TY - CHAP A1 - Lehmann, Mario A1 - Scholka, Jenny A1 - Wittenburg, Gretel A1 - Petto, Carola A1 - Geppert, Philipp A1 - Muschter, Antje A1 - Anderer, Ursula T1 - An easily accessible adult stem cell pool may be suitable for tissue engineering-based regeneration of articular cartilage defects T2 - eCM Conferences Open Access online periodical KW - adult stem cells KW - dental pulp stem cells KW - scaffold-free 3D culture KW - chondrogenic differentiation Y1 - 2016 UR - http://www.ecmjournal.org/journal/supplements/vol031supp01/vol031supp01_P.htm SN - 2522-235X N1 - TERMIS-EU Conference, Uppsala, Sweden SP - P190 ER - TY - GEN A1 - Lutter, Anne-Helen A1 - Scholka, Jenny A1 - Richter, Heiko A1 - Anderer, Ursula T1 - Applying XTT, WST-1, and WST-8 to human chondrocytes: A comparison of membrane-impermeable tetrazolium salts in 2D and 3D cultures T2 - Clinical Hemorheology and Microcirculation KW - chondrogenic differentiation Y1 - 2017 U6 - https://doi.org/10.3233/CH-179213 SN - 1875-8622 SN - 1386-0291 VL - 67 IS - 3-4 SP - 327 EP - 342 ER - TY - GEN A1 - Lutter, Anne-Helen A1 - Scholka, Jenny A1 - Richter, Heiko A1 - Anderer, Ursula T1 - Comparison of membrane-impermeable tetrazolium salt assays in 2D and 3D cultures of human chondrocytes T1 - 5. Naturwissenschaftstag, Senftenberg, 2017 KW - WST-8, WST-1, XTT KW - 2D/3D cultures KW - chondrocyte Y1 - 2017 U6 - https://doi.org/10.13140/RG.2.2.10682.49600 ER - TY - CHAP A1 - Lehmann, Mario A1 - Martin, Frank A1 - Scholka, Jenny A1 - Petto, Carola A1 - Wittenburg, Gretel A1 - Sack, Ulrich A1 - Anderer, Ursula T1 - Human chondrocytes and dental pulp stem cells enhance their chondrogenic differentiation potential via a two-step fusion culture technique T2 - Tissue Engineering, Part A, Vol. 21, Suppl. 1 KW - in vitro chondrogenesis KW - dental pulp stem cells KW - scaffold-free 3D culture KW - two-step fusion technique Y1 - 2015 U6 - https://doi.org/10.1089/ten.tea.2015.5000.abstracts SP - S-268 EP - S-269 ER - TY - GEN A1 - Lutter, Anne-Helen A1 - Liedtke, Victoria A1 - Ecke, Annemarie A1 - Scholka, Jenny A1 - Muschter, Antje A1 - Hentschel, Erik A1 - Becker, Roland A1 - Anderer, Ursula T1 - Direct co-culture of human chondrogenic microtissues with osteoblast-like Saos-2 cells positively influence the differentiation potential of cartilage cells in 3D culture Y1 - 2018 U6 - https://doi.org/10.1007%2Fs10354-018-0664-9.pdf N1 - Austrian Bone Conference – ABC 2018, November 23–24, 2018, Vienna, Austria ER - TY - GEN A1 - Lutter, Anne-Helen A1 - Liedtke, Victoria A1 - Scholka, Jenny A1 - Muschter, Antje A1 - Becker, Roland A1 - Anderer, Ursula T1 - Co-culture of human chondrogenic microtissues with osteoblast-like Saos-2 cells or HFF-1 fibroblasts influence the differentiation potential of spheroids T2 - Journal of Cellular Biotechnology N2 - BACKGROUND: Tissue engineering has become a major field of research in biotechnology and biomedicine. As a consequence, cell-based therapeutic approaches are entering the hospitals, especially for skeletal regeneration. Traumatic injuries of cartilage are treated with autologous cell suspensions or in vitro generated cartilage tissues, but there is actually no therapy available for degenerative cartilage defects. However, Osteoarthritis (OA) is a major public health problem in the world affecting 240 million people globally. OBJECTIVE: To develop suitable in vitro tissues, the properties of chondrogenic spheroids should be optimized via coculture with cells naturally occurring as joint neighbours. METHODS: Human chondrocytes were isolated from condyles and propagated in monolayer culture. Scaffold-free spheroids were generated and co-cultured with joint-specific partner cells (osteoblast-like osteosarcoma cells, fibroblasts). Morphology and differentiation was analyzed using histochemistry (Alcian blue, Safranin O) and immunohistochemistry for cartilage markers (collagen type II, Sox9, proteoglycan), proliferation-associated protein (Ki67) and markers of connective tissue (collagen type I and actin). RESULTS: The provision of a more natural microenvironment in vitro via co-culture of chondrocyte-based aggregates with osteoblast-like Saos-2 cells enhanced the differentiation potential of chondrogenic spheroids towards hyaline cartilage. CONCLUSIONS: The study showed the positive influence of Saos-2 cells on the differentiation potential of human chondrocytes in co-culture. Y1 - 2018 U6 - https://doi.org/10.3233/JCB-189005 SN - 2352-3697 SN - 2352-3689 VL - 4 IS - 1/2 SP - 31 EP - 42 ER - TY - GEN A1 - Ecke, Annemarie A1 - Lutter, Anne-Helen A1 - Scholka, Jenny A1 - Hansch, Anna A1 - Becker, Roland A1 - Anderer, Ursula T1 - Tissue Specific Differentiation of Human Chondrocytes Depends on Cell Microenvironment and Serum Selection T2 - Cells KW - Chondrocytes KW - Cartilage Y1 - 2019 U6 - https://doi.org/10.3390/cells8080934 SN - 2073-4409 VL - 8 IS - 8 ER - TY - GEN A1 - Ecke, Annemarie A1 - Scholka, Jenny A1 - Hansch, Anna A1 - Becker, Roland A1 - Anderer, Ursula T1 - Agar overlay technique vs. v-bottom plate: differentiation of human cartilage-like microtissues depend on plate format. T2 - Osteologie KW - microtissue KW - cartilage Y1 - 2019 U6 - https://doi.org/10.1055/s-0039-1680058 SN - 2567-5818 VL - 28 IS - 4 PB - Georg Thieme Verlag KG CY - Stuttgart ; New York ER - TY - GEN A1 - Lutter, Anne-Helen A1 - Ecke, Annemarie A1 - Scholka, Jenny A1 - Waldeck, Peter A1 - Tretter, Tim A1 - Anderer, Ursula T1 - Differentiation of human chondrocytes under the influence of microalgae-based exopolysaccharides T2 - 38. Jahrestagung der Deutschen Gesellschaft für Klinische Mikrozirkulation und Hämorheologie : Programm, 22. und 23. November 2019, Braunschweig N2 - BACKGROUND: Exopolysaccharides are formed by microalgae in response to adverse conditions and are found mainly as a coating around the cells. The protective function of EPS plays an important role in the survival of microalgae in polluted habitats. So far, the particular biological and physico-chemical properties of EPS have not been adequately investigated and their effect on cells such as human chondrocytes has not been tested. Microalgae in particular offer an ideal platform for the production of dietary supplements or drugs that enable new therapeutic approaches. METHODS: To investigate the influence of EPS on human chondrocytes, cytotoxicity and differentiation potential were analyzed in 3D cultures. RESULTS: We were able to demonstrate a positive, but donor-specific influence of EPS on the differentiation of human chondrocytes. Furthermore, the EPS showed no cytotoxic effects in a range of up to 250 µg EPS per ml medium. CONCLUSION: EPS have the potential to positively influence the differentiation of human chondrocytes. Further experiments are necessary to analyze the donor-specific effect. Y1 - 2019 UR - https://www.orbera.de/referenzen/DGKMH_2019/pdf_DGKMH_2019.pdf ER - TY - GEN A1 - Ecke, Annemarie A1 - Scholka, Jenny A1 - Hansch, Anna A1 - Becker, Roland A1 - Anderer, Ursula T1 - Human serum or FCS: Differentiation of human cartilage-like microtissues depends on serum selection T2 - DGZ 2018 International Meeting of the German Society for Cell Biology: “Concepts of Cell Organisation and Dynamics” N2 - Introduction: Regeneration of articular cartilage defects is limited. Hence, cell-based therapies for the treatment were developed using e.g. chondrocytes isolated from patient biopsies. For cell culture fetal calf serum (FCS) is often used as media supplement to establish engineering processes for transplants in regenerative medicine. However, the usage of xenogeneic serum could mask the intrinsic behavior of human cells in vitro. This could be particularly important for the dedifferentiation process during cell expansion and redifferentiation in 3D-culture. The aim of this study was to compare the development of microtissues derived from chondrocytes cultivated in presence of FCS with those cultivated in human serum (HS). Material and Methods: Human articular chondrocytes isolated from knee joints were expanded in medium with HS or FCS. Scaffold-free microtissues were generated using the agar overlay technique. After four and eight weeks the macroscopic appearance (reflected-light microscopy) was documented and the size of the microtissues was determined. The differentiation degree was evaluated via histology to visualize typical glycosamino¬glycans (Safranin O, Alcian blue) and immunohisto¬chemistry (IH) to detect cartilage-specific markers (collagen type II, proteoglycans, COMP, Sox9) on cryosections. Results and Conclusion: Cells from all donors in both medium compositions generated microtissues – but with distinct differences in the differentiation levels. Microtissues cultured in medium with FCS showed a higher differentiation level compared to microtissues in HS. This is evidenced by a higher amount of matrix resulting in bigger microtissues (diameter in FCS is 42% larger compared to HS) and a more cartilage-like composition of the matrix (not any/little colouring in HS versus intense staining in FCS). IH analysis showed the same pattern. Not any/less positivity for collagen type II, proteoglycan, COMP, and Sox9 in HS versus moderate-to-high positivity in FCS-cultured microtissues. Absolute values vary from donor to donor whereas an individualized expression profile only became visible in microtissues cultured in HS medium. HS should be preferred for the engineering of cartilage-like microtissues, as it rather enables a "human-based" situation in vitro. Supported by „Gesundheitscampus Brandenburg“ and „Ministerium für Wissenschaft, Forschung und Kultur“, state of Brandenburg, Germany. KW - cartilage-like microtissues KW - serum type Y1 - 2018 UR - https://www.researchgate.net/publication/338863739_HUMAN_SERUM_OR_FCS_DIFFERENTIATION_OF_HUMAN_CARTILAGE-LIKE_MICROTISSUES_DEPENDS_ON_SERUM_SELECTION ER - TY - GEN A1 - Ecke, Annemarie A1 - Scholka, Jenny A1 - Hansch, Anna A1 - Becker, Roland A1 - Anderer, Ursula T1 - Differentiation of human articular chondrocytes in microtissues depends on the cell aggregation technique KW - chondrogenic microtissues KW - differentiation KW - aggregation techniques Y1 - 2020 UR - https://www.ariabstracts.org/abstracts/2020/Collection1/c1.html ER - TY - GEN A1 - Anderer, Ursula A1 - Scholka, Jenny A1 - Hansch, Anna A1 - Becker, Roland A1 - Ecke, Annemarie T1 - Influence of the cell aggregation technique on the differentiation of human articular chondrocytes in microtissues T2 - Osteologie / Osteology KW - microtissue generation KW - chondrogenic differentiation KW - cell therapy KW - cartilage regeneration Y1 - 2022 U6 - https://doi.org/10.1055/s-0042-1755855 SN - 1019-1291 VL - 31 IS - 03 SP - 201 EP - 202 PB - Georg Thieme Verlag CY - Stuttgart ER - TY - GEN A1 - Martin, Frank A1 - Neubert, Annemarie A1 - Lutter, Anne-Helen A1 - Scholka, Jenny A1 - Hentschel, Erik A1 - Richter, Heiko A1 - Anderer, Ursula T1 - MTS, WST-8, and ATP viability assays in 2D and 3D cultures: Comparison of methodologically different assays in primary human chondrocytes T2 - Clinical Hemorheology and Microcirculation N2 - BACKGROUND: Tissue engineering enables the production of three-dimensional microtissues which mimic naturally occurring conditions in special tissues. These 3D culture systems are particularly suitable for application in regenerative medicine or experimental pharmacology and toxicology. Therefore, it is important to analyse the cells in their 3D microenvironment with regard to viability and differentiation. Tetrazolium assays (WST-8 and MTS) are still the methods of choice for estimating the number of living, metabolically active cells, with WST-8 being cell-impermeable compared to MTS. In contrast to these methods, the ATP assay is an endpoint method based on the luciferase-induced reaction of ATP with luciferin after cell lysis. OBJECTIVE:We compared three methodologically different proliferation/toxicity assays (MTS, WST-8, ATP) in monolayer (2D) and 3D culture systems to improve the technically challenging determination of the number of viable cells. METHODS: Chondrocytes were isolated from human articular cartilage. Three different test systems (MTS, WST-8, ATP) were applied to monolayer cells (2D, varying cell numbers) and spheroids (3D, different sizes) in 96-well plates. The intracellular ATP concentration was determined by luciferase-induced reaction of ATP with luciferin using a luminometer. Formazan formation was measured spectrophotometrically after different incubation periods. Evaluation was performed by phase contrast microscopy (toxicity), correlation of cell count and ATP concentration or absorption signal (Gompertz function) and propidium iodide (PI) staining to proof the cell lysis of all cells in spheroids. RESULTS: In 2D culture, all three assays showed a good correlation between the number of seeded cells and the ATP concentration or absorption data, whereas the MTS-assay showed the lowest specificity. In 3D culture, the spheroid sizes were directly related to the number of cells seeded. The absorption data of the WST-8 and MTS assay correlated only for certain spheroid size ranges, whereas the MTS-assay showed again the lowest specificity. Only the measured intracellular ATP content showed a linear correlation with all spheroid sizes ranging from 100–1000 µm. The WST-8 assay revealed the second-best sensitivity which allows the measurement of spheroids larger than 240 µm. Phase contrast observation of monolayer cells showed toxic effects of MTS after 6 h incubation and no signs of toxicity of WST-8. Staining with propidium iodide showed complete lysis of all cells in a spheroid in the ATP assay. CONCLUSION: Among tetrazolium-based assays, WST-8 is preferable to MTS because of its non-toxicity and better sensitivity. When determining the number of viable cells in the 2D system, caution is advised when using the ATP assay because of its two-phase slope of the correlation graph concerning cell number and intracellular ATP. In 3D systems of human chondrocytes, the ATP-assay is superior to the other two test systems, as the correlation graph between cell number and intracellular ATP is biphasic. Since differentiation processes or other metabolic events can influence the results of proliferation and toxicity assays (determination of viable cells), this should be taken into account when using these test systems. KW - ATP assay KW - MTS KW - WST-8 KW - Human chondrocytes KW - Spheroid KW - 3D culture KW - 2D culture Y1 - 2024 UR - https://journals.sagepub.com/doi/abs/10.3233/CH-248101 U6 - https://doi.org/10.3233/CH-248101 SN - ISSN 1386-0291 VL - 88 SP - 3 EP - 19 PB - IOS Press ER -