Finnish Red Cross Blood Service and Faculty of Biociences, Department of Biological and Environmental Sciences, Division of Genetics, University of Helsinki, Finland Transcriptome and Glycome Profiling of Human Hematopoietic Cd133+ and Cd34+ Cells

................................................................................. 8 REVIEW OF THE LITERATURE ........................................................ 10 Hematopoietic stem cells .............................................................. 10 Phenotypic characterization ...................................................... 11 Functional characterization ....................................................... 14 Cord blood-derived hematopoietic stem cells ................................... 15 Collection and banking ............................................................. 15 Cord blood transplantation ........................................................ 16 Ex vivo expansion ................................................................... 21 Transcriptome profi ling ................................................................. 22 Methods for transcriptome profi ling ............................................ 22 Gene expression profi ling of murine hematopoietic stem cells ........ 26 Gene expression profi les of human hematopoietic stem cells ......... 28 Glycosylation .............................................................................. 29 N-glycans and N-glycan synthesis .............................................. 29 Glycosylation in hematopoiesis .................................................. 33 Effect of glycosylation on homing............................................... 33 AIMS OF THE STUDY .................................................................... 35 MATERIALS AND METHODS ........................................................... 36 Ethics .................................................................................... 36 Cells ...................................................................................... 36 Methods ................................................................................. 37 Gene expression reanalysis ....................................................... 39 Annotation ............................................................................. 40 RESULTS AND DISCUSSION .......................................................... 41 CD133+ and CD34+ cells have similar gene expression (studies I and II) ..................................................................... 41 Novel molecular markers for HSCs could not be found (study II) .... 42 Differential gene expression between CD133+ and CD34+ cells suggests that CD133+ cells are more primitive (studies I and II) .... 43 CD133+ and CD34+ cells demonstrate similar total colony forming potential (studies I and II) ........................................................ 44 Reanalysis of the original gene expression data focuses on the similarities between CD133+ and CD34+ cells .............................. 45 CD133+ and CD34+ cells have divergent roles in hematopoietic regeneration (studies I and II) .................................................. 49 N-glycosylation-related gene expression is similar in CD133+ and CD34+ cells and correlates with the N-glycome profi le (study III) ... 52 Characteristic mannosylation pattern of CD133+ and CD34+ cells (study III) .............................................................................. 52 Differential expression of N-acetylglucosaminyl-transferase genes and the enrichment of biantennary complex type N-glycans in CD133+ cells (study III) ............................................................ 54 Despite the differential expression of genes coding for β1,4-galactosyltransferases there are no differences in the β1,4-galactosylation of CD133+ and CD133cells (study III) ..................................... 55 CD133+ and CD34+ cell specifi c biosynthesis of N-glycan terminal epitopes (study III) ................................................................. 55 CONCLUSIONS ............................................................................ 58 ACKNOWLEDGEMENTS ................................................................. 60 REFERENCES .............................................................................. 62