Bcl-2 enhances neurogenesis and inhibits apoptosis of newborn neurons in adult rat brain following a transient middle cerebral artery occlusion.

To determine whether Bcl-2 could influence adult neurogenesis and prevent apoptosis of newborn neurons, we injected Bcl-2 expressing plasmid into the lateral ventricle of rat brain immediately following a 30-min occlusion of the middle cerebral artery (MCAO). We found that Bcl-2 increased neural progenitor cells (BrdU+-DCX+) in the ipsilateral striatum, newborn immature neurons (BrdU+-Tuj-1+) and newborn mature neurons (BrdU+-MAP-2+) in the ipsilateral striatum and frontal cortex at 1 to 4 weeks following MCAO. Bcl-2 overexpression promoted development of newborn neurons into GABAergic and cholinergic neurons in the ipsilateral striatum. Moreover, Bcl-2 significantly decreased the apoptosis of newborn neurons, determined by double staining of Tuj-1 and activated caspase-3 (Tuj-1+-Casp+). These results indicate that overexpression of Bcl-2 in adult rat brain enhances neurogenesis and survival of newborn neurons. Increasing neurogenesis and preventing the death of newborn neuron may be a strategy to aid in the repair of adult brain after stroke.

[Protective effects of transplantation of hCNTF-transfected cell line on axotomy-induced degeneration of retinal ganglion cells].

OBJECTIVE: To investigate the protective effects of transplantation of cell line stably expressing and secreting human ciliary neurotrophic factor (hCNTF) on the degeneration of retinal ganglion cells (RGCs) after optic nerve transection. METHODS: Plasmid encoding hCNTF was transfected into human lung fibroblast (HLF) cell line, then the stably transfected clones were selected with methopterin. In adult SD rats, RGCs were labeled with retrograde axonal tracer fluorogold (FG) injected to their targets including dorsal lateral geniculate nuclei (dLGN) and superior colliculi (SC). Seven days later, the optic nerve was transected alone or in combination with transplantation of HLF cells. Five, 14, 17, 21 and 28 days after axotomy, the retinas were mounted and examined under fluorescence microscope to observe the RGCs. RESULTS: Compared to the controls, the density of RGC in axotomy group decreased by 67.44% and 82.73% on the 14th and 28th day, respectively. In the eyes with hCNTF-transfected HLF cells transplantation, RGC density was higher than that of the axotomy group on the 5th, 17th, 21st day after axotomy (P < 0.05). On the 5th day, the morphology of RGC in the hCNTF group remained the same as the controls, whereas the morphology of RGC in the axotomy alone group began to change. CONCLUSIONS: hCNTF administered at the time of optic nerve transection can protect RGC from degeneration, increasing the numbers of surviving RGCs and delaying the death of RGCs.

[Expression of transient receptor potential channel 4 in striatum and hippocampus of rats is increased after focal cerebral ischemia].

This paper was designed in middle cerebral artery occlusion (MCAO) model of rats, to explore the role of transient receptor potential channel 4 (TRPC4) as Ca(2+) selective channel by detecting the changes of the expression of TRPC4 in different parts of cerebral tissues under the condition of focal cerebral ischemia. The rats were sacrificed after MCAO surviving time 6 h, 12 h, 1 d, 3 d. As determined by Western blot, the expressions of TRPC4 in striatum and hippocampus of 12 h, 1 d, 3 d groups were significant higher than that in the control group (P<0.05). Immunohistochemical staining showed that the TRPC4 immunoreactive substances were present in the membrane of neurons. Compared with the control group, immunostaining positive cells increased in hippocampus and striatum of cerebral ischemia groups. The TRPC4 immunostaining positive cells increased significantly in 1d-group and 3d-group (P<0.05). It suggests that as a Ca(2+) selective channel, the variance of the expression of TRPC4 may play a role in acute and delayed neuronal injury in focal cerebral ischemia.

Role of vascular endothelial growth factor in neuronal DNA damage and repair in rat brain following a transient cerebral ischemia.

The antisense knockdown technique and confocal laser scanning microscopic analysis were used to elucidate vascular endothelial growth factor (VEGF) induction and its effect on DNA damage and repair in rat brain following a transient middle cerebral artery occlusion. Immunohistochemical study and in situ hybridization showed that the expression of VEGF and its mRNA was enhanced in the ischemic core and penumbra of ischemic brain. Western blot analysis further illustrated that VEGF induction was time-dependently changed in these areas. Double-staining analysis indicated that VEGF-positive staining existed in the neuron, but not in the glia, and it colocalized with excision repair cross-complementing group 6 (ERCC6) mRNA, a DNA repair factor. VEGF antisense oligodeoxynucleotide infusion reduced VEGF induction and resulted in an enlargement of infarct volume of the brain caused by ischemia. Moreover, it also increased the number of DNA damaged cells and lessened the induction of ERCC6 mRNA in ischemic brains. These results suggest that the induction of endogenous VEGF in ischemic neurons plays a neuroprotective role probably associated with the expression of ERCC6 mRNA.

[The expression of nestin in ischemia-injured brain of adult rat].

Immunohistochemistry and double immunofluorescent labeling techniques combined with confocal laser scanning microscope analysis were used to investigate the characteristic spatial induction profile of nestin following a transient middle cerebral artery occlusion in adult rat brain. The results showed that nestin was induced in ischemic core at 1 day after reperfusion. In addition to ischemic core, the expression of nestin increased in peri-ischemic I, II and III regions at 3 days and 1 week, then it decreased and narrowed along the rim of ischemic core 2 weeks after reperfusion. Double immunofluorescent labeling showed that nestin positive cells were mostly co-stained with GFAP,a astrocyte marker, in peri-ischemic I region 3 days after reperfusion. At 2 weeks, however nestin cells showed a long process and the cells double stained with nestin and NSE,a neuonal specific marker,increased in the ischemic brain. The results suggest that cerebral ischemia induces nestin expression in damaged neurons which might favor the neuroprotection against ischemic damage.

Melatonin attenuates MPTP-induced dopaminergic neuronal injury associated with scavenging hydroxyl radical.

To clarify the relationship between melatonin's hydroxyl radical (*OH) scavenging ability and its protective effect in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced neuronal injury, in the present study, the salicylate trapping method combined with high-performance liquid chromatography (HPLC)-electrochemical detection were used to measure the contents of dihydroxybenzoic acid (DHBA) and dopamine (DA) in brain tissues of C57BL/6 mice. Immunocytohistochemistry was used to detect tyrosine hydroxylase (TH)-like positive staining neurons. Results show that MPTP treatment induced an increase in the content of DHBA and decrease in the level of DA as well as the number of TH positive stained neurons in the mouse brain. However, melatonin dose-dependently inhibited the increase of DHBA levels in ventral midbrain tissues, the decrease of DA content and the loss of dopaminergic neurons. Moreover, the relationship between the changes of DHBA and DA levels in the brain of mice following MPTP and melatonin treatment showed a statistically significant negative correlation. Present results suggest that melatonin can ameliorate MPTP-induced dopaminergic neuronal lesions probably, at least partially, because of its inhibition of *OH generation.