Translational cell based therapies to repair the heart

Cardiovascular disease comprising of Coronary Artery Disease (CAD) and Valvular Heart Disease (VHD) represents the leading disease in western societies accounting for the death of numerous patients. CAD may lead to heart failure (HF) and despite the therapeutic options for HF which evolved over the past years, the incidence of HF is continuously increasing with a higher percentage of aged people. Similarly, an increase of VHD can be observed and although valve replacement represents the most common therapy strategy for VHD, approximately 30% of the treated patients are affected from prosthesis-related problems within 10 years. While mechanical valves require lifelong anticoagulation treatment, bioprosthetic valves present with continuous degeneration without the ability to grow, repair or remodel. The concept of regenerative medicine comprising of cell-based therapies, bio engineering technologies and hybrid solutions has been proposed as a promising next generation approach to address CAD and VHD. While myocardial cell therapy has been suggested to have a beneficial effect on the failing myocardium, heart valve tissue engineering has been demonstrated to be a promising concept to generate living, autologous heart valves with the capability to grow and to remodel which may be particularly beneficial for children. Although these regenerative strategies have shown great potential in experimental studies, the translation into a clinical setting has either been limited or has been too rapid and premature leaving many key questions unanswered. The aim of this thesis was the systematic development of translational, cell-based bio engineering concepts addressing CAD (part A) and VHD (part B) with a particular focus on minimally invasive, transcatheter-based implantation techniques. In the setting of myocardial regeneration, in the second chapter the intrinsic regenerative potential of the heart is investigated. Myocardial samples were harvested from all four chambers of the human heart and were assessed for resident stem/progenitor cell populations. The results demonstrated that BRCP+ cells can be detected within the human heart and that they were more abundant than their c-kit+ counterparts. In the non-ischemic heart they were preferentially located in the atria while following ischemia, their numbers were increased significantly in the left ventricle. There were no c-kit+/BCRP+ co-expressing stem/progenitor cell populations suggesting that these two markers are expressed by two distinct cell populations in the human heart. Although these results provided a valuable snapshot at cardiac progenitor cells after acute ischemia, the data also indicated that the absolute numbers of cells acquiring a myocardial phenotype are rather low and further effort is needed to upscale such cells into clinically relevant numbers. In chapter three, it is demonstrated that human bone marrow and adipose tissue derived mesenchymal stem cells can be efficiently isolated via minimally invasive procedures and expanded to clinically relevant numbers for myocardial cell therapy. Thereafter, these cells were tested in a uniquely developed intrauterine, fetal, preimmune ovine myocardial infarction model for the evaluation of human cell fate in vivo. After the successful intrauterine induction of acute myocardial infarction, the cells were intramyocardially transplanted and tracked using a multimodal imaging approach comprising MRI, Micro CT as well as in vitro analysis tools. The principal feasibility of intra-myocardial stem-cell transplantation following intra-uterine induction of myocardial infarction in the preimmune fetal sheep could be demonstrated suggesting this as a unique platform to evaluate human cell-fate in a relevant large animal-model without the necessity of immunosuppressive therapy. In chapter four, adipose tissue derived mesenchymal stem cells (ATMSCs) were processed to generate three dimensional microtissues (3D-MTs) prior to transplantation to address the important issue of cell retention and survival. Thereafter, the ATMSCs based 3D-MTs were transplanted into the healthy and infarcted porcine myocardium using a catheter-based, 3D electromechanical mapping guided approach. The previously used MRI based tracking concept was successfully translated into this preclinical model allowing for the in vivo monitoring of 3D-MTs. To address Valvular Heart Disease (part B), in chapter five, marrow stromal derived cells were used to develop a unique autologous, cell-based engineered heart valve in situ tissue engineering concept comprising of minimally-invasive techniques for both, cell harvest and valve implantation. Autologous marrow stromal derived cells were harvested, seeded onto biodegradable scaffolds and integrated into self-expanding nitinol stents, before they were transapically delivered into the pulmonary position of non-human primates within the same intervention while avoiding any in vitro bio-reactor period. The results of these experiments demonstrated the principal feasibility of generating marrow stromal cell-based, autologous, living tissue engineered heart valves (TEHV) and the transapical implantation in a one-step intervention. In chapter six, this concept was then successfully applied to the high-pressure system of the systemic circulation. After detailed adaption of the TEHV and stent design to the anatomic conditions of an orthotopic aortic valve, marrow stromal cell-based TEHV were implanted into the orthotopic aortic position. The implantation was successful and valve functionality was confirmed using fluoroscopy and trans-esophageal echocardiography. While displaying an ideal opening and closing behaviour with a sufficient co-aptation and a low pressure gradient, there were no signs of coronary occlusion or mal-perfusion. In conclusion, the results of this thesis represent a promising portfolio of translational concepts for cardiovascular regenerative medicine addressing CAD and VHD. In particular, it was demonstrated that mesenchymal stem cells / multipotent stromal derived cells represent a clinically relevant cell source for both myocardial regeneration and heart valve tissue engineering. It was shown that the preimmune fetal sheep myocardial infarction model represents a unique platform for the in vivo evaluation of human stem cells without the necessity of immunosuppressive therapy. Moreover, the concept of transcatheter based intramyocardial transplantation of mesenchymal stem cell-based 3D-MTs was introduced to enhance cellular retention and survival. Finally, in the setting of VHD it could be shown that marrow stromal cell based issue engineered heart valves can successfully generated and transapically implanted into the pulmonary and aortic position within a one-step intervention

The effects of the cardiopulmonary bypass on the gut microbiome and its contribution to postoperative SIRS [Die Effekte der Herz-Lungen-Maschine auf das intestinale Mikrobiom und die Relation zum postoperativen SIRS]

Cardiopulmonary bypass (CPB) made cardiac surgery possible. Despite its almost 70 years of existence and countless design improvements, it still represents one of the most invasive interventions on the human body’s physiological integrity. The adverse effects of CPB present as systemic inflammatory response syndrome (SIRS), which in its most severe form with an incidence between 10% and 20% causes metabolic and immunological mayhem, accounting in many cases for uncontrollable hemodynamic, respiratory, and coagulative instability that may result in high rates of morbidity and mortality. Interestingly, the alterations of the intestinal microbiome during CPB and their role in immune regulation have not been thoroughly investigated. Our scientific efforts aim to identify compositional and metabolic shifts in the microbiome after CPB using metagenomics and metabolomics, to correlate these findings to the postoperative clinical outcomes of the patients, and to reveal a possible mechanistic link to the etiology SIRS. This could generate novel translational and therapeutic approaches for amelioration of SIRS after CPB-assisted cardiac surgery based on microbiome modulation. Furthermore, the detection of specific baseline microbiome compositions prone to SIRS susceptibility may provide a tool for risk stratification

In situ heart valve tissue engineering using a bioresorbable elastomeric implant - From material design to 12 months follow-up in sheep

The creation of a living heart valve is a much-wanted alternative for current valve prostheses that suffer from limited durability and thromboembolic complications. Current strategies to create such valves, however, require the use of cells for in vitro culture, or decellularized human- or animal-derived donor tissue for in situ engineering. Here, we propose and demonstrate proof-of-concept of in situ heart valve tissue engineering using a synthetic approach, in which a cell-free, slow degrading elastomeric valvular implant is populated by endogenous cells to form new valvular tissue inside the heart. We designed a fibrous valvular scaffold, fabricated from a novel supramolecular elastomer, that enables endogenous cells to enter and produce matrix. Orthotopic implantations as pulmonary valve in sheep demonstrated sustained functionality up to 12 months, while the implant was gradually replaced by a layered collagen and elastic matrix in pace with cell-driven polymer resorption. Our results offer new perspectives for endogenous heart valve replacement starting from a readily-available synthetic graft that is compatible with surgical and transcatheter implantation procedures

Off-the-shelf human decellularized tissue-engineered heart valves in a non-human primate model

Heart valve tissue engineering based on decellularized xenogenic or allogenic starter matrices has shown promising first clinical results. However, the availability of healthy homologous donor valves is limited and xenogenic materials are associated with infectious and immunologic risks. To address such limitations, biodegradable synthetic materials have been successfully used for the creation of living autologous tissue-engineered heart valves (TEHVs) invitro. Since these classical tissue engineering technologies necessitate substantial infrastructure and logistics, we recently introduced decellularized TEHVs (dTEHVs), based on biodegradable synthetic materials and vascular-derived cells, and successfully created a potential off-the-shelf starter matrix for guided tissue regeneration. Here, we investigate the host repopulation capacity of such dTEHVs in a non-human primate model with up to 8 weeks follow-up. After minimally invasive delivery into the orthotopic pulmonary position, dTEHVs revealed mobile and thin leaflets after 8 weeks of follow-up. Furthermore, mild-moderate valvular insufficiency and relative leaflet shortening were detected. However, in comparison to the decellularized human native heart valve control - representing currently used homografts - dTEHVs showed remarkable rapid cellular repopulation. Given this substantial in situ remodeling capacity, these results suggest that human cell-derived bioengineered decellularized materials represent a promising and clinically relevant starter matrix for heart valve tissue engineering. These biomaterials may ultimately overcome the limitations of currently used valve replacements by providing homologous, non-immunogenic, off-the-shelf replacement constructs. © 2013 Elsevier Ltd

Off-the-shelf human decellularized tissue-engineered heart valves in a non-human primate model

Heart valve tissue engineering based on decellularized xenogenic or allogenic starter matrices has shown promising first clinical results. However, the availability of healthy homologous donor valves is limited and xenogenic materials are associated with infectious and immunologic risks. To address such limitations, biodegradable synthetic materials have been successfully used for the creation of living autologous tissue-engineered heart valves (TEHVs) invitro. Since these classical tissue engineering technologies necessitate substantial infrastructure and logistics, we recently introduced decellularized TEHVs (dTEHVs), based on biodegradable synthetic materials and vascular-derived cells, and successfully created a potential off-the-shelf starter matrix for guided tissue regeneration. Here, we investigate the host repopulation capacity of such dTEHVs in a non-human primate model with up to 8 weeks follow-up. After minimally invasive delivery into the orthotopic pulmonary position, dTEHVs revealed mobile and thin leaflets after 8 weeks of follow-up. Furthermore, mild-moderate valvular insufficiency and relative leaflet shortening were detected. However, in comparison to the decellularized human native heart valve control - representing currently used homografts - dTEHVs showed remarkable rapid cellular repopulation. Given this substantial in situ remodeling capacity, these results suggest that human cell-derived bioengineered decellularized materials represent a promising and clinically relevant starter matrix for heart valve tissue engineering. These biomaterials may ultimately overcome the limitations of currently used valve replacements by providing homologous, non-immunogenic, off-the-shelf replacement constructs. © 2013 Elsevier Ltd

Relative concentrations of haemostatic factors and cytokines in solvent/detergent-treated and fresh-frozen plasma

BACKGROUND: /st> Indications, efficacy, and safety of plasma products are highly debated. We compared the concentrations of haemostatic proteins and cytokines in solvent/detergent-treated plasma (SDP) and fresh-frozen plasma (FFP). METHODS: /st> Concentrations of the following parameters were measured in 25 SDP and FFP samples: fibrinogen (FBG), factor (F) II, F V, F VII, F VIII, F IX, F X, F XIII, von Willebrand factor (vWF), D-Dimers, ADAMTS-13 protease, tumour necrosis factor-α (TNF-α), interleukin (IL)-1β, IL-6, IL-8, and IL-10. RESULTS: /st> Mean FBG concentrations in SDP and FFP were similar, but in FFP, the range was larger than in SDP (P<0.01). Mean F II, F VII, F VIII, F IX, and F XIII levels did not differ significantly. Higher concentrations of F V (P<0.01), F X (P<0.05), vWF (P<0.01), and ADAMTS-13 (P<0.01) were found in FFP. With the exception of F VIII and F IX, the range of concentrations for all of these factors was smaller (P<0.05) in SDP than in FFP. Concentrations of TNF-α, IL-8, and IL-10 (all P<0.01) were higher in FFP than in SDP, again with a higher variability and thus larger ranges (P<0.01). CONCLUSIONS: /st> Coagulation factor content is similar for SDP and FFP, with notable exceptions of less F V, vWF, and ADAMTS-13 in SDP. Cytokine concentrations (TNFα, IL-8, and IL-10) were significantly higher in FFP. The clinical relevance of these findings needs to be established in outcome studie