Glia Fibrillary Acidic Protein Research Articles

Histomorphological Investigation of the Eye of the Tree Squirrel: A Preliminary Study

Abstract Squirrels are diurnal rodents with high visual acuity including unique properties well-suited for their natural environment. This study was conducted to explore some ocular microscopic features of tree-harbouring squirrels in the University of Ibadan, Ibadan, Nigeria. Two male squirrels were cage-trapped within the University premises. Light microscopic analysis was carried on paraffin-embedded eye samples harvested from the animals. The densely compacted stromal fibres, 351 ± 52.5 µm thick, formed the thickest part of the cornea, and the basement membrane of the corneal epithelium, 63.8 ± 13.0 µm thick, was notably positive with Periodic Acid Schiff (PAS) stain. Strong pigmentation was present at the choroid as well as the iridal and ciliary epithelia. The multiple layering of the retinal structure exhibited densely packed ganglion cells at the ganglion cell layer which together with the nerve fibre layer was observed to be thinnest at the more peripheral portion but becomes thicker towards the optic disc. Strongly positive glia fibrillary acidic protein (GFAP+) cells with their abundant fibrous processes were demonstrated immunohistochemically at the retinal nerve fibre layer and the optic nerve. Histological features of the retinal cellular components of the tree squirrels investigated has thus highlighted the structural adaptation of these animal species to their environmental arboreal habitat and diurnal lifestyle. Findings from this study, while further noted to be similar to that in human, showed that African tree squirrels represent promising rodent model for human retinal/ocular research.

Peroxisome proliferator-activated receptor gamma agonist pioglitazone alleviates hemorrhage-induced thalamic pain and neuroinflammation

BackgroundThalamic pain frequently occurs after stroke and is a challenging clinical issue. However, the mechanisms underlying thalamic pain remain unclear. Neuroinflammation is a key determining factor in the occurrence and maintenance of hemorrhage-induced thalamic pain. Pioglitazone is an agonist of peroxisome proliferator-activated receptor gamma (PPARγ) and shows anti-inflammatory effects in multiple diseases. The present work focused on exploring whether PPARγ is related to hemorrhage-induced thalamic pain. MethodsImmunostaining was conducted to analyze the cellular localization of PPARγ and co-localization was evaluated with NeuN, ionized calcium-binding adapter molecular 1 (IBA1), and glia fibrillary acidic protein (GFAP). Western blot analyses were used to evaluate MyD88, pNF-κB/NF-κB, pSTAT6/STAT6, IL-1β, TNF-α, iNOS, Arg-1, IL-4, IL-6, and IL-10 expression. Behavioral tests in mice were conducted to evaluate continuous pain hypersensitivity. ResultsWe found that pioglitazone appeared to mitigate the contralateral hemorrhage-induced thalamic pain while inhibiting inflammatory responses. Additionally, Pioglitazone induced phosphorylation of STAT6 and suppressed the phosphorylation NF-κB in our model of thalamic pain. These effects could be partially reversed with the PPARγ antagonist GW9662. ConclusionThe PPARγ agonist pioglitazone can mitigate mechanical allodynia by suppressing the NF-κB inflammasome while activating the STAT6 signal pathway, which are well-known to be associated with inflammation.

Suppression of neuronal apoptosis and glial activation with modulation of Nrf2/HO-1 and NF-kB signaling by curcumin in streptozotocin-induced diabetic spinal cord central neuropathy.

Diabetes is a global disease, commonly complicated by neuropathy. The spinal cord reacts to diabetes by neuronal apoptosis, microglial activation, and astrocytosis, with a disturbance in neuronal and glial Nuclear factor erythroid 2-related factor/Heme oxygenase-1 (Nrf2/HO-1) and Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB) signaling. Curcumin, a bioactive natural substance, showed neuroprotective role in many diseases. However, its role in the treatment of the diabetic central neuropathy of spinal cord and the underlying mechanisms still need clarification. The present study tried to evaluate the role of curcumin in diabetes-induced central neuropathy of the spinal cord in rats. Twenty rats were divided into three groups; group 1: a negative control group; group 2: received streptozotocin (STZ) to induce type I diabetes, and group 3: received STZ + Curcumin (150 mg/kg/day) for eight weeks. The spinal cords were examined for histopathological changes, and immunohistochemical staining for Glia fibrillary acidic protein (GFAP); an astrocyte marker, Ionized calcium-binding adaptor molecule 1 (Iba1), a microglial marker, neuronal nuclear protein (NeuN); a neuronal marker, caspase-3; an apoptosis marker, Nrf2/HO-1, NF-kB, and oxidative stress markers were assessed. Curcumin could improve spinal cord changes, suppress the expression of Iba1, GFAP, caspase-3, and NF-kB, and could increase the expression of NeuN and restore the Nrf2/HO-1 signaling. Curcumin could suppress diabetic spinal cord central neuropathy, glial activation, and neuronal apoptosis with the regulation of Nrf2/HO-1 and NF-kB signaling.

Neuroprotective potential of plant derived parenchymal stem cells extract on environmental and genetic models of Parkinson disease through attenuation of oxidative stress and neuroinflammation.

Parkinson's disease (PD) is a neurodegenerative disorder characterized by both motor and non-motor features. The current treatment regimen for PD are dopamine enhancers which have been reported to worsen the disease prognosis after long term treatment, thus, the need for better treatment options. This study sought to investigate the protective action of Double Stem Cell® (DSC), a blend of stem cells extracts from Swiss apples (Malus Domestica) and Burgundy grapes (Vitis vinifera) on 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced Parkinsonism in mice and genetic model of PD in Drosophila melanogaster. Male albino mice were pretreated with MPTP (4 × 20mg/kg, i.p., two hourly in 8h), twelve hours before administration of DSC (8, 40, or 200mg/kg, p.o.). Thereafter, behavioural, biochemical and immunohistochemical assays were carried out. The impact of vehicle or DSC supplementation on α-synuclein aggregation was evaluated in Drosophila melanogaster using the UAS-Gal4 system, female DDC-Gal4 flies were crossed with male UAS-α-synuclein, the progenies were examined for fecundity, locomotion, memory, and lifespan. MPTP-induced motor deficits in open field test (OFT), working memory impairment (Y-maze test (YMT)) and muscle incoordination (rotarod test) were ameliorated by DSC (8, 40 or 200mg/kg) through dose-dependent and significant improvements in motor, cognitive and motor coordination. Moreso, MPTP exposure caused significant increase in lipid peroxidation and decrease in antioxidant enzymes activities (glutathione, catalase and superoxide dismutase) in the midbrain which were attenuated by DSC. MPTP-induced expression of microglia (iba-1), astrocytes (glia fibrillary acidic protein; GFAP) as well as degeneration of dopamine neurons (tyrosine hydroxylase positive neurons) in the substantia nigra (SN) were reversed by DSC. Supplementation of flies feed with graded concentration of DSC (0.8, 4 or 20mg/ml) did not affect fecundity but improved climbing activity and lifespan. Findings from this study showed that Double Stem Cell improved motor and cognitive functions in both mice and Drosophila through attenuation of neurotoxin-induced oxidative stress and neuroinflammation.

The effects of social defeat stress on hippocampal glial cells and interleukin-6 in adolescence and adulthood.

Adolescent social stress has been associated with the vulnerability to developing psychopathological disorders in adulthood that are accompanied by brain inflammatory processes. The purpose of this study is to investigate the dynamic changes of the hippocampal neuroinflammatory mediators, including microglia, astrocyte, and interleukin-6 (IL-6) levels in mice experiencing social defeat stress during adolescence. Adolescent mice were divided into the control group and stress group. Mice in the stress group were exposed to chronic intermittent social defeat for a total of 12 days, and control mice were reared in normal conditions. The hippocampal microglia, astrocyte, and IL-6 levels were measured 24 h and 3 weeks after the end of stress exposure. Microglia activation characterized by increased ionized calcium-binding adapter molecule 1 positive cell numbers or staining area in the CA1 and CA3 regions of the hippocampus were observed 24 h after the end of stress, which did not last into the adulthood. No short-term or long-term alterations of the number of hippocampal CA1 and CA3 glia fibrillary acidic protein astrocytes were found in mice experiencing adolescent social defeat, whereas IL-6 levels were only increased 3 weeks after the end of stress. These data suggested that exposure to chronic social defeat stress led to short-term and long-term neuroinflammatory changes in the hippocampus.

The protective effect of biochanin A against rotenone-induced neurotoxicity in mice involves enhancing of PI3K/Akt/mTOR signaling and beclin-1 production

Rotenone is an insecticide that generates oxidative stress in the CNS and induces locomotor dysfunction and neurodegeneration in rodents. Biochanin A [BioA] is an isoflavone with antioxidant and anti-inflammatory actions. The antioxidant and the modulatory action of BioA on PI3K/Akt/mTOR signaling and autophagy were tested in rotenone-Parkinsonian mice. Mice were allocated into; Group I: oil control group, Group II: rotenone group [1-mg/kg/48h, subcutaneously], group III: rotenone and BioA [10-mg/kg]. Rotenone injection resulted in locomotor disturbances in mice, degeneration in dopaminergic neurons [tyrosine hydroxylase-immunoreactive cells], low striatal dopamine, increased malondialdehyde and decreased level of glutathione. Neuroinflammation was evidenced by upregulation of astrocytes [glia fibrillary acidic protein, GFAP] and elevated levels of cytokines. The phosphorylation of PI3K/Akt/mTOR and the autophagy-related protein, beclin-1, were decreased significantly as indicated by Western blot analysis. BioA treatment enhanced locomotor activity and afforded nigral neuroprotection. The mechanism by which BioA produced this effect includes increased antioxidant defenses, lessened proinflammatory cytokines, increased phosphorylation of PI3K/Akt/mTOR proteins and upregulated beclin-1. Importantly, BioA suppressed the striatal astrocyte marker [GFAP]. Overall, the currents study highlighted that BioA activates PI3K/Akt/mTOR signaling and enhances beclin-1 leading to neuroprotection for nigral dopaminergic neurons.

Activation of pro-apoptotic cells, reactive astrogliosis and hyperphosphorylation of tau protein in trimethyltin-induced hippocampal injury in rats

Neurodegenerative diseases cause neural cells to lose both the functional and sensory abilities as a result of genetic factors, proteopathies and mitochondrial dysfunction. Neurodegeneration forms the basis of most neurodegenerative disorders for example Alzheimer’s disease, Huntington’s diseases, and Parkinson’s diseases. The mechanism that underlines the process of neurodegeneration is not well understood. Understanding the process and mechanism involved in neurodegeneration might offer a better therapeutic approach to positively manage cases of neurodegenerative diseases. Therefore, this study’s target was to create an animal model to study neurodegeneration. Sixteen adult male Wistar rats were used in the study and divided into two groups. Control (0.2 mL of normal saline (NS)), and trimethyltin-treated (TMT, 8 mg/kg stat dose only). These animals underwent perfusion with 4% paraformaldehyde, brain excision and analysis of p53 antigen, GFAP and Bielshowsky on these tissues. The results showed that animals in the control group showed presence of activated p53 antigen, reactive astrogliosis, neurofibrillary tangles, and amyloid plaques within the cytoplasm of the hippocampal cells. Cornus Ammonis (CA2) and (CA3) showed more of the trimethylrtin injury than CA1 and CA4. This study thus revealed that, intra-peritoneal administration of single dose of 8mg/kg of trimethyltin can offer an attractive disease model to study some neurodegenerative diseases.
 Keywords: p53 antigen, Bielshowsky, Glia fibrillary acidic protein, Trimethyltin, Hippocampus,

Ficus exasperata Vahl leaves extract attenuates motor deficit in vanadium-induced parkinsonism mice.

Medicinal herbs have played significant roles in the treatment of various diseases in humans and animals. Sodium metavanadate is a potentially toxic environmental pollutant that induces oxidative damage, neurological disorder, Parkinsonism and Parkinson-like disease upon excessive exposure. This study is designed to investigate the impact of saponin fraction of Ficus exasperata Vahl leaf extract (at 50 and 100 mg/kg body weight for 14 days at different animal groupings) on vanadium treated mice. Animals were randomly grouped into five groups. Control (normal saline), NaVO3 (10 mg/kg for 7 days), withdrawal group, NaVO3+Vahl (low dose) and NaVO3+Vahl (high dose). The animals were screened for motor coordination using rotarod and PBTs and a post mortem study was conducted by quantitatively assessing the markers of oxidative stress such as lipid peroxidation, catalase, glutathione activities, and also through immunohistochemistry via glia fibrillary acidic protein, tyrosine hydroxylase and dopamine transporter to study the integrity of astrocytes and dopaminergic neurons of the substantia nigra (SNc). Vanadium-exposed group showed a decreased motor activity on the neurobehavioural tests as well as an increase in markers of oxidative stress. Saponin fraction of F. exasperata Vahl leaves extract produced a statistically significant motor improvement which may be due to high antioxidant activities of saponin, thereby providing an ameliorative effect on the histoarchitecture of the SNc. It can be inferred that the saponin fraction of F. exasperata Vahl leaves extract to possesses ameliorative, motor-enhancing and neurorestorative benefit on motor deficit in vanadium-induced parkinsonism mice.

Long-term culturing of porcine nodose ganglia

BackgroundNeuronal cell cultures are widely used in the field of neuroscience. Cell dissociation allows for the isolation of a desired cell type, yet the neuronal complexity that distinguishes the nervous system is often lost as a result. Thus, culturing neural tissues in ex vivo format provides a physiological context that more closely resembles the in vivo environment. New methodWe developed a simple method to culture nodose ganglia neurons from neonatal pigs long-term in ex vivo format using an in-house media formulation derived from commercially available components. ResultsGanglia were cultured for six and twelve months. mRNA expression of nestin was stable across time. Vasoactive intestinal peptide and tachykinin showed statistically insignificant increases and decreases in mRNA expression, respectively. mRNA expression of glia fibrillary acidic protein decreased, whereas myelin basic protein showed no statistically significant differences, over time. Immunofluorescence studies of sectioned ganglia demonstrated neurofilament-positive cell bodies, glia fibrillary acidic protein and myelin basic protein at all time points. A significant decrease in cell nuclei density and fragmented DNA were noted. Comparison with existing method(s)There are currently no methods that describe short-term or long-term culturing of porcine nodose ganglia. Further, the media formulation we developed is new and not previously reported. ConclusionsThe simple procedure we developed for culturing nodose ganglia will enable both short-term and long-term investigations aimed at understanding peripheral ganglia in vitro. It is also possible that the methods described herein can be applied to other models, different developmental stages, and potentially other neural tissues.

Pregabalin affords retinal neuroprotection in diabetic rats: Suppression of retinal glutamate, microglia cell expression and apoptotic cell death

Pregabalin is the first drug to receive FDA approval for treating diabetic neuropathic pain. This study investigated the neuroprotective effect of pregabalin in an experimental model of diabetic retinopathy and tested some possible mechanisms underlying the putative neuroprotective effect. Male Wistar rats received streptozotocin (45 mg/kg) to induce type 1 diabetes mellitus. After two weeks, a course of pregabalin (3, 10 and 30 mg/kg) has been launched for five consecutive weeks. Retinal expression of interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) was estimated by real-time PCR and retinal glutamate content was also estimated. Further, retinal caspase-3 immunoblotting and DNA fragmentation assays determined the degree of apoptosis. Pregabalin improved histopathological abnormalities in diabetic retinas and suppressed the diabetes-enhanced retinal expression of IL-1β, TNF-α, CD11b (a surface marker for microglia) while attenuated expression of caspase-3 and DNA fragmentation versus the diabetic group. In addition, diabetic rats treated with pregabalin displayed reductions in retinal glutamate, nitric oxide and malondialdehyde (MDA) and enhanced reduced glutathione (GSH) content versus the diabetic controls. Furthermore, pregabalin enhanced the histopathological picture and reduced fibrosis in the optic nerve of diabetic rats in addition to suppression of the content of the glia fibrillary acidic protein. The findings provide the first evidence demonstrating that pregabalin alleviates retinal neuroinflammation, apoptosis and oxidative stress in an experimental type 1 diabetes mellitus. Therefore, pregabalin might serve as a potential therapy for retinopathy after adequate clinical research.

Inhibitory effect of valproate sodium on pain behavior in diabetic mice involves suppression of spinal histone deacetylase 1 and inflammatory mediators

Anti-epileptic medications are included in the international guidelines for managing neuropathic pain. Valproate sodium (VPS) was recently described as “the forgotten analgesic” and has been reported to relief pain in various models of neuropathic pain. Some studies reported anti-inflammatory and histone deacetylase 1 (HDA1) inhibitory properties for sodium valproate. The aim of the current study was to investigate the modulatory effect of VPS on pain behavior and inflammatory reactions in alloxan-induced diabetic neuropathy focusing on HDA1 inhibition and glia reactivity. 28 Male Swiss albino mice were allocated into four groups, (1) vehicle group, (2) alloxan-diabetic group, (3 & 4) alloxan+VPS (25 or 50 mg/kg) groups. VPS was given daily for 5 weeks by oral gavage. Pain behavior demonstrated increased allodynia (von-Frey filaments) and hyperalgesia (hot-plate test) in alloxan-diabetic mice that was reduced significantly by at least one of VPS doses. Sciatic nerves in diabetic mice showed increased histopathology score, increased silver staining for the nerves-indicating myelopathy- and a decrease in immunostaining for nerve growth factor. Spinal cord of diabetic mice showed greater histopathologic score, increased CD11b and glia fibrillary acidic protein (GFAP) immunostaining than vehicle treated mice. Molecular investigations highlighted greater content of spinal histone deacetylases, tumor necrosis factor-α (TNF-α) and interlukin-1β (IL1β) that were favorably modified by VPS. Overall, the current data confirmed that the pain killing and anti-inflammatory activity of VPS is at least partly mediated through inhibition of spinal HDA1 and glia reactivity. These findings support the view of inviting antiepileptics for treating neuropathies.

Retinal Neuron Is More Sensitive to Blue Light-Induced Damage than Glia Cell Due to DNA Double-Strand Breaks.

Blue light is a major component of visible light and digital displays. Over-exposure to blue light could cause retinal damage. However, the mechanism of its damage is not well defined. Here, we demonstrate that blue light (900 lux) impairs cell viability and induces cell apoptosis in retinal neurocytes in vitro. A DNA electrophoresis assay shows severe DNA damage in retinal neurocytes at 2 h after blue light treatment. γ-H2AX foci, a specific marker of DNA double-strand breaks (DSBs), is mainly located in the Map2-posotive neuron other than the glia cell. After assaying the expression level of proteins related to DNA repair, Mre11, Ligase IV and Ku80, we find that Ku80 is up-regulated in retinal neurocytes after blue light treatment. Interestingly, Ku80 is mainly expressed in glia fibrillary acidic protein (GFAP)-positive glia cells. Moreover, following blue light exposure in vivo, DNA DSBs are shown in the ganglion cell layer and only observed in Map2-positive cells. Furthermore, long-term blue light exposure significantly thinned the retina in vivo. Our findings demonstrate that blue light induces DNA DSBs in retinal neurons, and the damage is more pronounced compared to glia cells. Thus, this study provides new insights into the mechanisms of the effect of blue light on the retina.

Acetylcholine suppresses the increase of glia fibrillary acidic protein expression via acetylcholine receptors in cAMP-induced astrocytic differentiation of rat C6 glioma cells

Astrocytes, the most common glial cells in the central nervous system, maintain neuronal functions and have roles in neurological diseases. Acetylcholine (ACh) is one of the most essential neurotransmitters, and ACh receptor (AChR) ligands were recently reported to influence astrocyte functions. However, the functions of ACh, the only endogenous agonist of AChR, in astrocytogenesis and in the expression of astrocytic marker genes have not been known. We previously demonstrated that the inhibition of acetylcholine esterase (AChE) suppressed the differentiation of rat glioma C6 cells, an astrocyte differentiation model, and we observed a suppressive effect of ACh agonists on astrocyte differentiation. Our present study revealed that in the cAMP-induced differentiation of C6 cells, an AChR antagonist alleviated the expression of glia fibrillary acidic protein (GFAP) that had been suppressed by dichlorvos (DDVP), an organophosphate and an AChE inhibitor. Our findings also demonstrated a direct effect of ACh on the GFAP expression, and that muscarinic AChR is involved in the suppressive effect of ACh on the GFAP expression in differentiation-induced C6 cells. This is the first report indicating that ACh the only endogenous agonist for AChRs functions as a mediator of astrocyte differentiation.

Interaction of Human Enterochromaffin Cells with Human Enteric Adenovirus 41 Leads to Serotonin Release and Subsequent Activation of Enteric Glia Cells.

Human adenovirus 41 (HAdV-41) causes acute gastroenteritis in young children. The main characteristics of HAdV-41 infection are diarrhea and vomiting. Nevertheless, the precise mechanism of HAdV-41-induced diarrhea is unknown, as a suitable small-animal model has not been described. In this study, we used the human midgut carcinoid cell line GOT1 to investigate the effect of HAdV-41 infection and the individual HAdV-41 capsid proteins on serotonin release by enterochromaffin cells and on enteric glia cell (EGC) activation. We first determined that HAdV-41 could infect the enterochromaffin cells. Immunofluorescence staining revealed that the cells expressed HAdV-41-specific coxsackievirus and adenovirus receptor (CAR); flow cytometry analysis supported these findings. HAdV-41 infection of the enterochromaffin cells induced serotonin secretion dose dependently. In contrast, control infection with HAdV-5 did not induce serotonin secretion in the cells. Confocal microscopy studies of enterochromaffin cells infected with HAdV-41 revealed decreased serotonin immunofluorescence compared to that in uninfected cells. Incubation of the enterochromaffin cells with purified HAdV-41 short fiber knob and hexon proteins increased the serotonin levels in the harvested cell supernatant significantly. HAdV-41 infection could also activate EGCs, as shown in the significantly altered expression of glia fibrillary acidic protein (GFAP) in EGCs incubated with HAdV-41. The EGCs were also activated by serotonin alone, as shown in the significantly increased GFAP staining intensity. Likewise, EGCs were activated by the cell supernatant of HAdV-41-infected enterochromaffin cells.IMPORTANCE The nonenveloped human adenovirus 41 causes diarrhea, vomiting, dehydration, and low-grade fever mainly in children under 2 years of age. Even though acute gastroenteritis is well described, how human adenovirus 41 causes diarrhea is unknown. In our study, we analyzed the effect of human adenovirus 41 infection on human enterochromaffin cells and found it stimulates serotonin secretion in the cells, which is involved in regulation of intestinal secretion and gut motility and can also activate enteric glia cells, which are found in close proximity to enterochromaffin cells in vivo This disruption of gut barrier homeostasis as maintained by these cells following human adenovirus 41 infection might be a mechanism in enteric adenovirus pathogenesis in humans and could indicate a possible serotonin-dependent cross talk between human adenovirus 41, enterochromaffin cells, and enteric glia cells.

Effects of therapeutic hypothermia on white matter injury from murine neonatal hypoxia-ischemia.

BackgroundTherapeutic hypothermia (TH) is the standard of care for neonates with hypoxic-ischemic encephalopathy but is not fully protective in the clinical setting. Hypoxia-ischemia (HI) may cause white matter injury (WMI), leading to neurological and cognitive dysfunction.MethodsP9 mice were subjected to HI as previously described. Pups underwent 3.5 hours of systemic hypothermia or normothermia. Cresyl violet and Perl’s iron stain for histopathological scoring of brain sections was completed blindly on all brains. Immunocytochemical (ICC) staining for myelin basic protein (MBP), microglia (Iba1), and astrocytes (GFAP) was performed on adjacent sections. Volumetric measurements of MBP coverage were used for quantitative analysis of white matter.ResultsTH provided neuroprotection by injury scoring for the entire group (n=44) (p<0.0002). ICC analysis of a subset of brains showed that the lateral caudate was protected from WMI (p<0.05). Analysis revealed decreased GFAP and Iba1 staining in hippocampal regions, mostly CA2/CA3. GFAP and Iba1directly correlated with injury scores of normothermic brains.ConclusionsTH reduced injury and qualitative data suggest that hippocampus and lateral caudate are protected from HI. Mildly injured brains may better show the benefits of TH. Overall these data indicate regional differences in WMI susceptibility and inflammation in a P9 murine HI model.

Analgesic effect and related mechanism of peripheral acupoints electroacupuncture on superficial partial-thickness burn rats

Objective: To observe the analgesic effect and related mechanism of peripheral acupoints electroacupuncture on superficial partial-thickness burn rats. Methods: Eighty SD rats were divided into sham injury group (SI), pure burn group (PB), electroacupuncture group (E), and sham electroacupuncture group (SE) according to the random number table, with 20 rats in each group. Right posterior leg of rats in group SI were sham injured, while superficial partial-thickness scald (hereinafter referred to as burn) model was reproduced on the right posterior leg of rats in the latter three groups. Electroacupuncture of peripheral acupoints of right posterior leg of rats (equivalent to Zusanli point and Sanyinjiao point of human) in group E were performed from post injury hour (PIH) 12 on, while rats in group SE were treated with sham electroacupuncture, with 30 min each time, one time a day for 3 days. Before injury and at PIH 12, 24, 36, 48, 60, and 72, the threshold of mechanical pain of 5 rats in each group was tested, and the threshold of heat pain of another 5 rats in each group was tested. At PIH 48, brain tissue of 5 rats in each group was obtained to observe the morphology and distribution of astrocytes with positive expression of glia fibrillary acidic protein (GFAP) in periaqueductal gray (PAG) area by immunohistochemical staining, and the number of astrocytes was calculated. At the same time, brain tissue of the rest 5 rats in each group was obtained to determine the expression of GFAP of astrocytes in PAG area with Western blotting. Data were possessed with analysis of variance of repeated measurement, one-way analysis of variance, and SNK test. Results: (1) Compared with that in group SI, the threshold of mechanical pain of rats in groups PB and SE had no significant change before injury and at PIH 12 (with P values above 0.05), but was significantly decreased from PIH 24 to 72 (with P values below 0.05); while the threshold of mechanical pain of rats in group E was significantly decreased from PIH 36 to 72 (with P values below 0.05). The threshold of mechanical pain of rats in group E was significantly higher than that in groups PB and SE at PIH 24 (with P values below 0.05). (2) Compared with that in group SI, the threshold of heat pain of rats in groups PB and SE had no significant change before injury (with P values above 0.05), but was significantly decreased from PIH 12 to 72 (with P values below 0.05); while the threshold of heat pain of rats in group E was significantly decreased from PIH 12 to 60 (with P values below 0.05). The threshold of heat pain of rats in group E was significantly higher than that in groups PB and SE from PIH 24 to 48 (with P values below 0.05). (3) The distribution of astrocytes with positive expression of GFAP in PAG area of rats in group SI was diffuse. The cell volume was small with cell body unobvious, and the projections were sparse, fine and short. The distribution of astrocytes with positive expression of GFAP in PAG area of rats in group PB was relatively concentrated. The cell body was hypertrophy and swelling, and the projections were increased and extended. The morphology and distribution of astrocytes with positive expression of GFAP in PAG area of rats in groups SE and E was similar to that in group PB. The numbers of astrocytes with positive expression of GFAP in PAG area of rats in groups SI, PB, E, and SE were 44±4, 39±4, 27±4, and 36±5, respectively. The number of astrocytes with positive expression of GFAP in PAG area of rats in group PB was significantly less than that in group SI (P<0.05), but similar to that in group SE (P>0.05). The number of astrocytes with positive expression of GFAP in PAG area of rats in group E was significantly less than that in groups PB and SE (with P values below 0.05). (4) The expressions of GFAP of astrocytes in PAG area of rats in groups SI, PB, E, and SE were 1.11±0.16, 0.66±0.15, 0.34±0.06, and 0.56±0.09, respectively. The expression of GFAP of astrocytes in PAG area of rats in group PB was significantly lower than that in group SI (P<0.05), but similar to that in group SE (P>0.05). The expression of GFAP of astrocytes in PAG area of rats in group E was significantly lower than that in groups PB and SE (with P values below 0.05). Conclusions: Electroacupuncture of peripheral acupoints can release the pain followed superficial partial-thickness burn in rats at early stage, and the possible mechanism is that it reduces the activation of astrocytes in PAG area.

Influence of autophagy on epithelial-mesenchymal transition and invasion of glioma cells

Objective To investigate the influence of autophagy on epithelial-mesenchymal transition (EMT) and invasion of glioma cells. Methods Autophagy of glioma cell line, SHG-44, was induced by starvation in Hank’s balanced salt solution (HBSS). The change in markers expression and invasion of SHG-44 cells was observed. The small interfering RNA (siRNA) of Atg3 was constructed and transfected into SHG-44 cells to inhibit autophagy under starvation. The change in the markers expression and invasion after transfection was compared to investigate the role of autophagy in EMT and invasion of SHG-44 cells. Results HBSS significantly induced autophagy of SHG-44 cells. The expression of epithelial marker, glia fibrillary acidic protein (GFAP) was downregulated whereas the expression of mesenchymal markers, Vimentin and matrix metalloproteinase (MMP)-2 was notably upregulated. The invasion number of cells was also significantly increased (18 100±1 500 vs. 68 750±3 350, t=864.345, P=0.001). After transfection of siRNA-Atg3, the starvation-induced autophagy of SHG-44 cells was obviously inhibited. In contrast to the control cells, there was no notable change in epithelial and mesenchymal markers expression, and the invasion number of SHG-44 cells was also significantly reduced (68 750±3 350 vs. 25 450±1 450, t=565.441, P=0.001). Conclusion Autophagy significantly induces EMT of glioma cells and promote their invasion. Key words: Glioma; Autophagy; Epithelial-mesenchymal transition; Invasion

3D Poly(Lactic-co-glycolic acid) Scaffolds for Treating Spinal Cord Injury.

In this paper, poly(lactic-co-glycolic acid) (PLGA) was used to fabricate spinal cord scaffolds using low temperature deposition manufacturing (LDM) technology. The PLGA scaffolds were characterized as having good porosity, hydrophilicity and considerable biodegradability. The effects of the PLGA scaffolds on cell proliferation and cytotoxicity were evaluated by culturing Schwann cells (SCs) on the surfaces of the scaffolds. The results showed that the SCs spread and proliferated well on the PLGA scaffolds. Histological assessment including Glia fibrillary acidic protein (GFAP) staining, Nissl staining, Luxol fast blue (LFB) staining and Bielschowsky silver staining showed that the spinal cord recoveries considerably improved with the PLGA scaffolds, indicating that the PLGA scaffolds exhibited potential for applications in the management of spinal cord injuries.

Roles of glutathione in antioxidant defense, inflammation, and neuron differentiation in the thalamus of HIV-1 transgenic rats.

Inflammation and oxidative stress in the brain are major causes of HIV-associated neurocognitive disorders. Previously we have reported high content of glutathione (GSH) in the thalamus of rats with F344 genetic background. In this study, we investigated the changes of GSH metabolism and GSH-dependent antioxidant enzymes in the rat thalamus in response to HIV-1 transgenesis, and their associations with oxidative stress, inflammation, and neuronal development. Male HIV-1 transgenic (HIV-1Tg) rats and wild type F344 rats at 10 months were used in this study, with 5 rats in each group. Parameters measured in this study included: total and oxidized GSH, glutathione peroxidase (GPx), glutathione-S-transferase (GST), gamma-glutamylcysteine synthetase (GCS), gamma-glutamyl transferase (GGT), cysteine/cystine transporters, 4-hydroxynonenal (HNE), interleukin 12 (IL12), neuronal nuclei (NeuN), microtubule-associated protein (MAP2), and glia fibrillary acidic protein (GFAP). The levels of total GSH, oxidized GSH (GSSG) and MAP2 protein, and enzymatic activities of GCS, GPx and GST were significantly higher in HIV-1Tg rats compared with F344 rats, but the ratio of GSSG/GSH, activity of GGT and levels of HNE, NeuN protein and GFAP protein did not change. HIV-1Tg rats showed a lower level of IL12 protein. GSH positively correlated with GCS, GST and MAP2, GSSG/GSH ratio positively correlated with HNE and IL12, the activities of GPx, GST and GCS positively correlated with each other, and negatively correlated with HNE. These findings suggest an important role of the GSH-centered system in reducing oxidative stress and neuroinflammation, and enhancing neuron differentiation in the thalamus of HIV-1Tg rats.

Complete Freund’s adjuvant–induced acute inflammatory pain could be attenuated by triptolide via inhibiting spinal glia activation in rats

BackgroundInflammatory pain is one of the most common clinical symptoms, mechanical allodynia and thermal hypersensitivities are associated with proinflammatory cytokines, and proinflammatory cytokine antagonists could alleviate the hypersensitivity. Previous studies showed that a traditional Chinese medicine ingredient, triptolide could inhibit inflammatory cytokines; however, it was still unknown whether triptolide had beneficial effects on treating inflammatory pain. Materials and methodsThe effects of triptolide on Complete Freund's Adjuvant–induced acute inflammatory pain were investigated using behavioral tests. The activation of spinal glia was morphologically observed by immunofluorescent histochemistry. The levels of OX42, glia fibrillary acidic protein, and phosphorylated extracellular signal–regulated kinase in the spinal cord were detected by Western blot, and the messenger RNA levels of interleukin 1β, interleukin 6, and tumor necrosis factor alpha were detected by real-time polymerase chain reaction. ResultsThese results demonstrate that the triptolide effectively attenuates inflammatory pain induced by Complete Freund's Adjuvant, the underlying mechanism may regulate the phosphorylated extracellular signal–regulated kinase signaling pathway and inhibit the spinal glia activation, and then downregulate the proinflammatory cytokines; the triptolide may be clinically useful as a drug of anti-inflammatory pain. ConclusionsIn the present study, we first reported that repeated systemic administration of triptolide could safely prevent and reverse inflammatory pain. The triptolide may serve as a new potential compound for developing safe therapeutics for patients suffering inflammatory pain.