Number Of NeuN-positive Cells Research Articles

Ischemia-reperfusion injury after spinal cord decompressive surgery-An invivo rat model.

Although decompression surgery is the optimal treatment for patients with severe degenerative cervical myelopathy (DCM), some individuals experience no improvement or even a decline in neurological function after surgery, with spinal cord ischemia-reperfusion injury (SCII) identified as the primary cause. Spinal cord compression results in local ischemia and blood perfusion following decompression is fundamental to SCII. However, owing to inadequate perioperative blood flow monitoring, direct evidence regarding the occurrence of SCII after decompression is lacking. The objective of this study was to establish a suitable animal model for investigating the underlying mechanism of spinal cord ischemia-reperfusion injury following decompression surgery for degenerative cervical myelopathy (DCM) and to elucidate alterations in neurological function and local blood flow within the spinal cord before and after decompression. Twenty-four Sprague-Dawley rats were allocated to three groups: the DCM group (cervical compression group, with implanted compression material in the spinal canal, n = 8), the DCM-D group (cervical decompression group, with removal of compression material from the spinal canal 4 weeks after implantation, n = 8), and the SHAM group (sham operation, n = 8). Von Frey test, forepaw grip strength, and gait were assessed within 4 weeks post-implantation. Spinal cord compression was evaluated using magnetic resonance imaging. Local blood flow in the spinal cord was monitored during the perioperative decompression. The rats were sacrificed 1 week after decompression to observe morphological changes in the compressed or decompressed segments of the spinal cord. Additionally, NeuN expression and the oxidative damage marker 8-oxoG DNA were analyzed. Following spinal cord compression, abnormal mechanical pain worsened, and a decrease in forepaw grip strength was observed within 1-4 weeks. Upon decompression, the abnormal mechanical pain subsided, and forepaw grip strength was restored; however, neither reached the level of the sham operation group. Decompression leads to an increase in the local blood flow, indicating improved perfusion of the spinal cord. The number of NeuN-positive cells in the spinal cord of rats in the DCM-D group exceeded that in the DCM group but remained lower than that in the SHAM group. Notably, a higher level of 8-oxoG DNA expression was observed, suggesting oxidative stress following spinal cord decompression. This model is deemed suitable for analyzing the underlying mechanism of SCII following decompressive cervical laminectomy, as we posit that the obtained results are comparable to the clinical progression of degenerative cervical myelopathy (DCM) post-decompression and exhibit analogous neurological alterations. Notably, this model revealed ischemic reperfusion in the spinal cord after decompression, concomitant with oxidative damage, which plausibly underlies the neurological deterioration observed after decompression.

Buyang Huanwu decoction promotes angiogenesis and improves hemorheological parameters after cervical spinal cord injury

ObjectiveTo explore the effects of Buyang Huanwu decoction (BYHWD) on vascular neogenesis and hemorheological parameters following cervical spinal cord injury (SCI). MethodsAn acute cervical SCI model was established using 84 female Sprague–Dawley rats. Functional recovery of the rats was evaluated using the forelimb locomotor scale score, forelimb grip strength test, and Basso-Beattie-Bresnahan score. The animals were subsequently euthanized at days 7 and 28 postoperatively. The gross morphology, neuronal survival, and myelin sheath in the injured area were evaluated using hematoxylin and eosin (HE), Nissl, and luxol fast blue (LFB) staining, respectively. Immunofluorescence staining was used to observe CD31 expression 7 days post-injury. Furthermore, the expression of CD31, neuronal nuclear protein (NeuN), and myelin basic protein (MBP) were evaluated 28 days post-injury. Additionally, vascular endothelial growth factor A (VEGFA) and VEGF receptor-2 (VEGFR-2) expression was evaluated using western blotting. Whole-blood viscosity, plasma viscosity, and red blood cell aggregation were measured using a hemorheometer. ResultsFrom postoperative days 3–28, motor function in the BYHWD group began to recover considerably compared to the SCI group. BYHWD effectively restored spinal cord histopathology. In addition, the number of NeuN-positive cells, and fluorescence intensity of CD31at 7 and 28 days and MBP significantly increased in the BYHWD group compared with the SCI group (all P < 0.05). Moreover, this decoction significantly upregulated the expression of VEGFA and VEGFR-2 (all P < 0.05). BYHWD improved the hemorheology results (i.e., except erythrocyte aggregation index in the low-dose group), revealing statistically significant differences compared with the SCI group (all P < 0.05). ConclusionBYHWD effectively promoted angiogenesis, improved hemorheological parameters, and protected neurons and myelin sheaths, ultimately promoting the recovery of neurological function after cervical SCI in rats. These findings suggest that BYHWD promotes vascular neogenesis through the VEGFA/VEGFR-2 pathway.

Thrombopoietin exerts a neuroprotective effect by inhibiting the suppression of neuronal proliferation and axonal outgrowth in intrauterine growth restriction rats

Chronic hypoxia in utero causes intrauterine growth restriction (IUGR) of the fetus. IUGR infants are known to be at higher risk for neurodevelopmental disorders, but the mechanism is unclear. In this study, we analyzed the structure of the cerebral cortex using IUGR model rats generated through a reduced uterine perfusion pressure operation. IUGR rats exhibited thinner cerebral white matter and enlarged lateral ventricles compared with control rats. Expression of neuron cell markers, Satb2, microtubule-associated protein (MAP)-2, α-tubulin, and nestin was reduced in IUGR rats, indicating that neurons were diminished at various developmental stages in IUGR rats, from neural stem cells to mature neurons. However, there was no increase in apoptosis in IUGR rats. Cells positive for Ki67, a marker of cell proliferation, were reduced in neurons and all glial cells of IUGR rats. In primary neuron cultures, axonal elongation was impaired under hypoxic culture conditions mimicking the intrauterine environment of IUGR infants. Thus, in IUGR rats, chronic hypoxia in utero suppresses the proliferation of neurons and glial cells as well as axonal elongation, resulting in cortical thinning and enlarged lateral ventricles. Thrombopoietin (TPO), a platelet growth factor, inhibited the decrease in neuron number and promoted axon elongation in primary neurons under hypoxic conditions. Intraperitoneal administration of TPO to IUGR rats resulted in increases in the number of NeuN-positive cells and the area coverage of Satb2. In conclusion, suppression of neuronal proliferation and axonal outgrowth in IUGR rats resulted in cortical thinning and enlargement of lateral ventricles. TPO administration might be a novel therapeutic strategy for treating brain dysmaturation in IUGR infants.

Neuroprotective Effects of Krypton Inhalation on Photothrombotic Ischemic Stroke.

This is the first in vivo study to investigate the neuroprotective effects of krypton on focal cerebral ischemia. The aim of the study was to analyze the effect of 2 h of inhalation of a krypton-oxygen mixture (Kr 70%/O2 30%) on the recovery of neurological functions and the degree of brain damage in rats after photoinduced ischemic stroke (PIS) and to investigate the possible mechanisms responsible for this neuroprotection. Experiments were performed on male Wistar rats weighing 250-300 g (n = 32). Animals were randomized into four groups. Two groups (n = 20) underwent photoinduced ischemic stroke, followed by 2 h of inhalation of krypton-oxygen mixture consisting of Kr 70%/O2 30% or a nitrogen-oxygen breathing mixture consisting of N2 70%/O2 30%, followed by neurological examinations on days 3 and 7. The other two groups (n = 12) received only gas mixtures of the same concentration and exposure duration as in those in the PIS groups, then Western blot analysis of the potential molecular mechanisms was performed. The results of the study show that treatment with the krypton-oxygen mixture consisting of Kr 70%/O2 30% improves the neurological status on day 7 of observation, reduces the lesion volume according to the MRI examination and the number of Iba-1- and caspase-3-positive cells in the damaged area, promotes the activation of neoangiogenesis (an increase in the von Willebrand factor), and reduces the penumbra area and the number of NeuN-positive cells in it on day 14 of observation. Inhalation of the krypton-oxygen mixture also significantly increases the levels of phosphorylated AKT kinase (protein kinase B) and glycogen synthase kinase 3b (pGSK3b) and promotes the expression of transcription factor Nrf2, which was accompanied by the lowered expression of transcription factor NFkB (p50). Thus, we showed pronounced neuroprotection induced by krypton inhalation after stroke and identified the signaling pathways that may be responsible for restoring neurological functions and reducing damage.

Photobiomodulation Promotes Hippocampal Neurogenesis and Improves Cognitive Function and Anti-Inflammatory Injury in Rats With Chronic Cerebral Hypoperfusion

To investigate the effect of photobiomodulation (PBM) on hippocampal neurogenesis, cognitive function, and inflammatory injury in rats with chronic cerebral hypoperfusion. Bilateral ovariectomy (OVX) was performed on female Sprague-Dawley (SD) rats. One week later, the rats were randomly assigned to three groups, Sham surgery (or Sham) group, bilateral common carotid artery occlusion (BCCAO) group, and PBM intervention (or BCCAO+PBM) group. There were 8 rats in each group. In the BCCAO group, chronic cerebral hyporeperfusion was induced by permanent ligation of bilateral common carotid arteries and no PBM was given. Rats in the Sham group underwent the same surgical procedure except for the occlusion of the two carotids arteries and no PBM was given. In addition to the BCCAO surgery, rats in the BCCAO+PBM group received 808 nm laser therapy (5 min each time at a laser dose of 20 mW/cm 2) of the frontal cortex every other day for 1 month. Between 86 and 90 days after BCCAO, Morris water maze (MWM) was used to observe the spatial learning and memory function of the rats. The rats were sacrificed on day 90 and immunofluorescence staining and Western blot were performed thereafter. Immunofluorescence staining was used to determine the expression of 5-bromodeoxyuracil nucleoside (BrdU), a cell proliferation marker, glial fibrillary acidic protein (GFAP), an astrocyte marker, doublecortin (DCX), a specific marker of newborn neuron precursor cells, NeuN, a marker of mature neurons, and Iba1, a microglia marker, in the hippocampal dentate gyrus (DG) region. Western blot was performed to analyze the protein expressions of inflammasome components, NLRP3, ASC, cleaved caspase-1, and Iba1 in the hippocampus. In the latency trial of MWM test, BCCAO+PBM rats spent shorter periods of time finding the underwater platform than the BCCAO rats did. In the probe trial, after the platform that was original placed in a quadrant was removed, the BCCAO+PBM rats spent longer periods of time exploring the quadrant than the BCCAO animals did ( P<0.05). Compared with BCCAO rats, BCCAO+PBM rats showed significant decrease in the immunofluorescence intensities of GFAP and Iba1 ( P<0.01). PBM intervention significantly increased the number of BrdU-positive cells in the hippocampal DG region compared with those of Sham and BCCAO groups ( P<0.05). Furthermore, the number of NeuN positive cells showed no significant difference among the three groups, while in BCCAO+PBM group, the number of DCX-positive cells was significantly increased ( P<0.001) and the number of DCX +/NeuN + co-located cells was significantly increased compared to that of the BCCAO group ( P<0.001). Compared with those of the BCCAO group, Western blot results showed that the protein expression levels of Iba1, NLRP3, and cleaved caspase-1 in the BCCAO+PBM group were significantly decreased ( P<0.05), while the ASC protein expression level showed no significant difference. PBM can effectively improve the spatial learning and memory function in rats with chronic cerebral hypoperfusion, inhibit the activation of glial cells, reduce inflammatory damage mediated by NLRP3 inflammasome, and promote the regeneration of endogenous neural stem cells in the hippocampal DG region of rats.

Electroacupuncture inhibits neuron injury of SAMP8 mice by reducing inflammatory response

To observe the effect of electroacupuncture (EA) at "Baihui"(GV20) and "Shenshu" (BL23) on the pathological injury of neurons in SAMP8 mice and the anti-inflammatory effect on neuron repair, providing a new experimental basis for EA prevention and treatment of Alzheimer's disease. Twelve 7-month-old SAMP8 mice were randomly divided into model and EA groups, and 6 SAMR1 mice of the same age and genetic background were used as normal group. Mice in the EA group were needled at GV20 and bilateral BL23, and EA (1 mA, 2 Hz) was applied to bilateral BL23 for 15 min, once daily, 10 d as a course for a total of 4 courses, with an interval of 1 d. Mice in the normal and model groups were captured and fixed in the same way as the EA group. The spatial learning and memory ability was detected by Morris water maze test. Neuronal nuclear antigen (NeuN) positive expression and the number of NeuN-positive cells in dentate gyrus (DG) were detected by immunofluorescence staining. The protein expression levels of ionized calcium binding adapter molecule 1(Iba-1), tumor necrosis factor-α(TNF-α), interleukin (IL)-6 and IL-1β in hippocampus were detected by Western blot. The ultrastructure of nerve cells in DG was observed by transmission electron microscopy. Compared with the normal group, the average escape latency was prolonged(P<0.01), the number of platform crossing was significantly reduced (P<0.01), the average fluorescence intensity of NeuN and the number of NeuN-positive cells in hippocampus DG region decreased (P<0.05), the expression levels of Iba-1, TNF-α, IL-6 and IL-1β in hippocampus were increased (P<0.05) in the model group.Compared with the model group, the ave-rage escape latency was shortened (P<0.01), the number of platform crossing times was significantly increased (P<0.01), the average fluorescence intensity of NeuN and the number of NeuN-positive cells in hippocampus DG region increased (P<0.05), the expression levels of Iba-1, TNF-α, IL-6 and IL-1β in hippocampus were decreased (P<0.05) in the EA group. The morphology of nerve cells in the hippocampus DG region was normal， and the organelles in the cytoplasm were clear, complete and regularly distributed in the normal group. However, the morphology of nerve cells in the model group was seriously irregular, which was also irregular in EA group but somewhat relieved compared with model group. EA at GV20 and BL23 can improve the learning and memory ability of SAMP8 mice, which may be related to inhibiting the neuroinflammatory response, increasing the number of neurons and improving the ultrastructure of the DG region of the hippocampus to play the role of neuron protection.

Acupuncture ameliorates neurological function by suppressing microglia polarization and inflammatory response after cerebral ischemia in rats

To observe the effect of acupuncture on microglia polarization and inflammatory reaction in rats with cerebral ischemia-reperfusion injury (CIRI), so as to explore its mechanisms underlying improvement of CIRI. Thirty male SD rats were randomly divided into sham operation, model, and acupuncture groups, with 10 rats in each group. The CIRI model was established by occlusion of the middle cerebral artery (MCAO) for 1 h, followed by reperfusion. After modeling, rats in the acupuncture group received manual acupuncture stimulation of "Dazhui" (GV14), "Baihui"(GV20), "Shuigou" (GV26), bilateral "Zusanli" (ST36) and "Fengchi" (GB20) by twirling the needles rapidly for 10 s/acupoint every 10 min, with the needles retained for 20 min. The treatment was conducted once daily for successive 7 days. The neurological function was evaluated according to Longa's method. The state of CIRI was observed after Nissl staining, and the expression levels of Iba-1, iNOS, Arg1, BDNF, GDNF and NeuN in the ischemic cortex tissue were detected by immunofluorescence staining. The contents of TNF-α, IL-6 and IL-10 in the ischemic tissue were assayed by ELISA. The protein expression levels of BDNF, GDNF, TLR4, MyD88 and NF-κB in the ischemic tissues were detected by Western blot. The neurological deficit score on the 24 h and 7th day was considerably higher in the model group than in the sham operation group (P<0.01), and evidently lower on the 7th day in the acupuncture group than in the model group (P<0.01). The number of NeuN positive cells，the area of immunofluorescence dual labelling of Arg1, BDNF and GDNF positive staining, IL-10 content, BDNF and GDNF protein expressions were significantly decreased (P<0.01), and the immunofluorescence dual labelling area of Iba-1 and iNOS, TNF-α and IL-6 contents, the pretein expression levels of TLR4, MyD88 and NF-κB considerably increased (P<0.01) in the model group relevant to the sham operation group. In contrast to the model group, the acupuncture group had a significant increase in the number of NeuN positive cells, the immunofluorescence dual labelling area of Arg1, BDNF and GDNF positive staining, IL-10 content, and BDNF and GDNF protein expressions (P<0.05, P<0.01), and an evident decrease in Iba-1 and iNOS positive staining, contents of TNF-α and IL-6, and the protein expression levels of TLR4, MyD88 and NF-κB (P<0.01, P<0.05). Nissl staining showed a marked reduction in the number of neurons, the nucleus pyknosis and nissl bodies and loose arrangement of the neuronal cells in the model group, which was relatively milder in the acupuncture group. Acupuncture intervention can improve neurological function in CIRI rats, which may be related to its effects in regulating the polarization of microglia, reducing inflammatory reaction and increasing the secretion of neurotrophic factors in the brain, inhibiting TLR4/MyD88/NF-κB signaling pathway.

Compensatory Hippocampal Neurogenesis in the Absence of Cognitive Impairment Following Experimental Hippocampectomy in Adult Rats.

Temporal lobe epilepsy (TLE) is the commonest type of focal epilepsy in adult humans, and hippocampal sclerosis (HS) is the main pathological finding in this type of epilepsy. In refractory TLE, patients are indicated for unilateral resection of the affected hippocampus by a surgical procedure called hippocampectomy which generally does not cause any cognitive impairment. Once adult hippocampus is a region of endogenous neurogenesis, even in elderly people, we have hypothesized that a compensatory increase in hippocampal neurogenesis might occur in the remaining hippocampus after unilateral hippocampectomy. To test this hypothesis, we performed unilateral hippocampectomy in adult Wistar rats, which were perfused at 15 (G15) and 30 (G30) days post-surgery. Eighteen Wistar rats were randomly distributed in the following experimental groups: control (no surgery, N = 6), G15 (N = 6), and G30 (N = 6). Adjacent cortex and hippocampus of the left hemisphere were completely removed. Behavioral procedures were performed to address possible cognitive impairments. Brains were collected and fixed from animals belonging to all experimental groups. Gross histopathology was performed using thionine staining. Neuroblasts and mature neurons were immunolabeled using anti-doublecortin (DCX) and anti-NeuN antibodies, respectively. Numbers of DCX and NeuN positive cells were quantified for all experimental groups. Animals submitted to hippocampectomy did not present any cognitive impairment as evaluated by eight-arm radial maze behavioral test. The remaining hippocampus presented a higher number of DCX positive cells compared to control (p < 0.001, ANOVA-Tukey) at both G15 and G30. A higher number of NeuN positive cells were present in the granular layer of dentate gyrus at G30 compared to control and G15 (p < 0.001, ANOVA-Tukey). The data suggest that unilateral hippocampectomy induces compensatory neurogenic effect in the contralateral hippocampus. This may underlie the reported absence of significant cognitive impairment and parallels the findings in human patients submitted to unilateral hippocampectomy to treat refractory TLE.

Learning Deficits Accompanied by Microglial Proliferation After the Long-Term Post-Injection of Alzheimer's Disease Brain Extract in Mouse Brains.

Human tauopathy brain injections into the mouse brain induce the development of tau aggregates, which spread to functionally connected brain regions; however, the features of this neurotoxicity remain unclear. One reason may be short observational periods because previous studies mostly used mutated-tau transgenic mice and needed to complete the study before these mice developed neurofibrillary tangles. To examine whether long-term incubation of Alzheimer's disease (AD) brain in the mouse brain cause functional decline. We herein used Tg601 mice, which overexpress wild-type human tau, and non-transgenic littermates (NTg) and injected an insoluble fraction of the AD brain into the unilateral hippocampus. After a long-term (17-19 months) post-injection, mice exhibited learning deficits detected by the Barnes maze test. Aggregated tau pathology in the bilateral hippocampus was more prominent in Tg601 mice than in NTg mice. No significant changes were observed in the number of Neu-N positive cells or astrocytes in the hippocampus, whereas that of Iba-I-positive microglia increased after the AD brain injection. These results potentially implicate tau propagation in functional decline and indicate that long-term changes in non-mutated tau mice may reflect human pathological conditions.

Combined transplantation of neural stem cells and bone marrow mesenchymal stem cells promotes neuronal cell survival to alleviate brain damage after cardiac arrest via microRNA-133b incorporated in extracellular vesicles.

Neural stem cell (NSC) transplantation has prevailed as a promising protective strategy for cardiac arrest (CA)-induced brain damage. Surprisingly, the poor survival of neuronal cells in severe hypoxic condition restricts the utilization of this cell-based therapy. Extracellular vesicles (EVs) transfer microRNAs (miRNAs) between cells are validated as the mode for the release of several therapeutic molecules. The current study reports that the bone marrow mesenchymal stem cells (BMSCs) interact with NSCs via EVs thereby affecting the survival of neuronal cells. Hypoxic injury models of neuronal cells were established using cobalt chloride, followed by co-culture with BMSCs and NSCs alone or in combination. BMSCs combined with NSCs elicited as a superior protocol to stimulate neuronal cell survival. BMSCs-derived EVs could protect neuronal cells against hypoxic injury. Silencing of miR-133b incorporated in BMSCs-derived EVs could decrease the cell viability and the number of NeuN-positive cells and increase the apoptosis in the CA rat model. BMSCs-derived EVs could transfer miR-133b to neuronal cells to activate the AKT-GSK-3β-WNT-3 signaling pathway by targeting JAK1. Our study demonstrates that NSCs promotes the release of miR-133b from BMSCs-derived EVs to promote neuronal cell survival, representing a potential therapeutic strategy for the treatment of CA-induced brain damage.

Papaverine provides neuroprotection by suppressing neuroinflammation and apoptosis in the traumatic brain injury via RAGE- NF-B pathway

The receptor for advanced glycation end products (RAGE)- Nuclear Factor kappa B (NF-κB) signal pathway may represent a new target for the treatment of traumatic brain injury (TBI). The aim of the study is to investigate effects of papaverine on secondary signaling mechanisms through this pathway in mice TBI model.Immunohistochemically, while the number of RAGE and NF- κB positive cells, apoptotic cells increased, the number of NeuN positive cells reduced in TBI.Papaverine reduced the number of RAGE positive cells on glia and the number of NF- κB positive cells on both neuron and glia. At the same time, it decreased the number of microglia labeled with P2RY12 increased due to TBI. It also increased the NeuN positive cells and mitigated the brain edema. Results of this study showed that papaverine reduced TBI- induced neuroinflammation and apoptosis, also provided neuroprotection via the RAGE- NF-κB signal path, which is one of the possible mechanisms in TBI.

IGF-1 Protects Neurons in the Cortex and Subventricular Zone in a Periventricular Leucomalacia Model.

Chronic cerebral hypoperfusion affects early and mature neurons in the subventricular zone (SVZ) and cerebral cortex. Herein, we investigated the effects of insulin-like growth factor-1 (IGF-1), a neurogenesis-promoting agent, on neurons in these regions in periventricular leucomalacia (PVL) model rats. Following right carotid artery ligation, the rats were placed in a hypoxia chamber and injected with recombinant IGF-1 (0.1 and 1 μg/μl). Their brain sections were immunohistochemically analysed using anti-nestin and anti-NeuN antibodies. The numbers of early-neuronal cells in the SVZ and mature neurons in the cerebral cortex were higher and lower, respectively, in the PVL group than in the control group. The number of NeuN-positive cells was significantly higher in the IGF-treated group than in the PVL group. PVL increased the number of early neuronal cells in the SVZ, reducing the survival of mature neurons in the cerebral cortex; IGF-1 reversed these effects.

Effects of resveratrol and soy isoflavones on learning and memory ability and expression of glucose transporter in aging model rats

To investigate the effects of resveratrol combined with soy isoflavones on avoidance memory, number of neuron-specific nuclear protein(NeuN) positive cells and expressions of glucose transporter(GLUT)1 and GLUT3 in hippocampus of aging model rats. A total of 60 female SD rats were randomly divided into 6 groups including sham control group, aging model group, 80 mg/kg resveratrol group, 160 mg/kg soy isoflavones group, 80 mg/kg resveratrol +160 mg/kg soy isoflavones group, 0. 8 mg/kg estradiol valerate group. The aging model rats was induced by ovariectomy combined with intraperitoneal injection of 100 mg/kg D-galactose. Intragastric administration was performed once a day for 12 weeks. The avoidance task was measured by the shuttle box test. The NeuN expression were measured by the immunofluorescence. The genes and proteins expression of GLUT1 and GLUT3 in rat hippocampus were detected by real-time PCR and Western blot, respectively. Compared with the sham control group, the avoidance latency in the aging model group was prolonged, and the active avoidance response rate and the total avoidance response rate were decreased. The number of NeuN positive cells decreased and the expression levels of GLUT1 and GLUT3 genes and proteins were decreased(P&lt;0. 05). Compared with the aging model group, the escape latency significantly declined(P&lt;0. 01), but the rates of active avoidance response and total avoidance response increased, the number of NeuN positive cells increased significantly, the expression levels of GLUT1 and GLUT3 genes and proteins up-regulated in the rats of the three intervention groups(P&lt;0. 05 or P&lt;0. 01). Compared with the soy isoflavones group, the active avoidance response rate was increased in the combined group(P&lt;0. 05). In comparison with those for the resveratrol group, the avoidance latency was shortened and the active avoidance response rate was increased, the number of NeuN positive cells and the expression levels of GLUT3 gene and protein were significantly increased in the combined group(P&lt;0. 05). There was no significant difference between the combined intervention group and the estradiol valerate group(P&gt;0. 05). Resveratrol and soy isoflavones alone and in combination can improve the learning and memory ability of aging rat models. The mechanism may be related to up-regulating the expression of GLUT1 and GLUT3 genes and proteins in the hippocampus, promoting the transmembrane transport of glucose and reducing neuronal loss.

Regional Expression of Act-MMP3 Contributes to the Selective Loss of Neurons in Ganglion Cell Layers following Acute Retinal ischemia/Reperfusion Injury

ABSTRACTPurpose: Evidences suggest that during ischemia/reperfusion events, neuronal loss in ganglion cell layers (GCLs) occurs initially in the peripheral retinae followed by the central. However, which key molecule or factor mediates this selective loss needs elucidation. In the present study, we detected the regional expression of active matrix metalloproteinase 3 (Act-MMP3) in the central and peripheral rat retinae following acute retinal ischemia/reperfusion (RI/R) injury and explored the effects and mechanisms of this regional expression on the selective neuronal loss in GCLs.Methods: QPCR and Western Blotting were used to detect the expression of Act-MMP3 in the central part and peripheral part of the adult rat retinae. Immunofluorescence and double immunofluorescence were used to assess the number of NeuN-positive cells in the GCLs and Iba-1+CD 68-positive cells were determined. Additionally, the Linear-regression analysis was performed to test the correlation between the ODV of Act-MMP3 and the neuronal loss in the GCLs/Iba-1+CD 68 positive cells in retinae.Results: An evident up-regulation of active matrix metalloproteinase 3 (Act-MMP3) in peripheral retinae preceded to that in central region following acute RI/R. We found Act-MMP3 up-regulation to be associated with the selective neuronal loss in GCLs (central: r = 0.7566, p < .0001, r2 = 0.5724; peripheral: r = 0.8241, p < .0001, r2 = 0.6792). Suppressing Act-MMP3 ameliorated the selective neuronal loss in GCLs following acute RI/R. Furthermore, the activation of microglia in the peripheral retinae also preceded to that in the central and was found to be correlated with the regional expression of Act-MMP3 (Central: r = 0.8540, p < .0001, r2 = 0.7294; Peripheral: r = 0.7820, p < .0001, r2 = 0.6116). Suppressing Act-MMP3 ameliorated the microglia regional activation following acute RI/R.Conclusion: The regional expression of Act-MMP3 in the rat retinae may contribute to the selective neuronal loss in GCLs and microglia regional activation in acute RI/R.

Erythropoietin Reduces Neurodegeneration and Long-Term Memory Deficits Following Sevoflurane Exposure in Neonatal Rats.

Exposure to general anesthetics induces neural apoptosis and degeneration in the immature neonatal brain. Erythropoietin (EPO) has been shown to protect neonatal animals against hypoxic-ischemic injury and general anesthesia-induced developmental neurotoxicity. However, preventive strategy caused by EPO against neurotoxicity due to general anesthesia is still uncertain. This study examined the effects of EPO administration on brain cytology and cognitive function in adolescent rats exposed to 3% sevoflurane as neonates. Seven-day-old rats received intraperitoneal saline (EPO 0U group) or EPO (60, 120, or 600U) 30min before exposure to 3% sevoflurane with 21% oxygen for 4h. The rats only received 21% oxygen without EPO and sevoflurane as the sham group. The Morris water maze task was performed time-dependently among the groups, 3weeks post-anesthesia exposure. Escape latency and % quadrant in the EPO 600U group were significantly reduced and increased, respectively, compared with those in the EPO 0U group 6weeks post-exposure. In addition, freezing time in response to the conditioned stimulus and the number of NeuN-positive cells in the hippocampal CA1 region were significantly increased in the EPO 120 and 600U groups than in the EPO 0U group 6weeks after exposure. Moreover, the statistical parameter mapping of positive cell density was increased in the EPO-treated rats. These results support the observations that pretreatment with EPO reduced long-term cognitive deficits and neuronal degeneration in cortex and hippocampus induced by sevoflurane exposure with low oxygen concentration in neonatal rats.

Dexmedetomidine reduced sevoflurane-induced neurodegeneration and long-term memory deficits in neonatal rats

Exposure to sevoflurane and other inhalational anesthetics can induce neurodegeneration in the developing brain. Although dexmedetomidine (DEX) has provided neuroprotection against hypoxic ischemic injury, relatively little is known about whether it has the neuroprotective effects against anesthetic-induced neurodegeneration. This study examined whether DEX improves the long-term cognitive dysfunction observed after exposure of neonatal rats to 3% sevoflurane. Seven-day-old rats received intraperitoneal saline (DEX 0) or DEX (6.6, 12.5, 25 μg/kg) 30 min before exposure to 3% sevoflurane with 21% oxygen for 4 h (n = 10 per group). The pups in the control group received only DEX 25 μg/kg without anesthesia. The escape latency in the Morris water maze was significantly increased in the DEX 0 group compared with the sham and control group, and the escape latency, but not the swimming path length, was significantly shorter at post-natal day 47 in the DEX 25 than in the DEX 0 group. The percent time spent in the quadrant was significantly decreased in the DEX 0 group compared with the sham and control group, and the percent time spent in the quadrant was significantly increased in the DEX 25 group compared with the DEX 0 groups. The freezing times of the DEX 0 and 6.6 groups were significantly decreased compared with those in the sham, control and DEX 25 groups. The number of NeuN-positive cells in the CA1 region was significantly decreased in the DEX 0 and 6.6 groups compared with the sham, control and DEX 25 groups. These findings indicate pre-treatment with DEX may improve long-term cognitive function and ameliorate the neuronal degeneration induced by sevoflurane exposure in neonatal rats.

Abstract TP110: Preischemic Neuroprotective Effect of Minocycline and Sodium Ozagrelon Transient Cerebral Ischemic Rat Model

Background: We investigated the neuroprotective properties of single doses of minocycline and ozagrel when administered prior to stroke. Methods: Male Sprague-Dawley rats were assigned randomly to one of the following groups: (1) control (Con) group (n=10), (2) minocycline (Mino) group (n=10), (3) sodium ozagrel (SO) group (n=10). Rats were treated with a single dose of minocycline or ozagrel at 30 minutes before stroke. A middle cerebral artery occlusion (MCAO) was made at 30 minutes after drug administration and reperfusion was done. The rats were subjected to a neurobehavioral test at days 1, 3 and 7 after MCAO. The cerebral ischemic volume was quantified by MetaMorph imaging software after TTC staining. The neuronal cell survival and astrocytes expansion were assessed by the NeuN and GFAP immunohistofluorescence staining. Apoptosis was detected by the TUNEL assay. We statistically analyzed and compared the results with each other. Results: Mino and SO groups had neuroprotective effect and showed a better behavioral performance of adhesive removal and treadmill test at 7 days after stroke. Mino and SO groups also showed a smaller infarct volume than control group at 7 days after stroke. Immunohistofluorescence staining showed a higher number of NeuN positive cells, lower activated astrocytes in GFAP and a lower apoptosis in TUNEL staining. Conclusion: This study showed that single doses of minocycline and ozagrel prior to stroke had neuroprotective effects. These agents will be useful not only in post-stroke therapy but also in stroke prevention in several cerebrovascular procedures like carotid endarterectomy, bypass procedure, endovascular angioplasty, thromboembolectomy or thrombolysis

Does the Apparent Diffusion Coefficient Value Predict Permanent Cerebral Ischemia/Reperfusion Injury in Rats?

Variation in tissue damage after cerebral ischemia/reperfusion (I/R) can cause uncertainty in stroke-related studies, which can be reduced if the damage can be predicted early after ischemia by measuring the apparent diffusion coefficient (ADC). We investigated whether ADC measurement in the acute phase can predict permanent cerebral I/R injury. The middle cerebral artery occlusion model was established using the intraluminal suture method to induce 60 minutes of ischemia followed by reperfusion in rats. T2-weighted images and diffusion-weighted images were obtained at 30 minutes and 24 hours after ischemia. Neuronal cell survival was assessed by neuronal nuclei (NeuN) immunofluorescence staining. The correlation between relative ADC (rADC) values at 30 minutes and I/R injury at 24 hours after ischemia was analyzed. Magnetic resonance imaging results were confirmed by histologic analysis. The correlation between rADC values at 30 minutes and 24 hours was strong in the ischemic core and peri-infarct region but moderate in the anterior choroidal and hypothalamic region. Histologic analysis revealed that the correlation between rADC values at 30 minutes and the number of NeuN-positive cells at 24 hours was strong in the ischemic core and peri-infarct region but moderate in the anterior choroidal and hypothalamic region. Furthermore, there was a strong positive correlation between the sum of rADC values of three regions at 30 minutes and the infarct volume at 24 hours. ADC measurement in the acute phase can predict permanent cerebral I/R injury and provide important information for the evaluation of ischemic stroke.

The additional oxygen as a carrier gas during long-duration sevoflurane exposure ameliorate the neuronal apoptosis and improve the long-term cognitive function in neonatal rats

The effects of the oxygen concentration as a carrier gas and long duration anesthesia exposure on neuroapoptosis and cognitive impairments in the developing brain are not fully understood. This study shows that long-duration sevoflurane anesthesia with or without additional oxygen induces neuroapoptosis and long-term cognitive dysfunction in neonatal rats. Seven-day-old rats were exposed to sevoflurane anesthesia for 2, 4, and 6 h with 21% or 30% oxygen. The control group received 21% oxygen alone for 6 h. Post-anesthesia blood gas analysis resulted in hypoxia and hypercapnia. Moreover, PO2 and base excess in the 30% oxygen group were significantly higher than the 21% oxygen group. The numbers of caspase-3-positive cells in both cortical layer 3 and the CA1 region in the hippocampus in the 6 h anesthesia exposure group with 21% oxygen were increased compared with the 6 h anesthesia exposure with 30% oxygen and control groups. Cognitive function was assessed in an additional group of rats, and the brains were stained for NeuN 6 weeks post-anesthesia. Although the Morris water maze task was acquired equally by all rats 3 weeks post-anesthesia, the escape latency was significantly longer in the 6 h sevoflurane with 21% oxygen group than the 6 h with 30% oxygen groups 6 weeks post-exposure. No difference was found with regard to freezing time among the groups in the fear conditioning test. The number of NeuN-positive cells in the CA1 region of the hippocampus in the control group was increased compared with the other groups. These findings indicate that long-duration sevoflurane exposure with 30% oxygen as a carrier gas would ameliorate neuronal apoptosis and improve long-term cognitive function in neonatal rats.

Mechanisms underlying the promotion of functional recovery by deferoxamine after spinal cord injury in rats.

Deferoxamine, a clinically safe drug used for treating iron overload, also repairs spinal cord injury although the mechanism for this action remains unknown. Here, we determined whether deferoxamine was therapeutic in a rat model of spinal cord injury and explored potential mechanisms for this effect. Spinal cord injury was induced by impacting the spinal cord at the thoracic T10 vertebra level. One group of injured rats received deferoxamine, a second injured group received saline, and a third group was sham operated. Both 2 days and 2 weeks after spinal cord injury, total iron ion levels and protein expression levels of the proinflammatory cytokines tumor necrosis factor-α and interleukin-1β and the pro-apoptotic protein caspase-3 in the spinal cords of the injured deferoxamine-treated rats were significantly lower than those in the injured saline-treated group. The percentage of the area positive for glial fibrillary acidic protein immunoreactivity and the number of terminal deoxynucleotidyl transferase dUTP nick end labeling-positive cells were also significantly decreased both 2 days and 2 weeks post injury, while the number of NeuN-positive cells and the percentage of the area positive for the oligodendrocyte marker CNPase were increased in the injured deferoxamine-treated rats. At 14-56 days post injury, hind limb motor function in the deferoxamine-treated rats was superior to that in the saline-treated rats. These results suggest that deferoxamine decreases total iron ion, tumor necrosis factor-α, interleukin-1β, and caspase-3 expression levels after spinal cord injury and inhibits apoptosis and glial scar formation to promote motor function recovery.