Dopaminergic differentiation of human neural stem cells mediated by co-cultured rat striatal brain slices.
Properly committed neural stem cells constitute a promising source of cells for transplantation in Parkinson's disease, but a protocol for controlled dopaminergic differentiation is not yet available. To establish a setting for identification of secreted neural compounds promoting dopaminergic differentiation, we co-cultured cells from a human neural forebrain-derived stem cell line (hNS1) with rat striatal brain slices. In brief, coronal slices of neonatal rat striatum were cultured on semiporous membrane inserts placed in six-well trays overlying monolayers of hNS1 cells. After 12 days of co-culture, large numbers of tyrosine hydroxylase (TH)-immunoreactive, catecholaminergic cells could be found underneath individual striatal slices. Cell counting revealed that up to 25.3% (average 16.1%) of the total number of cells in these areas were TH-positive, contrasting a few TH-positive cells (<1%) in non-induced areas. The presence of dopamine in the conditioned culture medium was confirmed by HPLC analysis. Interestingly, not all striatal slice cultures induced TH-expression in underlying hNS1 cells. Common to TH-inductive cultures was, however, the presence of degenerating, necrotic areas, suggesting that factors released during striatal degeneration were responsible for the dopaminergic induction of the hNS1 cells. Ongoing experiments aim to identify such factors by comparing protein profiles of media conditioned by degenerating (necrotic) versus healthy striatal slice cultures.
Anwar
Mohammad Raffaqat
MR
Department of Anatomy and Neurobiology, Institute of Medical Biology, University of Southern Denmark, Odense C, Denmark.
Andreasen
Christian Maaløv
CM
Lippert
Solvej Kølvraa
SK
Zimmer
Jens
J
Martinez-Serrano
Alberto
A
Meyer
Morten
M
D000818:Animals; D000831:Animals, Newborn; D002452:Cell Count / Q000379:methods; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology*; D048429:Cell Size / Q000187:drug effects; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D018920:Coculture Techniques / Q000379:methods; D003342:Corpus Striatum / Q000502:physiology*; D017077:Culture Media, Conditioned / Q000494:pharmacology; D004298:Dopamine / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D006801:Humans; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000378:metabolism; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D009924:Organ Culture Techniques; D051381:Rats; D017207:Rats, Sprague-Dawley; D013234:Stem Cells / Q000187:drug effects / Q000502:physiology*; D013997:Time Factors; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism
D017077:Culture Media, Conditioned; D009419:Nerve Tissue Proteins; D007770:L-Lactate Dehydrogenase; D014446:Tyrosine 3-Monooxygenase; D004298:Dopamine