[Changes of Wnt-3 protein during the proliferation of endogenous neural stem cells in neonatal rats with hypoxic-ischemic brain damage after hyperbaric oxygen therapy].
OBJECTIVE
Previous studies suggest that hyperbaric oxygen (HBO) treatment promotes the proliferation of neurocytes in neonatal rats following hypoxic-ischemic brain damage (HIBD). The Wnt signaling pathway is associated with neurogenesis. This study examined whether HBO promoted neural stem cells (NSCs) proliferation after HIBD, and whether that the proliferation correlated with Wnt-3 protein expression.


METHODS
Seven-day-old Sprague-Dawley rats were randomly divided into three groups: normal control, hypoxia-ischemia (HI), and HI-HBO. HI was induced by the ligation of left common carotid artery, followed by a 2-hr exposure to 8% O2 in the latter two groups. HBO was administered 3 hrs after HI in the HI-HBO group for continuous 7 days (2 atmospheres absolute, once daily). The proliferating NSCs in the subventricular zone (SVZ) was examined by BrdU/nestin immunofluorescence and the expression of Wnt-3 protein in NSCs was examined by nestin/Wnt-3 immunofluorescence at 6 and 24 hrs and at 3, 7 and 14 days of HI. The cellular expressions of nestin and Wnt-3 protein were analyzed by laser scanning confocal microscopy. The linear regression analysis was used to evaluate the correlation between cellular Wnt-3 and nestin protein. The expressions of nestin and Wnt-3 protein in the ischemic cerebral hemisphere were analyzed with Western blotting.


RESULTS
The number of BrdU/nestin positive cells in the SVZ increased 3 hrs after HBO therapy, peaked at 7 days and remained at a higher level until 14 days after HBO therapy in the HI-HBO group compared with the normal control and the HIBD groups. The level of Wnt-3 protein in NSCs increased significantly 3 hrs after HBO therapy, peaked at 3 days and remained at high levels until 14 days after HBO therapy in the HI-HBO group compared with the normal control and the HIBD groups. The level of cellular nestin protein was closely correlated with the level of cellular Wnt-3 protein (r = 0.893, P < 0.05). The Western blotting analysis demonstrated increased Wnt-3 and nestin protein expressions in the ischemic cerebral hemispheres.


CONCLUSIONS
HBO treatment promotes the proliferation of NSCs in HIBD neonatal rats, which is correlated with the activation of Wnt signaling.
Wang
Xiao-Li
XL
Department of Pediatrics, Xiangya Hospital, Central South University, Changsha 410008, China.
Yang
Yu-Jia
YJ
Wang
Qing-Hong
QH
Xie
Min
M
Yu
Xiao-He
XH
Liu
Chen-Tao
CT
Wang
Xia
X
D000818:Animals; D000831:Animals, Newborn; D015153:Blotting, Western; D001973:Bromodeoxyuridine / Q000378:metabolism; D049109:Cell Proliferation; D005260:Female; D005455:Fluorescent Antibody Technique; D006931:Hyperbaric Oxygenation; D020925:Hypoxia-Ischemia, Brain / Q000378:metabolism / Q000473:pathology / Q000628:therapy*; D007381:Intermediate Filament Proteins; D008297:Male; D009419:Nerve Tissue Proteins; D064231:Nestin; D009474:Neurons / Q000166:cytology*; D051381:Rats; D013234:Stem Cells / Q000166:cytology*; D051153:Wnt Proteins / Q000032:analysis*; D060508:Wnt3 Protein
D007381:Intermediate Filament Proteins; D009419:Nerve Tissue Proteins; C577486:Nes protein, rat; D064231:Nestin; D051153:Wnt Proteins; D060508:Wnt3 Protein; D001973:Bromodeoxyuridine