[Investigation of neural stem cell-derived donor contribution in the inner ear following blastocyst injection].
BACKGROUND
Utilising the enormous proliferation and multi-lineage differentiation potentials of somatic stem cells represents a possible therapeutical strategy for diseases of non-regenerative tissues like the inner ear. In the current study, the possibility of murine neural stem cells to contribute to the developing inner ear following blastocyst injection was investigated.


METHODS
Fetal brain-derived neural stem cells from the embryonic day 14 cortex of male mice were isolated and expanded for four weeks in neurobasal media supplemented with bFGF and EGF. Neural stem cells of male animals were harvested, injected into blastocysts and the blastocysts were transferred into pseudo-pregnant foster animals. Each blastocyst was injected with 5-15 microspheres growing from single cell suspension from neurospheres dissociated the day before. The resulting mice were investigated six months POST PARTUM for the presence of donor cells. Brainstem evoked response audiometry (BERA) was performed in six animals. To visualize donor cells Lac-Z staining was performed on sliced cochleas of two animals. In addition, the cochleas of four female animals were isolated and genomic DNA of the entire cochlea was analyzed for donor contribution by Y-chromosome-specific PCR.


RESULTS
All animals had normal thresholds in brainstem evoked response audiometry. The male-specific PCR product indicating the presence of male donor cells were detected in the cochleas of three of the four female animals investigated. In two animals, male donor cells were detected unilateral, in one animal bilateral.


CONCLUSIONS
The results suggest that descendants of neural stem cells are detectable in the inner ear after injection into blastocysts and possess the ability to integrate into the developing inner ear without obvious loss in hearing function.
Klinik für Hals-, Nasen- und Ohrenheilkunde, Kopf- und Halschirurgie der Ruhr-Universität Bochum am St. Elisabeth-Hospital, Bochum. stefan.volkenstein@ruhr-uni-bochum.de
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S
Volkenstein
D
D
Brors
S
S
Hansen
R
R
Mlynski
T C
TC
Dinger
A M
AM
Müller
S
S
Dazert
D000818:Animals; D001309:Auditory Threshold / Q000502:physiology; D002454:Cell Differentiation / Q000235:genetics / Q000502:physiology; D002540:Cerebral Cortex / Q000166:cytology; D003051:Cochlea / Q000473:pathology / Q000502:physiology; D004624:Embryo Transfer / Q000379:methods*; D053595:Embryonic Stem Cells / Q000637:transplantation*; D016057:Evoked Potentials, Auditory, Brain Stem / Q000502:physiology*; D005260:Female; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008815:Mice, Inbred Strains; D008822:Mice, Transgenic; D009474:Neurons / Q000637:transplantation*; D016133:Polymerase Chain Reaction; D011247:Pregnancy; D018183:Transplantation Chimera