Glial Fibrillary Acidic Protein mRNA Expression Research Articles

Combined Effect of Maternal Separation and Early-Life Immune Activation on Brain and Behaviour of Rat Offspring.

Adversity during early life, a critical period for brain development, increases vulnerability and can have a lasting impact on the brain and behaviour of a child. However, the long-term effects of cumulative early-life stressors on brain and behaviour are not well known. We studied a 2-hit rat model of early-life adversity using maternal separation (MS) and immune activation (lipopolysaccharide (LPS)). Rat pups underwent MS for 15 (control) or 180 (MS) minutes per day from postnatal day (P)2-14 and were administered saline or LPS (intraperitoneal) on P3. Open-field (OFT) and object-place recognition tests were performed on rat offspring at P33-35 and P42-50, respectively. The pre-frontal cortex (PFC) and hippocampus were removed at the experimental endpoint (P52-55) for mRNA expression. MS induced anxiety-like behaviour in OFT in male and reduced locomotor activity in both male and female offspring. LPS induced a subtle decline in memory in the object-place recognition test in male offspring. MS increased glial fibrillary acidic protein (GFAP) and brain-derived neurotrophic factor expression in PFC and ionised calcium-binding adapter molecule-1 expression in male hippocampus. MS and LPS resulted in distinct behavioural phenotypes in a sex-specific manner. The combination of MS and LPS had a synergistic effect on the anxiety-like behaviour, locomotor activity, and GFAP mRNA expression outcomes.

HIV Nef Expression Down-modulated GFAP Expression and Altered Glutamate Uptake and Release and Proliferation in Astrocytes.

HIV infection of astrocytes leads to restricted gene expression and replication but abundant expression of HIV early genes Tat, Nef and Rev. A great deal of neuroHIV research has so far been focused on Tat protein, its effects on astrocytes, and its roles in neuroHIV. In the current study, we aimed to determine effects of Nef expression on astrocytes and their function. Using transfection or infection of VSVG-pseudotyped HIV viruses, we showed that Nef expression down-modulated glial fibrillary acidic protein (GFAP) expression. We then showed that Nef expression also led to decreased GFAP mRNA expression. The transcriptional regulation was further confirmed using a GFAP promoter-driven reporter gene assay. We performed transcription factor profiling array to compare the expression of transcription factors between Nef-intact and Nef-deficient HIV-infected cells and identified eight transcription factors with expression changes of 1.5-fold or higher: three up-regulated by Nef (Stat1, Stat5, and TFIID), and five down-regulated by Nef (AR, GAS/ISRE, HIF, Sp1, and p53). We then demonstrated that removal of the Sp1 binding sites from the GFAP promoter resulted in a much lower level of the promoter activity and reversal of Nef effects on the GFAP promoter, confirming important roles of Sp1 in the GFAP promoter activity and for Nef-induced GFAP expression. Lastly, we showed that Nef expression led to increased glutamate uptake and decreased glutamate release by astrocytes and increased astrocyte proliferation. Taken together, these results indicate that Nef leads to down-modulation of GFAP expression and alteration of glutamate metabolism in astrocytes, and astrocyte proliferation and could be an important contributor to neuroHIV.

Aquaporin 4 is not present in normal porcine and human lamina cribrosa.

Aquaporin 4 is absent from astrocytes in the rodent optic nerve head, despite high expression in the retina and myelinated optic nerve. The purpose of this study was to quantify regional aquaporin channel expression in astrocytes of the porcine and human mouse optic nerve (ON). Ocular tissue sections were immunolabeled for aquaporins 1(AQP1), 4(AQP4), and 9(AQP9), myelin basic protein (MBP), glial fibrillary acidic protein (GFAP) and alpha-dystroglycan (αDG) for their presence in retina, lamina, myelin transition zone (MTZ, region just posterior to lamina) and myelinated ON (MON). Semi- quantification of AQP4 labeling & real-time quantitative PCR (qPCR) data were analyzed in retina and ON tissue. Porcine and control human eyes had abundant AQP4 in Müller cells, retinal astrocytes, and myelinated ON (MON), but minimal expression in the lamina cribrosa. AQP1 and AQP9 were present in retina, but not in the lamina. Immunolabeling of GFAP and αDG was similar in lamina, myelin transition zone (MTZ) and MON regions. Semi-quantitative AQP4 labeling was at background level in lamina, increasing in the MTZ, and highest in the MON (lamina vs MTZ, MON; p≤0.05, p≤0.01, respectively). Expression of AQP4 mRNA was minimal in lamina and substantial in MTZ and MON, while GFAP mRNA expression was uniform among the lamina, MTZ, and MON regions. Western blot assay showed AQP4 protein expression in the MON samples, but none was detected in the lamina tissue. The minimal presence of AQP4 in the lamina is a specific regional phenotype of astrocytes in the mammalian optic nerve head.

Rapid GFAP and Iba1 expression changes in the female rat brain following spinal cord injury.

Evidence suggests that rapid changes to supporting glia may predispose individuals with spinal cord injury (SCI) to such comorbidities. Here, we interrogated the expression of astrocyte- and microglial-specific markers glial fibrillary acidic protein (GFAP) and ionized calcium binding adaptor molecule 1 (Iba1) in the rat brain in the first 24 hours following SCI. Female Sprague-Dawley rats underwent thoracic laminectomy; half of the rats received a mild contusion injury at the level of the T10 vertebral body (SCI group), the other half did not (Sham group). Twenty-four hours post-surgery the amygdala, periaqueductal grey, prefrontal cortex, hypothalamus, lateral thalamus, hippocampus (dorsal and ventral) in rats were collected. GFAP and Iba1 mRNA and protein levels were measured by real-time quantitative polymerase chain reaction and Western blot. In SCI rats, GFAP mRNA and protein expression increased in the amygdala and hypothalamus. In contrast, gene and protein expression decreased in the thalamus and dorsal hippocampus. Interestingly, Iba1 transcripts and proteins were significantly diminished only in the dorsal and ventral hippocampus, where gene expression diminished. These findings demonstrate that as early as 24 hours post-SCI there are region-specific disruptions of GFAP and Iba1 transcript and protein levels in higher brain regions. All procedures were approved by the University of Technology Sydney Institutional Animal Care and Ethics Committee (UTS ACEC13-0069).

Glial Fibrillary Acidic Protein as Biomarker Indicates Purity and Property of Auricular Chondrocytes.

Instead of the silicone implants previously used for repair and reconstruction of the auricle and nose lost due to accidents and disease, a new treatment method using tissue-engineered cartilage has been attracting attention. The quality of cultured cells is important in this method because it affects treatment outcomes. However, a marker of chondrocytes, particularly auricular chondrocytes, has not yet been established. The objective of this study was to establish an optimal marker to evaluate the quality of cultured auricular chondrocytes as a cell source of regenerative cartilage tissue. Gene expression levels were comprehensively compared using the microarray method between human undifferentiated and dedifferentiated auricular chondrocytes to investigate a candidate quality control index with an expression level that is high in differentiated cells, but markedly decreases in dedifferentiated cells. We identified glial fibrillary acidic protein (GFAP) as a marker that decreased with serial passages in auricular chondrocytes. GFAP was not detected in articular chondrocytes, costal chondrocytes, or fibroblasts, which need to be distinguished from auricular chondrocytes in cell cultures. GFAP mRNA expression was observed in cultured auricular chondrocytes, and GFAP protein levels were also measured in the cell lysates and culture supernatants of these cells. However, GFAP levels detected from mRNA and protein in cell lysates were significantly decreased by increases in the incubation period. In contrast, the amount of protein in the cell supernatant was not affected by the incubation period. Furthermore, the protein level of GFAP in the supernatants of cultured cells correlated with the in vitro and in vivo production of the cartilage matrix by these cells. The productivity of the cartilage matrix in cultured auricular chondrocytes may be predicted by measuring GFAP protein levels in the culture supernatants of these cells. Thus, GFAP is regarded as a marker of the purity and properties of cultured auricular chondrocytes.

From Gut to Brain: Alteration in Inflammation Markers in the Brain of Dextran Sodium Sulfate-induced Colitis Model Mice.

ObjectiveNeuropsychiatric manifestations like depression and cognitive dysfunction commonly occur in inflammatory bowel disease (IBD). In the context of the brain-gut axis model, colitis can lead to alteration of brain function in a bottom-up manner. Here, the changes in the response of the hypothalamic-pituitary-adrenal axis and inflammation-related markers in the brain in colitis were studied.MethodsDextran sodium sulfate (DSS) was used to generate a mouse model of colitis. Mice were treated with DSS for 3 or 7 days and sacrificed. We analyzed the gene expression of brain-derived neurotrophic factor (BDNF), cyclo-oxygenase 2 (COX-2), and glial fibrillary acidic protein (GFAP), and the expression of GFAP, in the hippocampus, hypothalamus, and amygdala. Additionally, the levels of C-reactive protein (CRP) and serum cortisol/corticosterone were measured.ResultsAlteration of inflammatory-related markers varied depending on the brain region and exposure time. In the hippocampus, COX-2 mRNA, GFAP mRNA, and GFAP expression were upregulated during exposure to DSS. However, in the hypothalamus, COX-2 mRNA was upregulated only 3 days after treatment. In the amygdala, BDNF and COX-2 mRNAs were downregulated. CRP and corticosterone expression increased with DSS treatment at day 7.ConclusionIBD could lead to neuroinflammation in a bottom-up manner, and this effect varied according to brain region. Stress-related hormones and serum inflammatory markers, such as CRP, were upregulated from the third day of DSS treatment. Therefore, early and active intervention is required to prevent psychological and behavioral changes caused by IBD, and region-specific studies can help understand the precise mechanisms by which IBD affects the brain.

Intrathecal injection of AG-490 reduces bone-cancer-induced spinal cord astrocyte reaction and thermal hyperalgesia in a mouse model

To investigate the role of spinal interleukin-6-Janus kinase 2 (IL-6-JAK2) signaling transduction pathway in regulating astrocytes activation during the maintenance of bone cancer pain (BCP). Methods: NCTC 2472 fibrosarcoma cells were injected into the femur marrow cavity in C3H/HeNCrlVr male mice to establish BCP model and they were replaced by the equal volume of α-MEM in the sham model. The paw withdrawal latency (PWL) was measured after inoculation of tumor cells. The lumbar enlargement of spinal cord (L3-L5) was isolated, and Real-time RT-PCR and Western blot were used to detect the expression of spinal glial fibrillary acidic protein (GFAP) and JAK2 mRNA and protein, respectively. The expression level of spinal GFAP mRNA indirectly reflect astrocytes activation level. Pain behaviors and spinal cord GFAP mRNA and protein expression were observed at the given time points after intrathecal administration of JAK2 antagonist AG-490. Results: The PWL at 10, 14, 21 d after operation in BCP model group were significantly shorter than that in the sham group (P<0.05); the spinal GFAP and JAK2 mRNA and protein levels were higher in the BCP model group in comparison to mice in the sham group (P<0.05); intrathecal injection of JAK2 agonist AG-490 (30 or 90 nmol) significantly alleviated PWL, and downregulated the expression of spinal GFAP mRNA and protein (P<0.05). Conclusion: The IL-6-JAK2 signaling pathway plays an important role in maintaining the BCP by regulating the expression of GFAP in the spinal cord. Intrathecal injection of AG-490 can reduce the BCP, and inhibit the activation of IL-6-JAK2 signaling pathway, which may be one of the mechanisms for spinal astrocyte activation.

Neuroprotective Properties of Nigella sativa (L.) Seeds Extract in Sprague Dawley Rats models

Neuroinflammation has been closely linked to neurodegeneration that leads to dysfunction of memory and learning where glial fibrillary acidic protein (GFAP) plays important role positively within hippocampus. In experimental neurodegeneration achieved by two-vessel occlusion (2VO) intervention, Nigella sativa (L.) seeds extract (50 mg/kg/day orally) has shown to have anti-neuroinflammatory and antioxidant properties. The brain hippocampal tissues were humanely collected at the end of 10th treatment week and preserved in Allprotect™ reagent at –80°C. Total RNA was extracted and purified by phenol/chloroform method using kits and reverse transcribed into cDNA and relatively quantified as per ΔΔCq. The NSSE treatment showed significantly (p&lt;0.001) different relative GFAP mRNA expression in the treated group as compared to that of untreated 2VO whilst it was insignificantly (p&gt;0.5) different to that of healthy control (HC). Prolonged or daily treatment with NSSE (Nigella sativa (L.) seeds extract) may provide moderate anti-neuroprotective activity within hippocampus.Dhaka Univ. J. Pharm. Sci. 17(1): 113-121, 2018 (June)

Abstract WP418: Mineralocorticoid Receptor Signaling is Associated With Neuroinflammation and Changes in Cognitive Function in Angiotensin II-Induced Hypertension

Introduction: Mineralocorticoid receptor (MR) activation facilitates hypertension-associated cerebral artery dysfunction and reduced perfusion that may increase the risk for vascular dementia. Hypothesis: Angiotensin II (AngII)-hypertension results in neuroinflammation and oxidative stress that will impair cognition in a MR-dependent manner. Methods: 16-17 week old male C57bl/6 mice were treated with AngII (800ng/kg/min) ± the MR antagonist, eplerenone (EPL;100mg/kg/day) for 4 weeks; Sham mice served as control. Data are presented as mean ± SEM; n=4-12 per group *=different from Sham. Results: AngII reduced astrocyte density and increased microglial density; the changes in microglia were prevented by EPL. AngII reduced the mRNA expression of brain-derived neurotrophic factor (BDNF), doublecortin, glial fibrillary acidic protein (GFAP), and synaptophysin (SYP); the reductions in BDNF, GFAP and SYP mRNA expression were prevented by EPL. We assessed cognitive function using the Barnes maze; AngII treated mice spent less time in the target quadrant than the sham or AngII+EPL mice. There was a trend toward there being an increase in the latency to reach the target quadrant in the AngII mice but the changes were not significant. Changes in nest building are associated with cognitive decline, nest building was impaired in AngII mice and these changes were prevented by EPL. There was a trend for oxidized proteins to be increased in AngII mice that may be prevented by EPL but these changes were not significant. Preliminary studies suggest that AngII changes the morphology of cortical neurons suggesting that the increased blood pressure may lead to unhealthy neurons. Conclusion: AngII-hypertension results in neuroinflammation and creates an environment where markers for neuronal support and synapse formation are impaired. These changes are regulated by MR signaling and may be associated with the declined cognitive function.

GFAP expression is influenced by astrocytoma grade and rs2070935 polymorphism.

Glial fibrillary acidic protein (GFAP) is an intermediate filament that provides mechanical support to astrocytes. Rs2070935 is a single nucleotide polymorphism (SNP) located in the promoter region of the GFAP gene. The aim of this pilot study is to investigate GFAP expression at mRNA, protein levels and rs2070935 polymorphism in 50 different grade human astrocytoma samples. GFAP expression at mRNA level was measured using quantitative reverse transcription polymerase chain reaction (qRT-PCR) with SYBR Green dye, whereas the translational activity of the following gene was detected using western blot assay. Furthermore, genotypes of rs2070935 were identified using qPCR with TaqMan probes. As a result, GFAP mRNA and protein expression was found to be declining with increasing astrocytoma grade (p < 0.05). A tendency was observed between increased GFAP mRNA expression and shorter grade IV astrocytoma patient survival (p = 0.2117). The rs2070935 CC genotype was found to be associated with increased GFAP translational activity in grade II astrocytoma (p = 0.0238). Possible links between rs2070935 genotypes and alternative splicing of GFAP were also observed. The rs2070935 AA genotype was found to be associated with poor clinical outcome for grade IV astrocytoma patients (p = 0.0007), although the following data should be checked in a larger sample size of astrocytoma patients.

The effect of netrin-1 on activation of Mueller cell in diabetes rats

Objective To observe the effect of netrin-1 on retinal Muller cells in diabetes mellitus (DM) rats. Methods Fifty Sprague-Dawley rats were randomly divided into the normal control group (group A), normal + balanced salt solution (BSS) group (group B), normal+netrin-1 group (group C), DM+BSS group (group D) and DM+netrin-1 group (group E), with 10 rats in each group. DM rats were induced by intraperitoneal injection of Streptozotocin (60 mg/kg). The expression level of glial fibrillary acidic protein (GFAP) on retinal Muller cells was determined by immunohistochemistry, the level of GFAP mRNA was analyzed by real-time fluorescence quantitative reverse transcription polymerase chain reaction. Results Immunohistochemistry showed that GFAP was distributed in retinal ganglion cells and retinal nerve fiber layer in group A, B and C. Compared to group B, GFAP staining was brighter in the group D. There were significant differences in the expression of GFAP protein and mRNA among groups A-E (F=203.43, 72.91; P=0.00, 0.00), they were higher in group D than group A (t=−26.01, 22.26; P=0.00, 0.00), and group E (t=−10.78, 3.93; P=0.00, 0.00). They were higher in group E than group A (t=7.00, −9.82; P=0.00, 0.00). There were no significant differences in between group A and group C (t=−0.29, 0.50; P=0.77, 0.62). Conclusion The expression of GFAP in Muller cells of DM rats could be decreased by injecting netrin-1 into vitreous. Key words: Diabetic retinopathy/physiopathology; Microglia/physiology; Nerve growth factors; Animal experimentation

Cellular stress response in human Müller cells (MIO-M1) after bevacizumab treatment

Bevacizumab, an anti-vascular endothelial growth factor (VEGF) agent, is widely used in the treatment of retinal vascular diseases. However, due to the essential role Müller cell derived-VEGF plays in the maintenance of retinal neurons and glial cells, cell viability is likely to be affected by VEGF inhibition. We therefore evaluated the effect of bevacizumab-induced VEGF inhibition on Müller cells (MIO-M1) in vitro. MIO-M1 cells were cultured for 12 or 24 h in media containing bevacizumab at 0.25 or 0.5 mg/mL. Controls were cultured in medium only. Cell viability was determined with the trypan blue exclusion test and MTT assay. Caspase-3, beclin-1, glial fibrillary acidic protein (GFAP) and vimentin content were quantified by immunohistochemistry. Gene expression was evaluated by real-time quantitative PCR. Treatment with bevacizumab did not reduce MIO-M1 cell viability, but increased metabolic activity at 24 h (0.5 mg/mL) and induced apoptosis and autophagy, as shown by the increased caspase-3 levels at 12 h (0.25 and 0.5 mg/mL) and the increased beclin levels at 24 h (0.5 mg/mL). Caspase-3 mRNA was upregulated at 12 h and downregulated at 24 h in cells treated with bevacizumab at 0.25 mg/mL. Bevacizumab treatment was also associated with structural protein abnormalities, with decreased GFAP and vimentin content and upregulated GFAP and vimentin mRNA expression. Although bevacizumab did not significantly affect MIO-M1 cell viability, it led to metabolic and molecular changes (apoptosis, autophagy and structural abnormalities) suggestive of significant cellular toxicity.

Increased expression of astrocyte markers in schizophrenia: Association with neuroinflammation.

While schizophrenia may have a progressive component, the evidence for neurodegenerative processes as indicated by reactive astrocytes is inconclusive. We recently identified a subgroup of individuals with schizophrenia with increased expression of inflammatory markers in prefrontal cortex, and hypothesized that this subgroup would also have reactive astrocytes. We measured glial fibrillary acidic protein (GFAP) mRNA by quantitative real-time reverse transcriptase polymerase chain reaction (RT-PCR) and protein levels by immunoblotting in grey matter homogenate from 37 individuals with schizophrenia and 37 unaffected controls. We examined the morphology of GFAP-positive astrocytes in immunostained sections of middle frontal gyrus. We tested if GFAP expression or astrocyte morphology were altered in people with schizophrenia with increased expression of inflammatory markers. We used RNA-Seq data on a subset of patients and controls (n=20/group) to ascertain whether mRNA transcripts associated with astrogliosis were elevated in the individuals with active neuroinflammation. GFAP (mRNA and protein) levels and astrocyte morphology were not significantly different between people with schizophrenia and controls overall. However, individuals with schizophrenia with neuroinflammation had increased expression of GFAP mRNA (t(33)=2.978, p=0.005), hypertrophic astrocyte morphology (χ(2)(2)=6.281, p=0.043), and statistically significant elevated expression of three mRNA transcripts previously associated with astrogliosis. We found clear evidence of astrogliosis in a subset of people with schizophrenia. We suggest that the lack of astrogliosis reported in previous studies may be due to cohort differences in aetiopathology, illness stage, treatment exposure, or a failure to examine subsets of people with schizophrenia.

Expression of dedifferentiation markers and multilineage markers in U251 glioblastoma cells with silenced EGFR and FGFR genes

Epithelial growth factor (EGF) and basic fibroblast growth factor (bFGF), and their receptors, epithelial growth factor receptor (EGFR) and bFGF receptor (bFGFR), are frequently overexpressed in high-grade gliomas. In the present study, the EGF and bFGF levels in U251 glioblastoma cell culture supernatants were determined by ELISA, and enhanced green fluorescent protein (EGFP)-labeled recombinant lentiviral expression vectors with small interfering RNA targeting the EGFR and bFGFR genes were constructed. The mRNA expression levels of EGFR, bFGFR, cluster of differentiation (CD)133, glial fibrillary acidic protein (GFAP), tubulin-β3 (TUBB3) and myelin basic protein (MBP) were determined using quantitative polymerase chain reactions in U251 cells prior to and following silencing of the EGFR and/or bFGFR genes. Prior to silencing, the U251 cells secreted EGF and bFGF, and expressed EGFR, bFGFR, CD133, GFAP, TUBB3 and MBP mRNA. Subsequent to silencing the EGFR and/or bFGFR gene, CD133 mRNA expression decreased and GFAP and TUBB3 mRNA expression increased. Silencing the EGFR and FGFR genes acted synergistically to downregulate CD133 expression. The downregulation of CD133 mRNA expression and the upregulation of GFAP and TUBB3 mRNA expression were not significantly different when blocking the EGFR and FGFR pathways. These results indicate that autocrine or paracrine EGF and/or FGF mechanisms exist in U251 cells. Knocking down the EGFR and/or FGFR genes downregulates CD133 mRNA expression and facilitates glial and neuronal differentiation in U251 cells.

Role of Telomerase Reverse Transcriptase in Glial Scar Formation After Spinal Cord Injury in Rats

The study aims to determine the expression of telomerase reverse transcriptase (TERT) in the glial scar following spinal cord injury in the rat, and to explore its relationship with glial scar formation. A total of 120 Sprague–Dawley rats were randomly divided into three groups: SCI only group (without TERT interference), TERT siRNA group (with TERT interference), and sham group. The TERT siRNA and SCI only groups received spinal cord injury induced by the modified Allen’s weight drop method. In the sham group, the vertebral plate was opened to expose the spinal cord, but no injury was modeled. Five rats from each group were sacrificed under anesthesia at days 1, 3, 5, 7, 14, 28, 42, and 56 after spinal cord injury. Specimens were removed for observation of glial scar formation using hematoxylin-eosin staining and immunofluorescence detection. mRNA and protein expressions of TERT and glial fibrillary acidic protein (GFAP) were detected by reverse-transcription (RT)-PCR and western blotting, respectively. Hematoxylin-eosin staining showed evidence of gliosis and glial scarring in the spinal cord injury zone of the TERT siRNA and SCI only groups, but not in the sham group. Immunofluorescence detection showed a significant increase in GFAP expression at all time points after spinal cord injury in the SCI only group (81 %) compared with the TERT siRNA group (67 %) and sham group (2 %). In contrast, the expression of neurofilament protein 200 (NF-200) was gradually reduced and remained at a stable level until 28 days in the SCI only group. There were no NF-200-labeled cells in the spinal cord glial scar and cavity at day 56 after spinal cord injury. NF-200 expression at each time point was significantly lower in the SCI only group than the TERT siRNA group, while there was no change in the sham group. Western blotting showed that TERT and GFAP protein expressions changed dynamically and showed a linear relationship in the SCI only group (r = 0.765, P < 0.01), while there was no obvious linear relationship in the sham group (r = 0.208, P = 0.121). RT-PCR results showed a dynamic expression of TERT and GFAP mRNA in the SCI only group, exhibiting a linear relationship (r = 0.722, P < 0.01), while there was no linear relationship in the sham group (r = 0.206, P = 0.180). Our data indicate that TERT has a dynamic expression in the spinal cord glial scar, which positively correlates to GFAP expression, and may be important for promoting glial scar formation.

Role of chemokine CXC ligand 13 in spinal cord in neuropathic pain in rats

Objective To evaluate the role of chemokine CXC ligand 13 (CXCL13) in spinal cord in neuropathic pain (NP) in rats.Methods One hundred and eight male Sprague-Dawley rats,weighing 150-200 g,were randomly divided into 4 groups (n =27 each):sham operation group (group S),group NP,small interference RNA (siRNA) negative control group (group NS) and CXCL13-siRNA group (group CS).The animals were anesthetized with intraperitoneal ketamine 50 mg/kg.NP was induced by ligation of L5 spinal nerve (SNL) in groups NP,NS and CS.L5 spinal nerve was only exposed but not occluded in group S.CXCL13-siRNA lentivirus and siRNA negative control lentivirus were injected intrathecally in groups CS and NS,respectively.Mechanical pain threshold was measured at 3,7 and 14 days after SNL.Then the rats were sacrificed and L4-6 segments of the spinal cord were obtained for determination of coexpression of CXCL13 and Neun (by immunofluorescence),activation of astrocytes,and expression of CXCL13 and glial fibrillary acidic protein (GFAP) protein (by Western blot) and mRNA (by RT-PCR) in spinal cord tissues.Results Compared with group S,mechanical pain threshold was significantly decreased and the expression of CXCL13 and GFAP protein and mRNA was up-regulated at each time point after operation in groups NP,NS and CS (P ＜ 0.05).Compared with group NP,mechanical pain threshold was significantly increased and the expression of CXCL13 and GFAP protein and mRNA was down-regulated at each time point after operation in group CS (P ＜ 0.05).There was no significant difference in the indexes mentioned above at each time point after operation between groups NP and NS (P ＞ 0.05).Conclusion CXCL13 is involved in the development and maintenance of NP in rats via activation of astrocytes. Key words: Chemokine CXCL13 ; Neuralgia; Spinal cord

Curcumin as a Potential Treatment for Alzheimer's Disease: A Study of the Effects of Curcumin on Hippocampal Expression of Glial Fibrillary Acidic Protein

Curcumin, an agent traditionally utilized for its preventative action against tumorigenesis, oxidation, inflammation, apoptosis and hyperlipemia, has also been used in the treatment of Alzheimer's disease (AD). Recent advances in the study of AD have revealed astrocytes (AS) as being key factors in the early pathophysiological changes in AD. Glial fibrillary acidic protein (GFAP), a marker specific to AS, is markedly more manifest during morphological modifications and neural degeneration signature during the onset of AD. Several studies investigating the functionality of curcumin have shown that it not only inhibits amyloid sedimentation but also accelerates the disaggregation of amyloid plaque. Thus, we are interested in the relationship between curcumin and spatial memory in AD. In this study, we intend to investigate the effects of curcumin in amyloid-β (Aβ(1-40)) induced AD rat models on both the behavioral and molecular levels, that is to say, on their spatial memory and on the expression of GFAP in their hippocampi. Our results were statistically significant, showing that the spatial memory of AD rats improved following curcumin treatment (p < 0.05), and that the expression of GFAP mRNA and the number of GFAP positive cells in the curcumin treated rats was decreased relative to the AD group rats (p < 0.05). Furthermore, the expression level of GFAP mRNA in hippocampal AS in the AD rats significantly increased when compared with that in the sham control (p < 0.05). Taken together, these results suggest that curcumin improves the spatial memory disorders (such disorders being symptomatic of AD) in Aβ(1-40)-induced rats by down regulating GFAP expression and suppressing AS activity.

Effect of Jiaji electroacupuncture in transected rat spinal cord

We evaluated the effect of Jiaji electroacupuncture on cell proliferation and the expression of markers of endogenous neural stem cell activation after complete spinal cord transection. Female Wistar rats were assigned to 4 groups (n = 24 each): a sham-operated group, a control group, a Jiaji electroacupuncture group, and a Jiaji electroacupuncture preconditioning group. Motor function was significantly improved in the acupuncture groups compared to the control group at 7 and 14 d. Numbers of bromodeoxyuridine (BrdU)-, nestin-, and glial fibrillary acidic protein (GFAP)-positive cells were significantly greater in the acupuncture groups than in the controls at each time point. Expression of nestin and GFAP mRNA was significantly higher in the acupuncture groups than in the controls at each time point. Thus, Jiaji electroacupuncture and preconditioning may promote the proliferation of endogenous neural stem cells after spinal cord transection.

Altered glial gene expression, density, and architecture in the visual cortex upon retinal degeneration

Genes encoding the proteins of cytoskeletal intermediate filaments (IF) are tightly regulated, and they are important for establishing neural connections. However, it remains uncertain to what extent neurological disease alters IF gene expression or impacts cells that express IFs. In this study, we determined the onset of visual deficits in a mouse model of progressive retinal degeneration ( Pde6b − mice; Pde6b + mice have normal vision) by observing murine responses to a visual task throughout development, from postnatal day (PND) 21 to adult ( N = 174 reliable observations). Using Q-PCR, we evaluated whether expression of the genes encoding two Type III IF proteins, glial fibrillary acidic protein (GFAP) and vimentin was altered in the visual cortex before, during, and after the onset of visual deficits. Using immunohistochemical techniques, we investigated the impact of vision loss on the density and morphology of astrocytes that expressed GFAP and vimentin in the visual cortex. We found that Pde6b − mice displayed 1) evidence of blindness at PND 49, with visual deficits detected at PND 35, 2) reduced GFAP mRNA expression in the visual cortex between PND 28 and PND 49, and 3) an increased ratio of vimentin:GFAP-labeled astrocytes at PND 49 with reduced GFAP cell body area. Together, these findings demonstrate that retinal degeneration modifies cellular and molecular indices of glial plasticity in a visual system with drastically reduced visual input. The functional consequences of these structural changes remain uncertain.

Minocycline reduces the injury-induced expression of prodynorphin and pronociceptin in the dorsal root ganglion in a rat model of neuropathic pain

A role of neuropeptides in neuropathic pain development has been implicated; however, the neuroimmune interactions that are involved in the underlying mechanisms may be more important than previously thought. To examine a potential role of relations between glia cells and neuropeptides in neuropathic pain, we performed competitive reverse-transcription polymerase chain reaction (RT-PCR) from the dorsal lumbar spinal cord and the dorsal root ganglion (DRG) after chronic constriction injury (CCI) in the rat sciatic nerve. The RT-PCR results indicated that complement component 1, q subcomponent (C1q) mRNA expression was higher than glial fibrillary acidic protein (GFAP) in the spinal cord 3 and 7 days post-CCI, suggesting that spinal microglia and perivascular macrophages are more activated than astrocytes. In parallel, we observed a strong upregulation of prodynorphin mRNA in the spinal cord after CCI, with no changes in the expression of proenkephalin or pronociceptin. Conversely, the expression of GFAP mRNA in the DRG was higher than C1q, which suggests that the satellite cells are activated shortly after injury, followed by the macrophages and polymorphonuclear leukocytes infiltrating the DRG. In the DRG, we also observed a very strong upregulation of prodynorphin (1387%) as well as pronociceptin (122%) and a downregulation of proenkephalin (47%) mRNAs. Interestingly, preemptive and repeated i.p. injection of minocycline reversed the activation of microglia/macrophages in the spinal cord and the trafficking of peripheral immune cells into the DRG, and markedly diminished the upregulation of prodynorphin and pronociceptin in the DRG. We thus provide novel findings that inhibition of C1q-positive cells by minocycline can diminish injury-induced neuropeptide changes in the DRG. This suggests that immune cells-derived pronociceptive factors may influence opioid peptide expression. Therefore, the injury-induced activation of microglia and leukocytes and the subsequent activation of neuropeptides involved in nociception processes are potential targets for the attenuation of neuropathic pain.