Vimentin expression is retained in erythroid cells differentiated from human iPSC and ESC and indicates dysregulation in these cells early in differentiation
Abstract Background Pluripotent stem cells are attractive progenitor cells for the generation of erythroid cells in vitro as have expansive proliferative potential. However, although embryonic (ESC) and induced pluripotent (iPSC) stem cells can be induced to undergo erythroid differentiation, the ma...
| Main Authors: | , , , , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
BMC
2019-04-01
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| Series: | Stem Cell Research & Therapy |
| Online Access: | http://link.springer.com/article/10.1186/s13287-019-1231-z |
| _version_ | 1828857616164454400 |
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| author | Kongtana Trakarnsanga Daniel Ferguson Deborah E. Daniels Rebecca E. Griffiths Marieangela C. Wilson Kathryn E. Mordue Abi Gartner Tatyana N. Andrienko Annabel Calvert Alison Condie Angela McCahill Joanne C. Mountford Ashley M. Toye David J. Anstee Jan Frayne |
| author_facet | Kongtana Trakarnsanga Daniel Ferguson Deborah E. Daniels Rebecca E. Griffiths Marieangela C. Wilson Kathryn E. Mordue Abi Gartner Tatyana N. Andrienko Annabel Calvert Alison Condie Angela McCahill Joanne C. Mountford Ashley M. Toye David J. Anstee Jan Frayne |
| author_sort | Kongtana Trakarnsanga |
| collection | DOAJ |
| description | Abstract Background Pluripotent stem cells are attractive progenitor cells for the generation of erythroid cells in vitro as have expansive proliferative potential. However, although embryonic (ESC) and induced pluripotent (iPSC) stem cells can be induced to undergo erythroid differentiation, the majority of cells fail to enucleate and the molecular basis of this defect is unknown. One protein that has been associated with the initial phase of erythroid cell enucleation is the intermediate filament vimentin, with loss of vimentin potentially required for the process to proceed. Methods In this study, we used our established erythroid culture system along with western blot, PCR and interegation of comparative proteomic data sets to analyse the temporal expression profile of vimentin in erythroid cells differentiated from adult peripheral blood stem cells, iPSC and ESC throughout erythropoiesis. Confocal microscopy was also used to examine the intracellular localisation of vimentin. Results We show that expression of vimentin is turned off early during normal adult erythroid cell differentiation, with vimentin protein lost by the polychromatic erythroblast stage, just prior to enucleation. In contrast, in erythroid cells differentiated from iPSC and ESC, expression of vimentin persists, with high levels of both mRNA and protein even in orthochromatic erythroblasts. In the vimentin-positive iPSC orthochromatic erythroblasts, F-actin was localized around the cell periphery; however, in those rare cells captured undergoing enucleation, vimentin was absent and F-actin was re-localized to the enucleosome as found in normal adult orthrochromatic erythroblasts. Conclusion As both embryonic and adult erythroid cells loose vimentin and enucleate, retention of vimentin by iPSC and ESC erythroid cells indicates an intrinsic defect. By analogy with avian erythrocytes which naturally retain vimentin and remain nucleated, retention in iPSC- and ESC-derived erythroid cells may impede enucleation. Our data also provide the first evidence that dysregulation of processes in these cells occurs from the early stages of differentiation, facilitating targeting of future studies. |
| first_indexed | 2024-12-13T01:40:13Z |
| format | Article |
| id | doaj.art-ab66fdf60c4c4d08878afa9f330cc664 |
| institution | Directory Open Access Journal |
| issn | 1757-6512 |
| language | English |
| last_indexed | 2024-12-13T01:40:13Z |
| publishDate | 2019-04-01 |
| publisher | BMC |
| record_format | Article |
| series | Stem Cell Research & Therapy |
| spelling | doaj.art-ab66fdf60c4c4d08878afa9f330cc6642022-12-22T00:03:47ZengBMCStem Cell Research & Therapy1757-65122019-04-0110111010.1186/s13287-019-1231-zVimentin expression is retained in erythroid cells differentiated from human iPSC and ESC and indicates dysregulation in these cells early in differentiationKongtana Trakarnsanga0Daniel Ferguson1Deborah E. Daniels2Rebecca E. Griffiths3Marieangela C. Wilson4Kathryn E. Mordue5Abi Gartner6Tatyana N. Andrienko7Annabel Calvert8Alison Condie9Angela McCahill10Joanne C. Mountford11Ashley M. Toye12David J. Anstee13Jan Frayne14School of Biochemistry, University of BristolSchool of Biochemistry, University of BristolSchool of Biochemistry, University of BristolBristol Institute for Transfusion Sciences, National Health Service Blood and Transplant (NHSBT)School of Biochemistry, University of BristolSchool of Biochemistry, University of BristolSchool of Biochemistry, University of BristolSchool of Biochemistry, University of BristolSchool of Biochemistry, University of BristolScottish National Blood Transfusion Service, Jack Copland Centre, Heriot Watt Research ParkScottish National Blood Transfusion Service, Jack Copland Centre, Heriot Watt Research ParkScottish National Blood Transfusion Service, Jack Copland Centre, Heriot Watt Research ParkSchool of Biochemistry, University of BristolBristol Institute for Transfusion Sciences, National Health Service Blood and Transplant (NHSBT)School of Biochemistry, University of BristolAbstract Background Pluripotent stem cells are attractive progenitor cells for the generation of erythroid cells in vitro as have expansive proliferative potential. However, although embryonic (ESC) and induced pluripotent (iPSC) stem cells can be induced to undergo erythroid differentiation, the majority of cells fail to enucleate and the molecular basis of this defect is unknown. One protein that has been associated with the initial phase of erythroid cell enucleation is the intermediate filament vimentin, with loss of vimentin potentially required for the process to proceed. Methods In this study, we used our established erythroid culture system along with western blot, PCR and interegation of comparative proteomic data sets to analyse the temporal expression profile of vimentin in erythroid cells differentiated from adult peripheral blood stem cells, iPSC and ESC throughout erythropoiesis. Confocal microscopy was also used to examine the intracellular localisation of vimentin. Results We show that expression of vimentin is turned off early during normal adult erythroid cell differentiation, with vimentin protein lost by the polychromatic erythroblast stage, just prior to enucleation. In contrast, in erythroid cells differentiated from iPSC and ESC, expression of vimentin persists, with high levels of both mRNA and protein even in orthochromatic erythroblasts. In the vimentin-positive iPSC orthochromatic erythroblasts, F-actin was localized around the cell periphery; however, in those rare cells captured undergoing enucleation, vimentin was absent and F-actin was re-localized to the enucleosome as found in normal adult orthrochromatic erythroblasts. Conclusion As both embryonic and adult erythroid cells loose vimentin and enucleate, retention of vimentin by iPSC and ESC erythroid cells indicates an intrinsic defect. By analogy with avian erythrocytes which naturally retain vimentin and remain nucleated, retention in iPSC- and ESC-derived erythroid cells may impede enucleation. Our data also provide the first evidence that dysregulation of processes in these cells occurs from the early stages of differentiation, facilitating targeting of future studies.http://link.springer.com/article/10.1186/s13287-019-1231-z |
| spellingShingle | Kongtana Trakarnsanga Daniel Ferguson Deborah E. Daniels Rebecca E. Griffiths Marieangela C. Wilson Kathryn E. Mordue Abi Gartner Tatyana N. Andrienko Annabel Calvert Alison Condie Angela McCahill Joanne C. Mountford Ashley M. Toye David J. Anstee Jan Frayne Vimentin expression is retained in erythroid cells differentiated from human iPSC and ESC and indicates dysregulation in these cells early in differentiation Stem Cell Research & Therapy |
| title | Vimentin expression is retained in erythroid cells differentiated from human iPSC and ESC and indicates dysregulation in these cells early in differentiation |
| title_full | Vimentin expression is retained in erythroid cells differentiated from human iPSC and ESC and indicates dysregulation in these cells early in differentiation |
| title_fullStr | Vimentin expression is retained in erythroid cells differentiated from human iPSC and ESC and indicates dysregulation in these cells early in differentiation |
| title_full_unstemmed | Vimentin expression is retained in erythroid cells differentiated from human iPSC and ESC and indicates dysregulation in these cells early in differentiation |
| title_short | Vimentin expression is retained in erythroid cells differentiated from human iPSC and ESC and indicates dysregulation in these cells early in differentiation |
| title_sort | vimentin expression is retained in erythroid cells differentiated from human ipsc and esc and indicates dysregulation in these cells early in differentiation |
| url | http://link.springer.com/article/10.1186/s13287-019-1231-z |
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