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AIM: To optimize the technique of intravenous injection of Evans blue and retinal preparations in mice, improving the accuracy and repeatability of staining experiment of retinal preparations.METHODS: C57BL/6 male mice were intravenous injected with 10g/L(1%)Evans Blue 0.3mL and circulated in vivo for 10 or 20min, and the eyes were removed after sacrificed and fixed in 4% paraformaldehyde for 20, 40 or 60min. When failure of intravenous injection, the experiment was remediated by intraperitoneal injection of 1% Evans Blue 0.3mL, circulated in vivo for 3h and fixed for 60min to observe morphology, distribution and leakage of the retinal vessels. Besides, we compared the morphology, distribution and leakage of the retinal vessels after intravenous injection with those after intraperitoneal injection to determine the optimal conditions for in vivo circulation time and retinal preparations.RESULTS: After intravenous injection, compared to the retinal vascular condition under 20min in vivo circulation time of Evans blue and 20 or 40min of fixation, with 10min of in vivo Evans blue circulation and 60min of fixation, the morphology of retinal vascular was more intact with less retinal vascular leakage, and the vascular branches are clear. When intravenous injection failed, remediated results from intraperitoneal injection showed that the morphology and distribution of retinal vessels were intact. There was no significant difference in morphology, distribution and leakage of the retinal vessels after 3h of intraperitoneal Evans blue circulation compared to 10min intravenous Evans blue circulation.CONCLUSION: This experiment optimizes the protocol, improves the accuracy and reproducibility of retinal preparations, and provides a reference for the study of related retinal vascular diseases.
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OBJECTIVE: To observe the distribution characters of Evans Blue (EB) exudation spots in the abdominal area in acute intestinal mucosal injury (AIMI) rats by using latitude and longitude grid counting and multivariate statistical analysis and to explore the feasibility of these methods. METHODS: Twenty-four SD rats were randomly divided into blank control, 7.5%, 10%, and 12.5% mustard oil groups (n＝6 rats in each group). The AIMI model was established by mustard oil enema, followed by injection of EB (0.1 mL/100g) into the tail vein. At 5, 10, 15, 20 and 25 h after EB injection, the rats under anesthesia were fixed in supine position for observing and photographing the abdominal subcutaneous EB exudation spots. The H．E. staining was used to observe histopathological changes of colonic mucosa. The longitude and latitude grids of the abdominal region were constructed (by taking the midpoint of the superior sternum as the origin) to determine the position of the blue spots. That the coordinate grids of the two regions can be connected geometrically is termed as "characteristic region". The data were processed by using multivariate statistical analysis. RESULTS: ① H．E. staining showed edema and inflammatory cell infiltration after colonic enema of different concentrations of mustard oil. ② Clustering analysis indicated that the distribution of exudation points in the "characteristic grid" had no temporal variation trend, and was not related with the concentration of mustard oil (P>0.05). ③ Factor analysis and contour analysis about the exudation spots of EB at 7.5% concentration showed that the "characteristic region Ⅱ" of different factors presented a tendency of time-dependent exudation, i．e. reduction of exudation degree along with time (P<0.05), and it is located near "Tianshu" acupoint. ④At 5 h after injection of EB, the 8 "characteristic regions" presented an EB-concentration-dependent tendency (reduction in exudation degree along with the increase of EB-concentration), among which the exudation degree of region C (near "Tianshu" acupoint) of the abdomen was higher (P<0.05). CONCLUSION: The multivariate statistical analysis method can be used to identify the abdominal "characteristic regions" of exudation spots of EB in rats with AIMI, and the characteristic region has acupoint sensitization characteristics related to the time and severity of mucosal injury.
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PURPOSE: Patients treated with propranolol, a nonselective β-adrenoceptor antagonist, develop severe anaphylaxis, but the mechanism remains unknown. We determined effects of β₁- and β₂-adrenoceptor antagonists on the anaphylaxis-induced increase in vascular permeability in mice. METHODS: In anesthetized ovalbumin-sensitized C57BL mice, mean arterial blood pressure (MBP) was measured, and Evans blue dye extravasation and hematocrit (Hct) were assessed at 20 minutes after antigen injection. The following pretreatment groups (n=7/group) were studied: (1) sensitized control (non-pretreatment), (2) propranolol, (3) the selective β₂-adrenoceptor antagonist ICI 118,551, (4) the selective β₁-adrenoceptor antagonist atenolol, (5) adrenalectomy, (6) the selective β₂-adrenoceptor agonist terbutaline, and (7) non-sensitized groups. RESULTS: The antigen injection decreased MBP, and increased Hct and vascular permeability in the kidney, lung, mesentery, and intestine, but not in the liver or spleen. Pretreatment with ICI 118,551, propranolol and adrenalectomy, but not atenolol, reduced the survival rate and augmented the increases in Hct and vascular permeability in the kidney, intestine, and lung as compared with the sensitized control group. Pretreatment with terbutaline abolished the antigen-induced alterations. Plasma epinephrine levels were increased significantly in the sensitize control mice. CONCLUSIONS: Blockade of β₂-adrenoceptor can deteriorate systemic anaphylaxis by augmenting hyperpermeability-induced increase in plasma extravasation by inhibiting beneficial effects of epinephrine released from the adrenal glands in anesthetized mice.
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Animals , Humans , Mice , Adrenal Glands , Adrenalectomy , Anaphylaxis , Arterial Pressure , Atenolol , Capillary Permeability , Epinephrine , Evans Blue , Hematocrit , Intestines , Kidney , Liver , Lung , Mesentery , Mice, Inbred C57BL , Plasma , Propranolol , Spleen , Survival Rate , TerbutalineABSTRACT
Necrosis is a form of cell death, which is related to various serious diseases such as cardiovascular disease, cancer, and neurodegeneration. Necrosis-avid agents (NAAs) selectively accumulated in the necrotic tissues can be used for imaging and/or therapy of related diseases. The aim of this study was to preliminarily investigate necrosis avidity of I-evans blue (I-EB) and its mechanism. The biodistribution of I-EB at 24 h after intravenous administration showed that the radioactivity ratio of necrotic to viable tissue was 3.41 in the liver and 11.82 in the muscle as determined by counting in model rats. Autoradiography and histological staining displayed preferential uptake of I-EB in necrotic tissues. nuclear extracts from necrotic cells exhibited 82.3% of the uptake in nuclei at 15 min, as well as 79.2% of the uptake at 2 h after I-EB incubation. The DNA binding study demonstrated that evans blue (EB) has strong binding affinity with calf-thymus DNA (CT-DNA) (=5.08×10 L/(mol/L)). Furthermore, the accumulation of I-EB in necrotic muscle was efficiently blocked by an excess amount of unlabeled EB. In conclusion, I-EB can not only detect necrosis by binding the DNA released from necrotic cells, but also image necrotic tissues generated from the disease clinically.
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Objective: To prepare chitosan neurotoxin nanoparticles (CS-NT-NP) and study its effect on the permeability of blood brain barrier and the serum levels of S100β by intranasal administration. Methods: A formamide extraction-ultraviolet spectrophotometry method was employed to determine the concentration of Evans blue (EB) in brain by different routes of administration and preparations. Qualitative analysis of fluorescence intensity and distribution of EB in brain tissue was performed by fluorescence microscopy. The serum S100β protein concentration was determined by ELISA. Results: Compared with the control group and NT group, CS-NT-NP could significantly increase the content of EB in the brain with time-effect relation and reached the peak at 120 min (P < 0.01); Compared with muscle injection and ip injection, intranasal administration could significantly increase the content of EB and reached peak time quickly. The results were consistent with the experimental results of qualitative analysis of fluorescence intensity and distribution of EB in brain tissue by fluorescence microscopy and S100β protein in serum. It was consistent with the experimental results of S100β protein. Conclusion: CS-NT-NP by intranasal administration can significantly increase the permeability of BBB, and further increase the drug concentration in the brain, which is a good carrier of macromolecular drugs into the brain.
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Objective To investigate the neuroprotective effects of Fasudil in cerebral I/R injury in mice.Methods 51 C57BL/6J mice was divided into two groups,CMC treated group (n=26) and Fasudil treated group (n=25),randomly.The mice were treated with Fasudil (10 mg/kg) or CMC (0.5％ CMC 10 ml/kg) separately.Then the treated mice were subjected to 60 min of focal ischemia and 18 h reperfusion.The infarct volume of brain was analyzed by TTC staining with MCID image system.BBB permeability was assessed by Evans blue extravasation and albumin leakage which was detected by immuno-blotting assay.The activity of MMP9 was analyzed by zymography.Results The infarct volume in CMC group ((99.07±6.53) mm3) was larger than that in Fasudil group ((57.02±8.93) mm3),the difference was statistically significant (P＜0.01).The activity of MMP9 in the mice treated with Fasudil was lower than that in CMC group.Compared with the CMC group(albumin (2.95±0.77),Evans blue (5.15±0.24)),the albumin and Evans blue content in the Fasudil treated group (albumin (1.04±0.18),Evans blue (1.96±0.31))reduced significanctly(all P＜0.01).Conclusion Fasudil protects I/R damage by inhibiting the activity of MMP9 to maintain blood-brain barrier permeability.
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Background Retinal vascular recanalization is key to the treatment of retinal vascular occlusive disease.Studies confirmed that urokinase by intravitreal injection inhibits the expression of occludin protein at tight junction complexes among retinal capillary endothelial cells.Objective This study was to observe the effects of urokinase via eye local injection on the outward permeability of blood-retinal barrier by detecting the concentration of intravitreal Evans blue (EB).Methods Sixty healthy Sprague-Dawley (SD) rats were randomly assigned to four groups,and the right eyes of the rats were used as experimental eyes.Urokinase of 4 μl (350 U) and the equal volume of PBS (0.01 mol/L) was intravitreally injected separately in the intravitreal urokinase group and the intravitreal PBS group,and 10 μl urokinase (1000U) and the equal volume of PBS was injected via retrobulbar tissue respectively as the retrobulbar urokinase group and the retrobulbar PBS group.Twenty-four hours after injection of drugs,0.5％ EB 4 μl was intravitreally injected.Four hours later,the rats were sacrificed and the right eyeballs were excised for the extraction and drying.EB was extracted from dried vitreous by formamide.Then,the concentration of EB in formamide was determined by a formamide extraction-ultraviolet spectrophotometry method to calculate the concentration of EB in vitreous.The use and care of experimental animals followed the Regulations for the Administration of Affair Concerning Experimental Animals by State Science and Technology Commission (2011 version).Results The rat vitreous body showed the light blue color in intravitreal urokinase group and the retinal vessels were visible under the microscope,and that in the retrobulbar urokinase group presented blue color.However,in the intravitreal and retrobulbar PBS group,rat vitreous exhibited the deeper blue color and retinas were invisible.Absorbance of EB in formamide was 0.181 ±0.008,0.450±0.017,0.330±0.009 and 0.436±0.012 in the intravitreal urokinase group,intravitreal PBS group,retrobulbar urokinase group and retrobulbar PBS group,respectively.The intravitreal EB concentrations in the intravitreal urokinase group were (0.266±0.014)g/L,which was lower than (0.667±0.026) g/L,(0.496±0.015) g/L and (0.657±0.017) g/L of the intravitreal PBS group,retrobulbar urokinase group and retrobulbar PBS group,showing significant different among the four groups (F =100.406,P＜0.01),and the intravitreal urokinase group showed the lowest value in comparison with other three groups (all at P＜0.01).Conclusions Local application of urokinase around eye can augment the outward permeability of blood-retinal barrier in rats.Intravitreal assay of EB after intravitreal injection is a feasible approach to the determination of outward permeability of blood-retinal barrier.
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Background Retina fixed flat-mount perfused by Evans blue (EB) is a common method for the evaluation of blood-retinal barrier (BRB).However,previous method is inconvenient for some laboratories because the retinal specimen can not be observed by gereral microscope rather than confocal laser scanning microscope after the fixation.Objective This study was to modify the preparing way of flat-mounted retina in order to obtain transparent specimen for the observation of rat retinal vessels and the evaluation of leakage under the ordinary fluorescence microscope.Methods Forty male SD rats were divided into the control group,diabetes mellitus (DM) 1-month group,DM 3-month group and DM 6-month group according to the random number table.Streptozotocinum (STZ) of 2％ dissolved in 0.05 mmol/L sodium citrate-hydrochloric acid buffer was intraperitoneally injected in SD rats to establish DM models,and the equal volume of solvent was injected in the same way in the control rats.One month,three months and six months after injection,EB of 30 g/L was injected via rat femoral vein in the dose of 45 mg/kg.Fifteen minutes after injection of EB,the rats were sacrificed and the retinas were isolated and cut radially to prepare the flat-mounted retinas in PBS immediately and then were dried till the specimens were transparent.The specimens were examined under the fluorescence microscope.The percentage of EB leakage was quantitatively calculated by IPP 6.0 software.All procedures were performed following approval of the institutional animal care and use committee of Tianjin Medical University.Results The retina morphology was normal in the control group,and EB filled the vessels,exhibiting the red fluorescence under the fluorescence microscope.Compared with the control group,retinal background fluorescence was enhanced slightly in the DM 1-month group,and focal leakage of the EB from capillaries and focal dilated vessels were found in the DM 3-month group,further,vascular caliber inequality,retinal hypoperfusion area and a larger number of hyperfluorescence areas were seen in the DM 6-month group.The percentage of leakage area was (0.05 ±0.02) ％,(0.27 ±0.06) ％,(1.17 ±0.18)％ and (4.77 ±0.66)％ in the control group,DM 1-month group,DM 3-month group and DM 6-month group,respectively,showing a significant difference among the four groups (F =795.800,P＜0.001),and the leakage area was obviously larger in the DM 3-month group and DM 6-month group than that in thecontrol group (q'=10.338,q'=43.475,both at P＜0.001).Conclusions Modified EB-perfused retinal wholemount method is easy and helpful for clear visualization of retinal vessel leakage induced by BRB breakdown in the diabetic rats under the common fluorescence microscope.
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The acute inflammatory response consists of three main vascular effects: vasodilatation and increased blood flow, increased vascular permeability, and leukocytosis into the injured tissues. All three events are induced relatively quickly, and, for all three, the pattern of response is complex but consistent. Evans blue dye is an alkaline stain, so there is an affinity for the alkali in the acidic nucleus. The use of Evans blue dye as an in vivo marker through vascular permeability, facilitates the investigation of the effect of pathological changes in various disorders mainly, immunological disorders, inflammatory disorders, cardiovascular diseases like atherosclerosis, myocardial infarction, cancers and others. Endothelials have pathophysiological roles in pulmonary hypertension, arterial hypertension, atherosclerosis, cerebral vasospasm and inflammatory processes. The present review discussed with role of evans blue in the assessment of vascular permeability for the various pharmacological activities which are helps for the future investigations
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Objective To explore the effect of Ulinastatin on blood brain barrier (BBB) and apoptosis of neural cells in septic rats.Methods Fifty-two clean level male Sprague-Dawley rats were randomly (random number table) divided into six groups:Sham groups at 6 h and 24 h,each group with six rats.Sepsis groups (CLP) and Ulinastatin treated groups (UTI) at 6 h and 24 h,each group with ten rats.In CLP and UTI groups,cecal ligation and puncture (CLP) were performed to induce sepsis.Sham group was only opened and closed abdomen.Ulinastatin (50 000 U/kg) was administered via femoral vein 1 h after CLP.The same volume of saline instead of Ulinastatin was administered in Sham and CLP groups.The neurological status was assessed by Neurological Deficit Scale Scores (NDSS) at 6 h and 24 h after CLP.Then the brain was harvested for HE staining and weighing water content.The BBB permeability was assayed by Evans Blue dye extravasations.Apoptosis of neural cells were detected by TUNEL immune fluorescence.Statistical analysis was performed with SPSS version 13.0,ANOVA was used for multiple groups comparison and t-test for paired comparison.Results The Neurological Deficit Scale Scores of UTI group was lower than Sham group (P ＜ 0.05) but higher than that of CLP group (P ＜ 0.05).Swelling,degeneration and edema were observed in cerebral cortex and hippocampal neurons in CLP group through light microscope,and were more serious than those in UTI group.Compared with UTI 24 h group,BBB permeability of CLP 24 h group significantly rose (P ＜ 0.05).The number of apoptosis of neural cells increased more in CLP group than it did in UTI group (P ＜ 0.05).Conclusions Ulinastatin could protect the cerebral tissue in septic rats by alleviating the damage of BBB and reducing the apoptosis of neural cells.
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Objective To investigate whether the macromolecular materials could enter cerebrospinal fluid and brain tissues in craniotomy with incision or non-incision of dura and arachnoid. Methods Adult male SD rats were randomly divided into three groups according to the random number table. The dura and arachnoid of rats in group A were cut open during craniotomy after general anesthesia; epidural craniotomy was done in rats in group B after general anesthesia; rats in group C (control group) were only generally anesthetized. All the rats were injected with Evans blue, a tracer used to detect the results, half an hour before each time point (1,3, 6, 12, 24, 72 hours and 1 week) via vein. The rats were executed at each time point to obtain the specimens of brain. The content of Evans blue in brain tissue was measured by fluorescence spectrophotometer for statistical analysis. The water content in the brain tissue was measured in a part of rats selected in groups A and B preoperatively and at postoperative 3 and 27 hours. Results It was found that some regions of the brain tissue were stained light blue in group A at 1,3, 6 and 24 hours. The blue was much lighter in brain tissue obtained at 72 hours in group A, and no blue stained at 1 week in group A . The contents of Evans blue in the brain tissues of rats in group A at 1,3, 6, 12, 24, 72 hours and 1 week were (18.07±1.25) μg/ml, (36.21±0.78) μg/ml, (25.73±1.14) μg/ml, (16.53±0.84) μg/ml, (23.34±1.91) μg/ml, (43.34±2.25) μg/ml and (25.27±1.88)μg/ml respectively, which were significantly higher than (3.15±0.45)μg/ml, (3.36±0.33)μg/ml, (2.98±0.54)μg/ml, (3.47±0.55)μg/ml, (3.54±0.37) μg/ml, (2.88± 0.42) μg/ml and (2.85±0.22) μg/ml respectively in group B and (2.97±0.37)μg/ml in group C (P<0.01). There was no significant difference in water content in brain tissue before and after operation (P>0.05). Conclusion After craniotomy with incision of dura and arachnoid, some macromolecular materials can enter the subarachnoid space and the brain parenehyma through blood-brain barrier of the wound of the scalp if the dura is sutured loosely.
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OBJECTIVES: The objective of this study was the standardization of a collection technique and staining in liquid-base that allies the pratical and cytological wealth, making possible a larger reproductibility and microscopic easiness. METHODS: Female wistar rats (n=20) were submitted to the daily vaginal collection in saline and fastened washed (ether/alcohol) and stained in suspension with a solution of Evans Blue 0.025%. The sample was pondered by centrifugation and observed under lens of 40 x. RESULTS: The stained smears allowed clear differentiation of the phases of hormonal cycle (diestrus, proestrus, estrus and metestrus); besides the differentiation of the cellular types in relation to its maturation degree having as parameters the cellular size, nucleus / cytoplasm relationship (NCR) and ink reaction. The study demonstrated the existence of three basic cellular patterns: cells with low NCR, accentuated cyanophily and small size; cells with increment in NCR, cyanophilic loss and larger volume cytoplasmatic and without nuclei keratinization cells in squamous aspect. CONCLUSION: The staining of the material allowed, besides the cytological classification, the quantification possibility that would result in a perfected accompaniment of the cycle estrous.
OBJETIVOS: O objetivo deste estudo foi à padronização de uma técnica de coleta e coloração em meio líquido que alie a praticidade e a riqueza citológica, possibilitando uma maior reprodutividade e facilidade microscópica. MÉTODOS: Ratas wistar (n=20) foram submetidas à coleta vaginal diária em salina e o lavado fixado (éter/álcool) e corado em suspensão com solução de azul de Evans 0,025%. A amostra foi concentrada por centrifugação e observado sob objetiva de 40 x. RESULTADOS: Os esfregaços corados permitiram nítida diferenciação das fases do ciclo hormonal (diestro, proestro, estro e metaestro); além da diferenciação dos tipos celulares em relação ao seu grau de maturação tendo como parâmetros o tamanho celular, relação núcleo / citoplasma (RNC) e reação tintorial. O estudo demonstrou a existência de três padrões celulares básicos: células com baixa RNC, acentuada cianofília e pequeno tamanho; células com acréscimo na RNC, perda de cianofilia e maior volume citoplasmático e células queratinizadas anucleadas em aspecto de escama. CONCLUSÃO: A coloração do material permitiu, além da classificação citológica, a possibilidade de quantificação o que resultaria em um acompanhamento mais acurado do ciclo estral.
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Objective To investigate the possible mechanism of the gap junctional influence on the change in permeability of the blood-brain barrier(BBB)after reperfusion subsequent to cerebral ischemia.Methods In the test laser scanning confocal microscope(LSCM)was used to investigate the change of Cx43 levels and distribution.The MCAO/R model was induced using intraluminal suture technique first described by Longa with a little modification.A total of 60 Wistar rats were divided into 4 groups:the sham-operation group,control group,octanol-treatment group and DMSO vehicle control group. Control group were further divided into seven subgroups at different time points of reperfusion after middle cerebral artery occlusion.To observe the change in permeability of BBB,Evans blue(EB)in the brain tissue was surveyed by the means of EB fluorescent quantitation.Octanol-treatment group and DMSO vehicle control group were done at the point of the peak of permeability of BBB.Octanol,the specific blocker for gap junctions(GJ)was used in an intervention study.To compare the amount of EB with the same point of groups,the influence of octanol on BBB permeability was investigated.Results At 3 h of reperfusion after cerebral ischemia for 2 h,the permeability of BBB began to increase,reached the peak at 24 h of reperfusion and was still elevated at 72 h.The Cx43 expression formed into bigger plague and remained linear disposition in the penumbra after reperfusion subsequent to cerebral ischemia.Octanol group was done at 24 h of reperfusion after cerebral ischemia.The amount of EB of octanol group((4.924?0.296)?g/g)was significantly lower than that of corresponding operation control group(5.543?0.506)?g/g.Conclusions (1)Cx43 expression is concentrated around vessels in brain.The Cx43 forms into bigger plague and the function maybe strengthens after reperfusion.Gap junction might aggravate the disruption of BBB.(2) Octanol,the specific blocker of gap junctions,could effectively prevent the permeability of BBB from increasing and has a protective effect on BBB.
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AIM: To study the protective effects of nimodipine liposomes for injection (NDLI) on injuries of total cerebral ischemia/reperfusion(I/R) in rats and anoxia in mice. METHODS: Acute anoxia in mice was produced by hypoxia under normal pressure and decapitation. In these two models the survival time and persistent time of gasping were observed. Ameliorated pulsinelli four-vessel occlusion method was used to make global brain ischemia model. The EEG, the time of righting reflex recovery and Evans blue content in the homogenate of the brain tissues were recorded. RESULTS: NDLI obviously prolonged the survival time and persistent time of gasping in mice subjected to acute anoxia, remarkably shortened the time of EEG recovery and righting reflex recovery, and reduced Evens blue content in the homogenate. CONCLUSION: NDLI has significantly protective effects on injuries of total cerebral I/R and anoxia.
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BACKGROUND AND OBJECTIVES: The role of neurogenic inflammation in the airways appears to protect the airways from various irritants in the inspired air. Enhanced NO production by inducible NO synthase (iNOS), which is expressed locally at the site of inflammation, has been implicated in the pathogenesis of inflammation. We studied to elucidate the role of NO in the pathogenesis of neurogenic inflammation in the nasal mucosa. MATERIAL AND METHOD: This study investigated the changes of microvascular leakage in the rat model of challenge/rechallenge with capsaicin, following the effects of a NOS inhibitor, NG-nitro-L-arginine methyl ester (L-NAME) and a substrate, L-arginine. And for more evident proofs, this study investigated the immunohistochemical localization of the expressed iNOS after capsaicin-challenge in the rat nasal mucosa. RESULTS: Capsaicin-rechallenge enhanced microvascular leakage in the nasal mucosa. Pretreatment with L-NAME inhibited the enhancement of neurogenic inflammation with capsaicin-rechallenge, and L-arginine reversed the inhibition of L-NAME. Strong immunohistochemical staining for iNOS was localized to inflammatory cells in the epithelial layer. CONCLUSION: Capsaicin or sensory neuropeptides released by capsaicin-challenge may induce the expression of iNOS in the nasal mucosa, and NO, produced by expressed iNOS, may mediate the enhancement of neurogenic inflammation with capsaicin-rechallenge.
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Animals , Rats , Arginine , Capsaicin , Evans Blue , Immunohistochemistry , Inflammation , Irritants , Models, Animal , Nasal Mucosa , Neurogenic Inflammation , Neuropeptides , NG-Nitroarginine Methyl Ester , Nitric Oxide Synthase , Nitric OxideABSTRACT
Objective To explore the dynamic changes of blood brain barrier(BBB)permeability after experimental subarachnoid hemorrhage(SAH)in rats.Methods Eighty female SD rats were divided into normal saline control group(n=10)and SAH group(n=70).The SAH model was induced by injecting 300 ?l of autologous arterial blood into the subarachnoid space near the circle of Willis via a cannula in an artificial hole between the olfactory bulb and frontal lobe.BBB permeability in cerebral cortex of SAH and normal saline rats was assessed at 6,12,24,36,48,60,and 72 h after SAH establishment by fluorescence spectrophotometer and fluorescence microscopy for Evans Blue(EB)extravasation.The ultrastructural changes in BBB were observed with transmission electron microscope.Results Compared with the control group,the changes of Evans Blue content and extravasation in cerebral cortex of SAH group peaked at 36 h(P0.05).The severe ultrastructural abnormality was found at 36 h after SAH.Conclusion The changes of BBB permeability develop at the acute stage of SAH,resulting from multiple factors together.The BBB after SAH possesses a self-repairable property.
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Objective To verify the effect of Evans blue dye on determining the retina blood vessel leakage. Methods Male Sprague Dawley rats were used in this study. The VEGF induced retinal blood vessel leakage was checked with Evans blue dye. Then the blood retina barrier breakdown of 1 week diabe tic animals was quantified with Evans blue. The dye was extracted from retina by formamide and the extraction was checked with spectrophotometer. Evans blue leakage was normalized against wet or dry retina weight. Results The retinal Evans blue content of eyes treated with VEGF was remarkably higher than that of the controls (n=17, P
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Objective To look for the neurological evidence of the referred pain of the chronic prostatitis and the relationship between the pain of the chronic prostatitis, bladder, and pelvic floor. Methods The distribution of plasma extravasation was determined with Evans blue dye after irritation of the prostate and bladder of the SD rat with capsaisin, and the distribution of dye extravasation was analyzed. Substance P expression within the dorsal horn of the spinal cord was determined by hybridization in situ after irritation of the prostate, bladder and superficial somatic region of the pelvic floor with formalin. Results Pain irritation of the prostate resulted in plasma extravasation in L_5 to S_2 dermatomes (mainly in L_6 to S_1). In rats receiving bladder irritation, the distribution of plasma extravasation showed a similar pattern to that observed in rats receiving prostatic irritation. Irritation of the prostate, bladder and pelvic floor resulted in similar substance P expression within the dorsal horn of the spinal cord (L_6 to S_1). Conclusions Our results strongly suggest that referred pain status in inflammation of the bladder and prostate is neurogenically mediated. There are significant overlaps of nociceptive neurons within the spinal cord, which receive nociceptive inputs from the prostate, bladder and pelvic floor. Pathological changes in the prostate, bladder and pelvic floor can result in similar pain sensations.
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Objective To investigate the influences of blast wave overpressure created by explosion on rat's blood-brain barrier (BBB),and to provide an experimental evidence for further elucidating the mechanism of craniocerebral blast injury,to help direct the clini- cal therapeutic strategy.Methods Twenty male Sprague-Dawley rats were randomly divided into four groups:15min,12h and 24h post- explosion groups and normal control group with five rats for each group.Rats were injured with blast wave produced by explosion.Evan 's blue(EB)was intravenously injected into the injured rats,and the overpressure-induced leakage of BBB was examined under the fluores- cence microscope and by quantitative observation with opacimeter.Results Extensive impairment of BBB could be seen all over the brain 15 min after the overpressure explosion,and bright red EB fluorescent patches appeared as a method distributing along the capillary vessels. These patches were more distinct near the cerebral cortex,and with the lapse of time,their number decreased gradually from the inner to the outer areas.The changes in EB contents in brain tissue matched with that of the BBB.The EB contents in brain tissues in 15min group were(330.4?53.6)?g/g,and it was much higher compared with that of other groups(P
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BACKGROUND AND OBJECTIVES: Nitric oxide is a labile neurotransmitter causing vasodilation by relaxing vascular smooth muscle. Endogenous nitric oxide is an important modulator of airway function, but its role in the regulation of airway microvascular leak remains unclear. The purpose of this study was to investigate modulatory action of nitric oxide on capsaicin-induced neurogenic plasma extravasation. MATERIALS AND METHODS: Neurogenic inflammation in rat nasal mucosa was induced by intranasal application of the capsaicin, 50 mM, 50 microliter. Rats were administered i.v. Nw-nitro-L-arginine (L-NNA, 1 to 10 mg/kg), as well as concurrent pretreatment with L-arginine (50 mg/kg). The amount of plasma extravasation was measured by measuring amount of extravasated Evans blue using spectrophotometer and by counting percent area density of Monastral blue-labeled blood vessels. RESULTS: In L-NNA(50 mg/kg) pretreated group, the amount of extravasated Evans blue and percent area density of Monastral blue-labeled blood vessels decreased in a dose-dependent manner (p < 0.01). This inhibition was reversed significantly by adding L-arginine. CONCLUSION: It is suggested that endogenous nitric oxide may have as modulatory role in neurogenic plasma extravasation.