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Animal-derived materials, e.g. cell culture supplements, feeder cells or basal membrane extracts, are still widely used in cell culture and in vitro experiments, although their drawbacks like unknown compo-sition or batch-to-batch variability are known. Although there are xeno-free alternatives available and the market of synthetic, chemically well-defined alternatives is constantly growing, researchers often lack the time and money to actively change an established method. With the call "Xeno-free in-vitro science" Charité 3R offers funding for replacing animal-derived components in your in vitro project, thus contributing to the reduction of animals used for scientific purposes and increasing scientific quality.
Affordable FCS-free human cell culture: Development of a chemically defined protocol to promote uniform cell culture conditions
Fetal calf serum (FCS) remains the most commonly used medium supplement for in vitro cell cultivation. Among other things, FCS contains important proteins such as growth factors and adhesion proteins that promote e.g., cell growth. Besides the questionable ethical justifiability, the use of FCS is also scientifically controversial e.g., in terms of batch-to-batch variations, lack of reproducibility and the risk of unknown infectious contents. However, the exact composition is still elusive. In their project "Affordable FCS-free human cell culture: Development of a chemically defined protocol to promote uniform cell culture conditions", Alexandra Damerau and Moritz Pfeiffenberger from the Department of Rheumatology and Clinical Immunology aim to identify essential protein groups by mass spectrometric analyses of FCS in comparison to valuable human alternatives such as platelet lysate and AB serum. Subsequently, we will evaluate and confirm the importance of these proteins for the cultivation of mesenchymal stromal cells with the ultimate goal of providing a chemically defined medium in order to ultimately replace the use of animal-derived cell culture supplements.
Xeno-free cell culture of primary cardiac fibroblasts – Development of a diagnostic and drug screening platform for heart failure
Heart failure (HF), one of the leading causes of death worldwide, is characterized by inflammation and fibrosis. Currently, there is a need to better assess/diagnose the inflammatory and fibrosis process in the heart and to develop a patient-specific model system to screen for potential anti-inflammatory/anti-fibrotic treatment options. As cardiac fibroblasts (CFs) regulate beyond tissue scarring also the cardiac inflammatory response, these cells present an ideal target. In the project "Xeno-free cell culture of primary cardiac fibroblasts - Development of a diagnostic and drug screening platform for heart failure", Isabell Matz from Sophie Van Linthout's lab at the Berlin Institute of Health Center for Regenerative Therapies (BCRT) will develop together with faculty members a xeno-free cell culture platform to be used for patient diagnosis and drug screening by cultivating patient-derived CFs in a xeno-free manner replacing fetal bovine serum with human serum and/or human platelet lysate. With this we will pave the way to exclude animal-derived supplements in the cell culture of patient-specific fibroblasts thereby reducing the linked animal suffering and improving experimental standardization and reproducibility to adapt our model to clinical application.
Approaching xeno-free cultivation and utilization of pulmonary pathogens in vitro
Most experimental models of bacterial infection and disease pathology necessitate the utilization of bacterial pathogens in a laboratory setting. Established methodology for bacterial cultivation, however, is largely unfavorable for animal welfare due to the substantial reliance on animal-derived products found in agar plates and liquid growth media. Hence, the ambition is to completely replace or dramatically reduce the use of animal-derived biomaterial used to cultivate bacterial pathogens. The aim of the project "Approaching xeno-free cultivation and utilisation of pulmonary pathogens in vitro" by Cengiz Gökeri and Peter Pennitz from the Department of Infectious Diseases and Respiratory Medicine is to compare various xeno-free and xeno-reduced alternatives to one another and to currently preferred methodology, with the outlook of developing cost-effective, comparable, reproducible and improved protocols. Our results will be actively promoted in conferences, publications and collaborative research networks to demonstrate the feasibility of reducing, replacing and refining of animals and animal-derived products.
Development of human-based hydrogels as a substitute for mouse-derived Matrigel for cancer research
MatrigelTM and other Basement Membrane Extracts (BMEs) are widely used for almost every organoid study. However, the use of BMEs come with several problems. First of all, BMEs are made from mice sarcomas. Due to their biology, BMEs are complex and poorly defined, subject to lot-to-lot variation, have poor control of mechanical properties, and their stiffness is not in the physiological range of tumor stiffness. The aim of the project "Development of human-based hydrogels as a substitute for mouse-derived Matrigel for cancer research" by Björn Papke from the Institute of Pathology and Karl Herbert Hillebrandt from the Department of Surgery at the CVK is to develop a better for cancer research suited human-based hydrogel derived from human decellularized liver. Developing such a hydrogel and the awareness of its importance for cancer research can shift the field to these better-suited human-based hydrogels.
Modeling barrier tissues without obstacles: Development of a xeno-free matrix for layer-by-layer in vitro formation of cellular and barrier tissues
In vitro generation of neo-tissues is still restricted to the self-assembly of cells into spheroids/organoids or limited to cell-seeding onto biomimetic scaffolds. Growth factors and extracellular matrix (ECM)-like substances are used to enhance cell-seeding and differentiation. The state-of-the-art ECM substitute is an animal-derived basement membrane matrix providing all properties for e.g., in vitro angiogenesis and formation of barrier tissues. In the project "Modelling barrier tissues without obstacles: Development of a xeno-free matrix for layer-by-layer in vitro formation of cellular and barrier tissues", Alexandra Damerau and Timo Gaber from the Department of Rheumatology and Clinical Immunology, aim to, in collaboration with researchers from Helsinki and Würzburg, replace any animal-derived ECM with xeno-free hydrogels functionalized with growth factors and capable to retain structure during manipulation for in vitro tissue formation. They want to test these hydrogels in angiogenesis assays and for barrier tissue formation such as synovial membrane and cartilage which will be included in arthritis models identifying new therapeutic targets and ultimately reducing the number of animal experiments.