FACS-Sorter mit UV-Laser
Final Report Abstract
The FACS-Sorter with UV laser was needed to found a new cell isolation core facility for the SFB/TRR57 “organ fibrosis: from mechanisms of injury to modulation of disease” at the universities of Aachen and Bonn. In the first funding period, the major aim of this core facility was to establish procedures that allow providing researchers of the Collaborative Research Centre with healthy viable primary cells of murine and rat livers and kidneys. Using conventional isolation procedures, we have set up a logistical and methodological platform that routinely provides primary hepatic stellate cells (HSC) and hepatocytes to all projects in Aachen and Bonn upon request. Moreover, we have introduced novel innovative FACS-based sorting protocols that further allow enrichment and characterization of highly purified hepatic stellate cells from murine livers using their autofluorescence in the UV channel and negativity for hematopoietic surface markers. Other protocols were established for the isolation of immune cells from blood, liver, kidney, spleen, bone marrow and peritoneal lavage as well as standard procedures for sorting labeled cells (e.g. GFP, RFP, LacZ). Magnetic bead separation techniques were introduced for the isolation of glomerular cells from mouse kidney. The purified cells were taken as starting material by multiple researchers of the SFB/TRR 57 (P01, P04, P05, P06, P07, P08, P09, P13, P14, P17, P18) to address specific questions on fibrogenesis as outlined in the individual projects. In particular, the highly purified HSC allowed addressing sophisticated questions dealing with cell-specific expression of miRNAs and candidate genes relevant for fibrogenesis as well as studies that analyzed issues of cell-cell interactions.
Publications
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(2009): Antifibrotic effects of CXCL9 and its receptor CXCR3 in livers of mice and humans. Gastroenterology, 137, 309-319
Wasmuth, H. E., Lammert, F., Zaldivar, M. M., Weiskirchen, R., Hellerbrand, C., Scholten, D., Berres, M. L., Zimmermann, H., Streetz, K. L., Tacke, F., Hillebrandt, S., Schmitz, P., Keppeler, H., Berg, T., Dahl, E., Gassler, N., Friedman, S. L, Trautwein, C.
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(2009): Hepatic recruitment of the inflammatory Gr1+ monocyte subset upon liver injury promotes hepatic fibrosis. Hepatology, 50, 261-274
Karlmark, K. R., Weiskirchen, R., Zimmermann, H. W., Gassler, N., Ginhoux, F., Weber, C., Merad, M., Luedde, T., Trautwein, C., Tacke, F.
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(2010): Analysis of antigen-presenting functionality of cultured rat hepatic stellate cells and transdifferentiated myofibroblasts. Biochem Biophys Res Commun, 396, 342-347
Bomble, M., Tacke, F., Rink, L., Kovalenko, E., Weiskirchen, R.
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(2010): Antagonism of the chemokine CCL5 ameliorates experimental liver fibrosis in mice. J Clin Invest, 120, 4129-4140
Berres, M. L., Koenen, R. R., Rueland, A., Zaldivar, M. M., Heinrichs, D., Sahin, H., Schmitz, P., Streetz, K. L., Gassler, N., Weiskirchen, R., Proudfoot, A., Weber, C., Trautwein, C., Wasmuth, H. E.
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(2010): TAK1 suppresses a NEMO-dependent but NF- kappaB-independent pathway to liver cancer. Cancer Cell, 17, 481-496
Bettermann, K., Vucur, M., Haybaeck, J., Koppe, C., Janssen, J., Heymann, F., Weber, A., Weiskirchen, R., Liedtke, C., Gassler, N., Müller, M., de Vos, R., Wolf, M. J., Boege, Y., Seleznik, G. M., Zeller, N., Erny, D., Fuchs, T., Zoller, S., Cairo, S., Buendia, M. A., Prinz, M., Akira, S., Tacke, F., Heikenwalder, M., Trautwein, C., Luedde, T.
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(2011): Expression of the megalin C-terminal fragment by macrophages during liver fibrogenesis in mice. BBA Mol Bas Dis, 1812, 1840- 1848
Pieper-Fürst, U., Hall, R., Huss, S., Fischer, H. P., Tacke, F., Weiskirchen, R., Hochrath, K., Lammert, F.
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(2011): Micro-RNA profiling reveals a role for miR-29 in human and murine liver fibrosis. Hepatology, 53, 209-218
Roderburg, C., Urban, G. W., Bettermann, K., Vucur, M., Zimmermann, H., Schmidt, S., Janssen, J., Koppe, C., Knolle, P., Castoldi, M., Tacke, F., Trautwein, C., Luedde, T.
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(2012): A novel, dual role of CCN3 in experimental glomerulonephritis: pro-angiogenic and anti-mesangioproliferative effects. Am J Pathol, 180, 1979-90
van Roeyen, C. R., Boor, P., Rong, S., Kunter, U., Borkham- Kamphorst, E., Fleckenstein, S., Perbal, B., Trautwein, C., Weiskirchen, R., Ostendorf, T., Floege, J.
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(2012): Cyclin E1 controls proliferation of hepatic stellate cells and is essential for liver fibrogenesis in mice. Hepatology, 56(3):1140-9
Nevzorova, Y. A., Bangen, J. M., Weiskirchen, R., Haas, U., Hu, W., Heymann, F., Kroy, D., Gassler, N., Sicinski, P., Tacke, F., Trautwein, C., Liedtke, C.
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(2012): Hepatic macrophage migration and differentiation critical for liver fibrosis is mediated by the chemokine receptor C-C motif chemokine receptor 8 in mice. Hepatology 55, 898-909
Heymann, F., Hammerich, L., Storch, D., Bartneck, M., Huss, S., Russeler, V., Gassler, N., Lira, S. A., Luedde, T., Trautwein, C., Tacke, F.
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(2012): Primary cultures of glomerular parietal epithelial cells or podocytes with proven origin. PLoS ONE, 7, e34907
Kabgani, N., Grigoleit, T., Schulte, K., Sechi, A., Sauer-Lehnen, S., Tag, C., Boor, P., Kuppe, C., Warsow, G., Schordan, S., Mostertz, J., Chilukoti, R., Homuth, G., Endlich, N., Tacke, F., Weiskirchen, R., Füllen, G., Endlich, K., Floege, J., Smeets, B., Moeller, M. J.
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(2012): Update on hepatic stellate cells: pathogenic role in liver fibrosis and novel isolation techniques. Expert Rev Gastroenterol Hepatol., 6, 67-80
Tacke, F., Weiskirchen, R.
