1. Manifestation of Novel Social Challenges of the European Union in the Teaching Material of Medical Biotechnology Master’s Programmes at the University of Pécs and at the University of Debrecen Identification number: TÁMOP-4.1.2-08/1/A-2009-0011

2. TISSUE REPAIR (1) Dr. Judit Pongrácz Three dimensional tissue cultures and tissue engineering – Lecture 17 Manifestation of Novel Social Challenges of the European Union in the Teaching Material of Medical Biotechnology Master’s Programmes at the University of Pécs and at the University of Debrecen Identification number: TÁMOP-4.1.2-08/1/A-2009-0011

3. TÁMOP-4.1.2-08/1/A-2009-0011 Stem cells in the bone marrow Hematopoetic stem cells (HSC) Mesenchymal stem cells (MSC) „Side population” Multipotent adult progenitor cells (MAPC)

4. TÁMOP-4.1.2-08/1/A-2009-0011 Therapeutic indications for stem cell therapy Cardiovascular and ischemic diseases Diabetes Hematopoietic diseases Liver diseases Orthopedics More than 25 000 hematopoietic SC transplantations are performed yearly

5. TÁMOP-4.1.2-08/1/A-2009-0011 Embryonic stem cell Advantages of Embryonic SC: Pluripotent easy to isolate highly productive in culture high capacity to integrate into fetal tissue Disadvantages: immune rejection Differentiation into inadequate cell types tumors induction Risk of contamination

6. TÁMOP-4.1.2-08/1/A-2009-0011 Germ stem cells Pluripotent Scarce harvesting source May develop embryonic teratoma cells Adult stem cells Advantages : Multipotent Greater differentiation potential Less likely induce immune rejection reactions May be stimulated by drugs Disadvantages Disadvantages : Scarce and difficult to isolate Grow slowly, differentiate poorly in culture Difficult to handle and produce in adequate amounts for transplantation

7. TÁMOP-4.1.2-08/1/A-2009-0011 Hemopoetic Cell Transplantation (HCT) Diseases treatable with HCT: Hemopoetic malignancies Autotransplantation Allogenic transplantation Hereditery immunodeficiencies Aplastic hematologic diseases BM-derived stem cells were detected in numerous organs after transplantation or injury (sex-mismatched transplantation)

8. TÁMOP-4.1.2-08/1/A-2009-0011 Hematopoetic stem cells Stem cell theory emerged in the 50’s Located in the BM CD34+, CD133+, c-kit+, CD38-, CD45- Worldwide databases available of BM donors

9. TÁMOP-4.1.2-08/1/A-2009-0011 3. Cryopreservation Blood or bone marrow is frozen to preserve it 4. Chemotherapy High dose chemotherapy and/or radiation is given to the patient General principles of stem cell therapy 2. Processing Blood or bone marrow is processed in the laboratory to purify and concentrate the stem cells 1. Collection Stem cells are collected from the patient’s bone marrow or blood 5. Reinfusion Thawed stem cells are reinfused into the patient

10. TÁMOP-4.1.2-08/1/A-2009-0011 Bone repair with stem cells Baseline G-CSF mobilization Monocyte HSPC Bone Osteolineage cells Blood Perivascular cells G-CSFR G-CSF VCAM-1 CXCL12 VLA-4 c-kit kitL CXCR4 C3a uPAR Blood Monocyte HSPC Bone

11. TÁMOP-4.1.2-08/1/A-2009-0011 Cartilage regeneration Cultured chondrocytes injected under patch Periosteal patch harvested from tibia Tissue culture of isolated stem cells in bioreactors TGF- 

12. TISSUE REPAIR (2) Dr. Judit Pongrácz Three dimensional tissue cultures and tissue engineering – Lecture 18 Manifestation of Novel Social Challenges of the European Union in the Teaching Material of Medical Biotechnology Master’s Programmes at the University of Pécs and at the University of Debrecen Identification number: TÁMOP-4.1.2-08/1/A-2009-0011

13. TÁMOP-4.1.2-08/1/A-2009-0011 Liver repair Functions of the liver: Metabolism Energy homeostasis, glycogen production and storage Detoxification Bile production Plasma protein synthesis In case of injury the intrinsic repair capacity may be insufficient. thelong-term therapeutic option istransplantation. Today the long-term therapeutic option for liver failure is transplantation.

14. TÁMOP-4.1.2-08/1/A-2009-0011 Causes of liver failure Toxic compounds Drugs Alcohol Chemicals Infectious diseases Hepatitis viruses Bacteria Parasites (malaria) Intrinsic causes Genetic Autoimmune (primary biliary cirrhosis)

15. TÁMOP-4.1.2-08/1/A-2009-0011 Liver transplantation Today the only long term therapy for liver failure Immuno-suppression needed Patient is prone to infections Serious side-effects of immunosuppressant drugs Worldwide shortage of donors

16. TÁMOP-4.1.2-08/1/A-2009-0011 Potential in cellular therapy of liver failure Less invasive than organ transplantation Can be repeated multiple times Limiting factor is the inability of –Producing a sufficiently large number of hepatocytes –Keep hepatocytes ready for use on- demand Expansion of existing hepatocytes Using stem cells to differentiate hepatocytes

17. TÁMOP-4.1.2-08/1/A-2009-0011 Using stem cells for liver regeneration BM stem cells Hematopoetic SC Mesenchymal SC Stem/progenitor cells in the liver Embryonic stem cells

18. TÁMOP-4.1.2-08/1/A-2009-0011 HSC and liver regeneration BM resident HSC contains a population expressing SC markers (CD34, c-kit) and a-fetoprotein (  FP, liver progenitor cell marker) When BMSC were cultured in the presence of hepatocyte growth factor (HGF) showed hepatocyte-like characteristics These experiments were done in rodents and humans

19. TÁMOP-4.1.2-08/1/A-2009-0011 MSC and liver regeneration MSC subpopulation multipotent adult progenitor cells (MAPC) Human MAPC differentiated into hepatocyte-like cells in the presence of HGF Substantial delay of the differentiation Therefore the potential of clinical usage is questionable

20. TÁMOP-4.1.2-08/1/A-2009-0011 Cultured HSC differentiate into hepatocytes Autologous cultured hepatocytes Patient Liver damage Blood or BM Isolation of Stem Cells Cell seeding/Inoculation Culture + HGF Cellular differentiation Hepatocytes Hepatocytes

21. TÁMOP-4.1.2-08/1/A-2009-0011 Human trials Cases: Liver cancer Hepatitis B or C Cirrhosis (alcoholic, drug or primer)BMSC: Unsorted mononuclear cells Sorted CD34+ or CD133+ cells Route of administration: Peripheral vein Portal vein Hepatic arteryResults: Mostly well tolerated Improvements in Child-Pugh score, albumin, AST, ALP, bilirubin, clotting

22. TÁMOP-4.1.2-08/1/A-2009-0011 Injury-induced differentiation of BMSC (animal models) Murine model: sex-mismatched donor BMSC transplantation in a hereditary lethal liver disease model (tyrosinemia, FAH-/-) 1/3 of the hepatocytes were of donor origin after 22 weeks In an induced liver cirrhosis model, 25% of hepatocytes were of donor origin after just 4 weeks

23. TÁMOP-4.1.2-08/1/A-2009-0011 Conclusions Clinical application is not well established and not ready for routine therapy Which cases? Which cells? Which administration route? Risks and benefits of autologous cellular therapy in liver failure?