Betulinic acidのラット静脈虚血進行抑制効果について

Background/Objectives: Betulinic acid is a component of a Chinese traditional herb that upregulates endothelial nitric oxide (NO) synthase (eNOS) and reduces NADPH expression. Increased production of reactive oxygen species (ROS) and reactive nitrogen species (RNS) after cerebral arterial ischemia is a major cause of neuronal damage. Local venous ischemia can arise that slowly damages surrounding brain tissue during surgical procedures. The present study investigated the effects on infarct size when betulinic acid was administered after inducing two-vein occlusion (2VO) with a slowly developing lesion in rats. Methods: We elicited 2VO in 18 male Wistar rats by the photochemical thrombosis of two adjacent cortical veins combined with KCL-induced cortical spreading depression (CSD). The rats were then randomized into groups (n=9 each) to receive either dimethyl sulfoxide (DMSO) vehicle (control) or betulinic acid (30mg/kg/day; n=9) by daily gavage for seven days, and then infarct volume and 3-nitrotyrosine expression were assessed. Results: Daily administration of betulinic acid for seven days significantly reduced infarct volume from 3.81 ± 0.7 to 1.90 ± 0.3mm³ (p=0.017). Physiological data and regional cerebral blood flow did not significantly differ between the two groups during the study. We found 3-nitrotyrosine expression within the border zone of the infarct area and cleaved caspase-3 only within the lesion at the ipsilateral hemisphere in both groups. However, 3-nitrotyrosine/caspase-3 expression did not significantly differ at any time between the groups. Conclusions: Betulinic acid reduced neuronal damage in a rat model of cerebral venous ischemia.博士（医学）・乙第1460号・令和2年6月30

Traumatic brain injury enhances the formation of heterotopic ossification around the hip : an animal model study

Introduction: The incidence of heterotopic ossification (HO) is at its highest when trauma of the hip or pelvis concurs with traumatic brain injury (TBI). The pathogenic mechanisms underlying the neurogenic enhancement of the formation of HO remain, however, poorly understood. Hence, the goal of the present study was to develop a novel small animal model that combines hip and brain trauma that can prove the enhancement of HO around the hip after TBI. Materials and methods: Forty male Wistar rats were divided into four groups, to undergo hip surgery alone (group 1), hip surgery + moderate TBI (group 2), hip surgery + severe TBI (group 3) and only severe TBI (group 4). The femoral canal was reamed up to 2 mm and a muscle lesion was made to simulate hip surgery. An established controlled cortical impact model was used to create a TBI. Twelve weeks after surgery, the hip with the proximal half of the femur and the pelvic bone was removed and subjected to micro-computed tomography (µCT) analysis. A quantitative analysis using a modified Brooker score as well as a quantitative analysis using a bone-to-tissue ratio was used. Results: No HO could be found in all the ten animals that did not undergo hip surgery (group 4). In the animals that did undergo surgery to the hip, no HO was found in only one animal (group 1). All the other animals developed HO. In this study, significantly more HO was found in animals that underwent an additional severe TBI. Conclusion: The newly developed rat model, with a combined hip and brain trauma, showed an enhancement of the HO formation around the hip after severe TBI

In-vivo time course of organ uptake and blood-brain-barrier permeation of poly(L-lactide) and poly(perfluorodecyl acrylate) nanoparticles with different surface properties in unharmed and brain-traumatized rats

Background: Traumatic brain injury (TBI) has a dramatic impact on mortality and quality of life and the development of effective treatment strategies is of great socio-economic relevance. A growing interest exists in using polymeric nanoparticles (NPs) as carriers across the blood-brain barrier (BBB) for potentially effective drugs in TBI. However, the effect of NP material and type of surfactant on their distribution within organs, the amount of the administrated dose that reaches the brain parenchyma in areas with intact and opened BBB after trauma, and a possible elicited inflammatory response are still to be clarified. Methods: The organ distribution, BBB permeation and eventual inflammatory activation of polysorbate-80 (Tw80) and sodiumdodecylsulfate (SDS) stabilized poly(L-lactide) (PLLA) and poly(perfluorodecyl acrylate) (PFDL) nanoparticles were evaluated in rats after intravenous administration. The NP uptake into the brain was assessed under intact conditions and after controlled cortical impact (CCI). Results: A significantly higher NP uptake at 4 and 24 h after injection was observed in the liver and spleen, followed by the brain and kidney, with minimal concentrations in the lungs and heart for all NPs. A significant increase of NP uptake at 4 and 24 h after CCI was observed within the traumatized hemisphere, especially in the perilesional area, but NPs were still found in areas away from the injury site and the contralateral hemisphere. NPs were internalized in brain capillary endothelial cells, neurons, astrocytes, and microglia. Immunohistochemical staining against GFAP, Iba1, TNFα, and IL1β demonstrated no glial activation or neuroinflammatory changes. Conclusions: Tw80 and SDS coated biodegradable PLLA and non-biodegradable PFDL NPs reach the brain parenchyma with and without compromised BBB by TBI, even though a high amount of NPs are retained in the liver and spleen. No inflammatory reaction is elicited by these NPs within 24 h after injection. Thus, these NPs could be considered as potentially effective carriers or markers of newly developed drugs with low or even no BBB permeation

The neuroprotective effect of a new serotonin receptor agonist, BAY X3702, upon focal ischemic brain damage caused by acute subdural hematoma in the rat

We tested the neuroprotective effect of a novel, high affinity serotonin (5-HT 1A) agonist, BAY X3702, in a rat model of acute subdural hematoma (ASDH). Animals were treated with 0.01 mg/kg ( n=8), 0.003 mg/kg ( n=8) BAY X3702 or vehicle ( n=4) 15 min before (i.v.) and after (continuous infusion) injection of 400 μl of autologous blood into the subdural space. The ischemic brain damage at 4 h after ASDH was 59.01±39 and 60.8±49 mm 3 for the low- and high-dose BAY X3702 group, respectively, which was significantly smaller compared to the vehicle-treated ASDH group (106.2±33 mm 3). The result indicates that this novel, high affinity 5-HT 1A agonist, BAY X3702, is neuroprotective in this model

Influence of inspired oxygen on glucose-lactate dynamics after subdural hematoma in the rat

The mechanisms causing brain damage after acute subdural hematoma (SDH) are poorly understood. A decrease in cerebral blood flow develops immediately after the hematoma forms, thus reducing cerebral oxygenation. This in turn may activate mitochondrial failure and tissue damage leading to ionic imbalance and possibly to cellular breakdown. The purpose of this study was to test whether a simple therapeutic measure, namely increased fraction of inspired oxygen (FiO2 100), and hence increased arterial and brain tissue oxygen tension, can influence brain glucose and lactate dynamics acutely after subdural hematoma in the rat. Twenty-five male Sprague-Dawley anesthetized rats were studied before, during and after induction of the SDH in two separate groups. The Oxygen group (n = 10) was ventilated with 100% oxygen immediately after induction of the SDH. The Air group (n = 10) was ventilated during the entire study with 21% oxygen. Brain microdialysate samples were analyzed for glucose and lactate. All rats were monitored with femoral arterial blood pressure catheters, arterial blood gas analysis, arterial glucose, lactate and end tidal CO2 (EtCO2). Five male Sprague-Dawley rats were sham operated to measure the effect of oxygen challenge on glucose-lactate dynamics without injury. Arterial oxygen tension in the Oxygen group was 371 +/- 30 mmHg and was associated with significantly greater increase in dialysate lactate in the first 30 min after induction of SDH. Dialysate glucose initially dropped in both groups, after SDH, but then reverted significantly faster to values above baseline in the Oxygen group. Changes in ventilatory parameters had no significant effect on dialysate glucose and lactate parameters in the sham group. Extracellular dialysate lactate and glucose are influenced by administration of 100% O2 after SDH. Dialysate glucose normalizes significantly quicker upon 100% oxygen ventilation. We hypothesize that increased neural tissue oxygen tension, in presence of reduced regional CBF, and possibly compromised mitochondrial function, after acute SDH results in upregulation of rate-limiting enzyme systems responsible for both glycolytic and aerobic metabolism. Similar changes have been seen in severe human head injury, and suggest that a simple therapeutic measure, such as early ventilation with 100% O2, may improve cerebral energy metabolism, early after SDH. Further studies to measure the generation of adenosine triphosphate (ATP) are needed to validate the hypothesis

The Importance of Thrombin in Cerebral Injury and Disease

There is increasing evidence that prothrombin and its active derivative thrombin are expressed locally in the central nervous system. So far, little is known about the physiological and pathophysiological functions exerted by thrombin in the human brain. Extra-hepatic prothrombin expression has been identified in neuronal cells and astrocytes via mRNA measurement. The actual amount of brain derived prothrombin is expected to be 1% or less compared to that in the liver. The role in brain injury depends upon its concentration, as higher amounts cause neuroinflammation and apoptosis, while lower concentrations might even be cytoprotective. Its involvement in numerous diseases like Alzheimer’s, multiple sclerosis, cerebral ischemia and haemorrhage is becoming increasingly clear. This review focuses on elucidation of the cerebral thrombin expression, local generation and its role in injury and disease of the central nervous system

The role of mitochondrial transition pore, and its modulation, in traumatic brain injury and delayed neurodegeneration after TBI

Following severe traumatic brain injury (TBI), a complex interplay of pathomechanism, such as exitotoxicity, oxidative stress, inflammatory events, and mitochondrial dysfunction occurs. This leads to a cascade of neuronal and axonal pathologies, which ultimately lead to axonal failure, neuronal energy metabolic failure, and neuronal death, which in turn determine patient outcome. For mild and moderate TBI, the pathomechanism is similar but much less frequent and ischemic cell death is unusual, except with mass lesions. Involvement of mitochondria in acute post-traumatic neurodegeneration has been extensively studied during the last decade, and there are a number of investigations implicating the activation of the mitochondrial permeability transition pore (mPTP) as a “critical switch” which determines cell survival after TBI. Opening of the mPTP is modulated by several factors occurring after a severe brain injury. Modern neuroprotective strategies for prevention of the neuropathological squeal of traumatic brain injury have now begun to address the issue of mitochondrial dysfunction, and drugs that protect mitochondrial viability and prevent apoptotic cascade induced by mPTP opening are about to begin phase II and III clinical trials. Cyclosporin A, which has been reported to block the opening of mPTP, showed a significant decrease in mitochondrial damage and intra-axonal cytoskeletal destruction thereby protecting the axonal shaft and blunting axotomy. This review addresses an important issue of mPT activation after severe head injury, its role in acute post-traumatic neurodegeneration, and the rationale for targeting the mPTP in experimental and clinical TBI studies

Lactate and glucose as energy substrates and their role in traumatic brain injury and therapy

Traumatic brain injury is a leading cause of disability and mortality worldwide, but no new pharmacological treatments are clinically available. A key pathophysiological development in the understanding of traumatic brain injury is the energy crisis derived from decreased cerebral blood flow, increased energy demand and mitochondrial dysfunction. Although still controversial, new findings suggest that brain cells try to cope in these conditions by metabolizing lactate as an energy substrate '‘on-demand ’ in lieu of glucose. Experimental and clinical data suggest that lactate, at least when exogenously administered, is transported from astrocytes to neurons for neuronal utilization, essentially bypassing the slow, catabolizing glycolysis process to quickly and efficiently produce ATP. Treatment strategies using systemically applied lactate have proved to be protective in various experimental traumatic brain injury studies. However, lactate has the potential to elevate oxygen consumption to high levels and, therefore, could potentially impose a danger for tissue-at-risk with low cerebral blood flow. The present review outlines the experimental basis of lactate in energy metabolism under physiological and pathophysiological conditions and presents arguments for lactate as a new therapeutical tool in human head injury