Different Tissue-Derived Stem Cells: A Comparison of Neural Differentiation Capability

Abstract

Background: Stem cells are capable of self-renewal and differentiation into a wide range of cell types with multiple clinical and therapeutic applications. Stem cells are providing hope for many diseases that currently lack effective therapeutic methods, including strokes, Huntington's disease, Alzheimer's and Parkinson's disease. However, the paucity of suitable cell types for cell replacement therapy in patients suffering from neurological disorders has hampered the development of this promising therapeutic approach.

Aim: The innovative aspect of this study has been to evaluate the neural differentiation capability of different tissue-derived stem cells coming from different tissue sources such as bone marrow, umbilical cord blood, human endometrium and amniotic fluid, cultured under the same supplemented media neuro-transcription factor conditions, testing the expression of neural markers such as GFAP, Nestin and Neurofilaments using the immunofluorescence staining assay and some typical clusters of differentiation such as CD34, CD90, CD105 and CD133 by using the cytofluorimetric test assay.

Results: Amniotic fluid derived stem cells showed a more primitive phenotype compared to the differentiating potential demonstrated by the other stem cell sources, representing a realistic possibility in the field of regenerative cell therapy suitable for neurodegenerative diseases.

Fig 1. Light Microscopy images at 40X magnification of BM-MSC at 2 D.I.V a), 6 D.I.V. b), 10 D.I.V, c), in presence of neural differentiation factors as reported in the section: ‘Materials and Methods’.

Fig 2. Expression of neural-like cells specific markers in differentiated BM-MSCs, at 10 D.I.V. evaluated by immunostaining for a) GFAP, b) Nestin, c) Neurofilaments, respectively.

Fig 3. Light Microscopy images at 40X magnification of CB-MSC at 2 D.I.V a), 6 D.I.V. b), 10 D.I.V, c), in presence of neural differentiation factors as reported in the section: ‘Materials and Methods’.

Fig 4. Expression of neural-like cells specific markers in differentiated CB-MSCs evaluated by immunostaining for a) GFAP, b) nestin c) Neurofilaments, respectively.

Fig 5. Light Microscopy images at 40X magnification of hE-MSC at 2 D.I.V a), 6 D.I.V. b), 10 D.I.V. c), in presence of neural differentiation factors as reported in the section: ‘Materials and Methods’.

Fig 6. Expression of neural-like cells specific markers in differentiated he-MSCs assessed by immunostaining for a) GFAP, b) Nestin, c) Neurofilaments respectively.

Fig 7. Light Microscopy images at 40X magnification of AF-MSC at 2 D.I.V a), 6 D.I.V. b), 10 D.I.V, c), in presence of neural differentiation factors as reported in the section: ‘Materials and Methods’d-e) Light Microscopy images at 100X magnification of AF-MSC at 10 D.I.V. in presence of neural differentiation factors as reported in the section: ‘Materials and Methods’.

Fig 8. Expression of neural-like cells specific markers in differentiated AF-MSCs at 10 D.I.V. assessed by immunostaining for a) GFAP, b) Nestin, c) Neurofilaments, respectively.

Fig 9. Na+ current traces evoked by a series 15mV voltage steps recorded from amniotic fluid differentiated neural-like stem cells.

Fig 10. Quantitative analysis in form of histograms of the appearance of neural cell types morphology for each stem cell sources, at 2, 6, 10 days of culture in vitro.

Fig 11. Immunostaining picture coupled with bright field image, obtained for the neural markers NESTIN, from AF-MSCs, 10 D.I.V. of neural differentiation treatment.

Fig 12. Representative flowcytometry diagrams of the CD34, CD90, CD105 and CD133 surface markers expression in mesenchymal stem cells from Bone Marrow.

The fluorescence intensity as number of counts and the distribution diagram of positive cells are reported in ordinate and in abscissa respectively. Data represent means +/- SE of 3 independent experiments.

Fig 13. Representative flowcytometry diagrams of the CD34, CD90, CD105, CD133 CD surface markers expression in mesenchymal stem cells from, Cord Blood, Human Endometrium, Amniotic Fluid.

The fluorescence intensity as number of counts and the distribution diagram of positive cells are reported in ordinate and in abscissa respectively. Data represent means +/- SE of 3 independent experiments.

Fig 14. Value percentage of the expression of CD 34/90/105/133/15/24/29/44 for each stem cell source analyzed.

Fig 15. mRNA expression levels for classical neural genes such as GFAP, NESTIN and NEUROFILAMENTS, investigated with RT-PCR on BM-MSCs, CB-MSCs, he-MSCs and AF-MSCs, after 2, 6 and 10 D.I.V. after neural trans-differentiation treatment.

Panel 1. Light Microscopy images at 40X magnification of BM-MSC (1a-c), CB-MSC (3a-c), hE-MSC (5a-c) AF-MSC (7a-c) at 2 D.I.V a), 6 D.I.V. b), 10 D.I.V, c), in presence of neural differentiation factors as reported in the section: ‘Materials and Methods’. Panel 2. Expression of neural-like cells specific markers in differentiated BM-MSC (2a-c), CB-MSC (4a-c), hE-MSC (6a-c) AF-MSC (8a-c) at 10 D.I.V. assessed by immunostaining for a) GFAP, b) Nestin, c) Neurofilaments, respectively.