Evans Blue Content Research Articles

Protective Effect of Protocatechuic Aldehyde on Cerebral Ischemia/Reperfusion Injury in Rats through Blood-Brain Barrier Protection.

Cerebral ischemia can lead to destruction of the blood-brain barrier (BBB), the main cause of cerebral edema and cerebral infarction. BBB damage is also one of the key factors affecting the result of drug therapy. We studied the protective effect of 5-day pretreatment with protocatechuic aldehyde (PAL) at doses of 10 and 20 mg/kg on BBB function and structure after middle cerebral artery occlusion/reperfusion (MCAO/R) in rats. The infarct volume, behavioral neurological deficit score, and Evans blue content in the brain were estimated. We also evaluated the content of nitric oxide (NO) and activities of inducible and neuronal NO synthases. Expression of aquaporin-4 (AQP-4), occludin, claudin-5, and MMP-3 in the brain tissues was estimated by Western blotting. The BBB ultrastructure was analyzed under an electron microscope. We revealed that PAL at both used doses significantly reduced the neurological deficit score, brain infarct volume, and Evans blue extravasation. Electron microscopy showed that PAL significantly improved the ultrastructure of BBB and alleviated its injury. Pretreatment with PAL increased expression of occludin and claudin-5 and reduced expression of AQP-4 and MMP-3. At the same time, the release of NO and activities of NO synthases were notably inhibited. Our results suggest that PAL can be a promising compound to attenuate cerebral ischemia resulting from occlusion/reperfusion injury via BBB protection.

465 Focused Ultrasound-Mediated Blood-Spinal Cord Barrier Permeabilization: A Promising Targeted Approach for Treating Chronic Spinal Cord Injury in Rat Models

INTRODUCTION: Focused ultrasound (FUS) with microbubble contrast agents can temporarily disrupt the blood-brain barrier (BBB), enabling drug delivery. FUS may also offer a minimally invasive method for delivering therapeutic agents to the affected area in chronic spinal cord injury (SCI) by transiently opening the blood-spinal cord barrier (BSCB). METHODS: Nine female Sprague-Dawley rats underwent a T9-T11 laminectomy with contusion SCI and were randomly allocated into FUS or control group. Three weeks later, FUS was administered intravenously with microbubbles. A pressure sweep was performed until ultra-harmonics were captured using a wideband hydrophone. We administered 2% Evans blue (EB) immediately after FUS confirmed the BSCB permeability. Transcardial perfusion was performed 2 hours later, and spinal cords were collected. Spectrophotometry was used to determine the EB concentration. Statistical analysis was performed using a student t-test. RESULTS: Five rats were treated with FUS while four were used as controls. Successful feedback control was achieved with FUS, and ultra-harmonics were observed in the FUS group at a mean peak pressure of 0.5 MPa. Macroscopic identification of EB in the spinal cord sections of animals exposed to FUS confirmed the localized opening of the BSCB. Spectrophotometry revealed an increase in EB concentration in the FUS group compared to controls: Mean EB concentration was 0.03 μg in the FUS group versus 0.016 μg in controls at 1-cm above the injury (p < 0.001); 0.27 μg versus 0.093 μg at the injury site (p < 0.001); and 0.25 μg versus 0.18 μg at 1-cm below the injury(p < 0.001). CONCLUSIONS: FUS increases the permeability of the BSCB in a chronic contusion SCI rat model. This suggests that FUS-mediated BSCB permeabilization may hold promise as a targeted approach for improving therapeutic outcomes in the treatment of chronic SCI.

Aloe vera Leaf Extract Reduced BBB Permeability and Improved Neurological Results after Traumatic Brain Injury: The Role of Oxidative Stress.

Recognizing the importance of medicinal plants and the absence of specific medications for traumatic brain injury (TBI) treatment, this study was conducted to evaluate the effects of an aqueous extract of Aloe vera on oxidative stress, blood-brain barrier (BBB) permeability, and neurological scores following TBI. Adult male rats were categorized into five groups: sham, TBI, vehicle, low-dose Aloe vera (LA), and high-dose Aloe vera (HA). We induced diffuse TBI using the Marmaro model and administered the aqueous Aloe vera leaf extract, as well as vehicle, via intraperitoneal injection half an hour after TBI. Neurological outcomes were assessed both before and several hours after TBI. Additionally, oxidative stress factors were measured 24 hr after TBI, and Evans blue content (a BBB permeability index) was determined 5 hr after TBI in both serum and brain. Both LA and HA reduced the increase in BBB permeability after TBI, with HA having a more pronounced effect than LA. Both Aloe vera doses decreased brain MDA levels, increased brain TAC, and lowered both serum and brain PC levels. The impact of Aloe vera on brain oxidative parameters was more significant than on serum. HA also counteracted the declining effects of TBI on neurological outcomes at 4 and 24 hr post-TBI. This study suggests that Aloe vera extract may reduce BBB permeability and improve neurological outcomes after TBI by decreasing oxidative factors and increasing antioxidant factors.

Hyperglycemia disrupted the integrity of the blood-brain barrier following diffuse axonal injury through the sEH/NF-κB pathway.

We aimed to investigate the role of soluble epoxide hydrolase for hyperglycemia induced-disruption of blood-brain barrier (BBB) integrity after diffuse axonal injury (DAI). Rat DAI hyperglycemia model was established by a lateral head rotation device and intraperitoneal injection of 50% glucose. Glial fibrillary acidic protein, ionized calcium-binding adapter molecule-1, β-amyloid precursor protein, neurofilament light chain, and neurofilament heavy chain was detected by immunohistochemistry. Cell apoptosis was examined by terminal deoxynucleotidyl transferase nick-end labeling (TUNEL) assay. The permeability of blood-brain barrier (BBB) was assessed by expression of tight junction proteins, leakage of Evans blue and brain water content. The soluble epoxide hydrolase (sEH) pathway was inhibited by 1-trifluoromethoxyphenyl-3-(1-propionylpiperidin-4-yl) urea (TPPU) and the nuclear transcription factor kappa B (NF-κB) pathway was inhibited by pyrrolidine dithiocarbamate and activated by phorbol-12-myristate-13-acetate in vivo and/or vitro, respectively. The inflammatory factors were detected by enzyme-linked immunosorbent assay. Hyperglycemia could exacerbate axonal injury, aggravate cell apoptosis and glial activation, worsen the loss of BBB integrity, increase the release of inflammatory factors, and upregulate the expression of sEH and NF-κB. Inhibition of sEH could reverse all these damages and protect BBB integrity by upregulating the expression of tight junction proteins and downregulating the levels of inflammatory factors in vivo and vitro, while the agonist of NF-κB pathway abrogated the protective effects of TPPU on BBB integrity in vitro. sEH was involved in mediating axonal injury induced by hyperglycemia after DAI by disrupting BBB integrity through inducing inflammation via the NF-κB pathway.

Systemic mapping of organ plasma extravasation at multiple stages of chronic heart failure.

Introduction: Chronic Heart failure (CHF) is a highly prevalent disease that leads to significant morbidity and mortality. Diffuse vasculopathy is a commonmorbidity associated with CHF. Increased vascular permeability leading to plasma extravasation (PEx) occurs in surrounding tissues following endothelial dysfunction. Such micro- and macrovascular complications develop over time and lead to edema, inflammation, and multi-organ dysfunction in CHF. However, a systemic examination of PEx in vital organs among different time windows of CHF has never been performed. In the present study, we investigated time-dependent PEx in several major visceral organs including heart, lung, liver, spleen, kidney, duodenum, ileum, cecum, and pancreas between sham-operated and CHF rats induced by myocardial infarction (MI). Methods: Plasma extravasation was determined by colorimetric evaluation of Evans Blue (EB) concentrations at 3days, ∼10weeks and 4months following MI. Results: Data show that cardiac PEx was initially high at day 3 post MI and then gradually decreased but remained at a moderately high level at ∼10weeks and 4months post MI. Lung PEx began at day 3 and remained significantly elevated at both ∼10weeks and 4months post MI. Spleen PExwas significantly increased at ∼10weeks and 4months but not on day 3 post MI. Liver PEx occurred early at day 3 and remain significantly increased at ∼10weeks and 4months post MI. For the gastrointestinal (GI) organs including duodenum, ileum and cecum, there was a general trend that PEx level gradually increased following MI and reached statistical significance at either 10weeks or 4months post MI. Similar to GI PEx, renal PEx was significantly elevated at 4months post MI. Discussion: In summary, we found that MI generally incites a timedependent PEx of multiple visceral organs. However, the PEx time window for individual organs in response to the MI challenge was different, suggesting that different mechanisms are involved in the pathogenesis of PEx in these vital organs during the development of CHF.

Mesenchymal Stem Cells Overexpressing FGF21 Preserve Blood-Brain Barrier Integrity in Experimental Ischemic Stroke.

Blood-brain barrier (BBB) disruption is a prominent pathophysiological mechanism in stroke. Transplantation of mesenchymal stem cells (MSCs) preserves BBB integrity following ischemic stroke. Fibroblast growth factor 21 (FGF21) has been shown to be a potent neuroprotective agent that reduces neuroinflammation and protects against BBB leakage. In this study, we assessed the effects of transplantation of MSCs overexpressing FGF21 (MSCs-FGF21) on ischemia-induced neurological deficits and BBB breakdown. MSCs-FGF21 was injected into the rat brain via the intracerebroventricular route 24 h after middle cerebral artery occlusion (MCAO) surgery. The behavioral performance was assessed using modified neurological severity scores and Y-maze tests. BBB disruption was measured using Evans blue staining, IgG extravasation, and brain water content. The levels of tight junction proteins, aquaporin 4, and neuroinflammatory markers were analyzed by western blotting and immunohistochemistry. The activity of matrix metalloproteinase-9 (MMP-9) was determined using gelatin zymography. At day-5 after MCAO surgery, intraventricular injection of MSCs-FGF21 was found to significantly mitigate the neurological deficits and BBB disruption. The MCAO-induced loss of tight junction proteins, including ZO-1, occludin, and claudin-5, and upregulation of the edema inducer, aquaporin 4, were also remarkably inhibited. In addition, brain infarct volume, pro-inflammatory protein expression, and MMP-9 activation were effectively suppressed. These MCAO-induced changes were only marginally improved by treatment with MSCs-mCherry, which did not overexpress FGF21. Overexpression of FGF21 dramatically improved the therapeutic efficacy of MSCs in treating ischemic stroke. Given its multiple benefits and long therapeutic window, MSC-FGF21 therapy may be a promising treatment strategy for ischemic stroke.

Multiomics reveal human umbilical cord mesenchymal stem cells improving acute lung injury via the lung-gut axis.

Acute lung injury (ALI) and its final severe stage, acute respiratory distress syndrome, are associated with high morbidity and mortality rates in patients due to the lack of effective specific treatments. Gut microbiota homeostasis, including that in ALI, is important for human health. Evidence suggests that the gut microbiota improves lung injury through the lung-gut axis. Human umbilical cord mesenchymal cells (HUC-MSCs) have attractive prospects for ALI treatment. This study hypothesized that HUC-MSCs improve ALI via the lung-gut microflora. To explore the effects of HUC-MSCs on lipopolysaccharide (LPS)-induced ALI in mice and the involvement of the lung-gut axis in this process. C57BL/6 mice were randomly divided into four groups (18 rats per group): Sham, sham + HUC-MSCs, LPS, and LPS + HUC-MSCs. ALI was induced in mice by intraperitoneal injections of LPS (10 mg/kg). After 6 h, mice were intervened with 0.5 mL phosphate buffered saline (PBS) containing 1 × 106 HUC-MSCs by intraperitoneal injections. For the negative control, 100 mL 0.9% NaCl and 0.5 mL PBS were used. Bronchoalveolar lavage fluid (BALF) was obtained from anesthetized mice, and their blood, lungs, ileum, and feces were obtained by an aseptic technique following CO2 euthanasia. Wright's staining, enzyme-linked immunosorbent assay, hematoxylin-eosin staining, Evans blue dye leakage assay, immunohistochemistry, fluorescence in situ hybridization, western blot, 16S rDNA sequencing, and non-targeted metabolomics were used to observe the effect of HUC-MSCs on ALI mice, and the involvement of the lung-gut axis in this process was explored. One-way analysis of variance with post-hoc Tukey's test, independent-sample Student's t-test, Wilcoxon rank-sum test, and Pearson correlation analysis were used for statistical analyses. HUC-MSCs were observed to improve pulmonary edema and lung and ileal injury, and decrease mononuclear cell and neutrophil counts, protein concentrations in BALF and inflammatory cytokine levels in the serum, lung, and ileum of ALI mice. Especially, HUC-MSCs decreased Evans blue concentration and Toll-like receptor 4, myeloid differentiation factor 88, p-nuclear factor kappa-B (NF-κB)/NF-κB, and p-inhibitor α of NF-κB (p-IκBα)/IκBα expression levels in the lung, and raised the pulmonary vascular endothelial-cadherin, zonula occludens-1 (ZO-1), and occludin levels and ileal ZO-1, claudin-1, and occludin expression levels. HUC-MSCs improved gut and BALF microbial homeostases. The number of pathogenic bacteria decreased in the BALF of ALI mice treated with HUC-MSCs. Concurrently, the abundances of Oscillospira and Coprococcus in the feces of HUS-MSC-treated ALI mice were significantly increased. In addition, Lactobacillus, Bacteroides, and unidentified_Rikenellaceae genera appeared in both feces and BALF. Moreover, this study performed metabolomic analysis on the lung tissue and identified five upregulated metabolites and 11 downregulated metabolites in the LPS + MSC group compared to the LPS group, which were related to the purine metabolism and the taste transduction signaling pathways. Therefore, an intrinsic link between lung metabolite levels and BALF flora homeostasis was established. This study suggests that HUM-MSCs attenuate ALI by redefining the gut and lung microbiota.

Effects of Gryllus bimaculatus and Oxya chinensis sinuosa extracts on brain damage via blood-brain barrier control and apoptosis in mice with pentylenetetrazol-induced epilepsy.

The demand for environmentally friendly foods with high nutritional value and low carbon emissions is increasing with the aging of the global population and the crisis of food resources. Edible insects are becoming increasingly well-known as such foods. This study evaluated the effects and mechanisms of Gryllus bimaculatus (Cricket) (Gb) and Oxya chinensis sinuosa (Grasshopper) (Ocs) extracts on epilepsy. A pentylenetetrazol (PTZ)-induced seizure mouse model was used for the study, and Gb and Ocs extracts were administered for 29 days on alternate days at concentrations of 8 g/kg and 16 g/kg. The integrity of the blood-brain barrier (BBB) and brain edema was measured using the perfusion of Evans blue dye and brain water content. Gb and Ocs extracts prevented BBB permeabilization and cerebral edema through increasing the expression of tight junction-associated proteins in the endothelial cells and reducing water content in PTZ-treated mice. Additionally, Gb and Ocs extracts protected neurons from oxidative stress and apoptosis in different brain areas. These protective effects were demonstrated through the restoration of the expression of neuronal nuclear protein and postsynaptic density protein-95, thus increasing the levels of glutathione and superoxide dismutase, decreasing lipid peroxidation, and recovering apoptosis-associated proteins, such as Bax, cleaved PARP, and cleaved caspase-3, in epileptic mice. In addition, Gb and Ocs extracts rescued PTZ-induced hyperexcitable neurons to control mice level, as supported by the restored expression of gamma-aminobutyric acid (GABA) transporter 1, the metabotropic glutamate receptors-GRM2/3, and BDNF. This study suggested that Gb and Ocs extracts are novel medicinal candidates that can help ameliorate epilepsy by improving BBB health and preventing oxidative stress-mediated apoptosis.

Evaluating the neuroprotective effects of progesterone receptors on experimental traumatic brain injury: The PI3K/Akt pathway.

Studies have confirmed the salutary effects of progesterone (P4) on traumatic brain injury (TBI). This study investigated the beneficial effects of P4 via its receptors on TBI, and also whether progesterone receptors (PRs) can modulate TBI through PI3K/Akt pathway. Marmarou method was utilized to induce diffuse TBI in ovariectomized rats. P4 (1.7mg/kg) or the vehicle (oil) was administered 30min after TBI induction. Moreover, RU486 (PR antagonist) and its vehicle (DMSO) were injected before TBI induction and P4 injection. Brain Evans blue content, brain water content (WC), various oxidative stress parameters, IL-1β levels, tumor necrosis factor-α (TNF-α), histopathological alterations, and also phosphorylated Akt (p-Akt) and PI3K expressions in the brain were assessed 24h after TBI. The veterinary comma scale (VCS) was measured before and after TBI at different times. The findings revealed that P4 caused an increase in VCS and a decrease in brain WC, oxidative stress, TNF-α and IL-1β levels. RU486 inhibited the beneficial effects of P4 on these indices. Moreover, RU486 prevented the reduction of brain edema, inflammation, and apoptosis caused by P4. Moreover, P4 following TBI increased the expression of PI3K/p-Akt protein in the brain. RU486 eliminated the effects of P4 on PI3K/p-Akt expression. According to these findings, PRs are acting as critical mediators for the neuroprotective properties of P4 on oxidative stress, pro-inflammatory cytokine levels, and neurological outcomes. PRs also play an important role in regulating the PI3K/p-Akt expression and nongenomic function of P4.

How different reproduction protocols can affect the germination of seeds: The case of three stenoendemic species on Mt. Olympus (NC Greece)

Mt. Olympus, the highest mountain in Greece, is a biosphere reserve and a magnet for countless visitors. In the wider area of Olympus, at least 1,700 species and subspecies of plants are recorded, 26 of which are endemic. Most of the endemics only occur in the high or subalpine zone, which is expected to be strongly affected by climate change or in specific microsites that might be affected by several other factors. Thus, the unique flora of the mountain will probably become extinct. Ex situ conservation can be utilized to prevent and even reverse this trend and preserve plant diversity for future generations. The aim of the present study was to develop reproduction protocols for the endemic species on Mt. Olympus, Centaurea incompleta, Centaurea litochorea and Viola striis-notata, to facilitate their mass production, either for ex situ conservation or reintroduction into their natural habitats, if necessary. Seeds of the target species were collected in summer 2021. In a sample of the collected seeds, the embryo viability was checked using sequentially 1% w/v tetrazolium solution and Evans blue solution concentration of 0.25% w/v. As for the germination tests, two treatments were used to terminate seed dormancy: (a) cold stratification at } 2 °C, and (b) imbibition in gibberellic acid (250 ppm) for 48 hours. The results showed that more than 75% of the embryos in the fertile seeds were viable. In the seed germination tests, treatment with gibberellic acid resulted in germination percentages for Centaurea incompleta and Viola striis-notata are equal to or very close to the seed viability percentage. In contrast, no treatment was successful for Centaurea litochorea, as the control germination percentage was higher.

Toxicological evaluation of a nonlethal riot control combinational formulation upon dermal application using animal models

Numerous adverse effects on human health have been reported in epidemiological studies of oleoresin capsicum (OC) and other riot control agents (RCAs). Importantly, the daunting risk of such RCAs can be neutralized by optimizing the desired concentration of such agents for mob dispersal. Hence, a nonlethal riot control combinational formulation (NCF) was prepared for dispersing rioters without imparting fatal outcomes. However, for desired utilization of NCF, it is essential to recognize its extent of potential toxicity. Therefore, the current investigation evaluated the dermal toxicity of NCF using experimental animals in compliance with the OECD guidelines. Additionally, few essential metal ions were analyzed and found non -significantly different in the test rats as compared to control rats. Moreover, abnormal dermal morphology and lesions ultrastructural tissue defects were not noticed as evinced by different studies like ultrasonography, histology, and scanning electron microscopy (SEM) respectively. Further, Doppler ultrasonography exhibited non-significantly different blood flow velocity in both groups, whereas miles test demonstrated a significantly increased Evans blue concentration in test rats compared to the control rats, which might be due to an initial increase in blood flow via an instant action of the NCF at the cutaneous sensory nerve endings. However, our results demonstrated NCF can produce initial skin irritating and sensitizing effects in guinea pigs and rabbits without the antecedence of acute toxicity (≤2000 mg/kg) in Wistar rats.

Effects of hydrogen gas on NOD-like receptor protein 3 inflammasomes in the cerebral cortex of rats with traumatic brain injury

To investigate the effect of hydrogen gas on NOD-like receptor protein 3 (NLRP3) inflammasomes in the cerebral cortex of rats with traumatic brain injury (TBI). 120 adult male Sprague-Dawley (SD) rates were randomly divided into 5 groups (n = 24): sham operation group (S group), TBI model group (T group), TBI+NLRP3 inhibitor MCC950 group (T+M group), TBI+hydrogen gas group (T+H group), TBI+hydrogen gas+MCC950 group (T+H+M group). TBI model was established by controlled cortical impact. NLRP3 inhibitor MCC950 (10 mg/kg) was intraperitoneally injected for 14 consecutive days before TBI operation in T+M and T+H+M groups. 2% hydrogen inhalation was given for 1 hour at 1 hour and 3 hours after TBI operation in T+H and T+H+M groups. At 6 hours after TBI operation, the pericontusional cortex tissues were obtained, the content of Evans blue (EB) was detected to evaluate the permeability of the blood-brain barrier. Water content in brain tissue was detected. The cell apoptosis was detected by TdT-mediated dUTP nick end labeling (TUNEL) and the neuronal apoptosis index was calculated. The expressions of Bcl-2, Bax, NLRP3, apoptosis-associated speck-like protein containing CARD (ASC) and caspase-1 p20 were detected by Western blotting. The levels of interleukins (IL-1β, IL-18) were detected by enzyme-linked immunosorbent assay (ELISA). Compared with the S group, the content of EB in cerebral cortex, water content in brain tissue, apoptosis index and the expressions of Bax, NLRP3, ASC, caspase-1 p20 in T group were significantly increased, the expression of Bcl-2 was down-regulated, the levels of IL-1β and IL-18 were increased [the content of EB (μg/g): 87.57±6.89 vs. 10.54±1.15, water content in brain tissues: (83.79±2.74)% vs. (74.50±1.19)%, apoptotic index: (62.66±5.33)% vs. (4.61±0.96)%, Bax/β-actin: 4.20±0.44 vs. 1, NLRP3/β-actin: 3.55±0.31 vs. 1, ASC/β-actin: 3.10±0.26 vs. 1, caspase-1 p20/β-actin: 3.28±0.24 vs. 1, Bcl-2/β-actin: 0.23±0.03 vs. 1, IL-1β (ng/g): 221.58±19.15 vs. 27.15±3.27, IL-18 (ng/g): 87.26±7.17 vs. 12.10±1.85, all P < 0.05]. Compared with the T group, the T+M, T+H and T+H+M groups had significant reductions in the content of EB and water content in brain tissue, apoptotic index of the cerebral cortex, the expressions of Bax, NLRP3, and caspase-1 p20 in the brain tissue and the levels of IL-1β and IL-18, significant increases in the expression of Bcl-2. However, there was no significant difference in ASC expression. Compared with the T+H group, the content of EB in the cerebral cortex, water content in brain tissue, and apoptotic index, and the expressions of Bax, NLRP3 and caspase-1 p20 were further down-regulated in T+H+M group, the expression of Bcl-2 was further up-regulated, the levels of IL-1β and IL-18 were further decreased [the content of EB (μg/g): 40.49±3.15 vs. 51.96±4.69, water content in brain tissue: (76.58±1.04)% vs. (78.76±1.16)%, apoptotic index: (32.22±3.44)% vs. (38.54±3.89)%, Bax/β-actin: 1.92±0.16 vs. 2.56±0.21, NLRP3/β-actin: 1.94±0.14 vs. 2.37±0.24, caspase-1 p20/β-actin: 1.97±0.17 vs. 2.31±0.19, Bcl-2/β-actin: 0.82±0.07 vs. 0.52±0.04, IL-1β (ng/g): 86.23±7.09 vs. 110.44±10.48, IL-18 (ng/g): 40.18±3.22 vs. 46.23±4.02, all P < 0.05], but there were no statistical significance in all the indicators between T+M group and T+H group. The mechanism by which hydrogen gas alleviates TBI may be related to inhibiting NLRP3 inflammasomes in the cerebral cortex of rats.

Systemic Mapping of Organ Plasma Extravasation Post Myocardial Infarction

Chronic Heart failure (CHF) is a highly prevalent disease that leads to significant morbidity and mortality. Diffuse vasculopathy is a common feature of the morbidity associated with CHF. Increased vascular permeability leading to plasma extravasation (PEx) occurs in surrounding tissues following endothelial dysfunction. Such micro- and macro-vascular complications develop over time and lead to edema, inflammation, and multi-organ dysfunction in CHF. However, a systemic examination of PEx in vital organs among different time windows of CHF has never been performed before. In the present study, we investigated time-dependent PEx in several major visceral organs including heart, lung, liver, spleen, kidney, duodenum, ileum, cecum and pancreas between sham-operated and CHF rats induced by myocardial infarction (MI). Plasma extravasation was determined by evaluating Evans Blue concentrations calorimetrically at fluorescence excitation wavelength of 620 nm (bandwidth 10 nm) and an emission wavelength of 680 nm (bandwidth 40 nm) at 3 days, 8-10 weeks and 4 months following MI. Data show that heart PEx level was initially high at day 3 post MI and then gradually decreased but still remained at a moderately high level at 8-10 weeks and 4 months post MI. Lung PEx began at day 3 and remained significantly high at both 8-10 weeks and 4 months post MI. Spleen PEx level was significantly high at 8-10 weeks and 4 months but not day 3 post MI. Liver PEx occurs early at day 3 and remain significantly high at 8-10 weeks and 4 months post MI. For the gastrointestinal (GI) organs including Duodenum, ileum and cecum, there was a general trend that PEx level gradually increased following MI and reached a statistical significance at 4 months post MI. Different with GI PEx, renal PEx level was initially high at day 3 but back to normal level at 8-10 weeks post MI. However, renal PEx was significantly elevated again at 4 months post MI. In summary, we found that MI generally incites a time-dependent PEx of multiple visceral organs. However, the PEx time window for individual organ in response to the MI challenge was different, suggesting that different mechanisms might be involved in the pathogenesis of PEx in these vital organs during CHF. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Protective effect of barium chloride pretreatment on lung in mice with acute respiratory distress syndrome

To explore whether barium chloride (BaCl2) preconditioning has the protective effect on lipopolysaccharide (LPS)-induced acute respiratory distress syndrome (ARDS) model in mice and the possible mechanism. Sixty 8-12 week old healthy C57BL/6 male mice were randomly divided into control group, ARDS model group and BaCl2 pretreatment group, with 20 mice in each group. The BaCl2 pretreatment group was continuously injected with BaCl2 (4 mg/kg through the tail vein) for 3 days before ARDS model establishment. ARDS model was established by intratracheally injecting (3 mg/kg) LPS. The control group was intratracheally given the same volume of 0.9% normal saline. On 24th hour after ARDS model establishment, some mice were sacrificed for obtaining fresh lung tissue. And the right lower lobe of the lung was separated for observing the pathological changes of lung tissue while the left lung tissue was used to measure the wet/dry weight ratio (W/D) of the lung. Some mice were sacrificed for observing pulmonary microvascular permeability at 2nd hours after injecting Evans blue (EB) through tail vein. The left mice were killed for alveolar lavage to measure the levels of tumor necrosis factor-α (TNF-α) via enzyme linked immunosorbent assay (ELISA). Comparing with the control group, ARDS model group showed typical ARDS pathological changes, which included the increased W/D ratio (4.951±0.161 vs. 3.449±0.299, P < 0.01) and the content of EB in the lung tissue (μg/g: 0.130±0.027 vs. 0.085±0.011, P < 0.01), the damaged alveolar wall structure, lung congestion and exudates in the alveoli, as well as amounts of inflammatory cells. The pathological score of lung injury (10.33±1.15 vs. 1.67±0.58) and the level of TNF-α in BALF (ng/L: 900.85±247.80 vs. 68.21±5.79) were significantly increased in the ARDS model group (both P < 0.01). Comparing with the ARDS model group, the lung W/D ratio (4.620±0.125 vs. 4.951±0.161) and the EB content in the lung tissue (μg/g: 0.108±0.011 vs. 0.130±0.027) of BaCl2 pretreatment group were significantly reduced (both P < 0.01). And the damaged pulmonary structural BaCl2 pretreatment group were significantly alleviated. In addition, the pulmonary pathological score (5.00±1.00 vs. 10.33±1.15) and the level of TNF-α in BALF (ng/L: 169.16±73.33 vs. 900.85±247.80) were significantly decreased (both P < 0.01). Barium chloride pretreatment can improve the lung histopathological changes of ARDS model mice induced by LPS by reducing the permeability of pulmonary capillaries and local inflammatory reaction.Barium chloride has the protective effect against LPS attack in mice model of ARDS.

Effect of ventilator-induced lung injury on blood-brain barrier permeability in rats

To observe the effect of ventilator-induced lung injury (VILI) on blood-brain barrier permeability in rats. Forty-eight healthy clean male Sprague-Dawley (SD) rats were randomly divided into sham operation (Sham) group, low tidal volume (LVT) mechanical ventilation group (LVT group), normal tidal volume (NVT) mechanical ventilation group (NVT group) and high tidal volume (HVT) mechanical ventilation group (HVT group) with 12 rats in each group. After anesthesia, rats in the Sham group were intubated and kept spontaneous breathing. The rats in different tidal volume (VT) groups were mechanically ventilated by endotracheal intubation with VT of 6 mL/kg (LVT group), 10 mL/kg (NVT group), and 20 mL/kg (HVT group), respectively. The inspiration-expiration ratio of the three groups was 1:1, the ventilation frequency was 40 times/min, and the ventilation time was 3 hours. At the end of the experiment, the bronchoalveolar lavage fluid (BALF) of rats was collected, and the levels of pro-inflammatory factors [tumor necrosis factor-α (TNF-α), interleukins (IL-1β and IL-6)] in BALF were detected by enzyme-linked immunosorbent assay (ELISA). The lung tissues of rats were collected, and the lung wet/dry weight (W/D) ratio was calculated. The pathological changes of lung tissues were observed under light microscopy after hematoxylin-eosin (HE) staining, and lung injury scores were performed. The brain tissue of rats was taken to measure the brain water content, and the Evans blue (EB) content of brain tissue was measured to reflect the permeability of the blood-brain barrier. The tight junction proteins in the brain tissues were detected by Western blotting. After 3 hours of mechanical ventilation, with the increase of VT, the degree of lung injury in VILI rats gradually increased. When VT reached 20 mL/kg, lung tissue structure was significantly injured, alveolar wall edema, alveolar congestion, lung interstitial thickening, a large number of inflammatory cells infiltrated, and the lung injury score, lung W/D ratio, and the levels of TNF-α, IL-1β and IL-6 in BALF were significantly higher than those in the Sham group [lung injury score: 10.6±1.1 vs. 1.4±1.0, lung W/D ratio: 6.6±0.8 vs. 3.7±0.6, TNF-α (ng/L): 832.9±97.9 vs. 103.8±23.3, IL-1β (ng/L): 68.9±14.1 vs. 15.7±2.6, IL-6 (ng/L): 70.8±16.4 vs. 20.3±5.4, all P < 0.05]. Lung injury in rats was accompanied by aggravating brain injury. When VT reached 20 mL/kg, brain water content and EB content in brain tissue were significantly higher than those in the Sham group [brain water content: (85.4±3.6)% vs. (68.7±2.7)%, EB content in brain tissue (μg/g): 887±78 vs. 97±14, both P < 0.05], and the protein expressions of claudin-5, occluding and zonula occluden-1 (ZO-1) in the brain tissue were significantly lower than those in the Sham group [claudin-5 protein (claudin-5/β-actin): 0.67±0.12 vs. 1.45±0.19, occludin protein (occludin/β-actin): 0.48±0.11 vs. 0.99±0.21, ZO-1 protein (ZO-1/β-actin): 0.13±0.03 vs. 0.63±0.12, all P < 0.05]. VILI can induce brain edema and increase blood-brain barrier permeability in rats, which may be related to the down-regulation of tight junction protein expression in the brain tissue.

Targeted Therapy of Tuberculous Meningitis in Rats with Methylprednisolone Composite Nanoparticles.

It was to investigate the targeted therapeutic effect of methylprednisolone (MPS) composite nanoparticles (NPs) on tuberculous meningitis (TBM) in rats. A total of 180 special pathogen-free (SPF) Sprague Dawley (SD) rats (male) were randomly and equally assigned to the normal control group, TBM infection group, and TBM treatment group. Those in the TBM infection group and the TBM treatment group were injected with Mycobacterium tuberculosis suspension via the tail vein. After the TBM model was established, rats in the TBM treatment group were injected intraperitoneally with methylprednisolone-nano sterically stabilized liposomes (MPS-NSSLs), and those in the normal control group were injected with an equal amount of normal saline. MPS-NSSLs were prepared, and their quality evaluation, encapsulation rate, drug-lipid ratio, and stability were detected. The particle size distribution of MPS-NSSLs was 95.4 ± 0.7 nm, showing a complete spherical structure, and the encapsulation rate was 91.24 ± 0.27 %, and the drug-lipid ratio was about 0.4. After 7 days of treatment, the water content of brain tissue in the TBM infection group was drastically superior to that in the control group (P<0.05); Evans blue (EB) content in the TBM infection group was dramatically superior to that in the control group (P<0.05). The TBM rat model was successfully established, and this model verified that MPS-NSSLs had the characteristics of high efficiency and low toxicity in the treatment of TBM rats.

Exaggerated blood pressure response to static exercise in hindlimb ischemia-reperfusion.

Peripheral artery disease (PAD) reduces the blood flow supply in the affected limbs as one of the significant cardiovascular concerns. Revascularization surgery in the femoral artery plays a central role in treating PAD. Exercise is also a rehabilitation strategy suggested for PAD patients to improve vascular functions. However, the effects of limb ischemia-reperfusion (IR), one of the most predominant complications in revascularization surgery, on exercise-induced arterial blood pressure (BP) response are poorly understood. In the present study, we determined 1) the blood flow status in the hindlimb muscles of rats (plantar muscle, red and white portions of gastrocnemius) with different time points of the hindlimb IR; and 2) the BP response to static muscle contraction in rats at different time points after the blood flow reperfusion procedure. Results of this study indicated that, compared with the Sham group, the blood flow in the hindlimb muscles evaluated by Evans blue concentration was significantly reduced at 6h of femoral artery occlusion (FAO 6h) (vs. sham control, p < 0.05). The decreased blood flow was gradually recovered after the blood flow reperfusion for 18 (IR 18h), 66 (IR 66h), and 114 (IR 114h) hours (p < 0.05 vs. FAO 6h for all IR groups). The response of mean arterial pressure was 20 ± 4mmHg in Sham rats (n = 7); 32 ± 10mmHg in IR 18h rats (n = 10); 27 ± 7mmHg in IR 66h rats (n = 13); 26 ± 4mmHg in IR 114h rats (n = 9) (p < 0.05 vs. Sham for all groups). No significant difference was observed in the peak-developed tension during muscle contraction among all the groups (p > 0.05). In conclusion, static exercise-induced BP response is exaggerated following IR. Whereas the BP response is not statistically significant but tends to decrease with a prolonged IR time, the exaggerated BP response remains through time points from post-IR 18h-114h.

Protective Effect of Nimodipine Against Valproic-acid Induced Biochemical and Behavioral Phenotypes of Autism.

Present study was designed to investigate behavioral and biochemical role of nimodipine in prenatal valproic acid (Pre-VPA) induced autism in rats. Valproic acid was utilized to induce autistic phenotypes in Wistar rats. The rats were assessed for social behavior. Hippocampus and prefrontal cortex (PFC) were utilized for various biochemical assessments, whereas cerebellum was used to assess blood brain barrier (BBB) permeability. Pre-VPA rats showed reduction social interaction. Pre-VPA administration were decreased PFC levels of interleukin- 10 (IL-10), and glutathione along with hippocampus cAMP response element-binding protein (CREB) and brain-derived neurotrophic factor (BDNF). Also, the animals have shown increase in PFC levels of IL-6, tumor necrosis factor-α, thiobarbituric acid reactive substance, Evans blue leakage and water content. Nimodipine countered Pre-VPA administered reduction in social interaction, CREB, BDNF, inflammation, oxidative stress, BBB permeability. Pre-VPA has induced autistic phenotype, which were attenuated by nimodipine in rats. Nimodipine and other calcium channel blockers should further investigate to check the management of autism.

Astragalus polysaccharide protects against blood-brain barrier damage in MCAO rats by inhibiting P2X7R channel

To investigate the protective effect of astragalus polysaccharide (APS) against blood-brain barrier in a rat model of middle cerebral artery occlusion (MCAO) and the role of P2X7R channel in the protective mechanism. In rat microglial cell models of oxygen and glucose deprivation (OGD) or ATP treatment, the formation of blood-brain barrier in vitro was assessed using the leak test, and the effect of APS on the permeability of the blood-brain barrier was determined using LC-MS. In 12 SD rats, MCAO model was established followed by treatment with intraperitoneal injection of normal saline (n= 6) or APS (45 mg/kg, n=6) for 3 consecutive days, with another 6 rats without MCAO receiving saline injections as the control group. The permeability of the blood-brain barrier of the rats was evaluated by determining Evans blue (EB) extravasation, and ATP content in the brain tissue was detected using ELISA; the expression levels of matrix metalloproteinase-9 (MMP-9) and P2X7R in the brain tissue were detected with Western blot. In the in vitro cell model of OGD or ATP treatment, APS treatment obviously promoted the repair of blood-brain barrier integrity. In the rat models, the EB content in the brain tissue and the blood-brain barrier permeability increased significantly in MCAO+saline group and MCAO+APS group as compared with those in the control group (P < 0.01). Compared with saline treatment, APS treatment significantly decreased EB content in the brain tissue and improved the blood-brain barrier permeability in the MCAO rats (P < 0.05). MCAO caused a significant reduction of ATP content and obviously increased the expression levels of MMP-9 and P2X7R in the brain tissue of the rats (P < 0.01), and these changes were significantly alleviated after APS treatment (P < 0.01 or 0.05). APS can protect the brain tissue of MCAO rats by stabilizing the internal environment, down-regulating the expression of MMP-9 and improving the permeability of blood-brain barrier under cerebral ischemia and hypoxia, and its mechanism may involve the inhibition of P2X7R channel.

Nitrate Increases Aluminum Toxicity and Accumulation in Root of Wheat

Aluminum (Al) toxicity inhibits root growth, while nitrogen is an essential nutrient for plant growth and development. To explore the effects of nitrate (N) on Al toxicity and accumulation in root of wheat, two wheat genotypes, Shengxuan 6 hao (SX6, Al-tolerant genotype) and Zhenmai 168 (ZM168, Al-sensitive genotype), were used in a hydroponic experiment with four treatments (control without N or Al, N, Al, and Al+N, respectively). The results showed that N increased the inhibition of root elongation and aluminum accumulation in root. The Al-sensitive genotype suffered more serious Al toxicity than the Al-tolerant genotype. Histochemical observation clearly showed that Al prefers binding on the root apex 7–10 mm zones, and the Al-sensitive genotype accumulated more Al in these zones. Compared with other treatments, the Al+N treatment had significantly higher O2−, superoxides dismutase (SOD), catalase (CAT), peroxidase (POD) activities, H2O2, Evans blue uptake, malondialdehyde (MDA), ascorbic acid (AsA), pectin, and hemicellulose 1 (HC1) contents in both genotypes. Under Al+N treatment, O2− activity, Evans blue uptake, MDA, and HC1 contents of SX6 were significantly lower than those of ZM168, but SOD, CAT, and POD activities and AsA content exhibited an opposite trend. Therefore, aluminum toxicity and accumulation in root of wheat seedlings were aggravated by nitrate.