Rat Model Of Hemorrhage Research Articles

Label-free imaging of hemoglobin degradation and hemosiderin formation in brain tissues with femtosecond pump-probe microscopy.

The degradation of hemoglobin in brain tissues results in the deposition of hemosiderin, which is a major form of iron-storage protein and closely related to neurological disorders such as epilepsy. Optical detection of hemosiderin is vitally important yet challenging for the understanding of disease mechanisms, as well as improving surgical resection of brain lesions. Here, we provide the first label-free microscopy study of sensitive hemosiderin detection in both an animal model and human brain tissues.Methods: We applied spectrally and temporally resolved femtosecond pump-probe microscopy, including transient absorption (TA) and stimulated Raman scattering (SRS) techniques, to differentiate hemoglobin and hemosiderin in brain tissues. The label-free imaging results were compared with Perls' staining to evaluate our method for hemosiderin detection.Results: Significant differences between hemoglobin and hemosiderin transient spectra were discovered. While a strong ground-state bleaching feature of hemoglobin appears in the near-infrared region, hemosiderin demonstrates pure excited-state absorption dynamics, which could be explained by our proposed kinetic model. Furthermore, simultaneous imaging of hemoglobin and hemosiderin can be rapidly achieved in both an intracerebral hemorrhage (ICH) rat model and human brain surgical specimens, with perfect correlation with Perls' staining.Conclusion: Our results suggest that rapid, label-free detection of hemosiderin in brain tissues could be realized by femtosecond pump-probe microscopy. Our method holds great potential in providing a new tool for intraoperative detection of hemosiderin during brain surgeries.

Activation of signal transducer and activator of transcription 3 signaling attenuates neurogenesis in intracerebral hemorrhage rat model

Activation of signal transducer and activator of transcription 3 signaling attenuates neurogenesis in intracerebral hemorrhage rat model

Acupuncture through Baihui (DU20) to Qubin (GB7) mitigates neurological impairment after intracerebral hemorrhage.

Inflammation plays an important role in nerve defects caused by intracerebral hemorrhage. Repairing brain damage by inhibiting the macrophage-inducible C-type lectin/spleen tyrosine kinase (Mincle/Syk) signaling pathway is a potential new target for treating cerebral hemorrhage. In this study, we aimed to determine whether acupuncture through Baihui (DU20) to Qubin (GB7) is an effective treatment for intracerebral hemorrhage through the Mincle/Syk signaling pathway. An intracerebral hemorrhage rat model was established by autologous blood infusion into the caudate nucleus. Acupuncture through Baihui to Qubin was performed for 30 minutes, once every 12 hours, for a total of three times. Piceatannol (34.62 mg/kg), a Syk inhibitor, was intraperitoneally injected as a control. Modified neurological severity score was used to assess neurological function. Brain water content was measured. Immunohistochemistry and western blot assay were used to detect immunoreactivity and protein expression levels of Mincle, Syk, and CARD9. Real-time polymerase chain reaction was used to determine interleukin-1β mRNA levels. Hematoxylin-eosin staining was performed to observe histopathological changes. Our results showed that acupuncture through Baihui to Qubin remarkably improved neurological function and brain water content, and inhibited immunoreactivity and expression of Mincle, Syk, CARD9, and interkeukin-1β. Moreover, this effect was similar to piceatannol. These findings suggest that acupuncture through Baihui to Qubin can improve neurological impairment after cerebral hemorrhage by inhibiting the Mincle/Syk signaling pathway.

Nephroprotection through the Akt/eNOS pathway by centrally administered erythropoietin in a rat model of fixed-volume hemorrhage

AimsThis study was designed to investigate the protective effects of centrally administered erythropoietin (EPO) on brain oxidative stress and inflammatory markers to protect the kidneys during hemorrhagic shock (HS). Main methodsAnimals were assigned into three groups (n=6). Sham rats were subjected to cannulation of femoral artery and vein as well as stereotaxic surgery. In HS group, 50% of total blood volume was withdrawn and resuscitation was started 2h later. In EPO group, stereotaxic surgery in lateral ventricle was performed one week before induction of HS for administration of EPO (2IU) just before resuscitation. Plasma samples, kidney and brain tissues were allocated after a further 3h in all animals. Key findingsThere was a significant increase in survival rate in the EPO group (69.3%) compared to the HS group (35.7%). Brain EPO administration significantly attenuated the rises in BUN, plasma Cr and NGAL, brain and renal MDA content and also increased SOD activity in the kidney and brain compared to the HS group. Brain, plasma and kidney TNF-α and IL-6 levels were significantly reduced by EPO compared to HS group. EPO increased the phosphorylation of Akt on Ser473 and eNOS mRNA expression in the kidney tissue compared to the HS group. SignificanceIn conclusion, centrally administered EPO reduced pro-inflammatory and oxidative stress indices in the kidney and reduced apoptosis by activation of the Akt/eNOS signaling pathway. Hence, it can be hypothesized that EPO may play a major role in the central regulation of renal system as a neuromodulator.

Glycyrrhizin suppresses the expressions of HMGB1 and ameliorates inflammative effect after acute subarachnoid hemorrhage in rat model

High-mobility group box 1 (HMGB1), a nuclear protein that has endogenous cytokine-like activity, is involved in early brain injury after subarachnoid hemorrhage (SAH) by mediating inflammatory response. This study was conducted to investigate the effect of glycyrrhizin as an inhibitor of HMGB1 in a rat SAH model. Experimental SAH was induced by using autologous blood injection to prechiasmatic cistern. 15 mg/kg glycyrrhizin was administered immediately after SAH induction, and then administered once at 6, 12 and 18 h. All the rats were sacrificed at 24 h after neurological assessment and frontal brain tissue was taken for assay. Blood–brain barrier (BBB) permeability was determined by Evans blue (EB) extravasation. The expression of HMGB1 were detected by immunofluorescence, western blot and quantitative real-time PCR. Inflammatory mediators (TNF-α, IL-1β) were measured using specific ELISA. Fluoro-Jade C staining and TUNEL staining was performed for the quantitative assessment of neuronal injury. We found the use of glycyrrhizin significantly improved neurological scores, reduced HMGB1-positive cells, down-regulated mRNA and protein levels of HMGB1, inhibited BBB permeability, and attenuated neuronal cell death and apoptosis after SAH. The up-regulations of inflammation-related molecules (TNF-α, IL-1β) in SAH rats were suppressed by glycyrrhizin treatment. These findings suggest that glycyrrhizin is a potential candidate for the treatment of inflammatory brain injury after SAH.

Spatial-temporal expression of NDRG2 in brain tissues in a rat model of intracerebral hemorrhage: A pilot study.

N-myc downstream regulated gene 2 (NDRG2) was a member of the N-myc down regulated gene family which belongs to the alpha/beta hydrolase superfamily and played important roles in cell death. To date, the expression and effects of NDRG2 in brain after intracerebral hemorrhage (ICH) are unclear. In this study, we investigated the spatial-temporal expression of NDRG2 in brain tissues in a rat model of ICH. The expression levels of NDRG2 were tested in 3h, 6h, 12h, 24h, 48h, 72h, and 7d after ICH by western blot analysis. The results showed that the NDRG2 levels were increased and peaked at 24h after ICH, and then declined subsequently. Meanwhile, we also examined the NDRG2 cellular localization in brain tissues by immunofluorescence analysis with NeuN and GFAP (biomarker of neuron and astrocytes respectively). The results demonstrated that NDRG2 was mainly expressed in astrocytes, but not neurons, after ICH. Additionally, the results of double staining indicated that the rate of NDRG2- and TUNEL -positive cells was significantly higher in the brain tissues in rats after ICH. The roles of NDRG2 in ICH needed further investigation and inhibiting the expression of NDRG2 may have potential therapeutic effects in ICH.

Xenon Reduces Neuronal Hippocampal Damage and Alters the Pattern of Microglial Activation after Experimental Subarachnoid Hemorrhage: A Randomized Controlled Animal Trial.

The neuroprotective properties of the noble gas xenon have already been demonstrated using a variety of injury models. Here, we examine for the first time xenon's possible effect in attenuating early brain injury (EBI) and its influence on posthemorrhagic microglial neuroinflammation in an in vivo rat model of subarachnoid hemorrhage (SAH). Sprague-Dawley rats (n = 22) were randomly assigned to receive either Sham surgery (n = 9; divided into two groups) or SAH induction via endovascular perforation (n = 13, divided into two groups). Of those randomized for SAH, 7 animals were postoperatively ventilated with 50 vol% oxygen/50 vol% xenon for 1 h and 6 received 50 vol% oxygen/50 vol% nitrogen (control). The animals were sacrificed 24 h after SAH. Of each animal, a cerebral coronal section (-3.60 mm from bregma) was selected for assessment of histological damage 24 h after SAH. A 5-point neurohistopathological severity score was applied to assess neuronal cell damage in H&E and NeuN stained sections in a total of four predefined anatomical regions of interest. Microglial activation was evaluated by a software-assisted cell count of Iba-1 stained slices in three cortical regions of interest. A diffuse cellular damage was apparent in all regions of the ipsilateral hippocampus 24 h after SAH. Xenon-treated animals presented with a milder damage after SAH. This effect was found to be particularly pronounced in the medial regions of the hippocampus, CA3 (p = 0.040), and dentate gyrus (DG p = 0.040). However, for the CA1 and CA2 regions, there were no statistical differences in neuronal damage according to our histological scoring. A cell count of activated microglia was lower in the cortex of xenon-treated animals. This difference was especially apparent in the left piriform cortex (p = 0.017). In animals treated with 50 vol% xenon (for 1 h) after SAH, a less pronounced neuronal damage was observed for the ipsilateral hippocampal regions CA3 and DG, when compared to the control group. In xenon-treated animals, a lower microglial cell count was observed suggesting an immunomodulatory effect generated by xenon. As for now, these results cannot be generalized as only some hippocampal regions are affected. Future studies should assess the time and localization dependency of xenon's beneficial properties after SAH.

5-HT1a activation in PO/AH area induces therapeutic hypothermia in a rat model of intracerebral hemorrhage.

Therapeutic hypothermia is widely applied as a neuroprotective measure on intracerebral hemorrhage (ICH). However, several clinical trials regarding physical hypothermia encountered successive failures because of its side-effects in recent years. Increasing evidences indicate that chemical hypothermia that targets hypothalamic 5-HT1a has potential to down-regulate temperature set point without major side-effects. Thus, this study examined the efficacy and safety of 5-HT1a stimulation in PO/AH area for treating ICH rats. First, the relationship between head temperature and clinical outcomes was investigated in ICH patients and rat models, respectively. Second, the expression and distribution of 5-HT1a receptor in PO/AH area was explored by using whole-cell patch and confocal microscopy. In the meantime, the whole-cell patch was subsequently applied to investigate the involvement of 5-HT1a receptors in temperature regulation. Third, we compared the efficacy between traditional PH and 5-HT1a activation-induced hypothermia for ICH rats. Our data showed that more severe perihematomal edema (PHE) and neurological deficits was associated with increased head temperature following ICH. 5-HT1a receptor was located on warm-sensitive neurons in PO/AH area and 8-OH-DPAT (5-HT1a receptor agonist) significantly enhanced the firing rate of warm-sensitive neurons. 8-OH-DPAT treatment provided a steadier reduction in brain temperature without a withdrawal rebound, which also exhibited a superior neuroprotective effect on ICH-induced neurological dysfunction, white matter injury and BBB damage compared with physical hypothermia. These findings suggest that chemical hypothermia targeting 5-HT1a receptor in PO/AH area could act as a novel therapeutic manner against ICH, which may provide a breakthrough for therapeutic hypothermia.

ErbB4 protects against neuronal apoptosis via activation of YAP/PIK3CB signaling pathway in a rat model of subarachnoid hemorrhage

Neuronal apoptosis is a central pathological process in subarachnoid hemorrhage (SAH)-induced early brain injury. Previous studies indicated that ErbB4 (EGFR family member v-erb-b2 avian erythroblastic leukemia viral oncogene homolog 4) is essential for normal development and maintenance of the nervous system. In this study, we explored the neuroprotective effects of ErbB4 and its downstream YAP (yes-associated protein)/PIK3CB signaling pathway in early brain injury after SAH in a rat model using the endovascular perforation method. Rats were neurologically evaluated with the Modified Garcia Scale and beam balance test at 24h and 72h after SAH. An ErbB4 activator Neuregulin 1β1 (Nrg 1β1), ErbB4 siRNA and YAP siRNA were used to explore this pathway. The expression of p-ErbB4 and YAP was significantly increased after SAH. Multiple immunofluorescence labeling experiments demonstrated that ErbB4 is mainly expressed in neurons. Activation of ErbB4 and its downstream signals improved the neurological deficits after SAH and significantly reduced neuronal cell death. Inhibition of ErbB4 reduced YAP and PIK3CB expression, and aggravated cell apoptosis. YAP knockdown reduced the PIK3CB level and eliminated the anti-apoptotic effects of ErbB4 activation. These findings indicated that ErbB4 plays a neuroprotective role in early brain injury after SAH, possibly via the YAP/PIK3CB signaling pathway.

RETRACTED: Effect of microRNA-129-5p targeting HMGB1-RAGE signaling pathway on revascularization in a collagenase-induced intracerebral hemorrhage rat model

This article has been retracted: please see Elsevier Policy on Article Withdrawal (http://www.elsevier.com/locate/withdrawalpolicy). This article has been retracted at the request of the Editor-in-Chief. An Expression of Concern for this article was previously published while an investigation was conducted (see related editorial: https://doi.org/10.1016/j.biopha.2022.113812). This retraction notice supersedes the Expression of Concern published earlier. Concern was raised about the reliability of the Western blot data in Figure 2A, which contain suspected image duplications within the β-actin blot, and appear to represent a distinct phenotype as found in many other publications, as detailed here: https://pubpeer.com/publications/83FD53A8F4C5B60E2187CBF9F29B01; and here: https://docs.google.com/spreadsheets/d/1r0MyIYpagBc58BRF9c3luWNlCX8VUvUuPyYYXzxWvgY/edit#gid=262337249. Independent analysis confirmed these findings and also identified additional suspected image duplications within Figures 3 and 4A. The journal requested the corresponding author comment on these concerns and provide the associated raw data. The authors did not respond to this request and therefore the Editor-in-Chief decided to retract the article.

3% NaCl adenosine, lidocaine, Mg2+ (ALM) bolus and 4 hours "drip" infusion reduces noncompressible hemorrhage by 60% in a rat model.

Noncompressible torso hemorrhage is the leading cause of potentially survivable trauma in far-forward combat environments. Our aim was to examine the effect of small-volume 3% NaCl adenosine, lidocaine, and Mg (ALM) bolus and 0.9% NaCl/ALM "drip" on survivability and cardiac/gut/kidney function in a rat model of hepatic hemorrhage and shock. Male Sprague-Dawley rats (428 ± 4 g) were anesthetized and randomly assigned to one of five groups (n = 16): (1) Sham, (2) No treatment, (3) Saline controls, (4) ALM therapy, and (5) Hextend. Animals were ventilated, instrumented with single or double laparotomy for tissue probe insertion, and hemorrhage induced by liver resection. After 15 minutes, a single 3% NaCl ± ALM bolus (0.7 ml/kg) was injected IV (phase 1) and after 60 minutes, 4 hours 0.9% NaCl ± ALM stabilization "drip" (0.5 ml/kg/h) was administered (phase 2), with 1-hour monitoring. Mortality for Shams (no resection) was 0% (25%); No treatment, 87.5% (100%); Saline controls, 37.5% (75%); ALM therapy, 0% (25%), and Hextend, 87.5% (100%) (double laparotomy in parentheses). Hextend-treated animals died during the first 20 minutes of phase 2. A single ALM bolus during phase 1 led to a 2.4-fold higher cardiac output and improved hemodynamics. 3% NaCl ALM bolus increased tissue pO2 and flow in gut and kidney during phase 1 and, during ALM "drip" in phase 2, tissue pO2 decreased but flow continued to rise, indicating increased tissue O2 extraction and delivery. During phase 2, CO, ejection fraction, and fractional shortening decreased and were erratic in all groups except ALM treatment. ALM therapy led to up to 60% less bleeding over 6 hours compared to Saline controls and 75% less bleeding than Hextend. Small-volume ALM therapy significantly reduced mortality and internal bleeding compared to Saline controls or Hextend-treated rats. Hextend increased mortality, severe bleeding, and microvascular-organ injury.

CM352 Reduces Brain Damage and Improves Functional Recovery in a Rat Model of Intracerebral Hemorrhage.

BackgroundIntracerebral hemorrhage (ICH) is an acute neurological disorder with high mortality and no effective treatment. In addition to the initial bleeding event, rebleeding and hematoma expansion are associated with poor outcome in these patients. We studied the effectiveness of the new antifibrinolytic agent CM352, a short‐half‐life matrix metalloproteinase inhibitor, for achieving early hemostasis and improving functional recovery in a rat model of collagenase‐induced ICH.Methods and Results ICH was induced by striatal injection of collagenase, and 1 hour later, rats received an intravenous injection of saline (n=6) or CM352 (1 mg/kg, n=6). Hematoma (basal and after 3 and 24 hours) and lesion (14 days) volumes were quantified on T2‐weighted (T2) magnetic resonance images. Neurological and functional recovery was evaluated by using Bederson score and a cylinder test (basal, 24 hours, and 14 days). Early treatment (1 hour) with CM352 was efficient reducing hematoma expansion at 3 hours (P<0.01) and, more markedly, at 24 hours (P<0.01). Decreased bleeding after antifibrinolytic treatment was accompanied by reduced interleukin‐6 levels at 3 hours (P<0.05) and smaller lesion volume at 14 days (P<0.01). CM352 drastically reduced sensorimotor impairment (cylinder test) after ICH in rats at 24 hours (P<0.01) and 14 days (P<0.01). Similarly, it also attenuated neurological deficit (Bederson scale) at 24 hours (P<0.01) and 14 days (P<0.01). Interestingly, late (3 hours) CM352 administration also resulted in reduced lesion size and better functional outcome.Conclusions CM352, a new antifibrinolytic agent and matrix metalloproteinase inhibitor, effectively prevented hematoma growth and reduced lesion size in ICH in association with improved functional and neurological recovery.

Roflumilast Reduces Cerebral Inflammation in a Rat Model of Experimental Subarachnoid Hemorrhage.

Roflumilast, a selective inhibitor for PDE4, is approved by FDA as an anti-inflammation drug for treatment of chronic obstructive pulmonary disease (COPD). This study investigates the effects of roflumilast on cerebral inflammation in the rat SAH model. Here, we show that subcutaneous administration of roflumilast (3mg/kg) significantly improved the neurological deficits. Measurement of evans blue extravasation and brain water content revealed a significant reduction of blood-brain barrier permeability and brain edema. Importantly, roflumilast treatment remarkably decreased levels of IL-1β, IL-6, and TNF-α and the number of apoptotic neurons in the brain after SAH. These results indicate that roflumilast is effective in treating cerebral inflammation following SAH.

Electroacupuncture improves neurobehavioral function and brain injury in rat model of intracerebral hemorrhage.

Acupuncture has been widely used as a treatment for stroke in China for a long time. Recently, studies have demonstrated that electroacupuncture (EA) can accelerate intracerebral hemorrhage (ICH)-induced angiogenesis in rats. In the present study, we investigated the effect of EA on neurobehavioral function and brain injury in ICH rats. ICH was induced by stereotactic injection of collagenase type I and heparin into the right caudate putamen. Adult ICH rats were randomly divided into the following three groups: model control group (MC), EA at non-acupoint points group (non-acupoint EA) and EA at Baihui and Dazhui acupoints group (EA). The neurobehavioral deficits of ICH rats were assessed by modified neurological severity score (mNSS) and gait analysis. The hemorrhage volume and glucose metabolism of hemorrhagic foci were detected by PET/CT. The expression levels of MBP, NSE and S100-B proteins in serum were tested by ELISA. The histopathological features were examined by haematoxylin-eosin (H&E) staining. Apoptosis-associated proteins in the perihematomal region were observed by immunohistochemistry. EA treatment significantly promoted the recovery of neurobehavioral function in ICH rats. Hemorrhage volume reduced in EA group at day 14 when compared with MC and non-acupoint EA groups. ELISA showed that the levels of MBP, NSE and S100-B in serum were all down-regulated by EA treatment. The brain tissue of ICH rat in the EA group was more intact and compact than that in the MC and non-acupoint groups. In the perihematomal regions, the expression of Bcl-2 protein increased and expressions of Caspase-3 and Bax proteins decreased in the EA group vs MC and non-acupoint EA groups. Our data suggest that EA treatment can improve neurobehavioral function and brain injury, which were likely connected with the absorption of hematoma and regulation of apoptosis-related proteins.

Tranexamic Acid Attenuates The Loss of Lung Barrier Function in a Rat Model of Polytrauma And Hemorrhage With Resuscitation.

Severe trauma, hemorrhage, and resuscitation can lead to a trauma-related acute lung injury that involves rapid infiltration of immune cells and platelets. This infiltration involves exymatic degradation of matrix proteins, including plasmin, and causes loss of barrier function. Since tranexamic acid (TXA) inhibits plasminogen/ plasmin binding to target substrates, it may attenuate loss of barrier function after severe trauma, hemorrhage, and resuscitation. Sprague-Dawley rats were subjected to polytrauma (laparotomy, and trauma to intestines, liver, right leg skeletal muscle, and right femur fracture), then bled 40% of their blood volume. One hour after completion of polytrauma and hemorrhage, resuscitation was begun with fresh whole blood (FWB) or FWB with prior bolus administration of TXA (10 mg/kg in 0.2 mL). Polytrauma, hemorrhage, and resuscitation with FWB led to an elevation in lung water content that was significantly reduced with TXA administration. Polytrauma and hemorrhage led to rise in the number of neutrophils/monocytes and platelets in the lungs, and a rise in myeloperoxidase (MPO), neutrophil elastase and complement C5a content. While resuscitation with FWB significantly reduced the cellular infiltrate and MPO, FWB/TXA further reduced the levels of neutrophil/monocytes, neutrophil elastase, and complement C5a. Polytrauma and hemorrhage led to rise in lung plasmin activity that was significantly reduced with either FWB or FWB/TXA resuscitation. Severe trauma and hemorrhage leads to increases in lung water content, and immune cell, platelets, MPO, elastase, and C5a content in lung tissue, all markers of inflammation and acute lung injury. The addition of TXA to FWB resuscitation markedly attenuated the rise in these parameters suggesting its utility in treating acute lung injury.

Tissue Inflammatory Responses in an Anesthetized Rat Hemorrhage Model Associated with Endothelial Dysfunction

We have demonstrated endothelial dysfunction such as reduction in glycocalyx thickness and increased permeability in venules in cremaster muscle following hemorrhage and fluid resuscitation with a crystalloid or a colloid in a rat model. The present study further characterizes the model by measuring indices of inflammation and oxidative and nitrosative stress in tissues from these animals. Anesthetized Sprague‐Dawley rats (n=4–5/group) were bled 40% of their blood volume, subjected to a 30 min shock period and resuscitated with limited volumes of normal saline (NS) or 5% albumin (A), or no treatment (Hem). At 1hr after resuscitation, rats were euthanized and liver, kidney, lung and skeletal muscle were collected and assayed for cytokines and select indices of nitrosative and oxidative stress. In liver and kidney, IL‐1β, IL‐6 and TNF‐α levels were 2–3‐fold higher in the NS than the A group and about 2‐fold higher than the Hem group. IL‐10 levels were also significantly higher in the NS group than the other groups. Similar responses were observed in lung from these groups, while in muscle, only IL‐1β and IL‐6 levels were significantly higher. In contrast, no significant differences were observed in all tissues in nitric oxide, total antioxidant or reduced glutathione concentrations nor xanthine oxidase activity among the groups. In this model of hemorrhagic shock, NS induced the highest inflammatory response, while 5% albumin was associated with the least response. Interestingly, these current observations were related to the degree of endothelial dysfunction observed previously. Taken together these data suggest that the endothelial dysfunction observed in hemorrhagic shock and fluid resuscitation is associated with an inflammatory response without a major nitrosative or oxidative stress component.Support or Funding InformationFunded by the US Army Medical Research Materiel Command

Cerebral hemorrhage therapy by targeting VEGF and HGF in a preclinical trial in rats

Cerebral hemorrhage is the most common type of human cerebrovascular disease and frequently causes paralysis, vegetative state and mortality. The modulatory actions of vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) are vital in the human nervous system. The present study investigated the association between cerebral hemorrhage and the expression of VEGF and HGF in a rat model of cerebral hemorrhage. The therapeutic potential of cerebral hemorrhage was also evaluated using targeted drugs for VEGF and HGF in the cerebral hemorrhage rat model. Behavioral and preclinical changes and the survival rates of rats were assessed after treatment with VEGF receptor (VEGFR) and HGF receptor (HGFR). The results of Tarlov scores demonstrated that movement of limbs and coordination when walking were significantly improved in moderate and severe hemorrhage lesions in the VEGFR plus HGFR‑treated group and mainly alleviated in primary hemorrhage lesions compared with rats in the single VEGFR or HGFR‑treated groups and the control group (**P<0.01). Decreasing expression levels of VEGF and HGF were observed in the neural tissue of animals treated with VEGFR plus HGFR compared with the control group (**P<0.01). These preclinical observations indicated that VEGF and HGF serve a function in the pathological injury and repair of cerebral tissue in rats with cerebral hemorrhages. The therapeutic benefits of VEGFR plus HGFR suggested that VEGFR plus HGFR may be candidate drugs for cerebral hemorrhage, and thus offer a promising treatment for clinicians and doctors.

MR-angiography allows defining severity grades of cerebral vasospasm in an experimental double blood injection subarachnoid hemorrhage model in rats.

ObjectiveMagnetic resonance (MR) imaging has been used for the detection of cerebral vasospasm (VSP) related infarction in experimental subarachnoid hemorrhage (eSAH) in rats. Conventional angiography is generally used to visualize VSP, which is an invasive technique with a possible increase in morbidity and mortality. In this study we evaluated the validity of MR-angiography (MRA) in detecting VSP and its feasibility to define VSP severity grades after eSAH in rats.MethodsSAH was induced using the double-hemorrhage model in 12 rats. In two rats, saline solution was injected instead of blood (sham group). MR was performed on day 1, 2 and on day 5. T1-, T2-, T2*-weighted and time-of-flight MR sequences were applied, which were analyzed by two blinded neuroradiologists. Vessel narrowing of 25–50% was defined as mild, 50–75% as moderate and >75% as severe VSP.ResultsWe performed a total of 34 MRAs in 14 rats. In 14 rats, MRA was performed on day 2 and day 5. In six rats MRA was additionally performed on day1 before the blood injection. A good visualization of cerebral vessels was possible in all cases. No VSP was seen in the sham group neither on day 2 nor on day 5. We found vasospasm on day 2 in 7 of the 14 rats (50%) whereas all 7 rats had mild and one rat had additionally moderate and severe vasospasm in one vessel, respectively. On day 5 we found vasospasm in 8 of the 14 rats (60%) whereas 4 rats had severe vasospasm, 1 rat had moderate vasospasm and 3 rats demonstrated mild vasospasm. In 4 of the 14 rats (30%) an ischemic lesion was detected on day 5. Three of these rats had severe vasospasm and one rat had mild vasospasm. Severe vasospasm on day 5 was statistically significant correlated with the occurrence of ischemic lesions (Fisher’s Exact test, OR 19.5, p = 0.03).ConclusionsMRA is a noninvasive diagnostic tool, which allows a good visualization of the cerebral vasculature and provides reproducible results concerning the detection of VSP and the differentiation into three severity grades in rats. Future studies are needed to directly compare MRA with conventional angiography.

Therapeutic Benefits of Mesenchymal Stromal Cells in a Rat Model of Hemoglobin-Induced Hypertensive Intracerebral Hemorrhage.

Previous studies have shown that bone marrow mesenchymal stromal cell (MSC) transplantation significantly improves the recovery of neurological function in a rat model of intracerebral hemorrhage. Potential repair mechanisms involve anti-inflammation, anti-apoptosis and angiogenesis. However, few studies have focused on the effects of MSCs on inducible nitric oxide synthase (iNOS) expression and subsequent peroxynitrite formation after hypertensive intracerebral hemorrhage (HICH). In this study, MSCs were transplanted intracerebrally into rats 6 hours after HICH. The modified neurological severity score and the modified limb placing test were used to measure behavioral outcomes. Blood–brain barrier disruption and neuronal loss were measured by zonula occludens-1 (ZO-1) and neuronal nucleus (NeuN) expression, respectively. Concomitant edema formation was evaluated by H&E staining and brain water content. The effect of MSCs treatment on neuroinflammation was analyzed by immunohistochemical analysis or polymerase chain reaction of CD68, Iba1, iNOS expression and subsequent peroxynitrite formation, and by an enzyme-linked immunosorbent assay of pro-inflammatory factors (IL-1β and TNF-α). The MSCs-treated HICH group showed better performance on behavioral scores and lower brain water content compared to controls. Moreover, the MSC injection increased NeuN and ZO-1 expression measured by immunochemistry/immunofluorescence. Furthermore, MSCs reduced not only levels of CD68, Iba1 and pro-inflammatory factors, but it also inhibited iNOS expression and peroxynitrite formation in perihematomal regions. The results suggest that intracerebral administration of MSCs accelerates neurological function recovery in HICH rats. This may result from the ability of MSCs to suppress inflammation, at least in part, by inhibiting iNOS expression and subsequent peroxynitrite formation.

Limited Resuscitation With Fresh or Stored Whole Blood Corrects Cardiovascular and Metabolic Function in a Rat Model of Polytrauma and Hemorrhage.

We have recently shown that human whole blood stored at 4°C maintains hemostatic and platelet function. In this study, we compared restoration of hemodynamic, metabolic and hemostatic function after limited resuscitation with rat fresh whole blood, rat stored whole blood, or Lactated Ringers in traumatized rats. Rat whole blood was stored for 10 days at 4°C for evaluation of hemostatic function. Polytrauma was performed on isoflurane-anesthetized Sprague-Dawley rats (350-450 g) by damage to the intestines, liver, right leg skeletal muscle, and right femur fracture, followed by 40% hemorrhage. At 1 h, rats were resuscitated (20%) with either fresh whole blood (FWB), stored whole blood, 4°C for 7 days (SWB), Lactated Ringers (LR), or nothing. Blood samples were taken before and 2 h after trauma and hemorrhage to evaluate metabolic and hemostatic function. Whole blood stored for 10 days showed a significant prolongation in prothrombin time (PT) and activated partial thromboplastin time (aPTT), and fall in fibrinogen concentration, but no change in Maximum Clot Firmness or speed of clot formation. Platelet function was maintained until day 7 in storage, than fell significantly. Polytrauma and hemorrhage in rats led to a fall in arterial pressure, plasma bicarbonate, fibrinogen, and platelet function, and a rise in plasma lactate, PT, aPTT, and creatinine. Resuscitation with either FWB or 7 day SWB, but not LR, returned arterial pressure, plasma lactate and plasma bicarbonate to levels similar to control, but had no effect on the fall in fibrinogen or platelet function, or the rise in PT, aPTT, or creatinine. Hemostatic and platelet function of rat whole blood stored at 4°C is preserved for at least 7 days in vitro. Low volume resuscitation with SWB or FWB, but not LR, restores hemodynamic and metabolic function, but not the coagulopathy after severe trauma and hemorrhage.