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Neonatal hypoxic-ischemic encephalopathy (HIE) always results in severe neurologic dysfunction, nevertheless effective treatments are limited and the underlying mechanism also remains unclear. In this study, we firstly established the neonatal HIE model in the postnatal day 7 SD rats, Zea-Longa score and TTC staining were employed to assess the neurological behavior and infarct volume of the brain after cerebral hypoxia-ischemia(HI). Afterwards, protein chip was adopted to detect the differential proteins in the right cortex, hippocampus and lung, ultimately, PDGF was noticed. Then, immunohistochemistry, immunofluorescence double staining ofNeuN/PDGF, and western blot were used to validate the expression level of PDGF in the cortex and hippocampus at 6 hours (h), 12 h and 24 h after HI. To determine the role of PDGF, the primary cortical neurons were prepared and performed PDGF shRNA administration. The results showed that HIE induced a severe behavioral dysfunction and brain infarction in neonatal rats, and the expression of PDGF in right cortex and hippocampus was remarkably increased after HI. Whereas, suppressing PDGF resulted in a significant loss of neurons and inhibition of neurite growth. Moreover, the protein level of P-PI3K and P-AKT signaling pathways were largely decreased following PDGF-shRNA application in the cortical neurons. In conclusion, PDGF suppression aggravated neuronal dysfunction, and the underlying mechanism is associated with inhibiting the phosphorylation of P-PI3K and P-AKT. Together, PDGF regulating PI3K and AKT may be an important panel in HIE events and therefore may provide possible strategy for the treatment of HIE in future clinic trail. Refereed/Peer-reviewed

Background The limited neuronal differentiation of the endogenous or grafted neural stem cells (NSCs) after brain injury hampers the clinic usage of NSCs. Panax notoginseng saponins (PNS) were extensively used for their clinical value, such as in controlling blood pressure, blood glucose, and inhibiting neuronal apoptosis and enhancing neuronal protection, but whether or not it exerts an effect in promoting neuronal differentiation of the endogenous NSCs is completely unclear and the potential underlying mechanism requires further exploration. Methods Firstly, we determined whether PNS could successfully induce NSCs to differentiate to neurons under the serum condition. Mass spectrometry and quantitative polymerase chain reaction (Q-PCR) were then performed to screen the differentially expressed proteins (genes) between the PNS + serum and serum control group, upon which dihydropyrimidinase-like 2 (DPYSL2), a possible candidate, was then selected for the subsequent research. To further investigate the actual role of DPYSL2 in the NSC differentiation, DPYSL2-expressing lentivirus was employed to obtain DPYSL2 overexpression in NSCs. DPYSL2-knockout rats were constructed to study its effects on hippocampal neural stem cells. Immunofluorescent staining was performed to identify the differentiation direction of NSCs after 7 days from DPYSL2 transfection, as well as those from DPYSL2-knockout rats. Results Seven differentially expressed protein spots were detected by PD Quest, and DPYSL2 was found as one of the key factors of NSC differentiation in a PNS-treated condition. The results of immunostaining further showed that mainly Tuj1 and GFAP-positive cells increased in the DPYSL2-overexpressed group, while both were depressed in the hippocampal NSCs in the DPYSL2-knockout rat. Conclusions The present study revealed that the differentiation direction of NSCs could be enhanced through PNS administration, and the DPYSL2 is a key regulator in promoting NSC differentiation. These results not only emphasized the effect of PNS but also indicated DPYSL2 could be a novel target to enhance the NSC differentiation in future clinical trials.

Traumatic brain injury (TBI) is a common disease that usually causes severe neurological damage, and current treatment is far from satisfactory. The neuroprotective effects of neural stem cell (NSC) transplantation in the injured nervous system have largely been known, but the underlying mechanisms remain unclear, and their limited sources impede their clinical application. Here, we established a rat model of TBI by dropping a weight onto the cortical motor area of the brain and explored the effect of engrafted NSCs (passage 3, derived from the hippocampus of embryonic 12- to 14-d green fluorescent protein transgenic mice) on TBI rats. Moreover, RT-PCR and Western blotting were employed to investigate the possible mechanism associated with NSC grafts. We found rats with TBI exhibited a severe motor and equilibrium dysfunction, while NSC transplantation could partly improve the motor function and significantly reduce cell apoptosis and increase B-cell lymphoma–extra large (Bcl-xL) expression at 7 d postoperation. However, other genes including Bax, B-cell lymphoma 2, Fas ligand, and caspase3 did not exhibit significant differences in expression. Moreover, to test whether Bcl-xL could be used as a therapeutic target, herpes simplex virus (HSV) 1 carrying Bcl-xL recombinant was constructed and injected into the pericontusional cortices. Bcl-xL overexpression not only resulted in a significant improvement in neurological function but also inhibits cell apoptosis, as compared with the TBI rats, and exhibits the same effects as the administration of NSC. The present study therefore indicated that NSC transplantation could promote the recovery of TBI rats in a manner similar to that of Bcl-xL overexpression. Therefore, Bcl-xL overexpression, to some extent, could be considered as a useful strategy to replace NSC grafting in the treatment of TBI in future clinical practices.

Long-term memory formation requires gene expression and new protein synthesis. MicroRNAs (miRNAs), a family of small non-coding RNAs that inhibit target gene mRNA expression, are involved in new memory formation. In this study, elevated miR-151-5p (miR-151) levels were found to be responsible for hippocampal contextual fear memory formation. Using a luciferase reporter assay, we demonstrated that miR-151 targets APH1a, a protein that has been identified as a key factor in γ-secretase activity, namely APH1a. Blocking miR-151 can upregulate APH1a protein levels and subsequently impair hippocampal fear memory formation. These results indicate that miR-151 is involved in hippocampal contextual fear memory by inhibiting APH1a protein expression. This work provides novel evidence for the role of miRNAs in memory formation and demonstrates the implication of APH1a protein in miRNA processing in the adult brain.

Spinal cord injury (SCI) often causes neurological deficits with poor recovery; the treatment, however, is far from satisfaction, and the mechanisms remain unclear. Using immunohistochemistry and western blotting analysis, we found α-synuclein (SNCA) was significantly up-regulated in the spinal caudal segment of rats subjected to spinal cord transection at 3 days post-operation. Moreover, the role of SNCA on neuronal growth and apoptosis in vitro was determined by using overexpressing and interfering SNCA recombined plasmid vectors, and the underlying mechanism was detected by QRT-PCR and western blotting. Spinal neurons transfected with SNCA-shRNA lentivirus gave rise to an optimal neuronal survival, while it results in cell apoptosis in SNCA-ORF group. In molecular level, SNCA silence induced the up-regulation of CNTF and down-regulation of Caspase7/9. Together, endogenous SNCA plays a crucial role in spinal neuronal survival, in which the underlying mechanism may be linked to the regulation both apoptotic genes (Caspase7/9) and CNTF. The present findings therefore provide novel insights into the role of SNCA in spinal cord and associated mechanism, which may provide novel cue for the treatment of SCI in future clinic trials.

Alterations in adult neurogenesis have been regarded as a major cause of cognitive impairment in Alzheimer's disease (AD). The underlying mechanism of neurogenesis deficiency in AD remains unclear. In this study, we reported that Integrin-linked Kinase (ILK) protein levels and phosphorylation were significantly decreased in the hippocampus of APP/PS1 mice. Increased ILK expression of dentate gyrus (DG) rescued the hippocampus-dependent neurogenesis and memory deficits in APP/PS1 mice. Moreover, we demonstrated that the effect of ILK overexpression in the hippocampus was exerted via AKT-GSK3β pathway. Finally, we found that Fluoxetine, a selective serotonin reuptake inhibitor, could improve the impaired hippocampal neurogenesis and memory by enhancing ILK-AKT-GSK3β pathway activity in APP/PS1 mice. Thus, these findings demonstrated the effects of ILK on neurogenesis and memory recovery, suggesting that ILK is an important therapeutic target for AD prevention and treatment.

It has been reported that oligodendrocyte precursor cells (OPCs) may be used to treat contusive spinal cord injury (SCC), and may alter microRNA (miRNA/miR) expression following SCC in rats. However, the association between miRNA expression and the treatment of rats with SCC with OPC transplantation remain unclear. The present study transplanted OPCs into the spinal cord of rats with SCC and subsequently used the Basso, Beattie and Bresnahan (BBB) score to assess the functional recovery and pain scores. An miRNA assay was performed to detect differentially expressed miRNAs in the spinal cord of SCC rats transplanted with OPCs, compared with SCC rats transplanted with medium. Quantitative polymerase chain reaction was used to verify significantly altered miRNA expression levels. The results demonstrated that OPC transplantation was able to improve motor recovery and relieve mechanical allodynia in rats with SCC. In addition, through a miRNA assay, 45 differentially expressed miRNAs (40 upregulated miRNAs and 5 downregulated miRNAs) were detected in the spinal cord of rats in the OPC group compared with in the Medium group. Differentially expressed miRNAs were identified according to the following criteria: Fold change >2 and P

Neuroregeneration and apoptosis are two important pathophysiologic changes after spinal cord injury (SCI), but their underlying mechanisms remain unclear. MicroRNAs (miRNAs) play a crucial role in the regulation of neuroregeneration and neuronal apoptosis, research areas that have been greatly expanded in recent years. Here, using miRNA arrays to profile miRNA transcriptomes, we demonstrated that miR-127-3p was significantly down-regulated after spinal cord transection (SCT). Then, bioinformatics analyses and experimental detection showed that miR-127-3p exhibited specific effects on the regulation of neurite outgrowth and the induction of neuronal apoptosis by regulating the expression of the mitochondrial membrane protein mitoNEET. Moreover, knockdown of MitoNEET leaded to neuronal loss and apoptosis in primary cultured spinal neurons. This study therefore revealed that miR-127-3p, which targets mitoNEET, plays a vital role in regulating neurite outgrowth and neuronal apoptosis after SCT. Thus, modificatioin of the mitoNEET expression, such as mitoNEET activition may provide a new strategy for the treatment of SCI in preclinical trials.

Spinal cord injury (SCI) often causes severe functional impairment with poor recovery. The treatment, however, is far from satisfaction, and the mechanisms remain unclear. By using proteomics and western blot, we found spinal cord transection (SCT) resulted in a significant down-regulation of α-synuclein (SNCA) in the motor cortex of SCT rats at 3 days post-operation. In order to detect the role of SNCA, we used SNCA-ORF/shRNA lentivirus to upregulate or knockdown SNCA expression. In vivo, SNCA-shRNA lentivirus injection into the cerebral cortex motor area not only inhibited SNCA expression, but also significantly enhanced neurons' survival, and attenuated neuronal apoptosis, as well as promoted motor and sensory function recovery in hind limbs. While, overexpression SNCA exhibited the opposite effects. In vitro, cortical neurons transfected with SNCA-shRNA lentivirus gave rise to an optimal neuronal survival and neurite outgrowth, while it was accompanied by reverse efficiency in SNCA-ORF group. In molecular level, SNCA silence induced the upregulation of Bcl-2 and the downregulation of Bax, and the expression of NGF, BDNF and NT3 was substantially upregulated in cortical neurons. Together, endogenous SNCA play a crucial role in motor and sensory function regulation, in which, the underlying mechanism may be linked to the regulation of apoptosis associated with apoptotic gene (Bax, Bcl2) and neurotrophic factors expression (NGF, BDNF and NT3). These finds provide novel insights to understand the role of SNCA in cerebral cortex after SCT, and it may be as a novel treatment target for SCI repair in future clinic trials.

Synaptosomal-associated protein 25 kDa (SNAP-25) is localized on the synapse and participates in exocytosis and neurotransmitter release. Decreased expression of SNAP-25 is associated with Alzheimer's disease and attention deficit/hyperactivity disorder. However, the expression of SNAP-25 in spinal cord contusion injury is still unclear. We hypothesized that SNAP-25 is associated with sensory and locomotor functions after spinal cord injury. We established rat models of spinal cord contusion injury to detect gene changes with a gene array. A decreased level of SNAP-25 was detected by quantitative real time-polymerase chain reaction and western blot assay at 1, 3, 7, 14 and 28 days post injury. SNAP-25 was localized in the cytoplasm of neurons of the anterior and posterior horns, which are involved in locomotor and sensory functions. Our data suggest that reduced levels of SNAP-25 are associated with sensory and locomotor functions in rats with spinal cord contusion injury.

The role of sodium channel voltage-gated beta 2 (SCN2B) in brain aging is largely unknown. The present study was therefore designed to determine the role of SCN2B in brain aging by using the senescence-accelerated mice prone 8 (SAMP8), a brain senescence-accelerated animal model, together with the SCN2B transgenic mice. The results showed that SAMP8 exhibited impaired learning and memory functions, assessed by the Morris water maze test, as early as 8 months of age. The messenger RNA (mRNA) and protein expressions of SCN2B were also upregulated in the prefrontal cortex at this age. Treatment with traditional Chinese anti-aging medicine Xueshuangtong (Panax notoginseng saponins, PNS) significantly reversed the SCN2B expressions in the prefrontal cortex, resulting in improved learning and memory. Moreover, SCN2B knockdown transgenic mice were generated and bred to determine the roles of SCN2B in brain senescence. A reduction in the SCN2B level by 60.68% resulted in improvement in the hippocampus-dependent spatial recognition memory and long-term potential (LTP) slope of field excitatory postsynaptic potential (fEPSP), followed by an upregulation of COX5A mRNA levels and downregulation of fibroblast growth factor-2 (FGF-2) mRNA expression. Together, the present findings indicated that SCN2B could play an important role in the aging-related cognitive deterioration, which is associated with the regulations of COX5A and FGF-2. These findings could provide the potential strategy of candidate target to develop antisenescence drugs for the treatment of brain aging.

The neuroprotective effects of olfactory ensheathing cells (OECs) after transplantation have largely been known in the injured nervous system. However, the underlying mechanisms still must be further elucidated. We explored the effects of OEC transplantation on the recovery of neurophysiologic function and the related anti-apoptosis mechanism in acute traumatic brain injury.The OECs from neonatal Sprague-Dawley rats were isolated, identified and labeled and then were immediately transplanted into the regions surrounding the injured brain site that is resulted from free-weight drop injury.Nerve growth factor and it's recepor, p75 was expressed in cultured OECs. Transplanted OECs survived, migrated around the injury site and significantly improved the neurological severe scores compared with the control group (P 0.05). OEC transplantation significantly increased the number of GAP-43-immunopositive fibers and synaptophysin-positive vesicles (P 0.05) but significantly decreased the number of apoptotic cells (P 0.05). On the molecular level, the expression of Bad in the OEC transplantation group was significantly downregulated (P 0.05).OEC transplantation could effectively improve neurological deficits in TBI rats; the underlying mechanism may be related with their effects on neuroprotection and regeneration induction, which is associated with the downregulation of the apoptotic molecule Bad.

Transplantation of neural stem cells (NSCs) into lesioned spinal cord demonstrated a beneficial effect for neural repair, the underlying mechanism, however, remains to be elusive. Here, we showed that NSCs, possessing the capacity to differentiate toward into neurons and astrocytes, exhibit a neuroprotective effect by anti-apoptosis mechanism in spinal cord hemi-transected rats despite it did not improve behavior. Intravenous NSCs injection substantially upregulated the level of BDNF mRNA but not its receptor TrkB in hemisected spinal cord, while caspase-7, a downstream apoptosis gene of caspase-3, has been largely down-regulated. TUNEL staining showed that the number of apoptosis cells in injured spinal cord decreased significantly, compared with seen in rats with no NSCs administration. The present finding therefore provided crucial evidence to explain neuroprotective effect of NSCs grafts in hemisected spinal cord, which is associated with BDNF upregulation and caspase-7 downregulation.