Successful elimination of non-neural cells and unachievable elimination of glial cells by means of commonly used cell culture manipulations during differentiation of GFAP and SOX2 positive neural progenitors (NHA) to neuronal cells.
BACKGROUND
Although extensive research has been performed to control differentiation of neural stem cells - still, the response of those cells to diverse cell culture conditions often appears to be random and difficult to predict. To this end, we strived to obtain stabilized protocol of NHA cells differentiation - allowing for an increase in percentage yield of neuronal cells.


RESULTS
Uncommitted GFAP and SOX2 positive neural progenitors - so-called, Normal Human Astrocytes (NHA) were differentiated in different environmental conditions to: only neural cells consisted of neuronal [MAP2+, GFAP-] and glial [GFAP+, MAP2-] population, non-neural cells [CD44+, VIMENTIN+, FIBRONECTIN+, MAP2-, GFAP-, S100beta-, SOX2-], or mixture of neural and non-neural cells.In spite of successfully increasing the percentage yield of glial and neuronal vs. non-neural cells by means of environmental changes, we were not able to increase significantly the percentage of neuronal (GABA-ergic and catecholaminergic) over glial cells under several different cell culture testing conditions. Supplementing serum-free medium with several growth factors (SHH, bFGF, GDNF) did not radically change the ratio between neuronal and glial cells--i.e., 1,1:1 in medium without growth factors and 1,4:1 in medium with GDNF, respectively.


CONCLUSIONS
We suggest that biotechnologists attempting to enrich in vitro neural cell cultures in one type of cells - such as that required for transplantology purposes, should consider the strong limiting influence of intrinsic factors upon extracellular factors commonly tested in cell culture conditions.
Witusik
Monika
M
Medical University of Lodz, Department of Molecular Pathology and Neuropathology, 8/10 Czechoslowacka str, 92-216 Lodz, Poland. monikawitusik@interia.pl
Piaskowski
Sylwester
S
Hulas-Bigoszewska
Krystyna
K
Zakrzewska
Magdalena
M
Gresner
Sylwia M
SM
Azizi
S Ausim
SA
Krynska
Barbara
B
Liberski
Pawel P
PP
Rieske
Piotr
P
D001253:Astrocytes / Q000187:drug effects / Q000502:physiology; D015415:Biomarkers / Q000032:analysis / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods; D002454:Cell Differentiation / Q000187:drug effects*; D019070:Cell Lineage / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000032:analysis; D005346:Fibroblast Growth Factors / Q000494:pharmacology; D005353:Fibronectins / Q000032:analysis; D005904:Glial Fibrillary Acidic Protein / Q000032:analysis; D024002:HMGB Proteins / Q000032:analysis; D006801:Humans; D018960:Hyaluronan Receptors / Q000032:analysis; D008869:Microtubule-Associated Proteins / Q000032:analysis; D009414:Nerve Growth Factors / Q000032:analysis / Q000494:pharmacology; D009419:Nerve Tissue Proteins / Q000032:analysis / Q000187:drug effects; D009457:Neuroglia / Q000166:cytology* / Q000502:physiology*; D009474:Neurons / Q000166:cytology* / Q000502:physiology*; D064568:S100 Calcium Binding Protein beta Subunit; D009418:S100 Proteins / Q000032:analysis; D055748:SOXB1 Transcription Factors; D013234:Stem Cells / Q000166:cytology / Q000187:drug effects* / Q000502:physiology*; D014157:Transcription Factors / Q000032:analysis; D014746:Vimentin / Q000032:analysis
D015415:Biomarkers; C497483:CD44 protein, human; D016895:Culture Media, Serum-Free; D004268:DNA-Binding Proteins; D005353:Fibronectins; D005904:Glial Fibrillary Acidic Protein; D024002:HMGB Proteins; D018960:Hyaluronan Receptors; C489788:MAP2 protein, human; D008869:Microtubule-Associated Proteins; D009414:Nerve Growth Factors; D009419:Nerve Tissue Proteins; D064568:S100 Calcium Binding Protein beta Subunit; D009418:S100 Proteins; C527782:SOX2 protein, human; D055748:SOXB1 Transcription Factors; D014157:Transcription Factors; D014746:Vimentin; D005346:Fibroblast Growth Factors