OBJECTIVETo study the change in ultrastructure of C6 glioma cells after photodynamic therapy (PDT), to compare morphological differences in necrosis and apoptosis before and after PDT treatment, and to evaluate the effect of photodynamic therapy on the blood brain tumor barrier (BTB) of C6 glioma.

METHODSThe model was produced by transplanting C6 glioma cells cultured in vitro using Peterson method into the caudate nuclei of Wister rats. The experiment group received PDT for two weeks after the operation. The sub-cellular structure, blood-brain-barrier (BBB) and BTB in both groups were observed under electron microscope.

RESULTSApoptosis in different phases and necrosis could be observed in some C6 glioma cells. Swelling occurred on the ultrastructure of cellular organs such as mitochondria and endoplasmic reticulum in most of the cells. Damage to the BTB, reduction of the number of cellular organs in endothelial cells of the capillary blood vessels, stretch of the tight junction, and enlargement of the gaps between endothelial cells were also seen in the experiment group. Meanwhile, limited impact on the normal sub-cellular structures and BBB was observed.

CONCLUSIONPDT could induce apoptosis and necrosis of C6 glioma cells due to the damage to the ultrastructure of mitochondria and endoplasmic reticulum. The weakened function of C6 glioma BTB initiated by PDT makes it possible to perform a combined therapy of PDT and chemotherapy for glioma.

Animals ; Blood-Brain Barrier ; Brain Neoplasms ; drug therapy ; ultrastructure ; Cell Line, Tumor ; Glioma ; drug therapy ; ultrastructure ; Photochemotherapy ; Rats

OBJECTIVETo study the changes of serum neuron specific enolase in rats with septic shock.

METHODSThe model of septic shock was set up by injection of lipopolysaccharide (LPS, from Escherichia coil O55: B5) at a dose of 25 mg/kg through femoral vein. Twenty male Wistar rats were randomly divided into 2 groups: normal control group (LPS was substituted by same volume of normal saline solution) and septic shock group. Six hours after the septic shock model formed, whole blood was taken for measuring the serum neuron specific enolase (NSE). The brains of the rats were taken for histopathological examination.

RESULTSThe serum NSE of septic shock group was significantly higher than that of control group [(10.0781 +/- 0.526) microg/L vs. (3.7188 +/- 0.602) microg/L, P < 0.05]. Neurons were severely damaged 6 hours after injection of LPS. Neuronal necrosis and the damage of blood-brain barrier were seen by light and electron microscope in septic shock group but not in the control group.

CONCLUSIONNSE in serum increased when septic encephalopathy occurred, which indicated that NSE might become a marker of neural damage in septic shock.

Animals ; Biomarkers ; blood ; Blood Pressure ; Blood-Brain Barrier ; ultrastructure ; Brain ; cytology ; pathology ; Cell Death ; Disease Models, Animal ; Male ; Microscopy, Electron ; Neurons ; pathology ; ultrastructure ; O Antigens ; toxicity ; Phosphopyruvate Hydratase ; blood ; Rats ; Rats, Inbred BB ; Shock, Septic ; blood ; enzymology ; pathology

OBJECTIVETo investigate the effects of aluminum on the integrity of blood brain barrier in juvenile rats.

METHODSThe 40-day old Sprague-Dawley (SD) rats were exposed to aluminium chloride by intraperitoneal injection, at a dose of Al3+ 0 mg/kg, 2.5 mg/kg, 5 mg/kg, 10 mg/kg, respectively. Morris water amaze system was used to test the learning and memory ability. The Evans blue content in brain was analyzed after injection. The ultrastructure's change of the blood brain barrier (BBB) was observed with transmission electron microscope. The expression of occluding protein in BBB was determined by Western blot method.

RESULTSAs compared with control group, the permeability of BBB in mid-level Al and high-level Al was enhanced (P <0.01), the expression of occluding protein was descended (P <0.01). The ultrastructures of the BBB were changed. No differences between every group on learning and memory ability (P>0.05).

CONCLUSIONSShort time and low dose of Al might not change the ability of learning and memory in juvenile rats, however the permeability and ultrastructures of the BBB might be significantly changed.

Aluminum ; toxicity ; Animals ; Blood-Brain Barrier ; drug effects ; ultrastructure ; Brain ; metabolism ; Capillary Permeability ; drug effects ; Maze Learning ; Rats ; Rats, Sprague-Dawley ; Toxicity Tests

OBJECTIVE: To explore the mechanism that mediates the effect of soybean isoflavones (SI) against cerebral ischemia/reperfusion (I/R) injury in light of the regulation of regional cerebral blood flow (rCBF), ferroptosis, inflammatory response and blood-brain barrier (BBB) permeability. METHODS: A total of 120 male SD rats were equally randomized into sham-operated group (Sham group), cerebral I/R injury group and SI pretreatment group (SI group). Focal cerebral I/R injury was induced in the latter two groups using a modified monofilament occlusion technique, and the intraoperative changes of real-time cerebral cortex blood flow were monitored using a laser Doppler flowmeter (LDF). The postoperative changes of cerebral pathological morphology and the ultrastructure of the neurons and the BBB were observed with optical and transmission electron microscopy. The neurological deficits of the rats was assessed, and the severities of cerebral infarction, brain edema and BBB disruption were quantified. The contents of Fe²⁺, GSH, MDA and MPO in the ischemic penumbra were determined with spectrophotometric tests. Serum levels of TNF-α and IL-1βwere analyzed using ELISA, and the expressions of GPX4, MMP-9 and occludin around the lesion were detected with Western blotting and immunohistochemistry. RESULTS: The rCBF was sharply reduced in the rats in I/R group and SI group after successful insertion of the monofilament. Compared with those in Sham group, the rats in I/R group showed significantly increased neurological deficit scores, cerebral infarction volume, brain water content and Evans blue permeability (P < 0.01), decreased Fe²⁺ level, increased MDA level, decreased GSH content and GPX4 expression (P < 0.01), increased MPO content and serum levels of TNF-α and IL-1β (P < 0.01), increased MMP-9 expression and lowered occludin expression (P < 0.01). All these changes were significantly ameliorated in rats pretreated with IS prior to I/R injury (P < 0.05 or 0.01). CONCLUSION SI preconditioning reduces cerebral I/R injury in rats possibly by improving rCBF, inhibiting ferroptosis and inflammatory response and protecting the BBB.
Rats ; Male ; Animals ; Rats, Sprague-Dawley ; Matrix Metalloproteinase 9/metabolism* ; Soybeans/metabolism* ; Occludin/metabolism* ; Tumor Necrosis Factor-alpha/metabolism* ; Ferroptosis ; Blood-Brain Barrier/ultrastructure* ; Brain Ischemia/metabolism* ; Cerebral Infarction ; Reperfusion Injury/metabolism* ; Isoflavones/therapeutic use* ; Infarction, Middle Cerebral Artery

OBJECTIVETo observe the counteraction of Gastrodia elata and E-gelatin on the effect of subchronic lead poisoning on the ability of learning and memory and the ultrastructure in hippocampus.

METHODSSubchronic lead acetate exposure was given to rats (0.2 g x kg(-1) x d(-1)). Single and combined administration of Gastrodia elata (4 g x kg(-1) x d(-1)) and E-gelatin (1 g x kg(-1) x d(-1)) were conducted at the same time. Pb concentration in blood, and the ability of learning and memory (Y-maze test) of rats were measured. Ultrastructure of CA3 pyramidal cells in hippocampus under transmission electron microscope was observed.

RESULTSBlood Pb concentrations in each group (Pb group: 690.6 micro g/L, Pb + Gastodiae eleta group: 688.8 micro g/L, Pb + E-gelatin group: 663.8 micro g/L, Pb + combined group: 667.2 micro g/L) were higher than that in the control (28.24 micro g/L, P < 0.01). But there was no significant difference (P > 0.05) among these groups. In Y-maze test, the number of electric stroke in Pb group is higher than that in control (P < 0.01). Gastrodia elata or E-gelatin used singly could significantly reduce the number of electric stroke of lead-exposed groups (P < 0.05 in the first month and P < 0.01 in the second and third month). And the effect of combined use of them was more efficient than single use (P < 0.01). Under electron microscope, no anomaly was seen in the pyramidal cells of CA3 area in hippocampus of control group. But there was significant anomaly such as neucleus separation in the cells of Pb exposed group. In Pb-Gastrodia elata group, there were some stress response phenomena such as the occurrence of huge mitochondria while in Pb-E-gelatin group, the anomaly was mild. The cells in the CA3 area in hippocampus of the pb-Gastrodia-E-gelatin group were almost normal, and also showed some stress response phenomena.

CONCLUSIONGastrodia elata and E-gelatin may protect the neurons in CA3 area of hippocampus against damage induced by lead to improve the ability of learning and memory, and they have synergistic effect.

Animals ; Blood-Brain Barrier ; drug effects ; Drugs, Chinese Herbal ; pharmacology ; Gelatin ; pharmacology ; Hippocampus ; drug effects ; physiology ; ultrastructure ; Lead ; blood ; toxicity ; Male ; Maze Learning ; drug effects ; Memory ; drug effects ; Microscopy, Electron ; Rats ; Rats, Wistar

OBJECTIVETo study the changes of blood-brain barrier-tight junction (BBB-TJ) proteins ZO-1, occludin and actin following hypoxia-ischemia (HI) in order to explore the possible mechanism of permeability increasing of blood-brain barrier (BBB) induced by HI.

METHODSBBB models were established by co-culture of cell ECV304 and astrocytes (AS) in vitro, then randomly assigned to control and HI groups. Transmission electron microscope was used to observe the changes of BBB-TJ. The distribution of actin was determined by direct-immunofluorescence microscope. Definite permeability of BBB models by 125I-BSA was detected by gamma events-per-unit-time meter. Expression of actin, ZO-1 and occludin was detected by Western blot.

RESULTSAfter 10-day culture, endothelial cells connected tightly, with plenty of TJ which was smooth, continuous and of high density, in the BBB models. After 5 hrs of HI, the TJ was opened with intercellular gaps formation. The direct immunofluorescence showed that the peripheral filament bands became blurred, the cell-cell junction loosened and fissure appeared in the HI group. The permeability of 125I-BSA in the HI group increased significantly compared with the control group (P<0.01). Expression of ZO-1 decreased markedly, while expression of actin and occludin was not different in the HI group compared with the control group.

CONCLUSIONSThe changes in occludin distribution and decreased expression of ZO-1 lead the reorganization of BBB-actin protein, which may be one of the mechanisms of permeability increasing of BBB following HI.

Actins ; analysis ; physiology ; Animals ; Blood-Brain Barrier ; Female ; Hypoxia ; metabolism ; Ischemia ; metabolism ; Male ; Membrane Proteins ; analysis ; physiology ; Occludin ; Permeability ; Phosphoproteins ; analysis ; physiology ; Rats ; Rats, Sprague-Dawley ; Tight Junctions ; ultrastructure ; Zonula Occludens-1 Protein

AIMA method of coculture of brain capillary endothelial cells (BCECs) and astrocytes of rats was used to evaluate nanoparticle's blood-brain barrier (BBB) transcytosis and toxicity at the endothelial tight junction.

METHODSA lipophilic fluorescent probe, 6-coumarin, was incorporated in poly (ethyleneglycol)-poly (lactide) nanoparticle using double emulsion/solvent evaporation method. BCECs and astrocytes were firstly isolated from brain of newborn rats and characterized by their morphology and immunocytochemistry staining, separately. Subsequently, a coculture model with BCECs on the top of micro-porous membrane of cell culture insert and astrocytes on the bottom side was established. The permeability of 14C-labeled sucrose and nanoparticle were determined, separately.

RESULTSThe mean weight-based diameter of 6-coumarin loaded nanoparticles was (102.4 +/- 6.8) nm, with zeta potential of (-16.81 +/- 1.05) mV. BCECs were positive for factor VIII staining and glial fibrillary acidic protein was expressed in astrocytes. The transendothelial electrical resistance reached up to (313 +/- 23) omega x cm2. The tight junction between BCECs in the coculture model could be visualized by both scanning electron microscopy and transmission electron microscopy. The unchanged paracellular transport of sucrose proved that nanoparticle with concentration lower than 200 microg x mL(-1) did not impact the integrity of BBB endothelial tight junctions. The permeability of 10 microg x mL(-1) 6-coumarin labeled nanoparticle was 0.29 x 10(-3) cm x min(-1).

CONCLUSIONThis in vitro experimental model of rat BBB was close to resemble the in vivo situation for examination of the permeability of nanoparticle and toxicity evaluation.

Animals ; Animals, Newborn ; Astrocytes ; metabolism ; ultrastructure ; Biological Transport ; Blood-Brain Barrier ; Brain ; blood supply ; cytology ; Capillaries ; cytology ; Cell Membrane Permeability ; Coculture Techniques ; Coumarins ; administration & dosage ; pharmacokinetics ; toxicity ; Endothelial Cells ; metabolism ; ultrastructure ; Factor VIII ; metabolism ; Glial Fibrillary Acidic Protein ; metabolism ; Nanoparticles ; Polyesters ; Polyethylene Glycols ; Rats ; Rats, Sprague-Dawley ; Sucrose ; pharmacokinetics

OBJECTIVETo explore the effects of paeoniflorin on blood brain barrier and pathological changes in brain ischemia.

METHODMice were divided into sham operation group, model group, positive control group-Deng zhanhua tablet group and three different dose (high, middle, low-dose) groups of paeoniflorin. The neurological symptoms of rats were observed. The SOD of ischemic brain tissue, MDA BBB and EAA contents were determined. The ultrastructure on the brain cells was inspected by transmission electron microscope.

RESULTPaeoniflorin had the protetive effects on 4VO model of total cerebral ischemia. Paeoniflorin could obviously increase SOD content, reduce MDA content in rat brain-tissue and alleviate oxidative stress damage by cerebral ischemia on rat brain. Paeoniflorin could improve pathological changes of cell nuclear, perikaryon, mitochondria and myelin sheath, which was the morphologic basis of the protective effect on ischemia. Paeoniflorin could alleviate the incrense of EAA content caused by and hypoxia and inhibit the excitatory neural toxicity by EAA.

CONCLUSIONPaeoniflorin has the protection effect on the brain edema after cerebral ischemia, the oxidative stress damage on brain tissue, the ultrastructure lesions of cells and the BBB. The protective mechanism may be related to inhibiting intracellular calcium overload, anti-free radical and reducing EAA content.

Animals ; Behavior, Animal ; drug effects ; Benzoates ; isolation & purification ; pharmacology ; Blood-Brain Barrier ; drug effects ; pathology ; ultrastructure ; Brain ; blood supply ; drug effects ; metabolism ; Brain Ischemia ; pathology ; physiopathology ; prevention & control ; Bridged-Ring Compounds ; isolation & purification ; pharmacology ; Glucosides ; isolation & purification ; pharmacology ; Male ; Malondialdehyde ; metabolism ; Microscopy, Electron ; Monoterpenes ; Neuroprotective Agents ; isolation & purification ; pharmacology ; Paeonia ; chemistry ; Plants, Medicinal ; chemistry ; Random Allocation ; Rats ; Rats, Wistar ; Superoxide Dismutase ; metabolism

BACKGROUNDNeuroprotective strategies following cardiopulmonary resuscitation (CPR) are an important focus in emergency and critical care medicine. Matrix metalloproteinases (MMPs), especially MMP9 attracted much attention because of its function in focal brain ischemia/reperfusion injury. In the focal cerebral ischemia model in rats, SB-3CT can suppress the expression of MMP9, relieving brain edema, and there was no studies on global cerebral ischemia-reperfusion injury after CPR.

METHODSOne hundred and twenty rats were randomly assigned to sham-operated (n = 40), resuscitation treatment (n = 40), and resuscitation control (n = 40) groups. Sham-operated group rats were anesthetized only and intubated tracheally, while the resuscitation treatment and resuscitation control groups also received cardiac arrest by asphyxiation. In the resuscitation treatment group, SB-3CT was injected intraperitoneally after restoring spontaneous circulation (ROSC), defined as restoration of supraventricular rhythm and mean arterial pressure (MAP) > or = 60 mm Hg for more than 5 minutes. The resuscitation control group also implemented ROSC without injection of SB-3CT. The rats were executed and samples were taken immediately after death, then at 3, 9, 24, and 48 hours (n = 8). Brain tissue expression of MMP9 protein, MMP9 mRNA, water content, Evans blue content, TNF-alpha, IL-1, and IL-6 was measured, and the brain tissue ultramicrostructure studied with electron microscopy.

RESULTSIn the resuscitation control group, brain tissue expression of MMP9 protein and mRNA, water content, Evans blue content, TNF-alpha, IL-1, and IL-6 were significantly elevated at 3 hours, and peaked at 24 hours after resuscitation, when compared with the sham-operated group (P < 0.05). Tissue ultramicrostructure also changed in the resuscitation control group. By contrast, although all these indexes were increased in the resuscitation treatment group compared with the sham-operated group (P < 0.05), they were lower than in the resuscitation control group (P < 0.05).

CONCLUSIONSExpression of MMP9 protein and mRNA, water content, Evans blue content, TNF-alpha, IL-1, and IL-6 increased in rat brain tissue after CPR, indicating disruption of the blood-brain barrier and excess inflammatory reaction. MMP9 expression was reduced with SB-3CT, resulting in reduced brain injury.

Animals ; Blood-Brain Barrier ; drug effects ; Brain ; immunology ; ultrastructure ; Cardiopulmonary Resuscitation ; Cytokines ; analysis ; Heterocyclic Compounds, 1-Ring ; pharmacology ; Inflammation ; prevention & control ; Male ; Matrix Metalloproteinase 9 ; analysis ; genetics ; Matrix Metalloproteinase Inhibitors ; Neuroprotective Agents ; pharmacology ; RNA, Messenger ; analysis ; Rats ; Rats, Sprague-Dawley ; Sulfones ; pharmacology

OBJECTIVETo research the effects of moschus, borneol, styrax and benzoinum on the structure and function of blood brain barrier in cerebral ischemia-reperfusion injury model rats.

METHODFocal middle cerebral artery occlusion (MCAO) was introduced as an in vivo ischemic model in rats. After 2 h MCAO, nylon suture was pulled up 1 cm to give blood reperfusion. After 22 h reperfusion, all animals were decapitated. The ultramicrostructure of blood brain barrier of ischemia hemisphere side in fronto-parietal cortex region by transmission electron microscope, and the content of VEGF and MMP-9 in ischemia side brain tissue were measured by ELISA.

RESULTIn model and solvent group rats, the capillary endothelium cells, astro-glial cells and nerve cells in ischemia hemisphere side in fronto-parietal region were emerged in different degree compared with sham-operated groups, which exhibited tight junction between endothelial cells being opened, basal lamina being dissolved, and permeability increasing, and cellularedema. In borneol (0.2 g x kg(-1)) group rats, the structure of three kinds of cells were nearly normal, which tight junction structure was clear, rough endoplasmic reticulum and polyribosome could be found in cytoplasm. In moschus (66.6 mg x kg(-1)) group rats, the structure of capillary endothelium cells and astrocytes were nearly normal as well as the basal lamina, but the electrons in neurons was maldistribution. In styrax (1.332 g x kg(-1)) group rats, astrocytes were nearly normal, while capillary endothelial cells and neurons exhibited oedema in different degrees. And the basal lamina was discontinuous, augmentation of cell spaces in endothelial cells increased the permeability, some endoplasmic reticulum broadened and ribosome ablated. In benzoinum (1.0 g x kg(-1)) group rats, oedema of capillary endothelial cells and astrocytes was significant, basal lamina broke. Meanwhile endoplasmic reticulum broadened as vacuole, the number of ribosome in rough endoplasmic reticulum decreased, crista mitochondriales in some neurons disappeared as vacuole which hint oedema happened. Results also showed that borneol decrease the level of VEGF in ischemia side brain tissue significantly, while has little influence on the level of MMP-9. Moschus showed the tendency to decrease the level of VEGF and MMP-9 in ischemia side brain tissue.

CONCLUSIONAromatic resuscitation drugs showed the protection effect on blood brain barrier in cerebral ischemia-reperfusion injury rats, which the protection effect of moschus and borneol were better than that of styrax and benzoinum. The mechanism of protection effect maybe related to decrease the level of VEGF and MMP-9.

Animals ; Benzoin ; pharmacology ; Blood-Brain Barrier ; drug effects ; metabolism ; ultrastructure ; Bornanes ; pharmacology ; Brain Ischemia ; drug therapy ; metabolism ; Disease Models, Animal ; Fatty Acids, Monounsaturated ; pharmacology ; Infarction, Middle Cerebral Artery ; drug therapy ; metabolism ; Male ; Matrix Metalloproteinase 9 ; drug effects ; metabolism ; Neuroprotective Agents ; pharmacology ; Plant Extracts ; pharmacology ; Rats ; Reperfusion Injury ; drug therapy ; metabolism ; Styrax ; chemistry ; Vascular Endothelial Growth Factor A ; drug effects ; metabolism