TY - GEN A1 - Hüttner, Sören A1 - Henze, Henriette A1 - Elster, Dana A1 - Koch, Philipp A1 - Anderer, Ursula A1 - Eyss, Björn von A1 - Maltzahn, Julia von T1 - A dysfunctional miR-1-TRPS1-MYOG axis drives ERMS by suppressing terminal myogenic differentiation T2 - Molecular Therapy N2 - Rhabdomyosarcoma is the most common pediatric soft tissue tumor, comprising two major subtypes: the PAX3/7-FOXO1 fusion-negative embryonal and the PAX3/7-FOXO1 fusion-positive alveolar subtype. Here, we demonstrate that the expression levels of the transcriptional repressor TRPS1 are specifically enhanced in the embryonal subtype, resulting in impaired terminal myogenic differentiation and tumor growth. During normal myogenesis, expression levels of TRPS1 have to decrease to allow myogenic progression, as demonstrated by overexpression of TRPS1 in myoblasts impairing myotube formation. Consequentially, myogenic differentiation in embryonal rhabdomyosarcoma in vitro as well as in vivo can be achieved by reducing TRPS1 levels. Furthermore, we show that TRPS1 levels in RD cells, the bona fide model cell line for embryonal rhabdomyosarcoma, are regulated by miR-1 and that TRPS1 and MYOD1 share common genomic binding sites. The myogenin (MYOG) promoter is one of the critical targets of TRPS1 and MYOD1; we demonstrate that TRPS1 restricts MYOG expression and thereby inhibits terminal myogenic differentiation. Therefore, reduction of TRPS1 levels in embryonal rhabdomyosarcoma might be a therapeutic approach to drive embryonal rhabdomyosarcoma cells into myogenic differentiation, thereby generating postmitotic myotubes. Y1 - 2023 U6 - https://doi.org/10.1016/j.ymthe.2023.07.003 SN - 1525-0024 SN - 1525-0016 VL - 31 IS - 9 SP - 2612 EP - 2632 ER - TY - GEN A1 - Hiemann, Rico A1 - Roggenbuck, Dirk A1 - Sack, Ulrich A1 - Anderer, Ursula A1 - Conrad, Karsten T1 - Die HEp-2-Zelle als Target für multiparametrische Autoantikörperanalytik – Automatisierung und Standardisierung/The HEp-2 cell as target for multiparametric autoantibody analyses: automation and standardisation T2 - Journal of Laboratory Medicine N2 - Trotz Einführung von Multiplex-Assays ist die Bestimmung von Autoantikörpern (AAK) mittels indirekter Immunfluoreszenz (IIF) nach wie vor ein wesentlicher Bestandteil der Autoimmundiagnostik. Das Screening auf nichtorganspezifische AAK wird in der Routinediagnostik von autoimmunen rheumatischen Erkrankungen mittels indirekter Immunfluoreszenz an HEp-2-Zellen durchgeführt. Leider differieren die mit diesem Test erhaltenen Befunde z. T. recht erheblich auf Grund von objektiven (z. B. HEp-2-Zellfixierung) und subjektiven (z. B. Erfahrungswerte) Faktoren. Die Intra- und Interlaborvarianzen sind daher relativ hoch. Erste Ergebnisse von Evaluierungsstudien eines neu entwickelten Systems zur automatischen Bildanalyse (AKLIDES®) zeigen einen Weg in Richtung Standardisierung (bzw. Harmonisierung) des HEp-2-Zell-Assays auf, in dem zunächst subjektive Fehlinterpretationen vermieden werden können. Eine weitere Harmonisierung erfordert die Weiterentwicklung der Software-Algorithmen zur Mustererkennung sowie neue Kalibrationssysteme. KW - Autoantikörper KW - Automatisierung KW - HEp-2 KW - Immunfluoreszenz Y1 - 2011 U6 - https://doi.org/10.1515/JLM.2011.053 SN - 1439-0477 VL - 35 IS - 6 SP - 351 EP - 361 ER - TY - GEN A1 - Hahm, Daniel A1 - Anderer, Ursula T1 - Establishment of HEp‐2 cell preparation for automated analysis of ANA fluorescence pattern T2 - Cytometry. Part A N2 - AbstractBackgroundIdentification of antinuclear antibodies (ANAs) has large clinical importance for the assessment of autoimmune diseases. HEp‐2 cell preparations on microscopic slides are commonly used as antigenic substrate. Methods used for cell preparation are important for ANA pattern analysis; however, these methods differ widely and are mostly not specified.MethodsHEp‐2 cells were fixed using acetic acid‐ethanol, methanol‐acetone, acetone, formaldehyde, paraformaldehyde, or glutaraldehyde. Morphological analysis was done after haematoxylin‐eosin staining and DAPI‐staining of cell nuclei.ResultsThe results demonstrate a high variability of cell and nuclear morphology depending on the used fixatives. Aldehyde fixatives conserved the cell structures best, acetone fixatives revealed remarkable changes.ConclusionsAfter selecting appropriate fixation procedures to preserve nuclear structures further experiments are necessary to find out which fixation procedure preserves the disease‐linked antigens the best way and are, therefore, suitable to be used in ANA‐testing of AABs. © 2006 International Society for Analytical Cytology KW - Cell Biology KW - Histology KW - Pathology and Forensic Medicine KW - Autoimmune diagnostic KW - Antinuclear antibodies KW - HEp-2 cells Y1 - 2006 U6 - https://doi.org/10.1002/cyto.a.20223 SN - 1552-4922 VL - 69A IS - 3 SP - 178 EP - 181 ER - TY - GEN A1 - Hiemann, Rico A1 - Hilger, Nadja A1 - Michel, Jörg A1 - Nitschke, Jörg A1 - Böhm, Alexander A1 - Anderer, Ursula A1 - Weigert, Martin A1 - Sack, Ulrich T1 - Automatic Analysis of Immunofluorescence Patterns of HEp‐2 Cells T2 - Annals of the New York Academy of Sciences N2 - The standard screening test for the recognition of autoimmune diseases is the proof of autoantibodies in serum of patients by indirect immunofluorescence (IIF) based on HEp‐2 cells. Manual evaluation of this test is very subjective, slow, and there are no objective parameters as guidelines available. Interlaboratory tests showed occasionally large deviations in the test evaluation resulting in a high variance of results. The aim of this project is fast, objective, safe, and economical automatic analysis of HEp‐2 IIF patterns. Images of IIF patterns were completely and automatically captured using an inverse motorized fluorescence microscope. Thereby, device‐specific parameters were controlled automatically, too. For fast analysis of IIF patterns new algorithms of image processing were developed. Artifacts were recognized and excluded from analysis by the developed software. Analysis of more than 80,000 images clearly demonstrated full automatization and fast processing of IIF patterns. Additionally serum‐specific fluorescence could be easily distinguished from background. Even very weak but positive patterns can be recognized and used for diagnosis. A detailed separation into different basic patterns is possible. Objective, fast, and disease‐related economical analysis of HEp‐2 immunofluorescence patterns is feasible. The implemented software algorithms allowed a mathematical way of describing IIF patterns and can therefore be a useful tool for the needed standardization process. KW - Autoimmune diagnostics KW - HEp-2 cell KW - Immunofluorescence KW - Automatic analysis KW - Screening Y1 - 2007 U6 - https://doi.org/10.1196/annals.1398.042 SN - 0077-8923 VL - 1109 IS - 1 SP - 358 EP - 371 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 - 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 - 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 - 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 - Lutter, Anne-Helen A1 - Hempel, Ute A1 - Dieter, Peter A1 - Anderer, Ursula T1 - Multinuclearity is not mandatory for Bone Resorption Mononuclear Osteoclasts are able to resorbe Extracellular Bone Matrix in vitro T2 - Bone-Tec, Munich, 2017 KW - mononuclear osteoclast Y1 - 2017 UR - https://www.researchgate.net/publication/327860900 ER - TY - GEN A1 - Lutter, Anne-Helen A1 - Hempel, Ute A1 - Dieter, Peter A1 - Anderer, Ursula T1 - The biomarker-based ODEM-assay affects cellular characteristics of osteoclasts in vitro Tissue culture polystyrene vs Osteoblast-Derived Extracellular Matrix T1 - 6th Munich Biomarker Conference, Munich, 2016 Y1 - 2016 U6 - https://doi.org/10.13140/RG.2.2.20195.96806 ER -