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1,2 Gundula Schulze‐Tanzil, VMD Research group “Bioreconstruction” 1 Institute of Anatomy, Paracelsus Medical University, Salzburg and Nuremberg, Germany.

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Presentation on theme: "1,2 Gundula Schulze‐Tanzil, VMD Research group “Bioreconstruction” 1 Institute of Anatomy, Paracelsus Medical University, Salzburg and Nuremberg, Germany."— Presentation transcript:

1 1,2 Gundula Schulze‐Tanzil, VMD Research group “Bioreconstruction” 1 Institute of Anatomy, Paracelsus Medical University, Salzburg and Nuremberg, Germany 2 Department of Orthopaedic, Trauma and Reconstructive Surgery, Charité- Universitätsmedizin Berlin, Campus Benjamin Franklin, Germany 1

2 Education: 1990‐ School of veterinary medicine, Freie Universität Berlin VMD Degree, Freie Universität Berlin Postdoctoral Fellow, Institute of Anatomy, Freie Universität Berlin Scientific assistant, Centrum of Anatomy, Charité University of Medicine, CCM, Berlin Dep. of Trauma and Reconstructive Surgery, CBF/Centrum of Anatomy, CCM, Charité University of Medicine, Berlin Leader of the research laboratory of experimental surgery, AG Bioreconstruction Dep. of Trauma and Reconstructive Surgery, CBF, Charité University 2005‐ ‐2014 of Medicine, Berlin Habilitation: Anatomy and Cell Biology Professorship at the Institute of Anatomy, Paracelsus Medical University, Salzburg and Nuremberg, Germany Teaching Activities: Dissection courses in gross anatomy, courses in histology, seminars and lessions in anatomy and cell biology Scholarships: 1996‐ ‐2008 Doctoral scholarship, Freie Universität Berlin Rahel‐Hirsch habilitation Scholarship, Charité Berlin 2

3 Ad hoc reviewer for Acta Biomaterialia, Annals of Anatomy, Arthritis Rheumatism, Arthritis Research Therapy, Berliner Münchner Tierärztliche Wochenschrift, BMC Musculoskeletal Disorders, Biomaterials, Biotechnology Progress, Biotechnology Bioengineering, Bone, Cell Tissue Research, Cell Biology and Toxicology, Cells Tissues Organs, Clinical Medicine Insights: Arthritis Musculoskeletal Disorders, Clinical Experimental Immunology, Connective Tissue Research, Cytokine, Drug Dev reviews, European Cells Materials, European J Histochemistry, Experimental Biology and Medicine, Fibrogenesis and Tissue Repair, Folia Biologica, Gene Therapy, Histochemistry Cell Biology, Histology Histopathology, International J Molecular Science, J Biochips Tissue Chips, J Cellular Biochemistry, J Tissue Engineering Regenerative Medicine, J Biomedicine and Biotechnology, J Biomed Mater Res Part A, J Immunol Methods, J Orthop Surg Res, J Nanomedicine, Materials Letters, Medicinal Chemistry, Osteoarthritis Cartilage, Oxid Antioxid Med Sci, PLOSone, Review Material Letters, The Lancet, Tissue Engineering Part A. 3

4 Scientific interests cartilage and tendon/ligament biology osteoarthritis (in vitro and in vivo models) tendon injury and healing (in vitro and in vivo models) interplay of anti‐ and proinflammatory cytokines in tendon and cartilage, co‐culture models cell‐ and biomaterial based cartilage and tendon repair: cartilage and tendon tissue engineering tendon/ligament ageing comparative anatomy/cell biology of tendons and cartilage subtypes 4

5 Cell biology ●characterization of human and animal-derived primary cells: chondrocytes, tenocytes, ligament cells, synovial fibroblasts, PBMCs and neutrophils ●three-dimensional cultures (pellet, organoid, spheroids, hydrogel and scaffold cultures, statical and dynamical seeding of biomaterials and self prepared cell-freed extracellular matrices ●interaction of musculoskeletal cells in co-cultures ●immunohistochemistry/-cytochemistry, immunofluorescence microscopy ●confocal laser scanning microscopy ●flow cytometry ●histology/histopathology of musculoskeletal tissues ●transmission-/scanning electron microscopy ●ELISA, assays (cytokines, proliferation, DMMB-assay, cell vitality, cytotoxicity, caspase activity, TUNEL…) ●SDS-PAGE and Western Blot Analyses ●RT PCR and Real Time (RTD)-PCR ●transfection of primary cells: adenoviral, lipofection, Electroporation ●gene silencing using siRNA Techniques 5

6 in the minipig and rabbi Animal models ●matrix assisted autologous chondrocytes transplantation (chondral vs. Osteochondral defects) Lohan A et al., 2014 ●partial Achilles tendon defect model in the rabbit (Stoll C et al., 2011, Biomaterials) ●nude mice xenograft model: in vivo Chondrogenesis and tendogenesis Lohan A et al., 2011: Histochem Cell Biol ●osteoarthritis model in the rat (surgically induced: MMT) Techniques t gross anatomical preparation techniques of human and animal-derived samples 6

7 Some recent publications (2014) Mrosewski I, Jork N, Gorte K, Conrad C, Wiegand E, Kohl B, Ertel W, John T, Oberholzer A, Kaps C, Schulze- Ta nzil G (2014). Regulation of OA associated key mediators by TNFα and IL-10: Effects of IL-10 overexpression in human synovial fibroblasts and a synovial cell line. Cell Tissue Res 357(1): The Phantom 5 Consortium (2014). A promoter level mammalian expression atlas. Nature 27;507(7493): Gröger D, Kerschnitzki M, Weinhart M, Schneider T, Kohl B, Wagermaier W, Schulze-Tanzil G, Fratzl P, Haag R (2014). Selectivity in Bone Targeting with Different Polyanionic Dendritic Dye Conjugates. Advanced Healthcare Materials 3(3): Lohan A, Marzahn U, El Sayed K, Haisch A, Müller RD, Kohl B, Stölzel K, Ertel W, John T, Schulze-Tanzil G (2014). Osteochondral cartilage defect repair using autologous orthotopic and heterotopic chondrocytes in the rabbit model. Ann Anat. 196(5): Girke G, Kohl B, Busch C, John T, Godkin O, Ertel W, Schulze-Tanzil G (2014). Tenocyte activation and regulation of complement factors in response to in vitro cell injury. Mol Immunol. 60(1): Stölzel K, Schulze-Tanzil G, Olze H, Schwarz S, Feldmann EM, Rotter N (2014). Immortalised human mesenchymal stem cells undergo chondrogenic differentiation in alginate and PGA/PLLA scaffolds. Cell Tissue Bank May 16 Hoyer M, Meier C, Breier A, Hahner J, Heinrich G, Drechsel N, Meyer M, Rentsch C, Garbe LA, Ertel W, Lohan A, Schulze-Tanzil G. In vitro characterization of self-assembled anterior cruciate ligament cell spheroids for ligament tissue engineering. Histochem Cell Biol Sep 26. Jagielski M, Wolf J, Marzahn U, Völker A, Lemke M, Meier C, Ertel W, Godkin O, Arens S, Schulze-Tanzil G (2014). The influence of IL-10 and TNFα on chondrogenesis of human mesenchymal stromal cells in three- dimensional cultures. Int J Mol Sci. 15(9): Hoyer M, Drechsel N, Meyer M, Meier C, Hinüber C, Breier A, Hahner J, Heinrich G, Rentsch C, Garbe LA, Ertel W, Schulze-Tanzil G, Lohan A. (2014). Embroidered polymer-collagen hybrid scaffold variants for ligament tissue engineering. Mater Sci Eng C Mater Biol Appl. 1;43:

8 Cartilage tissue engineering to characterize /compare the quality of tissue engineered cartilage produced by heterotopic chondrocytes seeded on polyglycolic acid (PGA) scaffolds in vitro and in vivo porcine auricular, nasoseptal articular chondrocytes dynamic culture 3 weeks → analysis Implanted ino nude mice 1, 6, 12 weeks → analysis Lohan A et al., 2011 Histochem Cell Biol. 136(1):57-69

9 Chondrogenesis by heterotopic chondrocytes in vivo 1 week6 weeks 12 weeks art i c ular explantation auricu lar n on-se ed ed nas o septal resorbed Lohan A et al., 2011 Histochem Cell Biol. 136(1):57-69

10 articular Chondrogenesis by heterotopic chondrocyte mono‐ and cocultures auriculararticular / auricularnasoseptalarticular / nasoseptal lig h t micr oscopy 100 µm live / death assay 100 µm alcian b l u e stai ning 20 µm articular / nasoseptal articular / auricular cel l tracking El Sayed K et al., 2013 J Tissue Eng Regen Med 7, µm

11 Cartilage repair in vivo intact articular cartilage chondral full thickness defect repair tissue after 6 month gl y c os a m inog l y ca n staining (r ed ) HE Lohan A et al., 2013, 195(5):

12 influence of autologous leukocytes on tenocytes? PBMCs inflammation tenocyte response neutrophils transwell system: indirect co-culture ●to cytokines? ●to leukocytes? ●to complement activation? ●to cell injury? Achilles tenocytes 10 ng/mL TNFα, 24 h 4x10 6 rabbit leukocytes 10 5 rabbit tenocytes MMP * 30 re l ativ e gene expression IL-6 30 IL-1β TNFα µm * ** * Al-Sadi O et al., Muscles Ligaments coco TNF  PBMC neutro. coco TNFTNF PBM C neu tro. coco TNF  PBMC neutro. coco Tendons J (3): TNFTNF PBMCPBMC neu tro.

13 tenocyte response to cell injury scratch assay computer guided reproducible plotting system „healing“ after injury 4 h8 h 16 h24 h green: vital, red: dead cells human Hamstring tenocytes Girke G et al., 2014 Mol Immunol 60(1):14-22.

14 tenocyte response to cell injury 10 tenocyte response 8 relative g ene expression * ●to cytokines? ●to leukocytes? ●to complement activation? ●to cell injury? 6 4 * * * 0 2 * * * c ontrol h 4 h4 h 24 h24 h 4 h4 h 24 h24 h 4 h4 h 24 h24 h 4 h4 h 24 h24 h 4 h4 h 24 h24 h 4 h4 h 24 h24 h 4 h4 h 24 h24 h C3aR C5aR CD46CD55 TNFαTNFα IL -1β MMP-1 human Hamstring tenocytes Girke G et al., 2014 Mol Immunol

15 decellularized tendon ECM scaffolds human tenocytes decellularization recellularization porcine AS tendon reseeded tendon ECM tendon repair? decellularized ECM HE Lohan et al., 2013, Schulze-Tanzil G et al., 2012 Connect Tissue Res. 54(4-5): Cells 1:1010.

16 tenocyte implantation in partial tendon defects (rabbit) rabbit Achilles Achilles tenocytes tendon expansion in rabbit tenocytes monolayer PGA-culture 1 week medial M. gastrocnemius tendon cell vitality implantation in tendon defect PGA-transplant empty defect implanted PGA 6 weeks healing? macroscopical and histological scoring green = vital red = dead Stoll C et al., 2011 Biomaterials 32:4806

17 tenocyte implantation in partial tendon defects (rabbit) co empty defect tenocytes + PGA PGA HE macroscopical score sulphat ed GAGs: bl u e * healt h y = 17points 0 5 empty PGA PGA+ cells histological score, 6 weeks hea l t h y = 20 po ints 15 elastic fibers Stoll C et al., 2011 Biomaterials 32: empty PGA PGA+ cells 0

18 Lapine anterior cruciate ligament (ACL) mean substance posterior cruciate ligament enthesis anterior cruciate ligament Dimensions: 15 mm x 3 mm x 4 mm Hoyer et al., Materials Science and Engineering 1;43:290-9.

19 embroidered scaffolds with collagen ● anterior cruciate ligament ( ACL) tissue engineering using embroidered scaffolds supplemented with collagen embroidery pattern PLA-CLPLA + PLA-CL PLA stitch length stitch angle loading axis ●Poly(lactic-co-ε-caprolacton)=PLA-CL, monofilament ●Poly-L-lactic acid = PLA, multifilament ●both materials combined Dimensions of the lapine ACL (15 mm x 3 mm x 4 mm) P(LA-CL) + collagen foam spheroid-based seeding vitality Hoyer et al., Materials Science and Engineering 1;43:290-9.

20 40 60 M Cellularity [nuclei/ µ m 2 ] ACL spheroid characterization HESafranin OAzAn da y s ABC Total sGAG *** N days [µg/1 0 6 cell s per s p heroid] µm s y da 7 da y s s t a tical D EF µm G HI 7 da ys d y na m ical ØØ sysy Total collagen O *** * days µm JK L [ µg /1 0 6 cell s p e r spheroid] da da y s statical 200µm statical Hoyer et al., Histochem Cell Biol. In press dynamical days

21 Collagen scaffolds seeded with ligament cells 7 d7 d green: vital, red: dead cells 21 Hoyer et al., Materials Science and Engineering 1;43:290-9.


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