétion de protéines en lien avec la réponse au stress ainsi que d’exosomes contenant des molécules impliquées dans la communication intercellulaire sans être liées à un changement de type cellulaire. De plus, les CSMh traitées semblent améliorer la guérison de plaie par activité paracrine et sont plus résistante à la sénescence oxydative. Ces résultats encourageants nous permettent d’envisager des stratégies plus poussées de pré-conditionnement cellulaire ex vivo de cellules CD133+ avant leur implantation. À terme, cela pourrait mener à une optimisation de la thérapie cellulaire afin d’en maximiser les bénéfices cliniques et d’en exploiter leur plein potentiel.Ischemic cardiomyopathy and heart failure are the second annual cause of mortality in Canada. Despite rigorous prevention strategies and drug regimens preventing progression, no therapeutic modality can currently reverse the pathologic modifications of the disease. In the last quarter century, numerous advances in the field of regenerative medicine and cell therapy have demonstrated promising effects. COMPARE-AMI, a phase II randomized controlled trial (RCT), evaluated the effect of intracoronary injection of CD133+ cells in acute myocardial infarction following percutaneous intervention. IMPACTCABG, also a phase II RCT, evaluated the effect of intramyocardial injection of CD133+ hematopoietic stem cells in chronic ischemic cardiomyopathy at the time of surgical revascularization. That being said, we believe that the CD133+ cells used in therapy have shared transcriptomic signatures that are responsible for their clinical effects. By classifying patients into responders and non-responders according to an improvement in ejection fraction, we evaluated, a posteriori, those expression patterns. Autologous CD133+ cells of patients classified as responders expressed highly conserved transcriptomic signatures that could be responsible for the clinical outcomes of patients. Notably, these signatures were related to cardioprotective mechanisms including angiogenesis, stress response regulation and cell survival. In order to harness the full potential of stem cell therapy, we attempted to reproduce the identified phenotypes by pharmacological intervention with HSP90 inhibitors which are known to mimic some of these effets. Conditioned human mesenchymal stem cells (hMSC) activated ‘RISK’-type signaling pathways and augmented exosome and protein secretion relating to the response to cellular stress; this activation was unrelated to a switch of cell type. Furthermore, treated hMSC seemed to favour improved wound healing by paracrine activity and were more resistant to oxidative senescence. These encouraging results allow us to develop novel, more advance, strategies of ex vivo cell preconditioning before implantation, including of CD133+ cells. Ultimately, we hope that optimisation of cells through this mechanism and others will allow us to unleash the full potential of stem cell therapy

Optimising cell therapy for treating heart failure

Cell therapy is a promising strategy for treating ischaemic chronic heart failure. However, its therapeutic efficacy has not been fully established. In addition, arrhythmia occurrence is a concern of this treatment. In this work, therapeutic benefits, arrhythmogenicity and underlying mechanisms, which were hypothesised to be modulated by cell-delivery route into the heart, were investigated with the aim of optimising cell therapy. Injection of either skeletal myoblasts or mononuclear bone marrow cells into the rat ischaemic chronically failing heart via either direct intramyocardial or retrograde intracoronary route similarly improved both cardiac function and physical activity over the 84 days analysed. Survival of the grafted cells in the myocardium was extremely poor and trans-differentiation or fusion of the grafted cells into cardiomyocytes or vessels were only rarely identified via either celldelivery route. Therefore, paracrine effects including increased neovascular formation and attenuated fibrosis in the myocardium were considered to play an important role in the therapeutic benefits of cell therapy using either cell-type. Of note, direct intramyocardial injection of either cell-type, but not retrograde intracoronary injection, produced spontaneous ventricular arrhythmias including ventricular tachycardia in the early periods following cell injection. Local heterogeneity in the myocardium induced by clusters of grafted cells, which also involved inflammatory response, was considered to be a cause of the arrhythmias following intramyocardial cell injection. In contrast, in the late periods, injection of skeletal myoblasts via either route, but not mononuclear bone marrow cells, caused latent ventricular tachycardia possibly via regression of connexin43 in the native myocardium. Efficiency of engraftment of mononuclear bone marrow cells in the myocardium following intracoronary injection was very poor in the normal heart, but was enhanced by induction of ischaemia-reperfusion prior to cell injection. Most of the enhanced cell-engraftment was dependent on P-selectin-mediated cellular interaction between donor cells and endothelium