BACKGROUND: Characterisation of stem cells by flow cytometry, their expansion and differentiation are presently of major interest for cell engineering as the basis of a therapeutic concept for transplantation. Haematopoietic stem cells (HSC) express CD34, the adhesion structure which binds 2L-selectin, CD117, a receptor for stem cell factor (SCF; c-kit ligand), and CD133, a transmembrane protein belonging to the family of mucoproteins. METHODS: The aim of the present investigation was the systematic investigation of proliferation and differentiation characteristics of umbilical cord blood stem cells (UCBSC) isolated by an immmunomagnetic separation system using CD133 antibody-coated microbeads and to evaluate the effects of different sera and various concentrations, as well as the effects of IL-3 and IL-6 on total cell expansion and differentiation of isolated CD133+ cells. Differentiation patterns were measured by flow cytometry. RESULTS: After the immmunomagnetic separation the yield of CD133+ cells was 0.45+/-0.17 x 10(6) cells/ml; the purity of isolated CD133+ cells was 95.79+/-1.86%. The majority of CD133+ cells coexpressed CD117. The most pronounced expansion during cultivation of 2 weeks was achieved in media supplemented with 12.5% horse serum plus 12.5% fetal calf serum (FCS) with stem cell factor and interleukine 3; the fold-expansion was 16.67+/-6.20. During the cultivation period, UCBSC were constantly loosing stem cell markers and differentiated towards myelo-monocyte lineage (granulocytes and/or monocytes). CONCLUSIONS: These in vitro results demonstrate that thorough investigation of various cultivation conditions is needed for successful expansion and differentiation of stem cells towards different lineages to be used therapeutically for replacement of damaged cells.
BACKGROUND: Characterisation of stem cells by flow cytometry, their expansion and differentiation are presently of major interest for cell engineering as the basis of a therapeutic concept for transplantation. Haematopoietic stem cells (HSC) express CD34, the adhesion structure which binds 2L-selectin, CD117, a receptor for stem cell factor (SCF; c-kit ligand), and CD133, a transmembrane protein belonging to the family of mucoproteins. METHODS: The aim of the present investigation was the systematic investigation of proliferation and differentiation characteristics of umbilical cord blood stem cells (UCBSC) isolated by an immmunomagnetic separation system using CD133 antibody-coated microbeads and to evaluate the effects of different sera and various concentrations, as well as the effects of IL-3 and IL-6 on total cell expansion and differentiation of isolated CD133+ cells. Differentiation patterns were measured by flow cytometry. RESULTS: After the immmunomagnetic separation the yield of CD133+ cells was 0.45+/-0.17 x 10(6) cells/ml; the purity of isolated CD133+ cells was 95.79+/-1.86%. The majority of CD133+ cells coexpressed CD117. The most pronounced expansion during cultivation of 2 weeks was achieved in media supplemented with 12.5% horse serum plus 12.5% fetal calf serum (FCS) with stem cell factor and interleukine 3; the fold-expansion was 16.67+/-6.20. During the cultivation period, UCBSC were constantly loosing stem cell markers and differentiated towards myelo-monocyte lineage (granulocytes and/or monocytes). CONCLUSIONS: These in vitro results demonstrate that thorough investigation of various cultivation conditions is needed for successful expansion and differentiation of stem cells towards different lineages to be used therapeutically for replacement of damaged cells.
Authors: Andreas J Flammer; Mario Gössl; Robert Jay Widmer; Martin Reriani; Ryan Lennon; Darrell Loeffler; Sarah Shonyo; Robert D Simari; Lilach O Lerman; Sundeep Khosla; Amir Lerman Journal: Eur Heart J Date: 2012-07-31 Impact factor: 29.983