Generation of human induced pluripotent stem cell-derived cardiomyocytes in 2D monolayer and scalable 3D suspension bioreactor cultures with reduced batch-to-batch variations
Sarkawt Hamad, Daniel Derichsweiler, Symeon Papadopolous, Filomain Nguemo, Tomo Saric, Agapios Sachinidis, Jürgen Hescheler, Bastiaan Boukens, Kurt Pfannkuche
Theranostics online date 2019-7-18; doi:10.7150/thno.32058; accepted
To read the authors manuscript follow this link
On March 21st the 7th "Forschungsbörse" took place at the Medical Faculty Cologne. This event is organized by the students and allows research groups of the faculty to present their current work
and discuss project opportunities with the students.
I had alot of interesting discussions with our highly motivated students that inquire for information on scientific projects and seek to inform themselves on current progress in cardiac tissue engineering and stem cell technologies.
(Foto: Dorothea Hensen, Medizinfoto Köln)
First Cologne Conference on Cardiac Repair
Save the date: Sept. 26-27, 2019
Organizers: Kurt Pfannkuche and Tomo Saric
Further infos will appear soon on our conference website
Sponsor: Fritz-Thyssen-Foundation, Cologne
Leitmarkt NRW: CardioPatch project selected for funding
Vaskularisierter, bioartifizieller Herzmuskel aus induziert-pluripotenten Stammzellen
We proudly announce that the CardioPatch initiative has been selected by the "Leitmarkt-Agentur" for funding.
In the frame of this project the CardioPatch consortium will employ a radically novel approach to generate vascularized engineered tissues for therapeutical approaches.
LightFab GmbH, Aachen
Miltenyi Biotech GmbH, Bergisch Gladbach
Taros Chemicals GmbH und Co KG, Dortmund
Fraunhofer-Institut für Lasertechnik ILT, Aachen (Dr. Elke Bremus-Koebberling, Dr. Nadine Nottrodt, Dr. Martin Wehner)
Universitätsklinik Köln (AG PD Dr. Kurt Pfannkuche)
Universität zu Köln, Institut für physikalische Chemie (AG Prof. Annette Schmidt)
Follow this link to see a list with all granted projects: Pressemitteilung
"In vitro grown micro-tissues for cardiac cell replacement therapy in vivo"
Raja Ghazanfar Ali Sahito, Xiaowu Sheng, Martina Maass, Nelly Mikhael,l Sarkawt Hamad, Carlos O. Heras-Bautista, Daniel Derichsweiler, Dimitry Spitkovsky, Frank Suhr, Markus Khalil, Konrad Brockmeier, Marcel Halbach, Tomo Saric, Jürgen Hescheler, Benjamin Krausgrill, Kurt Pfannkuche
Background/Aims: Different approaches have been considered to improve heart reconstructive medicine and direct delivery of pluripotent stem cell-derived cardiomyocytes (PSC-CMs) appears to be highly promising in this context. However, low cell persistence post-transplantation remains a bottleneck hindering the approach. Here, we present a novel strategy to overcome the low engraftment of PSC-CMs during the early post-transplantation phase into the myocardium of both healthy and cryoinjured syngeneic mice.
Methods: Adult murine bone marrow mesenchymal stem cells (MSCs) and PSC-CMs were co-cultured on thermo-responsive polymers and later detached through temperature reduction, resulting in the protease-free generation of cell clusters (micro-tissues) composed of both cells types. Micro-tissues were transplanted into healthy and cryo-injured murine hearts. Short term cell retention was quantified by real-time-PCR. Longitudinal cell tracking was performed by bioluminescence imaging for four weeks. Transplanted cells were further detected by immunofluorescence staining of tissue sections.
Results: We demonstrated that in vitro grown micro-tissues consisting of PSC-CMs and MSCs can increase cardiomyocyte retention by >10-fold one day post-transplantation, but could not fully rescue a further cell loss between day 1 and day 2. Neutrophil infiltration into the transplanted area was detected in healthy hearts and could be attributed to the cellular implantation rather than tissue damage exerted by the transplantation cannula. Injected PSC-CMs were tracked and successfully detected for up to four weeks by bioluminescence imaging.
Conclusion: This approach demonstrated that in vitro grown micro-tissues might contribute to the development of cardiac cell replacement therapies.
Read the full text:
Fig. 5. Histological observation of transplanted micro-tissues. Micro-tissues of low-dose MSC/iPS-CM were transplanted into cryoinjured cardiac tissue. Trichrome staining revealed fibrotic areas of myocardial damage after one week (A) and two weeks (E). Transplanted cells were stained with antibodies against luciferase (green) and nuclei were co-stained by Hoechst dye (blue). Tissue autofluorescence (red) was recorded to visualize the endogenous cardiac tissue. Panel A/B and C/D show results for two individual animals one week after cryoinjury and cell transplantation. Panel E/F show engrafted cells from one animal two weeks after cell transplantation. Panel G/H display magnified regions from the section shown in F.
CCCR: First Cologne Conference on Cardiac Repair: 26./27. September 2019
Together with Tomo Saric at the Institute of Neurophysiology, Cologne we are organizing the first Cologne Conference on Cardiac Repair. The Event will take place on the 26st and 27th Sept. 2019 at the Center of Physiology, Cologne. We gratefully aknowledge support of the Fritz-Thyssen-Foundation to organize the meeting. I am comming back with more detailed information on topics, invited and confirmed speakers soon. This will be a great chance for networking, I am happy to be part of it!
Carlos O.Heras Bautista*, Nelly Mikhael*, Jennifer Lam, Vaibhav Shinde, Alisa Katsen-Globa, Sabine Dieluweit,
Marek Molcanyi, Vladimir Uvarov, Peter Jütten, Raja G.A. Sahito, Francisco Mederos Henry, Alexander Piechot,
Konrad Brockmeier, Jürgen Hescheler, Agapios Sachinidis, Kurt Pfannkuche
Pathophysiological conditions, such as myocardial infarction and mechanical overload affect the mammalian heart integrity, leading to a stiffened fibrotic tissue. With respect to the pathophysiology of cardiac fibrosis but also in the limelight of upcoming approaches of cardiac cell therapy it is of interest to decipher the interaction of cardiomyocytes with fibrotic matrix. Therefore, we designed a hydrogel-based model to engineer fibrotic tissue in vitro as an approach to predict the behavior of cardiomyocytes facing increased matrix rigidity. Here, we generated pure induced pluripotent stem cell-derived cardiomyocytes and cultured them on engineered polyacrylamide hydrogels matching the elasticities of healthy as well as fibrotic cardiac tissue. Only in cardiomyocytes cultured on matrices with fibrotic-like elasticity, transcriptional profiling revealed a substantial up-regulation of a whole panel of cardiac fibrosis-associated transcripts, including collagen I and III, decorin, lumican, and periostin. In addition, matrix metalloproteinases and their inhibitors, known to be essential in cardiac remodeling, were found to be elevated as well as insulin-like growth factor 2. Control experiments with primary cardiac fibroblasts were analyzed and did not show comparable behavior. In conclusion, we do not only present a snapshot on the transcriptomic fingerprint alterations in cardiomyocytes under pathological conditions but also provide a new reproducible approach to study the effects of fibrotic environments to various cell types.
Mobility Grant for MSc. Sarkawt Hamad
Congratulations to Sarkawt for winning a mobility grant (IPaK: Promoting international students at the University of Cologne) supported by the DAAD and selected by the medical faculty, Cologne. The grant will allow Sarkawt to stay for 2 months in the laboratory of Dr. Carolina Galvez-Monton located at Badalona, Spain. The aim of this visit is to initate first experiments of cardiac cell replacement therapy using human induced pluripotent stem cell derived cardiomyocytes in a large animal model.
Novel project granted by the Elisabeth and Rudolf Hirsch-Foundation
We have just received the good news that the Elisabeth and Rudolf Hirsch Foundation gives support to the team thereby allowing us to conduct initial experiment of cardiac cell replacement therapy in a porcine model. This project is handled mainly by Daniel Derichsweiler and Msc. Sarkawt Hamad in the laboratory with support by Karina Neumann. Cell production technologies are established and experimental permissions were granted.
Collaboration News Gobio GmbH
Our group has supported the SME Gobio GmbH (Aarbergen, Germany) in the develop of a stem cell-based cell assays for toxicity measurements. The collaborative work was funded by the German Federal Ministry for Economic Affairs and Energy within a "ZIM project" and has now resulted in the application of patents in Europe and US
("Indicator stem cell line/living organism (non human) and a method for detection of the genotoxic potential of aquatic samples or aqueous solutions of test compounds.")
Networking Event Badalona - Spain
In preparation for the upcoming projects in the field of cardiac cell replacement therapy we have visited the IGTP Institute in Badalona, Spain to meet our collaboration partners Dr. Carolina Galvez-Monton and Prof. Bas Boukens (Amsterdam) for a brain storming. The novel facility at IGTP provides the best possible environment for our research with state of the art facilities and instrumentation.
Networking Event Tandojam - Cologne Initiative
In the follow up of our first online course in cell culture technologies for students at our partner University SAU in Tandojam, PK vicechancelor Prof. Memon is guest in Cologne to report on the ongoing initiative to establish a new institute for stem cell research. The building is now ready to use and we are planning the next steps.
Novel project granted by the German Research Foundation (DFG)
Our team has now received a novel grant to optimize the early retention of transplanted stem cell-derived cardiomyocytes in a rodent model of cardiac infarction. In the frame of the project our successful strategy of micro-tissue transplantation will be further developed to enhance the efficiency of the approach. Magnetic targeting is among the strategies that appear attractive to boost the cell transplantation success.
Generation of human induced pluripotent stem cell derived cardiomyocytes (HiPS-CMs) optimized
Sarkawt and Daniel from my team have made a hell of effort to optimize the differentiation of HiPS-CMs for reproducible generation of cardiac differentiation, high differentiation efficiencies of up to 95%, high yields of maximally 100 million cardiomyocytes per 100 ml suspension culture and controlled costs. Alltogether, we are now moving forward to plan preclinical studies in the porcine model together with Dr. Gálvez Montón in Badalona, Spain. Pall Corporation has just offered support for the bioreactor technology - hope to getting started soon.
Tandojam Laboratory of Regenerative Medicine
Our team supports the SAU to rise a own stem cell laboratory at Tandojam, Pakistan. This collaboration has arisen from a project ("KAZE" project) funded by the BMBF in the frame of scientific collaborations with partners in Pakistan in 2014. Raja is busy in networking and keeping everybody in the consortium busy. The Vice-chancellor of SAU Prof. Mujeeb-u-ddin Sahrai has visited Cologne on several occasions to keep the business rolling. Efforts have already payed-off and the Pakistan Ministry has just granted about 3.5 Mio. Euro to raise a stem cell laboratory at SAU. We are involved in the planning and help the SAU to get started in the field of stem cell technology. The online lecture series in April will be followed by further and more specific training measures in the future.
Online course on basic stem cell culturing techniques at Sindh Agricultural University (SAU)
Together with our colleagues at the Institute of Neurophysiology Cologne we have prepared an online course for cell culturing technologies for students at the SAU in Tandojam, Pakistan. The lecture was attended by nearly 50 persons from student to principal investigator level. We are planning further activities in the future to intensify our collaboration with the SAU in the field of Stem Cell Research. Many thanks to the Vice Chancellor Prof. Mujeeb-u-ddin Sahrai for his great support of the project.