9° ITALIAN CONGRESS SIGU "ITALIAN SOCIETY OF HUMAN GENETICS"

Venice, November 8-11, 2006

9° ITALIAN CONGRESS SIGU “ITALIAN SOCIETY OF HUMAN GENETICS”

Venice, November 8-11, 2006


AMNIOTIC FLUID MESENCHYMAL STEM CELLS AS CELL THERAPY?
Bolda F., Lanfranchi A., Mattarucchi E., Simoni G., Maggi F., Grati F., Pasquali F., Porta G., Porta F.
Centro di Terapia Cellulare e Genica delle Oncoemopatie Infantili, Laboratorio Cellule Staminali, Dipartimento di Pediatria, Ospedali Civili di Brescia, Brescia.
Dipartimento di Scienze Biomediche Sperimentali e Cliniche, Università dell’Insubria, Varese.
Laboratorio TOMA Advanced Biomedical Assays, Sezione di Genetica.
Introduction and targets:
Mesenchymal stem cells are multipotent precursors able to differentiate in osteogenic, cartilaginous and adipogenic tissues.
Osteogenesis Imperfecta is a genetic disease due to a defective production of type I collagen, the most important protein of the EM osteogenic tissue. Recent studies have reported that these cells can represent a good MSCs source and a target for genetic therapy in molecular defective cases.
The purpose of this study was to isolate MSCs from amniotic fluid and differentiate in osteoblasts.
Methods:
Cells derived from surnatant of amniocyte cultures and intended to karyotype analysis, were cultured in MSCs specific medium for 8 weeks, analyzed in flow citometry and RT-PCR to evaluate markers expression.
Results:
AFMSCs were successfully isolated and cultured. After 6, 7 and 8 weeks cells were screened by flow cytometry analysis demonstrating these cells are positive for SH3, SH4, CD90, CD44 and CD 29, weakly positive for CD105 but negative for CD45, CD34,CD25, CD31 and HLA-DR. Genetic expression levels are comparable to flow cytometry datas.
Osteogenic differentiation was induced during eighth week after increase of MSCs markers expression. After 3 weeks culture using specific stimulating factors were evaluated morphology and the capability of calcium salts deposition in EM using Alizarin red-S.
Conclusions:
The results obtained with immunoistochemistry and molecular analysis underline the presence of MSCs multipotent in amniotic fluid. This study also demonstrate the possibility to select, expand and differentiate MSCs from amniocytes cultures. AFMSC are a great potential for cellular and genetic therapy of genetic defects in connective tissues where an early intervention can be decisive.