Search

Aims General anesthesia has been widely applied in surgical or nonsurgical medical procedures, but the mechanism behind remains elusive. Because of shared neural circuits of sleep and anesthesia, whether serotonergic system, which is highly implicated in modulation of sleep and wakefulness, regulates general anesthesia as well is worth investigating. Methods Immunostaining and fiber photometry were used to assess the neuronal activities. Electroencephalography spectra and burst‐suppression ratio (BSR) were used to measure anesthetic depth and loss or recovery of righting reflex to indicate the induction or emergence time of general anesthesia. Regulation of serotonergic system was achieved through optogenetic, chemogenetic, or pharmacological methods. Results We found that both Fos expression and calcium activity were significantly decreased during general anesthesia. Activation of 5‐HT neurons in the dorsal raphe nucleus (DRN) decreased the depth of anesthesia and facilitated the emergence from anesthesia, and inhibition deepened the anesthesia and prolonged the emergence time. Furthermore, agonism or antagonism of 5‐HT 1A or 2C receptors mimicked the effect of manipulating DRN serotonergic neurons. Conclusion Our results demonstrate that 5‐HT neurons in the DRN play a regulative role of general anesthesia, and activation of serotonergic neurons could facilitate emergence from general anesthesia partly through 5‐HT 1A and 2C receptors., General anesthesia decreases the activities of serotonergic neurons in the dorsal raphe. Activation of DRN 5‐HT neurons reduced the depth of anesthesia and accelerated the emergence from general anesthesia, partially through the 5‐HT 1A and 2C receptors.

Background The neuropeptide orexin promotes arousal from general anaesthesia, however the neuronal circuits that mediate this effect have not been defined. We investigated whether orexinergic neurones modulate the basal forebrain (BF) and locus coeruleus (LC) in emergence from anaesthesia. Methods Hcrtcre rats were generated using a CRISPR/Cas9-based approach. Viruses encoding optogenetic probes were injected into the perifornical lateral hypothalamic (PeFLH) area, optogenetic fibres were embedded in the PeFLH, BF, or LC, and changes in anaesthesia state under 1.4 vol% or 0.8 vol% isoflurane were determined. Results In the PeFLH, 98.8% (0.4%) of orexin-A-positive cells expressed tdTomato, and 91.9% (2.2%) of tdTomato cells were orexin-A-positive. Under 1.4 vol% isoflurane anaesthesia, compared with control groups, burst suppression ratio was less, and emergence time was shorter in groups with optogenetic activation of orexinergic cell bodies in the PeFLH (923 [162] vs 493 [68] s, P=0.0003) or orexinergic terminals in the BF (937 (122) vs 674 (108) s, P=0.0049) or LC (913 [128] vs 742 [76] s, P=0.022). Optical stimulation of orexinergic terminals in the BF and LC also improved the movement scores of rats under 0.8 vol% isoflurane anaesthesia. Conclusions Activation of orexinergic terminals in the FB or LC mediates facilitation of emergence from anaesthesia by orexinergic neurones during isoflurane anaesthesia.

Astrocytes experience significant metabolic shifts in the "sensitive period" of neurological function recovery following cerebral ischemia. However, the changes in astrocyte lipid metabolism and their implications for neurological recovery remain unknown. In the present study, we employed a mouse middle cerebral artery occlusion model to investigate the changes in de novo lipogenesis and interleukin-33 (IL-33) production in astrocytes and elucidate their role in blood-brain barrier (BBB) repair in the subacute phase of cerebral ischemia. Neurological behavior evaluation was used to assess functional changes in mice. Pharmacological inhibition and astrocyte-specific downregulation of fatty acid synthase (FASN) were used to evaluate the role of de novo lipogenesis in brain injury. Intracerebroventricular administration of recombinant IL-33 was performed to study the contribution of IL-33 to BBB disruption. Extravasation of Evans blue dye, dextran and IgG were used to assess BBB integrity. Western blotting of tight junction proteins ZO-1, Occludin, and Claudin-5 were performed at defined time points to evaluate changes in BBB. It was found that de novo lipogenesis was activated, and IL-33 production increased in astrocytes at the subacute stage of cerebral ischemia injury. Inhibition of lipogenesis in astrocytes decreased IL-33 production in the peri-infarct area, deteriorated BBB damage and interfered with neurological recovery. In addition, supplementation of IL-33 alleviated BBB destruction and improved neurological recovery worsened by lipogenesis inhibition. These findings indicate that astrocyte lipogenesis increases the production of IL-33 in the peri-infarct area, which promotes BBB repair in the subacute phase of cerebral ischemia injury and improves long-term functional recovery.

Background Orexin can facilitate emergence after general anaesthesia via multiple neural pathways. Dopaminergic neurones in the ventral tegmental area (VTA) participate in behavioural arousal from anaesthesia. We investigated the regulation of dopaminergic VTA neurones by orexinergic neurones during emergence from general anaesthesia. Methods Orexins were microinjected into the VTA to determine the effects on isoflurane anaesthesia induction, emergence, and maintenance. Orexin receptors and dopaminergic neurones in the VTA were identified using immunofluorescence. Orexinergic terminals in the VTA were optogenetically regulated to detect the endogenous orexin-mediated regulation of dopaminergic neurones during anaesthesia in Hcrtcre rats. Results Injection of orexin-A (100 pmol) into the VTA reduced emergence time [from 949 (118) to 727 (101) s; P=0.0058] and reduced the electroencephalographic burst–suppression ratio (BSR) (26.6 [10.2]% vs 44.3 [6.8]%; P=0.0027) during isoflurane anaesthesia. The percentage of dopaminergic neurones that expressed either orexin-1 receptor or orexin-2 receptor was 73.4 (5.0)% and 74.4 (62.4)%, respectively. Optogenetic activation of orexinergic projections to the VTA reduced the BSR (from 40.5 [2.7]% to 22.4 [11.8]%; P=0.0019) and facilitated emergence (915 [89] vs 685 [68] s; P=0.0026), whereas optical inhibition prolonged the time to wakefulness (from 941 [92] to 1279 [250] s; P=0.011). Dopaminergic neurones in the VTA showed increased firing frequency (387 [78]% of control, P=0.005) after bath application of orexin-A. Conclusions Orexin promotes emergence from isoflurane anaesthesia through activation of dopaminergic neurones in the VTA.

Purpose Chemokine CX3CL1 and its receptor CX3CR1 in the lumbar spinal cord play crucial roles in pain processing. Electroacupuncture (EA) is recognized as an alternative therapy in pain treatment due to its efficacy and safety. However, the analgesic mechanism of EA remains unclear. The aim of this study was to investigate whether EA suppressed complete Freund's adjuvant (CFA)-induced pain via modulating CX3CL1-CX3CR1 pathway. Materials and methods Inflammatory pain was induced by intraplantar injection of CFA to the left hind paw of Sprague-Dawley rats. EA with 2 Hz for 30 mins was given to bilateral Zusanli acupoints (ST36) on the first and third day after CFA injection. Mechanical allodynia and thermal hyperalgesia were tested with von Frey tests and Hargreaves tests, respectively. The expressions of CX3CL1, CX3CR1 and p38 mitogen-activated protein kinase (MAPK) were quantified with Western blots. The release of IL-1β, IL-6 and TNF-α were evaluated with ELISA. Recombinant CX3CL1 or control IgG were then injected through intrathecal catheters in the EA-treated CFA model rats. The behavioral tests, p38 MAPK activation and cytokine release were then evaluated. Results EA significantly inhibited inflammatory pain induced by CFA for 3 days. Meanwhile, EA downregulated the expression of CX3CL1 but not CX3CR1 in the lumbar spinal cord of the CFA rats. Besides, activation of p38 MAPK and the release of pain-related cytokines (IL-1β, IL-6 and TNF-α) were inhibited by EA. Intrathecal injection of CX3CL1 largely reversed the analgesic effect of EA treatment and re-activated p38 MAPK signaling, and resulted in pro-inflammatory cytokines increase in acupuncture-treated rats. Conclusion Our findings indicate that EA alleviates inflammatory pain via modulating CX3CL1 signaling in lumbar spinal cord, revealing a potential mechanism of anti-nociception of EA in inflammatory pain.

Background Orexin, a neuropeptide derived from the perifornical area of the hypothalamus (PeFLH), promotes the recovery of propofol, isoflurane, and sevoflurane anesthesias, without influencing the induction time. However, whether the orexinergic system also plays a similar role in desflurane anesthesia, which is widely applied in clinical practice owing to its most rapid onset and offset time among all volatile anesthetics, has not yet been studied. In the present study, we explored the effect of the orexinergic system on the consciousness state induced by desflurane anesthesia. Methods The c-Fos staining was used to observe the activity changes of orexinergic neurons in the PeFLH and their efferent projection regions under desflurane anesthesia. Chemogenetic and optogenetic techniques were applied to compare the effect of PeFLH orexinergic neurons on the induction, emergence, and maintenance states between desflurane and isoflurane anesthesias. Orexinergic terminals in the paraventricular thalamic nucleus (PVT) were manipulated with pharmacologic, chemogenetic, and optogenetic techniques to assess the effect of orexinergic circuitry on desflurane anesthesia. Results Desflurane anesthesia inhibited the activity of orexinergic neurons in the PeFLH, as well as the neuronal activity in PVT, basal forebrain, dorsal raphe nucleus, and ventral tegmental area, as demonstrated by c-Fos staining. Activation of PeFLH orexinergic neurons prolonged the induction time and accelerated emergence from desflurane anesthesia but only influenced the emergence in isoflurane anesthesia, as demonstrated by chemogenetic and pharmacologic techniques. Meanwhile, optical activation of orexinergic neurons exhibited a long-lasting inhibitory effect on burst-suppression ratio (BSR) under desflurane anesthesia, and the effect may be contributed by the orexinergic PeFLH-PVT circuitry. The orexin-2 receptor (OX2R), but not orexin-1 receptor (OX1R), in the PVT, which had been inhibited most significantly by desflurane, mediated the proemergence effect of desflurane anesthesia. Conclusions We discovered, for the first time, that orexinergic neurons in the PeFLH could not only influence the maintenance and emergence from isoflurane and desflurane anesthesias but also affect the induction under desflurane anesthesia. Furthermore, this specific effect is probably mediated by orexinergic PeFLH-PVT circuitry, especially OX2Rs in the PVT.

Postmenopausal women have a higher incidence of stroke compared to the age-matched males, and the estrogen was thought to be the main cause of such difference. However, estrogen replacement therapy for the prevention of postmenopausal stroke shows controversial results and is widely disputed because of its serious side effects after chronic administration. Genistein (Gen), a natural phytestrogen with fewer side effects, has a protective effect against cerebral ischemia damage. However, whether Gen could effectively prevent postmenopausal stroke has not been elucidated. In the current study, reproductively senescent mice were treated with Gen (10 mg/kg) for 2 weeks before having transient cerebral ischemia insults. Neurological scores, infarct volumes, and cell apoptosis were evaluated 24 h after reperfusion. The levels of inflammatory factors and nod-like receptor protein 3 (NLRP3) inflammasome-related proteins were also examined. The results showed that Gen treatment reduced infarct volumes, improved neurological scores, attenuated apoptosis, and decreased inflammatory factor release. The expression of NLRP3 inflammasome-related proteins in microglia was downregulated by Gen. However, the overexpression of NLRP3 in microglia abrogated the Gen-induced inhibition of inflammatory factor release and reversed the neuroprotective effect of Gen. Taken together, the results suggest that Gen treatment could attenuate the acute injury induced by cerebral ischemia in reproductively senescent mice via the inhibition of the NLRP3 inflammasome in microglia, indicating that Gen could be a candidate drug for the treatment of stroke in postmenopausal women.

Studies have failed to translate more than 1000 experimental treatments from bench to bedside, leaving stroke as the second leading cause of death in the world. Thrombolysis within 4.5 hours is the recommended therapy for stroke and cannot be performed until neuroimaging is used to distinguish ischemic stroke from hemorrhagic stroke. Therefore, finding a common and critical therapeutic target for both ischemic and hemorrhagic stroke is appealing. Here, we report that the expression of myeloid differentiation protein 2 (MD2), which is traditionally regarded to be expressed only in microglia in the normal brain, was markedly increased in cortical neurons after stroke. We synthesized a small peptide, Trans-trans-activating (Tat)-cold-inducible RNA binding protein (Tat-CIRP), which perturbed the function of MD2 and strongly protected neurons against excitotoxic injury in vitro. In addition, systemic administration of Tat-CIRP or genetic deletion of MD2 induced robust neuroprotection against ischemic and hemorrhagic stroke in mice. Tat-CIRP reduced the brain infarct volume and preserved neurological function in rhesus monkeys 30 days after ischemic stroke. Tat-CIRP efficiently crossed the blood-brain barrier and showed a wide therapeutic index for stroke because no toxicity was detected when high doses were administered to the mice. Furthermore, we demonstrated that MD2 elicited neuronal apoptosis and necroptosis via a TLR4-independent, Sam68-related cascade. In summary, Tat-CIRP provides robust neuroprotection against stroke in rodents and gyrencephalic nonhuman primates. Further efforts should be made to translate these findings to treat both ischemic and hemorrhagic stroke in patients.

Background Previous clinical studies have shown that emergence from isoflurane anaesthesia takes longer in elderly patients compared with middle-aged patients. The current study investigated whether delayed emergence from anaesthesia in older age is associated with the age-related decrease in orexin receptors by using a rat model. Methods Adult and aged Sprague–Dawley rats were used to assess the time to emergence after 30 min isoflurane anaesthesia (1.4 vol%), and differences in the orexinergic systems, including the number of orexinergic neurones, plasma orexin concentrations, and expression of orexin-1 (OX1R) and orexin-2 receptors, compared using immunofluorescence, radioimmunoassay, western blot, and real-time polymerase chain reaction. The effects of OX1R expression on emergence time were determined by virus-mediated overexpression of OX1R using intra-cerebroventricular injection. Results The median (range) emergence time of aged rats was longer than that in adult rats [1082 (1010–1130) compared with 848 (829–938) s; P=0.0009]. Plasma orexin concentrations were higher in the aged group than the adult group [34 (33–37) and 25 (22–31) pg ml−1, respectively; P=0.04], but the number of orexinergic neurones was similar in both groups. Protein expression of OX1R was lower in the aged group compared with the adult group [0.47 (0.35–0.58) compared with 0.97 (0.86–1.32), respectively; P=0.002]. Overexpression of OX1R significantly shortened the emergence time in aged rats from [1120 (1040–1190) s] to [769 (576–928) s; P=0.03]. Conclusions Age-related decrease in OX1R expression is associated with delayed emergence from isoflurane anaesthesia in aged rats.

Background Sevoflurane preconditioning induces brain ischemic tolerance, but the mechanism remains poorly elucidated. Nitration is an important form of post-translational modification in pathological signaling. This study was to investigate the role of thioredoxin-1 (Trx-1) nitration in neuroprotection effect induced by sevoflurane preconditioning in a transient stroke model in rats. Methods Adult male Sprague–Dawley rats were preconditioned with 2% sevoflurane or vehicle oxygen exposure, 1 h per day, for 5 consecutive days. At 24 h after the last exposure, rats were subjected to focal brain ischemia induced by middle cerebral artery occlusion (MCAO) for 90 min, followed by 72-h reperfusion. Trx-1 expression and activity, as well as the content of nitrotyrosine at penumbra were detected at 24 h after preconditioning and 2, 8, 24, 72 h after MCAO. Nitrated Trx-1 was examined by immunoprecipitation at 8 h after MCAO. The role of Trx-1 nitration in ischemic tolerance was assessed by administration of nitrated human-Trx-1 prior to MCAO. Neurological scores, brain infarct volumes and TUNEL staining were evaluated at 24 h after reperfusion. Results Ischemic stroke decreased Trx-1 activity but not the expression in penumbra tissue. The content of nitrotyrosine was elevated after MCAO. Preconditioning with sevoflurane increased Trx-1 activity and reduced its nitration at 8 h after MCAO in comparison with vehicle preconditioning. The decrement of Trx-1 activity was correlated with its nitration level. Exogenous administration of nitrated human-Trx-1 reversed the brain ischemic tolerance of sevoflurane preconditioning, exacerbating brain infarct volume, neurobehavioral defects and apoptosis, while administration of human-Trx-1 had no effect on the sevoflurane preconditioning-induced neuroprotection. Conclusion Ischemic stroke reduces Trx-1 activity via post-translational nitrative modulation in rats. Sevoflurane preconditioning induces brain ischemic tolerance and anti-apoptosis by partially preserving Trx-1 activity via inhibiting nitration.

Social stress, a common stressor, causes multiple forms of physical and mental dysfunction. Prolonged exposure to social stress is associated with a higher risk of psychological disorders, including anxiety disorders and major depressive disorder (MDD). The orexinergic system is involved in the regulation of multiple motivated behaviors. The current study examined the regulatory effect of orexinergic projections from the lateral hypothalamic area (LHA) to the lateral habenula (LHb) in depression- and anxiety-like behaviors after chronic social defeat stress. When mice were defeated during social interaction, both orexinergic neurons in the LHA and glutamatergic neurons in the LHb were strongly activated, as indicated by the FosTRAP strategy. Infusion of orexin in the LHb significantly alleviated social avoidance and depression-like behaviors induced by chronic social defeat stress. Administration of an orexin receptor 2 antagonist in the LHb further aggravated the depressive phenotype. Photoactivation of orexinergic cell bodies in the LHA or terminals in the LHb relieved anxiety-like behaviors induced by chronic social defeat stress. Collectively, we identified the antidepressant and anxiolytic effects of the circuit from LHA orexinergic neurons to the LHb in response to chronic social stress, providing new evidence of the antidepressant properties of LHA orexin circuits.

Backgrounds Opinion remains controversial as to whether joint-saving surgery could be safely performed in patients with juxta-articular osteosarcoma. The aim of this study was to assess the validity of adjuvant cryosurgery in joint salvage surgery. Methods We evaluated the oncological and functional outcomes of patients who underwent joint-sparing surgery in which argon-based cryoablation was employed to aid partial epiphysis-preserved tumor resection for osteosarcoma around the knee (7 in proximal tibia, 5 in distal femur). Results The study included 12 patients (5 male, 7 female, mean age 15.8 years (11–24). At a mean follow-up of 48.4 months (38–61), lung metastasis occurred in three patients. Among these, one patient had died, one was alive with disease, and one had no evidence of disease after lobectomy. Histological examination of the resected specimens revealed no viable tumor at the osteotomy plane. There was no local recurrence in the residual epiphysis except one in soft tissue. The mean Musculoskeletal Tumor Society functional score was 92.7%. Conclusions Argon-based cryosurgery is a reliable method of sterilizing the tumor in the epiphysis thus allowing safe joint-saving tumor resection possible in patients with juxta-articular osteosarcoma.

Background Isoflurane preconditioning (IPC) induces cerebral ischaemic tolerance, but the mechanism remains poorly understood. The aim of this study was to determine changes in mitochondrial function in the brain after IPC, and whether the cannabinoid receptor 1 (CB1R) could be involved in the mechanism of mitochondrial protection mediated by IPC. Methods Adult male Sprague–Dawley rats were pretreated with isoflurane 2% for 1 h day−1, for 5 days consecutively, and then subjected to 120 min right middle cerebral artery occlusion. Cannabinoid receptor 1 expression in the cellular and mitochondrial membrane was measured. The CB1R agonist HU-210 was administered alone, or the antagonists AM251 and SR141716A were given to the animals before each preconditioning. Neurological scores, infarct volume, apoptosis, and mitochondrial function were examined after middle cerebral artery occlusion. Results Expression of CB1R on cellular and mitochondrial membranes was increased 6 h after preconditioning. Both IPC and HU-210 administration before middle cerebral artery occlusion improved neurological outcomes and reduced infarct volume. Isoflurane preconditioning increased the expression of the anti-apoptotic proteins Bcl-2 and Bcl-XL and reduced apoptosis in neurones. Isoflurane preconditioning and HU-210 also markedly preserved the activity of respiratory chain complexes, reduced mitochondrial radical generation, preserved mitochondrial membrane potential, and inhibited mitochondrial permeability transition pore opening. Cannabinoid receptor 1 antagonists abolished the improvement in mitochondrial function and the neuroprotective effects induced by IPC. Conclusions Our results indicate that IPC elicits brain ischaemic tolerance and mitochondrial protection by activating the CB1R, which provides a new mechanism for IPC-induced neuroprotection against cerebral ischaemia.

The collapse of the blood-brain barrier (BBB) is one of the fundamental pathophysiology changes during cerebral ischemia reperfusion injury. Resveratrol has been recently reported to reduce cerebral ischemic damage by regulating the matrix metalloproteinase-9 (MMP-9). But, more direct evidence for the explanation of the BBB protected by resveratrol against cerebral ischemia reperfusion is still lacking. Therefore, the present study was aimed to investigate the regulation of BBB integrity by resveratrol after cerebral ischemia reperfusion and to determine the role of the MMP-9 and its endogenous inhibitor TIMP-1 balance in this process. Cerebral ischemia was induced by middle cerebral artery occlusion in rats. The BBB function was evaluated by brain water content and the Evans blue dye extravasation; the activities of MMP-9 and TIMP-1 were detected by using gelatin zymography analysis, and cellular apoptosis was examined by TUNEL staining. We confirmed that resveratrol reduced the cerebral ischemia reperfusion damage, brain edema, and Evans blue dye extravasation. Moreover, we found that resveratrol improved the balance of MMP-9/TIMP-1 in terms of their expressions and activities. A TIMP-1 neutralizing antibody reversed those neuroprotective effects of resveratrol. In conclusion, resveratrol attenuated the cerebral ischemia by maintaining the integrity of BBB via regulation of MMP-9 and TIMP-1.

The neuroprotective properties for certain medical gases have been observed for decades, leading to extensive research that has been widely reported and continues to garner interest. Common gases including oxygen, hydrogen, carbon dioxide and nitric oxide, volatile anesthetics such as isoflurane, sevoflurane, halothane, enflurane and desflurane, non-volatile anesthetics such as xenon and nitrous oxide, inert gases such as helium and argon, and even gases classically considered to be toxic (e.g., hydrogen sulfide and carbon monoxide) have all been supported by the evidence alluding to their use as potential neuroprotective agents. A wide range of neural injury types such as ischemic/hemorrhagic, stroke, subarachnoid hemorrhage, traumatic brain injury, perinatal hypoxic-ischemic brain injuries, neurodegenerative disease as well as spinal cord ischemia have been used as platforms for studying the neuroprotective effects of these gases, yet until now, none of the gases has been widely introduced into clinical use specifically for protection against neural injury. Insufficient clinical data together with contradictory paradigms and results further hinders the clinical trials. However, pre-clinical models suggest that despite the various classes of gases and the broad range of injuries to which medical gases confer, protection, several underlying mechanisms for their neuroprotective properties are similar. In this review, we summarize the literature concerning the neuroprotective effect of each gas and its underlying mechanisms, extract common targets reported for the neuroprotective effects of different gases, highlight the conflicting observations from clinical trials and further discuss the possible hindrances impeding clinical applications in order to propose future research perspectives and therapeutic exploitations.

Background A wealth of evidence has demonstrated that sevoflurane preconditioning induces brain ischemic tolerance, but the mechanism remains poorly understood. This study was designed to investigate the role of canonical Notch signaling in the neuroprotection induced by sevoflurane preconditioning in a mouse model. Methods C57BL/6 mice were pretreated with 1-h sevoflurane exposure at a dose of 2.5% for 5 consecutive days. Twenty-four hours after the last exposure, all mice were subjected to focal cerebral ischemia by right middle cerebral artery occlusion for 60 min. Neurobehavioral scores, brain infarct volumes, and cellular apoptosis were determined at 72 h after reperfusion (n = 10 per group). The activation of Notch signaling was evaluated (n = 5 per group), and its role in ischemic tolerance was assessed by intraperitoneal administration of γ-secretase inhibitor DAPT (100 mg/kg, n = 10 per group) and conditional Notch-RBP-J knockout technique (n = 8 per group). Results Sevoflurane preconditioning reduced brain infarct volumes (42.5%), improved neurologic outcomes (P < 0.01 vs. control), and attenuated neuronal cell apoptosis (cells positive for terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate nick-end labeling reduced to 21.2%). The expression of Notch1 intracellular domain (1.35 folds) and the transcriptions of Hes1 (1.95 times) and Hes5 (1.48 times) were up-regulated. DAPT augmented the brain infarcts (1.64-fold) and decreased neurologic scores (P = 0.43 vs. sevoflurane) in sevoflurane-preconditioned mice. Brain infarct volumes, neurobehavioral scores, and apoptotic cell numbers showed no significance between Notch knockout mice with sevoflurane preconditioning and wild-type mice without preconditioning. Conclusions Sevoflurane preconditioning-induced protective effects against transient cerebral ischemic injuries are mediated by the activation of canonical Notch signaling pathway in mice.

Anesthetic preconditioning appears to be a viable strategy to treat ischemic cerebral injury. Here we investigated 1) whether the protection conferred by sevoflurane preconditioning sustains in time; 2) whether sevoflurane preconditioning diminishes mitochondrial dysfunction following cerebral ischemia; and 3) whether mitochondrial permeability transition pore plays a crucial role in the sevoflurane preconditioning.Laboratory investigation.University research laboratory.: Sprague-Dawley rats.Rats underwent 2 hrs of focal cerebral ischemia induced by middle cerebral artery occlusion. Preconditioning was elicited with sevoflurane (2.3%) for 60 mins at 24 hrs before ischemia. The involvement of mitochondrial permeability transition pore was determined with a mitochondrial permeability transition pore opener atractyloside and a specific mitochondrial permeability transition pore inhibitor cyclosporin A. In vitro study was performed on acutely isolated mitochondria subjected to calcium overload.Sevoflurane preconditioning significantly decreased the infarct size by 35.9% (95% confidence interval 6.5-28.4, p.001). This reduction of injury volume was associated with a long-term improvement of neurological function according to modified neurological severity score (F = 13.6, p = .001) and sticky-tape test (F = 29.1, p.001) for 42 days after ischemia. Furthermore, sevoflurane preconditioning markedly protected mitochondria, as indicated by preserved respiratory chain complex activities and membrane potential, lowered mitochondrial hydrogen-peroxide production, and attenuated mitochondrial permeability transition pore opening. Isolated mitochondria also demonstrated a reduced sensitivity to Ca-induced mitochondrial permeability transition pore opening after pre-exposure to sevoflurane in vitro (95% confidence interval 24.2-196.5,p = .006). Inhibiting mitochondrial permeability transition pore using cyclosporin A resulted in protective effects similar to those seen with sevoflurane preconditioning, whereas pharmacologically opening the mitochondrial permeability transition pore with atractyloside abrogated all the positive effects of sevoflurane preconditioning and cyclosporin A, including suppression of mitochondrial permeability transition pore opening, counteraction of mitochondria-dependent apoptotic pathway, and subsequent histological and behavioral improvements.Sevoflurane preconditioning protects mitochondria from cerebral ischemia/reperfusion injury and ameliorates long-term neurological deficits. Inhibition of mitochondrial permeability transition pore opening is a crucial step in mediating the neuroprotection of sevoflurane preconditioning.

Background Paraplegia is one of the most common complications following aortic aneurismal surgery. The present study examined the hypothesis that neuroprotection of sevoflurane postconditioning (PostC) against spinal cord reperfusion injury is associated with free radicals-mediated up-regulation of antioxidant enzymes in rabbits. Materials and Methods New Zealand White rabbits were subjected to sevoflurane PostC, oxygen (O2), or sham treatment at the onset of spinal cord reperfusion for 10min. Spinal cord ischemia was induced by clamping the aorta both below the left renal artery and above the aortic bifurcation. Forty-eight hours after reperfusion, the motor function of the lower limbs was evaluated and the lumbar spinal cord segment (L5-7) was harvested for histopathologic and biochemical analyses in the presence or absence of N-2-mercaptopropionylglycine, a potent oxygen free radical scavenger. Results Activities of superoxide dismutase and catalase in spinal cord tissue were increased in the sevoflurane PostC group 1h after reperfusion and retained a higher level 6 or 24h after reperfusion. Sevoflurane PostC produced a significant reduction of the malondialdehyde content at 6, 24, and 48h after reperfusion. A 20min continuous administration of N-2-mercaptopropionylglycine starting 10min before the beginning of PostC attenuated the neuroprotective effect against spinal cord ischemia, and reversed the increase of the two enzyme activities in spinal cord tissue induced by sevoflurane PostC. Conclusion An initial oxidative stress, as a trigger to up-regulate the antioxidant enzyme activities, plays an important role in the protection of sevoflurane PostC against spinal cord reperfusion injury.

This study investigates the effects of electroacupuncture (EA) preconditioning on blood-brain barrier (BBB) integrity and matrix metalloproteinase-9 (MMP-9) expression in subsequent ischemic hemisphere.Focal cerebral ischemia was induced by middle cerebral artery occlusion (MCAO) in rats. Animals were randomly divided into four groups: normal, sham-operated, MCAO and EA groups. In EA group, rats received electroacupuncture stimuli at the Baihui acupoint (GV 20) 30 minutes/day for 5 days. Twenty-four hours after last treatment, the MCAO was performed. The brain water content and BBB permeability were measured 24 hours after MCAO. MMP-9 expression and activity were measured at 6, 12 and 24 hours after MCAO.The results showed that the brain water content of ischemic hemisphere was lower in EA group (81.45 +/- 1.09%) compared with MCAO group (83.98 +/- 1.30%; p0.05). Similarly, the Evans blue content in EA group (4.90 +/- 1.77 microg/g) was lower compared with MCAO group (9.41 +/- 2.87 microg/g; p0.05). The protein expression and enzyme activity of MMP-9 increased and reached maximum at 24 hours after reperfusion. However, the protein expression was lower in EA group at 12 and 24 hours after reperfusion (p0.01, versus MCAO group), and enzyme activity was lower in EA group only at 24 hours (p0.01, versus MCAO group).EA preconditioning could attenuate brain edema and BBB disruption caused by subsequent cerebral ischemia. EA preconditioning could decrease MMP-9 expression and activity, which may be an important mechanism of cerebral ischemic tolerance.

Summary Background With the improved survival for patients with bone sarcomas, there is a trend to reconstruct intercalary femur defects using biologic techniques. This study examined whether the results of a frozen femur autograft with vascularised fibula are comparable to other reconstructive options in terms of the functional outcomes and the complications. Materials and methods Between 2008 and 2012, eight patients with bony sarcoma of the femur were subjected to reconstruction with a recycled frozen autograft combined with a vascularised fibula flap inside. The oncologic and functional results were analysed retrospectively. Results The mean follow-up was 48.7 months (37–71). The oncologic results were continuously disease free in five patients; there was no evidence of disease in one, one patient was alive with disease and another died of the disease. The average length of defect was 13.6 cm (9–21). Bone union was achieved in all cases. The mean time to bone union was 7.9 months (5–19) and to full weight bearing was 7.8 months (6–11). There was no infection or construct fracture in this series. Two complications were observed. One tumour recurrence in soft tissue was treated with reresection. One tibia fracture was successfully managed with cast immobilisation. The average Musculoskeletal Tumor Society functional score was 95% (27–30). The construct was intact in all patients. Conclusions Vascularised fibular flap combined with frozen autografts is a dependable and durable option for the reconstruction of large bony defects after femoral sarcoma resection. It had the merits of reliable bone union and low complications, which compare well with other biological reconstructions. Evidence Rating Scale for Therapeutic Studies Level IV, therapeutic study.

Background:Mechanism of sevoflurane preconditioning–induced cerebral ischemic tolerance is unclear. This study investigates the role of N-myc downstream–regulated gene-2 (NDRG2) in the neuroprotection of sevoflurane preconditioning in ischemic model both in vivo and in vitro.Methods:At 2 h after sevoflurane (2%) preconditioning for 1 h, rats were subjected to middle cerebral artery occlusion for 120 min. Neurobehavioral scores (n = 10), infarct volumes (n = 10), cellular apoptosis (n = 6), and NDRG2 expression (n = 6) were determined at 24 h after reperfusion. In vitro, cultural astrocytes were exposed to oxygen–glucose deprivation for 4 h. Cellular viability, cytotoxicity, apoptosis, and NDRG2 expression (n = 6) were evaluated in the presence or absence of NDRG2-specific small interfering RNA or NDRG2 overexpression plasmid.Results:Sevoflurane preconditioning decreased apoptosis (terminal deoxynucleotidyl transferase–mediated 2’-deoxyuridine 5’-triphosphate nick-end labeling–positive cells reduced to 31.2 ± 5.3% and cleaved Caspase-3 reduced to 1.42 ± 0.21 fold) and inhibited NDRG2 expression (1.28 ± 0.15 fold) and nuclear translocation (2.21 ± 0.29 fold) in ischemic penumbra. Similar effects were observed in cultural astrocytes exposed to oxygen–glucose deprivation. NDRG2 knockdown by small interfering RNA attenuated oxygen–glucose deprivation–induced injury (cell viability increased to 80.5 ± 4.1%; lactate dehydrogenase release reduced to 30.5 ± 4.0%) and cellular apoptosis (cleaved Caspase-3 reduced to 1.55 ± 0.21 fold; terminal deoxynucleotidyl transferase–mediated 2’-deoxyuridine 5’-triphosphate nick-end labeling–positive cells reduced to 18.2 ± 4.3%), whereas NDRG2 overexpression reversed the protective effects of sevoflurane preconditioning. All the data are presented as mean ± SD.Conclusion:Sevoflurane preconditioning inhibits NDRG2 up-regulation and nuclear translocation in astrocytes to induce cerebral ischemic tolerance via antiapoptosis, which represents one new mechanism of sevoflurane preconditioning and provides a novel target for neuroprotection.

Stroke has severe consequences in postmenopausal women. As replacement therapy of estrogen have various adverse effects and the undermined outcomes. Genistein, a natural phytoestrogen, has been suggested to be a potential neuroprotective agent for such stroke patients. However, the role of genistein and its underlying mechanism in ovariectomized mice has not yet been evaluated. In the present study, ovariectomized mice were treated with genistein (10 mg/kg) or vehicle daily for two weeks before developing transient cerebral ischemia (middle cerebral artery occlusion). The neurological manifestation was evaluated, and infarct volumes were demonstrated by 2,3,5-triphenyltetrazolium chloride staining at 24 h after reperfusion. In addition, phosphorylation of extracellular signal-regulated kinase (ERK) was detected by Western blotting and immunofluorescence staining, and cellular apoptosis was evaluated in the ischemic penumbra. We found that treatment with genistein reduced infarct volumes, improved neurological outcomes and attenuated cellular apoptosis at 24 h after reperfusion. ERK1/2 showed increased phosphorylation by genistein treatment after reperfusion, and an ERK1/2 inhibitor U0126 abolished this protective effect of genistein in terms of infarct volumes, neurological scores and cellular apoptosis. Our findings indicate that treatment with genistein can reduce the severity of subsequent stroke episodes, and that this beneficial function is associated with ERK activation.

Ginsenoside Rd (Rd), one of the main active ingredients in Panax ginseng, has been demonstrated to protect against ischemic cerebral damage in vitro and in vivo. In this study, we aimed to further define the preclinical characteristics of Rd. We show that Rd passes the intact blood-brain barrier and exerts protection in both transient and permanent middle cerebral artery occlusion (MCAO) in rats. In the dose-response study, Rd (10–50 mg/Kg) significantly reduced the infarct volume on postoperative days (PODs) 1, 3, and 7. This protection was associated with an improved neurological outcome for as many as 6 weeks after transient MCAO, as assessed by modified neurological severity score, modified sticky-tape test, and corner test. For comparison, Rd was significantly more effective than edaravone and slightly more effective than N-tert-butyl-alpha-phenylnitrone (PBN). In the therapeutic window study, Rd exhibited remarkable neuroprotection, even when administered for as many as 4 h after the recirculation of transient MCAO or after the onset of permanent MCAO. Furthermore, in female rats or 16-month-old male rats, the salutary effects of Rd were also observed. These findings suggest Rd is a promising neuroprotectant and provide support for future clinical studies to confirm whether Rd is beneficial in ischemic stroke.

Background Signal transducer and activator of transcription 3 (STAT3) activation in ischemic brain has been verified. However, the mechanism and the role of STAT3 activation after cerebral ischemia-reperfusion are poorly elucidated. In the present study, we sought to test the hypothesis that STAT3 activation after cerebral ischemia-reperfusion was related to reactive oxygen species (ROS) production. Methods Adult male Sprague-Dawley rats were subjected to focal cerebral ischemia induced by middle cerebral artery occlusion. STAT3 activation was evaluated by immunohistochemistry and Western blotting. Rats were subjected to permanent ischemia or ischemia-reperfusion to clarify the temporal profile of STAT3 activation. The role of ROS in inducing STAT3 activation was assessed by administration of the ROS scavenger dimethylthiourea (DMTU). The effects of DMTU and the STAT3 activation inhibitor AG490 administration on brain ischemic injuries were evaluated by neurologic behavior scores and brain infarct volumes. Results The activation of STAT3 after middle cerebral artery occlusion was significantly increased within peri-ischemia neurons and astrocytes. STAT3 activation mainly occurred in the reperfusion phase rather than in the ischemia phase. In addition, DMTU suppressed STAT3 activation in a dose-dependent manner, indicating that STAT3 activation may be a subsequent event after ROS production. DMTU and AG490 significantly reduced infarct sizes and improved neurologic outcomes. Conclusion In comparison with ischemia, reperfusion is a more powerful stimulus for STAT3 activation. ROS scavenging is closely correlated with an inhibition of STAT3 activation. Neuroprotective effects are achieved through ROS scavenging and down-regulation of STAT3 activation.

Age is the single most important risk factor for cerebral stroke. Unfortunately, the effect of age on ischemic brain damage is less clear. In this study, we sought to examine the potential influence of aging on the histologic and functional outcomes after ischemia. Juvenile (4 weeks of age), young adult (4 months of age), mid-aged (11-12 months of age), and aged (18-19 months of age) mice were subjected to transient middle cerebral artery occlusion. There was no remarkable difference of infarct volume on postoperative days 1 and 3. However, on postoperative day 7, aged mice exhibited significantly worsened infarct volume compared with juvenile and young mice. Intriguingly, the increase of infarct volume was most prominent in the striatal area rather than in cortex. Accordingly, aged mice displayed a slower and incomplete functional recovery after stroke. We further evaluated the effects of aging on the oxidative damage and mitochondrial dysfunction following ischemia. Brain tissues were assayed for lipid, DNA, and protein peroxidation products, mitochondrial enzyme activities, mitochondrial membrane potential, production of reactive oxygen species, and antioxidant activities. Aging was associated with declined mitochondrial function and antioxidant detoxification following ischemia, thereby inducing a deteriorated oxidative damage. Regional subanalyses demonstrated that, in accordance with infarct area, the pro-oxidant/antioxidant imbalance occurred more prominently in subcortical areas. Collectively, these findings suggest mitochondria-mediated oxidative damage may be involved in the age-related aggravated injury in subcortical areas. Mitochondrial protection could be a promising target for neuroprotective therapy, especially in the aged population.

BACKGROUND: It has been reported that sevoflurane preconditioning can induce neuroprotection, the mechanisms of which, however, are poorly elucidated. We designed the present study to examine the hypothesis that sevoflurane preconditioning could reduce cerebral ischemia– reperfusion injury through up-regulating antioxidant enzyme activities before ischemic injury by generating reactive oxygen species (ROS). METHODS: In preconditioning groups, adult male Sprague–Dawley rats were pretreated with 1 hour sevoflurane exposure at a dose of 1%, 2%, or 4% for 5 consecutive days. At 24 hours after the last exposure, all rats were subjected to focal brain ischemia induced by middle cerebral artery occlusion for 120 minutes followed by 72-hour reperfusion. The role of ROS in ischemic tolerance was assessed by administration of the free radical scavenger dimethylthiourea and antioxidant N-acetylcysteine before each preconditioning. Brain ischemic injury was evaluated by neurologic behavior scores and brain infarct volume calculation. Antioxidant enzyme activities (superoxide dismutase, catalase, and glutathione peroxidase [GSH-px]) of brain tissue and blood serum were tested at 24 hours after the last sevoflurane preconditioning. RESULTS: Sevoflurane preconditioning reduced infarct size and improved neurobehavioral outcome in a dose-dependent manner. The neuroprotective effects of sevoflurane preconditioning were abolished by dimethylthiourea and N-acetylcysteine. The activities of catalase and glutathione peroxidase (GSH-px) in the brain tissue were elevated by sevoflurane preconditioning before ischemic injury. The up-regulated activity of GSH-px in serum negatively correlated with brain infarct volume percentage. CONCLUSION: Sevoflurane preconditioning induces cerebral ischemic tolerance in a dose– response manner through ROS release and consequent up-regulation of antioxidant enzyme activity before ischemic injury in rats. Serum GSH-px activity could be developed as a marker to assess the effectiveness of sevoflurane preconditioning before ischemia.

Background and Purpose— Our previous study has demonstrated that the rapid tolerance to cerebral ischemia by electroacupuncture (EA) pretreatment was possibly mediated through an endocannabinoid system-related mechanism. The purpose of this study was to investigate whether activation of epsilon protein kinase C (εPKC) was involved in EA pretreatment-induced neuroprotection via cannabinoid receptor type 1 in a rat model of transient focal cerebral ischemia. Methods— The activation of εPKC in the ipsilateral brain tissues after EA pretreatment was investigated in the presence or absence of cannabinoid receptor antagonists. At 2 hours after the end of EA pretreatment, focal cerebral ischemia was induced by middle cerebral artery occlusion for 120 minutes in rats. The neurobehavioral scores, infarction volumes, neuronal apoptosis, and the expression of Bcl-2 and Bax were evaluated after reperfusion in the presence or absence of εPKC-selective peptide inhibitor (TAT-εV1–2) or activator (TAT–ψεRACK). Results— EA pretreatment enhanced εPKC activation. Systemic delivery of TAT–ψεRACK conferred neuroprotection against a subsequent cerebral ischemic event when delivered 2 hours before ischemia. Pretreatment with EA reduced infarct volumes, improved neurological outcome, inhibited neuronal apoptosis, and increased the Bcl-2 -to- Bax ratio after reperfusion, and the beneficial effects were attenuated by TAT-εV1–2. In addition, the blockade of cannabinoid receptor type 1, but not cannabinoid receptor type 2 receptor, reversed the increase in εPKC activation and neuroprotection induced by EA pretreatment. Conclusion— EA pretreatment may activate endogenous εPKC-mediated anti-apoptosis to protect against ischemic damage after focal cerebral ischemia via cannabinoid receptor type 1, which represents a new mechanism of EA pretreatment-induced rapid tolerance to focal cerebral ischemia in rats.

We previously found that ginsenoside Rd (Rd), one of the major active ingredients in Panax ginseng, protects neuronal cells from hydrogen peroxide and oxygen-glucose deprivation, an in vitro model of cerebral ischemia. In this study, we examined the protective effects of Rd in an animal model of focal cerebral ischemia. Rats administered with Rd or vehicle were subjected to transient middle cerebral artery occlusion (MCAO). Rd (50 mg/kg) significantly reduced the infarct volume by 52.8%. This reduction of injury volume was associated with an improvement in neurological function and was sustained for at least 2 weeks after the induction of ischemia. To evaluate the underlying mechanisms of Rd against stroke, brain tissues were assayed for mitochondrial enzyme activities, mitochondrial membrane potential (MMP), production of reactive oxygen species (ROS), energy metabolites, and apoptosis. Rd markedly protected mitochondria as indicated by preserved respiratory chain complex activities and aconitase activity, lowered mitochondrial hydrogen peroxide production, and hyperpolarized MMP. Microdialysis results illustrated that Rd significantly decreased the accumulation of lactate, the end product of anaerobic glycolysis, and increased pyruvate, the end product of aerobic glycolysis, hence inducing a lower lactate/pyruvate ratio. Additionally, in vitro studies further exhibited that Rd protected isolated mitochondria from calcium-induced damage by attenuating mitochondrial swelling, preserving MMP and decreasing ROS production. Moreover, Rd treatment reduced mitochondrial release of cytochrome c (CytoC) and apoptosis-inducing factor (AIF), thereby minimizing mitochondria-mediated apoptosis following ischemia. In conclusion, these findings demonstrated that Rd exerts neuroprotective effects in transient focal ischemia, which may involve an integrated process of the mitochondrial protection, energy restoration and inhibition of apoptosis.

Background We have previously reported that electroacupuncture (EA) pretreatment induced tolerance against cerebral ischemic injury, but the mechanisms underlying this effect of EA are unknown. In this study, we assessed the effect of EA pretreatment on the expression of α7 nicotinic acetylcholine receptors (α7nAChR), using the ischemia-reperfusion model of focal cerebral ischemia in rats. Further, we investigated the role of high mobility group box 1 (HMGB1) in neuroprotection mediated by the α7nAChR and EA. Methods Rats were treated with EA at the acupoint "Baihui (GV 20)" 24 h before focal cerebral ischemia which was induced for 120 min by middle cerebral artery occlusion. Neurobehavioral scores, infarction volumes, neuronal apoptosis, and HMGB1 levels were evaluated after reperfusion. The α7nAChR agonist PHA-543613 and the antagonist α-bungarotoxin (α-BGT) were used to investigate the role of the α7nAChR in mediating neuroprotective effects. The roles of the α7nAChR and HMGB1 release in neuroprotection were further tested in neuronal cultures exposed to oxygen and glucose deprivation (OGD). Results Our results showed that the expression of α7nAChR was significantly decreased after reperfusion. EA pretreatment prevented the reduction in neuronal expression of α7nAChR after reperfusion in the ischemic penumbra. Pretreatment with PHA-543613 afforded neuroprotective effects against ischemic damage. Moreover, EA pretreatment reduced infarct volume, improved neurological outcome, inhibited neuronal apoptosis and HMGB1 release following reperfusion, and the beneficial effects were attenuated by α-BGT. The HMGB1 levels in plasma and the penumbral brain tissue were correlated with the number of apoptotic neurons in the ischemic penumbra. Furthermore, OGD in cultured neurons triggered HMGB1 release into the culture medium, and this effect was efficiently suppressed by PHA-543,613. Pretreatment with α-BGT reversed the inhibitory effect of PHA-543,613 on HMGB1 release. Conclusion These data demonstrate that EA pretreatment strongly protects the brain against transient cerebral ischemic injury, and inhibits HMGB1 release through α7nAChR activation in rats. These findings suggest the novel potential for stroke interventions harnessing the anti-inflammatory effects of α7nAChR activation, through acupuncture or pharmacological strategies.