Unravelling Heterogeneity of Amplified Human Amniotic Fluid Stem Cells Sub-Populations
Francesca Casciaro,
Silvia Zia,
Mattia Forcato,
Manuela Zavatti,
Francesca Beretti,
Emma Bertucci,
Andrea Zattoni,
Pierluigi Reschiglian,
Francesco Alviano,
Laura Bonsi,
Matilde Yung Follo,
Marco Demaria,
Barbara Roda,
Tullia Maraldi
Affiliations
Francesca Casciaro
Department of Biomedical, Metabolic and Neural Sciences, University of Modena and Reggio Emilia, 41124 Modena, Italy
Silvia Zia
Stem Sel srl., 40127 Bologna, Italy
Mattia Forcato
Department of Life Sciences, University of Modena and Reggio Emilia, 41124 Modena, Italy
Manuela Zavatti
Department of Biomedical, Metabolic and Neural Sciences, University of Modena and Reggio Emilia, 41124 Modena, Italy
Francesca Beretti
Department of Biomedical, Metabolic and Neural Sciences, University of Modena and Reggio Emilia, 41124 Modena, Italy
Emma Bertucci
Department of Medical and Surgical Sciences for Mothers, Children and Adults, University of Modena and Reggio Emilia, Azienda Ospedaliero Universitaria Policlinico, 41124 Modena, Italy
Andrea Zattoni
Department of Chemistry “G. Ciamician”, University of Bologna, 40125 Bologna, Italy
Pierluigi Reschiglian
Department of Chemistry “G. Ciamician”, University of Bologna, 40125 Bologna, Italy
Francesco Alviano
Unit of Histology, Embryology and Applied Biology, Department of Experimental, Diagnostic and Specialty Medicine, University of Bologna, 40125 Bologna, Italy
Laura Bonsi
Unit of Histology, Embryology and Applied Biology, Department of Experimental, Diagnostic and Specialty Medicine, University of Bologna, 40125 Bologna, Italy
Matilde Yung Follo
Cellular Signalling Laboratory, Department of Biomedical and Neuromotor Sciences, University of Bologna, 40125 Bologna, Italy
Marco Demaria
European Research Institute for the Biology of Ageing (ERIBA), University Medical Center Groningen (UMCG), University of Groningen, 9713 Groningen, The Netherlands
Barbara Roda
Department of Chemistry “G. Ciamician”, University of Bologna, 40125 Bologna, Italy
Tullia Maraldi
Department of Biomedical, Metabolic and Neural Sciences, University of Modena and Reggio Emilia, 41124 Modena, Italy
Human amniotic fluid stem cells (hAFSCs) are broadly multipotent immature progenitor cells with high self-renewal and no tumorigenic properties. These cells, even amplified, present very variable morphology, density, intracellular composition and stemness potential, and this heterogeneity can hinder their characterization and potential use in regenerative medicine. Celector® (Stem Sel ltd.) is a new technology that exploits the Non-Equilibrium Earth Gravity Assisted Field Flow Fractionation principles to characterize and label-free sort stem cells based on their solely physical characteristics without any manipulation. Viable cells are collected and used for further studies or direct applications. In order to understand the intrapopulation heterogeneity, various fractions of hAFSCs were isolated using the Celector® profile and live imaging feature. The gene expression profile of each fraction was analysed using whole-transcriptome sequencing (RNAseq). Gene Set Enrichment Analysis identified significant differential expression in pathways related to Stemness, DNA repair, E2F targets, G2M checkpoint, hypoxia, EM transition, mTORC1 signalling, Unfold Protein Response and p53 signalling. These differences were validated by RT-PCR, immunofluorescence and differentiation assays. Interestingly, the different fractions showed distinct and unique stemness properties. These results suggest the existence of deep intra-population differences that can influence the stemness profile of hAFSCs. This study represents a proof-of-concept of the importance of selecting certain cellular fractions with the highest potential to use in regenerative medicine.
