الفهرس 
Introduction 1Only 14 pages are availabe for public view
 |  | from | 241 |  |  |
| | | from | 241 | | |
Abstract
Mesenchymal stem cells (MSCs) are multipotent adult stem cells
present in all tissues. As multipotent cells, MSCs can differentiate into
different tissues originating from mesoderm ranging from bone and
cartilage , to cardiac muscle .
Mesenchymal stem cells have been proposed as potential sources of stem
cells for regeneration of the CNS .
Thus, one of the goals of regenerative medicine is to ameliorate
irreversible destruction of brain tissue and spinal cord by harnessing the
power of stem cells to initiate neurogenesis in damaged areas of the brain
Accordingly, the source represented by bone marrow offers theoretically
unlimited therapeutically applications.
The aim of this work is to study neurogenesis using MSCs as
model of stem cells and then follow the cells as they form neurons.
The present study involved 28 patients, it was held at clinical
pathology department , Kasr El Eini medical school & EL Menofyia
University Hospital during the period from January 2010 to December
2011.
The samples were subjected to mononuclear cell separation by
ficoll-hypaque (1:2 ratios); The MNC suspensions were seeded at a
concentration of (1 x 106 cells / ml) in uncoated plastic tissue culture
flasks 25 cm2, incubated for 9 days in 5 ml of the fresh complete nutrient
medium. Then at day 9 the adherent cell layer with morphology of
SUMMARY
156
MSCs at the base of the flasks were harvested by trypsinization and
identified by flowcytometric analysis of CD44, Oct-3/4 and CD34.
The MSCs were differentiated into neural cells by a cocktail of
Retinoic acid (RA) , basic human Fibroblast Growth Factor (bFGF)
basic, basic Human Epidermal Growth Factor (bHEGF) and Insulin-like
Growth Factor I (IGF-I) and proved by morphology , nestin expression
and GFAP staining.
The results of this study revealed that the isolation success rate of
MSCs from bone marrow aspirate was from 95% to 98% identified by
morphology, CD44 expression, intracellular Oct 3/4 expression as
stemness marker and lack of specific cell surface markers of
hematopoietic cells like CD34. The expression of CD44 was ranging
from 60.30 to 98.9 of the cultured cells expressing the marker with mean
±SD (81.54±11.58).The expression of Oct3/4 was ranging between
29.5 to 89.8 of the cultured cells expressing the marker with mean ± SD
(56.12±17.37). The double expression of CD44 and oct3/4 was ranging
from 27 to 88.3 with mean ±SD (54.03±17.42). The expression of CD34
was ranging from 0.08 to 1.97 of the cultured cells expressing the marker
with mean ±SD (1.10 ± 0.47).
MSCs induced with neural inductive media show morphological
changes consistent with neurogenesis as compared to the symmetric
morphology of the uninduced cells, as shown by inverted microscope.
Induced MSCs showed high positive expression of nestin, ranged
between 15.8 to 65.4 % with mean ± SD 33.41± 15.58. In comparison to
non-induced MSCs which showed low positive expression of nestin
ranged between 3.53 to 15.80 % with mean ±SD 8.28 ± 3.62%.
SUMMARY
157
Induced cells showed positive staining with GFAP while uninduced
cells showed negative staining.
Regarding the growth curve pattern: the growth of induced cells
showed a plateau pattern, while the growth of uninduced cells showed a
linear pattern.