Role of microglia-mediated neuronal injury in neurodegenerative diseases / 中国组织工程研究

BACKGROUND: Microglia are resident immune cells of the central nervous system that normally perform sensing, housekeeping, and defense functions. In the context of neurodegenerative diseases, the dysfunction of microglia leads to or aggravates neuronal injury. OBJECTIVE: To investigate the mechanism of microglia-mediated neuronal injury in neurodegenerative diseases. METHODS: The first author searched for relevant articles published from January 2001 to January 2020 in PubMed, CNKI, Wanfang database, and VIP database with the key words of “microglia; neurodegenerative diseases; neuronal injury”. RESULTS AND CONCLUSION: In neurodegenerative diseases, microglia perform excessive sensing due to toxic substances during normal function, leading to increasing activation of microglia, accompanied with hyperfunction of housekeeping and intense neuroinflammation causing neuronal impairment. The dysregulation can also be manifested as dysfunction of sensing and housekeeping due to specific gene mutations, which bring about accumulation of toxic substances, aggravating the dysregulation of defense function and inducing apoptosis or necrosis of neurons as a result. Further exploration on the mechanism of microglia-mediated neuronal injury in neurodegenerative diseases may provide several targets for the treatment of neurodegenerative disease.

LncRNA MALAT1 aggravates oxygen-glucose deprivation/reoxygenation-induced neuronal endoplasmic reticulum stress and apoptosis via the miR-195a-5p/HMGA1 axis

BACKGROUND: This study aimed to investigate the potential role and molecular mechanism of lncRNA metastasis associated lung adenocarcinoma transcript 1 (MALAT1) in cerebral ischemia/reperfusion injury. RESULTS: Using an oxygen-glucose deprivation/reoxygenation (OGD/R) cell model, we determined that the expression of MALAT1 was significantly increased during OGD/R. MALAT1 knockdown reversed OGD/R-induced apoptosis and ER stress. Mechanistically, MALAT1 promoted OGD/R-induced neuronal injury through sponging miR-195a-5p to upregulating high mobility group AT-hook1 (HMGA1). CONCLUSIONS: Collectively, these data demonstrate the mechanism underlying the invovlvement of MALAT1 in cerebral ischemia/reperfusion injury, thus providing translational evidence that MALAT1 may serve as a novel biomarker and therapeutic target for ischemic stroke.

Neuroprotective effects and mechanism of ethanol extract of Cistanche tubulosa against oxygen-glucose deprivation/reperfusion / 中国中药杂志

To investigate the inhibitory effects and mechanism of Cistanche tubulosa ethanol extract( CTEE) against oxygen-glucose deprivation/reperfusion( OGD/R)-induced PC12 cells neuronal injury. In this study,OGD/R-induced PC12 cells were used to explore the neuroprotective effects of CTEE( 12. 5,25,50 mg·L-1) by detecting cell viability with MTT assay,apoptosis with AO/EB and Hoechst 33258,mitochondrial membrane potential changes with JC-1 staining,mitochondrial oxidative stress with MitoSOX staining,as well as the apoptosis-related protein expression( PARP,cleaved PARP,caspase-3,cleaved caspase-3,Bax,Bcl-2) with Western blot. RESULTS:: showed that CTEE effectively protected OGD/R-induced neuronal injury and increased the survival rate of PC12 cells.AO/EB and Hoechst 33258 staining showed that CTEE could effectively inhibit apoptosis. Moreover,JC-1 and MitoSOX staining results showed that CTEE decreased mitochondrial stress and mitochondrial membrane potential imbalance in PC12 cells in a concentration-dependent manner. Meanwhile,CTEE could obviously suppress the activation of key proteins in mitochondrial apoptosis pathway such as caspase-3 and PARP,and significantly inhibit the rise of Bax and down-regulation of Bcl-2. In conclusion,CTEE has obvious protective effects on OGD/R-induced PC12 cells neuronal injury,potentially via inhibiting mitochondrial oxidative stress and apoptosis-related signaling pathway.

The Mechanistic Basis for Photobiomodulation Therapy of Neuropathic Pain by Near Infrared Laser Light

Background and Objective Various irradiances have been reported to be beneficial for the treatment of neuropathic pain with near infrared light. However, the mechanistic basis for the beneficial outcomes may vary based on the level of irradiance or fluence rate used. Using in vivo and in vitro experimentalmodels, this study determined the mechanistic basis of photobiomodulation therapy (PBMT) for the treatment of neuropathic pain using a high irradiance. Study Design/Materials and Methods ln vitro experiments: Cultured, rat DRG were randomly assigned to control or laser treatment (L T) groups with different irradiation times (2, 5, 30, 60 or 120s). The laser parameters were: output power » 960 mW, irradiance » 300mW/cm2, 808 nm wavelength and spot size » 3cm diameter/ area » 7.07cm2, with different fluences according to irradiation times. Mitochondrial metabolic activity was measured with the MTS assay. The DRG neurons were immunostained using a primary antibody to ß-Tubulin III. ln vivo experiments: spared nerve injury surgery (SNI), an animal model of persistent peripheral neuropathic pain, was used. The injured rats were randomly divided into three groups (n » 5). 1) Control: SNI without LT, 2) Short term: SNI with LT on day 7 and euthanized on day 7, 3) Long term: SNI with LT on day 7 and euthanized on day 22. An 808 nm wavelength laser was used for all treatment groups. Treatment was performed once on Day 7 post-surgery. The transcutaneous treatment parameters were: output power: 10 W, fluence rate: 270 mW/cm2, treatment time: 120s. The laser probe was moved along the course of the sciatic/sural nerve during the treatment. Within 1 hour of irradiation, behavior tests were performed to assess its immediate effect on sensory allodynia and hyperalgesia caused by SNI. Results ln vitro experiments: Mitochondrial metabolism was significantly lower compared with controls for all LT groups. Varicosities and undulations formed in neurites of DRG neurons with a cell body diameter 30µm or less. ln neurites of DRG neurons with a cell body diameter of greater than 30µm, varicosities formed only in the 120s group. ln vivo experiments: For heat hyperalgesia, there was a statistically significant reduction in sensitivity to the heat stimulus compared with the measurements done on day 7 prior to LT. A decrease in the sensitivity to the heat stimulus was found in the LT groups compared with the control group on day 15 and 21. For cold allodynia and mechanical hyperalgesia, a significant decrease in sensitivity to cold and pin prick was found within 1 hour after L T. Sensitivity to these stimuli returned to the control levels after 5 days post-L T. No significant difference was found in mechanical allodynia between control and L T groups for all time points examined. Conclusion These in vitro and in vivo studies indicate that treatment with an irradiance/fluence rate at 270 m W/cm2 or higher at the level of the nerve can rapidly block pain transmission. A combination therapy is proposed to treat neuropathic pain with initial high irradiance/fluence rates for fast pain relief, followed by low irradiance/ fluence rates for prolonged pain relief by altering chronic inflammation.

Neurogenic stress cardiomyopathy: What do we need to know

The interaction between the heart and brain is complex and integral to the maintenance of normal cardiovascular function. Even in the absence of coronary disease, acute neuronal injury can induce a variety of cardiac changes. Recent neuroimaging data revealed a network including the insular cortex, anterior cingulate gyrus, and amygdala playing a crucial role in the regulation of central autonomic nervous system. Damage in these areas has been associated with arrhythmia, myocardial injury, higher plasma levels of brain natriuretic peptide, catecholamines, and glucose. Some patients after brain injury may die due to occult cardiac damage and functional impairment in the acute phase. Heart failure adversely influences acute stroke mortality. Troponin and NT-proBNP are elevated in acute brain injury patients, in response to the activated renin–angiotensin–aldosterone system and other neurohumoral changes, as a protective mechanism for sympathoinhibitory activity. Such patients have been shown to be associated with higher short- and long-term mortality. While thrombolysis, neuroprotection, and other measures, alone or in combination, may limit the cerebral damage, attention should also be directed toward the myocardial protection. Early administration of cardioprotective medication aimed at reducing increased sympathetic tone may have a role in myocardial protection in stroke patients. For a full understanding of the brain–heart control, the consequences of disruption of this control, the true incidence of cardiac effects of stroke, and the evidence-based treatment options further research are needed.

Protective effect of Angelica polysaccharide on retinal nerve cells in glaucoma rats and its mechanism / 吉林大学学报(医学版)

Objective: To investigate the protective effect of Angelica polysaccharide on the glaucoma-induced retinal neuronal injury, and to elucidate its mechanism. Methods: Eighty-four SD rats were randomly divided into control group, model group, positive drug group, low, middle and high doses of Angelica polysaccharide groups. The model of rat glaucoma was established by sclera sclerotherapy. The intraocular pressures (IOP) of rats in each group were measured. The superoxide dismutase (SOD) activities, malondialdehyde (MDA) contents and nitric oxide (NO) levels in retina tissue of the rats were measured by colorimetric assay. The expression levels of Caspase-3 mRNA and protein in retina tissue of the rats were observed by RT-PCR and Western blotting method. Results: Compared with control group, the IOP of the rats in model group at each time point were significantly increased (P<0. 05); compared with model group, the IOP of the rats in different doses of Angelia polysaccharide groups were significantly decreased (P<0. 05); compared with low dose of Angelica polysaccharide group, the IOP of the rats in high dose of Angelica polysaccharide group was significant decteased (P<0. 05). The levels of MDA and NO in the retina tissue of the rats in model group were higher than those in control group (P<0. 05), but the SOD activities were decreased (P<0. 05); compared with model group, the levels of MDA and NO in retinal tissue of the rats in different doses of Angelica polysaccharide group were significantly decreased (P<0. 05), and the SOD activities were significantly increased (P<0. 05); compared with low dose of Angelica polysaccharide group, the above indexed had significant differences in high dose of Angelica polysaccharide group (P<0. 05). Compared with model group, the expression levels of Caspase-3 mRNA and protein in different doses of Angelica polysaccharide groups were significantly decreased (P<0. 05); compared with low dose of Angelica polysaccharide group, the expression levels of Caspase-3 mRNA and protein were significantly decreased (P < 0. 05). Conclusion: Angelica polysaccharide has protective effect on the retinal tissue of glaucoma model rats, which can decrease the levels of MDA and NO in retina tissue and increase the activity of SOD and decrease the expression levels of Caspase-3 mRNA and protein in retina tissue. It may be one of the mechanisms of polysaccharide protecting in retinal tissue nerve cells.

Regulation of miR-133b on Methamphetamine-induced Neuronal Apoptosis in PC12 Cells / 中山大学学报(医学科学版)

[Objective]To investigate the expression changes of miR-133b in methamphetamine(MA)-induced neuro-nal injury in PC12 cells and its regulative effects on cellular apoptosis.[Methods]PC12 cells were cultured and divided into control group and MA treated group.In MA treated group,PC12 cells were insulted with 800μmol/L MA in culture medium. The cellular injury of PC12 cells was observed under microscope. The cellular apoptosis was detected by Hoechst33342/PI double staining,and the expression level of miR-133b was examined by real-time quantitative PCR(RT-PCR). Further-more,miR-133b mimic and inhibitors were transfected into PC12 cells to analyze miR-133b's function in MA-induced cell apoptosis.[Results]The data showed that 800 μmol/L MA could induce obvious cellular injury,cause neurite shortened and increase the cell apoptosis. The RT-PCR data showed that the expression of miR-133b of PC12 cells treated with MA de-creased significantly.The apoptosis rate of PC12 cells decreased after transfection of miR-133b mimic,while increased after transfection of miR-133b inhibitors.[Conclusions]High concentration of MA causes neuron damage and induces neuronal apoptosis,and also decreases the levels of miR-133b expression. Whereas,overexpression of miR-133b can reduce the apoptosis of cultured PC12 cells.Thus,miR-133b plays a crucial role in MA mediated neurotoxicity.This study provides a theoretical basis for elucidating the mechanism of MA-induced neurotoxicity and may provide a new strategy for treating MA addiction.

Protective effect of Angelica polysaccharide on retinal nerve cells in glaucoma rats and its mechanism / 吉林大学学报(医学版)

Objective:To investigate the protective effect of Angelica polysaccharide on the glaucoma-induced retinal neuronal injury,and to elucidate its mechanism.Methods:Eighty-four SD rats were randomly divided into control group,model group,positive drug group,low,middle and high doses of Angelica polysaccharide groups.The model of rat glaucoma was established by sclera sclerotherapy.The intraocular pressures (IOP) of rats in each group were measured.The superoxide dismutase (SOD) activities,malondialdehyde (MDA) contents and nitric oxide (NO) levels in retina tissue of the rats were measured by colorimetric assay.The expression levels of Caspase-3 mRNA and protein in retina tissue of the rats were observed by RT-PCR and Western blotting method.Results:Compared with control group,the IOP of the rats in model group at each time point were significantly increased (P＜0.05);compared with model group,the IOP of the rats in different doses of Angelia polysaccharide groups were significantly decreased (P＜0.05);compared with low dose of Angelica polysaccharide group,theIOP of the rats in high dose of Angelica polysaccharide group was significant decteased (P＜0.05).The levels of MDA and NO in the retina tissue of the rats in model group were higher than those in control group (P＜0.05),but the SOD activities were decreased (P＜0.05);compared with model group,the levels of MDA and NO in retinal tissue of the rats in different doses of Angelica polysaccharide group were significantly decreased (P＜0.05),and the SOD activities were significantly increased (P＜0.05);compared with low dose of Angelica polysaccharide group,the above indexed had significant differences in high dose of Angelica polysaccharide group (P＜0.05).Compared with model group,the expression levels of Caspase-3 mRNA and protein in different doses of Angelica polysaccharide groups were significantly decreased (P＜0.05);compared with low dose of Angelica polysaccharide group,the expression levels of Caspase-3 mRNA and protein were significantly decreased (P ＜ 0.05).Conclusion:Angelica polysaccharide has protective effect on the retinal tissue of glaucoma model rats,which can decrease the levels of MDA and NO in retina tissue and increase the activity of SOD and decrease the expression levels of Caspase-3 mRNA and protein in retina tissue.It may be one of the mechanisms of polysaccharide protecting in retinal tissue nerve cells.

Study on glycyrrhizin in reducing neuronal damage by inhibiting high mobility group protein 1 in immature rats with epilepsy / 中华实用儿科临床杂志

Objective To study the effect of glycyrrhizin(GL) on the gene expression of high mobility group protein 1 (HMGB1) in hippocampus and serum.To evaluate the effect on the expression of neuron-specific nuclear-binding protein (Neu-N) in the hippocampus CA1,CA3 regions in the chronic stage of an immature rat epilepsy model.Methods Fifty-two 21 day-old SD rats were randomly divided into control group,model group Ⅰ and model group Ⅱ according to the random table method.Model group Ⅰ was induced epilepsy by kainic acid (KA),and the model group Ⅱ was pretreated with GL by intraperitoneal injection at 30 min before KA injection.According to the different observation time points,each group was divided into 4 subgroups:3 h,12 h,24 h and 7 d.Model group Ⅱ was divided into 3 subgroups:10 mg/kg,50 mg/kg,100 mg/kg,according to the different doses of GL.There were 3 animals in each subgroup.Score was performed according to the Racine score,and quantitative real-time polymerase chain reaction and Western blot were applied to detect the mRNA and protein expression of HMGB1 in the acute phase.Enzyme-linked immunosorbent assay(ELISA) was applied to measure the expression of HMGB1 in blood;immunohistochemical was applied to measure the expression of Neu-N in hippocampus in the chronic phase(7 d).Results Compared with model group Ⅰ,seizure onset time was obviously prolonged in model group Ⅱ [(24.08 ± 1.98) min vs.(33.39 ± 2.66) min],and the difference was statistically significant (t =9.231,P ＜0.05);Comparing KA model group Ⅰ with control group,the gene expression of HMGB1 significantly increased,and reached a peak at the time of 12 h (H =10.532,P ＜ 0.05),but the protein expression of HMGB1 was changed obviously and there was no significant difference (H =5.227,P ＞0.05).The expression of HMGB1 in the serum also significantly increased,especially at 12 h (H =6.897,P ＜0.05).At the time of 12 h after KA injection,the gene expression of HMGB1 in the hippocampus was significantly decreased in model group Ⅱ compared with model group Ⅰ (H =10.721,P ＜0.05) (especially in the 100 mg/kg model group).Also,the expression of HMGB1 in the scrum was obviously decreased (H =6.967,P ＜ 0.05) (especially in the 100 mg/kg model group).At the time of 7 d after KA injection,hippocampal neuron loss in model group.Ⅰ was significantly reduced compared with control group (P ＜ 0.05),and hippocampal neuron loss in model group Ⅱ was evidently decreased compared with model group Ⅰ (P ＜ 0.05),(especially in the 100 mg/kg model group in CA1,50 mg/kg model group in CA3).Conclusions In the immature rat temporal lobe epilepsy model,GL may have neuroprotective by inhibiting the synthesis and release of HMGB1,inhibiting inflammation further to restrain the loss of neurons in the chronic phase.

Quality evaluation of Gualou Guizhi decoction based on chemical compositions and biological effects / 中国中药杂志

The paper was aimed to establish a quality evaluation model for Gualou Guizhi decoction based on the chemical compositions and biological effects. Ultra high performance liquid chromatograph-mass spectrometer was used to analyze and determine 24 kinds of chemical compositions in Gualou Guizhi decoction, and then, biological activity effect was quantitatively assessed in a zebrafish neuronal injury model which was induced by mycophenolate mofetil (MMF). As a result, the established method for quality evaluation of Gualou Guizhi decoction based on the chemical compositions and biological effects is feasible, stable and reliable, which can provide reference for quality control of compound Chinese medicines.

Recent Updates in Neuroprotective and Neuroregenerative Potential of Centella asiatica

Centella asiatica, locally well known in Malaysia as pegaga, is a traditional herb that has been used widely in Ayurvedic medicine, traditional Chinese medicine, and in the traditional medicine of other Southeast Asian countries including Malaysia. Although consumption of the plant is indicated for various illnesses, its potential neuroprotective properties have been well studied and documented. In addition to past studies, recent studies also discovered and/or reconfirmed that C. asiatica acts as an antioxidant, reducing the effect of oxidative stress in vitro and in vivo. At the in vitro level, C. asiatica promotes dendrite arborisation and elongation, and also protects the neurons from apoptosis. In vivo studies have shown that the whole extract and also individual compounds of C. asiatica have a protective effect against various neurological diseases. Most of the in vivo studies on neuroprotective effects have focused on Alzheimer’s disease, Parkinson’s disease, learning and memory enhancement, neurotoxicity and other mental illnesses such as depression and anxiety, and epilepsy. Recent studies have embarked on finding the molecular mechanism of neuroprotection by C. asiatica extract. However, the capability of C. asiatica in enhancing neuroregeneration has not been studied much and is limited to the regeneration of crushed sciatic nerves and protection from neuronal injury in hypoxia conditions. More studies are still needed to identify the compounds and the mechanism of action of C. asiatica that are particularly involved in neuroprotection and neuroregeneration. Furthermore, the extraction method, biochemical profile and dosage information of the C. asiatica extract need to be standardised to enhance the economic value of this traditional herb and to accelerate the entry of C. asiatica extracts into modern medicine.

Effects of endogenous histamine on transient forebrain ischemia-induced neuronal damage at late phase of reperfusion in mice / 中国病理生理杂志

AIM: To determine the effect of endogenous histamine on transient forebrain ischemia-induced neuronal injury at the late phase of reperfusion using histidine decarboxylase knockout ( HDC-KO) mice.METHODS:Wild-type (WT) and HDC-KO mice were subjected to bilateral common carotid artery occlusion for 30 min followed by 3 d or 15 d of reperfusion.At different time points after reperfusion, the body weight, mortality rate, learning and memory in fear conditioning test and hippocampal CA 1 neuronal density were evaluated .RESULTS: At 1 d after reperfusion , the body weight loss was observed in both WT and HDC-KO mice.At 4 d, 5 d, 6 d and 7 d after reperfusion, the increment in the body weight of the HDC-KO mice was significantly smaller than that of the WT mice .During the period between 8 d and 14 d after reperfusion, the mortality rate of the HDC-KO mice was higher than that of the WT mice (P<0.05).At 14 d after reperfusion , the HDC-KO mice exhibited more aggravated deficits in contextual and cue memory compared with the WT mice.Correspondingly , a more severe CA1 neuronal injury in the HDC-KO mice than that in the WT mice was ob-served at 15 d but not at 3 d after reperfusion (P<0.05).CONCLUSION:Endogenous histamine may attenuate learn-ing/memory deficits and neuronal injury at the late phase of ischemia /reperfusion.However, the involved mechanisms need to be further investigated .

Ulinastatin attenuates oxidation, inflammation and neural apoptosis in the cerebral cortex of adult rats with ventricular fibrillation after cardiopulmonary resuscitation

OBJECTIVE: The role of Ulinastatin in neuronal injury after cardiopulmonary resuscitation has not been elucidated. We aim to evaluate the effects of Ulinastatin on inflammation, oxidation, and neuronal injury in the cerebral cortex after cardiopulmonary resuscitation. METHODS: Ventricular fibrillation was induced in 76 adult male Wistar rats for 6 min, after which cardiopulmonary resuscitation was initiated. After spontaneous circulation returned, the rats were split into two groups: the Ulinastatin 100,000 unit/kg group or the PBS-treated control group. Blood and cerebral cortex samples were obtained and compared at 2, 4, and 8 h after return of spontaneous circulation. The protein levels of tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6) were assayed using an enzyme-linked immunosorbent assay, and mRNA levels were quantified via real-time polymerase chain reaction. Myeloperoxidase and Malondialdehyde were measured by spectrophotometry. The translocation of nuclear factor-κB p65 was assayed by Western blot. The viable and apoptotic neurons were detected by Nissl and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL). RESULTS: Ulinastatin treatment decreased plasma levels of TNF-α and IL-6, expression of mRNA, and Myeloperoxidase and Malondialdehyde in the cerebral cortex. In addition, Ulinastatin attenuated the translocation of nuclear factor-κB p65 at 2, 4, and 8 hours after the return of spontaneous circulation. Ulinastatin increased the number of living neurons and decreased TUNEL-positive neuron numbers in the cortex at 72 h after the return of spontaneous circulation. CONCLUSIONS: Ulinastatin preserved neuronal survival and inhibited neuron apoptosis after the return of spontaneous circulation in Wistar rats via attenuation of the oxidative stress response and translocation of nuclear factor-κB p65 in the cortex. In addition, Ulinastatin decreased the production of TNF-α, ...

Inhibitory Effect of Melatonin on Kainic Acid-Induced Hippocampal Neuronal Injury / 대한소아신경학회지

PURPOSE: It has been suggested that the pineal hormone melatonin(MEL) protects neurons in vitro from excitotoxicity mediated by kainate-sensitive glutamate receptors and from oxidative stress-induced DNA damage and apoptosis. The present study evaluated the antioxidatives and anti-inflammatory effect of melatonin on kainic acid(KA)-induced neuronal injury in the hippocampus in vivo. METHODS: 30 adult male Sprague-Dawley rats were divided into two equal groups. Control group was treated with KA only and test group was treated with KA and MEL. We injected 10 mg/kg KA intraperitoneal into rats. This results in selective neuronal injury accompanied by intense microglial activation and triggers DNA damage in the hippocampus. We tested the in vivo efficacy of MEL in preventing KA-induced neuronal injury and neuroinflammation in the hippocampus. MEL(2.5 mg/kg) was injected i.p. four times : 20 min before KA, immidiately after, and 1 and 2 h after the KA. Rats were sacrificed 72 h later and their hippocampi were examined for evidence of DNA damage (in situ dUTP-end-labeling, i.e. TUNEL staining), cell viability(H&E staining), microglial (isolectin-B4 histochemistry), astroglial responses(glial fibrillary acidic protein, GFAP immunohistochemistry), and lipid peroxidation(4-hydroxynonenal immunohistochemistry). RESULTS: The cumulative dose of 10 mg/kg MEL attenuates KA-induced neuronal death as well as microglial activation and lessens DNA breaks. CONCLUSION: A possible mechanism of MEL-provided neuroprotection lies in its antioxidant and anti-inflammatory action. Present data suggest that MEL holds potential for the treatment of acute brain pathologies such as epilepsy-associated brain damage, stroke, and brain trauma.

Ischemia-responsive Protein(irp94) Gene Expression in a Neuronal Cell Culture Model of Ischemia

BACKGROUND: The ischemia responsive protein 94 kDa(irp94) gene belongs to the heat shock protein 110 family and was isolated in 1999 from rat brain by transiently induced forebrain ischemia. The PC12 cell is the pheochromocytoma cell line of rat, which is differentiated to a sympathetic neuron-like cell by the stimulation of a nerve growth factor. This study is to determine whether irp94 is expressed when an ischemia-like condition is induced by ATP depletion in cultured PC12 cells in vitro. METHODS: PC12 cells were maintained as monolayer cultures in RPMI-1640 medium(Sigma) supplemented with 10% horse serum, 5% fetal bovine serum, 5 mg/ml transferrin, and 1 mg/ml insulin in a humidified 5% CO2 incubator at 37degrees C. The ATP depleting agent antimycin A was added at concentrations of 1, 2.5, and 5 microM to simulate ischemia, and 10 microgram/ml of tunicamycin, which is expected to express heat shock protein maximally, was used as a positive control. The cells were harvested after a 60-minute incubation, and the total RNA was extracted. The reverse transcription polymerase chain reaction(RT-PCR) was performed to use 501 bp irp94 cDNA as a molecular probe, and the expression of irp94 mRNA was analyzed by northern blotting. RESULTS: The irp94 mRNA expression was enhanced, compared to the negative control group, as the concentration of antimycin A was increased. CONCLUSION: This study suggests that irp94 mRNA expression is enhanced as the severity of ischemia is increased. Thus, it is possible to investigate the mechanism of ischemic neuronal injury indirectly by using this in-vitro model of neuronal ischemia.

RELATIONSHIP BETWEEN NEURONAL INJURIES AND REACTION OF ASTROCYTES FOLLOWING CEREBRAL ISCHEMIA REPERFUSION IN RAT / 神经解剖学杂志

The involvement of astrocytes and correlation between neuronal injury and astrocyte response were studied. Blockingmiddle cerebral artery and reperfusing o. 5～48 h, H-E staining, immunoccytochemistry single-and double-labeling, dotble label-ing combined with TUNEL and GFAP immunocytochemistry were used to investigate neuronal injury and astrocyte response.The is chemic area peaked at 24 h of reperfusion. The neurons presented irreversible degeneration at 6 h of reperfusion. At24 h,ischemic area in the preoptic area developed into infarcted area; astrocytes exhibited differential morphological features: reactive,malnourished and degenerative changes. At 48 h of reperfusion, the number of astrocytes began to go up. The astrocytes in is-chemic area didn't proliferate within 48 h. By contrast, a few astrocytes underwent apoptosis. In conclusion, these data indicatethat the reaction of astrocytes is closely connected with the extent of neuronal injuries. The reactive astrocytes imply that astro-cytes positively respond to the neuronal injuries, which might play a role in promoting neuronal survival.

The Effects of Continuous and Intermittent Interruption of the Cerebral Blood Flow on Brain Edema and Ischemic Neuronal Injury in Gerbils

Temporary occlusion of the cerebral blood flow is an effective maneuver to prevent and/or to control excessive bleeding during neurosurgical operations. Many studies have been reported employing single occlusion of various durations. However, there has been only a few studies examing the consequences of repeated occlusions on the development of cerebral edema and neuronal injury in the gerbil. Three separate episodes of 5-minute ischemia spaced at varied time interval was produced in Mongolian gerbils by occlusion of bilateral common carotid arteries. Quantitative estimates of cerebral edema and neuronal injury were obtained 24 hours after the third occlusion. The result was compared to that of single 15-minute occlusion. In gerbils with three 5-minute occlusions at 10-minute intervals, cerebral edema was not significant. However, the animals killed 24 hours after three 5-minute occlusions at 1-hour intervals or single 15-minute occlusion showed severe cerebral edema. Such animals showed significantly more neuronal injury than in animals with three 5-minute occlusions at 10-minute intervals. These results suggest that ischemic brain damage may be reduced with repeated vascular occlusions spaced at short intervals.