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 /1/A

HEMATOPOIETIC STEM CELLS AND TRANSDIFFERENTIATIO N Dr. Péter Balogh and Dr. Péter Engelmann Transdifferentiation and regenerative medicine – Lecture 6 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 /1/A

TÁMOP /1/A Issues of hemopoietic differentiation Development of hemopoietic systemDevelopment of hemopoietic systemEmbryonicPostnatal Regenerating hemopoiesisRegenerating hemopoiesis Use of HSCs in non-hemopoietic regenerative medicineUse of HSCs in non-hemopoietic regenerative medicine

TÁMOP /1/A Ontogeny of embryonic hemopoietic tissues Pro-definitivePrimitiveMeta-definitiveMeso-definitiveAdult- definitive E10.5E7.5E9.0E8.25 Hemangioblast Hemogenic Endothelium MyeloidLymphoid-MyeloidCFU-s Neonatal HSC HSC Yolk sac Allantois Chorion Allantois YS blood islands Placenta AGM Liver Placenta AGM Liver Yolk sac Embryo pSP Umbilicalartery Vitellineartery Umbilicalartery Vitellineartery

TÁMOP /1/A Evolution of hemopoietic tissues in rodents Yolk sac Onset of circulation ProPrimitiveMetaMesoAdult E7.5E8.5E9.5E10.5E11.5E12.5E15E13.5E14.5birthE11Para-aorticSplanchnopleura Allantois AGM Aortic clusters Placenta

TÁMOP /1/A Characteristics of murine embryonic HSCs (AGM/YS/FL) Leukocyte surface markers:Leukocyte surface markers: Ly-6A(Sca- 1), c-kit+, CD34+, CD45+, Shared endothelial markers:Shared endothelial markers: CD31+, VE- cadherin+ TF:TF: Runx1+ SCL+ Gata-2+

TÁMOP /1/A Transcriptional induction of eHSCs Intrinsic signals: TF Runx1:Runx1: promotes fetal transition of hemogenic endothelium into hemopoietic cells GATA-2:GATA-2: sequential promotion of mesodermal specification, hemangioblast formation and erythroid differentiation

TÁMOP /1/A Extrinsic regulation of eHSCs Extrinsic signals: Extrinsic signals: interactions with other germ layer elements Yolk sacYolk sac (endoderm and mesoderm) The chorio-allantoic placentaThe chorio-allantoic placenta (mesoderm and trophectoderm) AGM regionAGM region (dorsal ectoderm, mesoderm and ventral endoderm) –Ventralizing factors –Ventralizing factors – promote hemopoiesis (VEGF, bFGF, TGFβ and BMP4) –Dosalizing factors –Dosalizing factors – antagonize hemopoiesis (EGF and TGFα)

TÁMOP /1/A Hemopoietic differentiation models Classic dichotomy model Modified classic model Myeloid-based model Myeloid potential M M Erythroid potential E E Megakaryocyte potential m m T-cell potential T T B-cell potential B B (CMLP) (CLP) (CMEP) (CLP) (CMEP) (CMLP) (CMEP)

TÁMOP /1/A Transcriptional regulation of early hemopoietic commitmentHemangioblasts Hemogenic endothelium Apoptosis SCL AML-1 GATA-2 Lmo SCL AML-1 GATA-2 Lmo PU.1/GATA-3/Ikaros PU.1/GATA-1 CMP HSC CLP Notch1 Ikaros HoxB4 GATA-2 Notch1 Ikaros HoxB4 GATA-2 Bcl-2 p21

TÁMOP /1/A Transcriptional regulation of myeloid differentiationMonocyte Neutrophil Eosinophil Erythrocyte Megakaryocyte GMP CMP HSC EMP PU.1 & GATA-1 PU.1 GATA-1/FOG ICSBP, PU.1 C/EBP , GATA-1 GATA-1 GATA-1, 2 C/EBP 

TÁMOP /1/A Transcriptional regulation of lymphoid differentiationCLP Pro-T PU.1 IL-7R  PU.1 IL-7R  GM-CSFR  Notch1 Pu.1, E2A PU.1 Pax5? EBF Pax5? EBF Pax5 Early pro-B Late pro-B Pre-pro-B V-D-JHD-JH B cell Monocyte HSC

TÁMOP /1/A HSC HSC HSC HSC Endothel Fibroblast Osteoblast G-CSF, GM-CSF TPO TGF  Steady-state and activated haemopoiesisEPO EPO IL-1  TNF  Macrophage Bacterial infection Inflammation Anemia Hypoxia HSC HSC HSC HSC SCF, FLT-3I, TPO Endothel G-CSF, GM-CSF Fibroblast Osteoblast Blood vessel

TÁMOP /1/A Human hemopoietic potential Intraembryonic sources and potential: D19:D19: HPP in embryo in the absence of detectable CD34+ hematopoietic cells, and spanned both lymphoid and myeloid lineages D24:D24: in the splanchnopleural mesoderm D27:D27: aorta with CD34+ progenitors Yolk sac: Yolk sac: only myelopoiesis starting at around the 3rd week

TÁMOP /1/A Other potential uses of hemopoietic stem cells Regenerative medicine in parenchymal tissues: muscle, neural tissues, liver, etc. Sources: adult or embryonic (umbilical vein mononuclear cells) Experimental settings: use of genetically marked cells or inducible Cre-Lox transgenic animals, and their detection in damaged/regenerating tissues

TÁMOP /1/A Summary Hemopoiesis is established in successive waves at various anatomical locations, where hemopoietic activities at different host tissues result in diverse cellular progeny. The hemopoietic committment is under the combined effects of endogenous programing and external signals, where latter elements may alter the steady-state hemopoiesis. (a)(b) (c)Hemopoietic stem cells may promote the regeneration of non-hemopoietic tissues by (a) promoting vascular repair, (b) tissue repair and (c) possible transdifferentiation.