DEPARTAMENTO DE CIÊNCIAS DA VIDA ... - Estudo Geral

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I.4 – Objectives 29
Chapter II 31
II – Materials and Methods 33
II.1 - Materials 33
II.1.1 - Cell culture 33
II.1.2 – Polyacrylamide hydrogels 33
II.1.3 - Immunocytochemistry 33
II.1.4 – Biological material 34
II.1.4.1 - Umbilical Cord Samples 34
II.1.4.2 - human Mesenchymal Stem Cells (hMSCs) 34
II.2 - Methods 34
II.2.1 – Isolation of Mesenchymal Stem Cells (MSCs) from
Umbilical Cord Fragments (UCFs) in Tissue Culture Plates
(TCPs) and Polyacrylamide hydrogels 34
II.2.2 - Cell culture 35
II.2.3 - Cryopreservation of MSCs 35
II.2.4 - Phenotypic characterization of UCM-MSCs 35
II.2.5 - Preparation of polyacrylamide hydrogels 36
II.2.6 Differentiation protocols 38
II.2.6.1 adapted from Engler, et al. 2006 38
II.2.7 - Rheological characterization of polyacrylamide hydrogels 38
II.2.8 - Immunocytochemistry 39
II.2.9 - Statistical analysis 39
Chapter III 41
III – Results 43
III.1 – Rheological Characterization of Polyacrylamide Hydrogels 43
III.2 – Cell Adhesion to Polyacrylamide Hydrogels functionalized with Collagen I
(COL-1) 44
III. 3 – hMSCs Proliferation Assay 46
III.4 – Isolation and Proliferation of MSCs from Human Umbilical Cord Isolation
of hMSC from the Wharton Jelly on Polyacrylamide Hydrogels 47
III.5 - Immunophenotypic characterization of UCM-MSC 48