Role of macrophage autophagy in postoperative pain and inflammation in mice.

BACKGROUND: Postoperative pain and inflammation are significant complications following surgery. Strategies that aim to prevent excessive inflammation without hampering natural wound-healing are required for the management of postoperative pain and inflammation. However, the knowledge of the mechanisms and target pathways involved in these processes is lacking. Recent studies have revealed that autophagy in macrophages sequesters pro-inflammatory mediators, and it is therefore being recognized as a crucial process involved in regulating inflammation. In this study, we tested the hypothesis that autophagy in macrophages plays protective roles against postoperative pain and inflammation and investigated the underlying mechanisms. METHODS: Postoperative pain was induced by plantar incision under isoflurane anesthesia in mice lacking macrophage autophagy (Atg5flox/flox LysMCre +) and their control littermates (Atg5flox/flox). Mechanical and thermal pain sensitivity, changes in weight distribution, spontaneous locomotor activity, tissue inflammation, and body weight were assessed at baseline and 1, 3, and 7 days after surgery. Monocyte/macrophage infiltration at the surgical site and inflammatory mediator expression levels were evaluated. RESULTS: Atg5flox/flox LysMCre + mice compared with the control mice exhibited lower mechanical and thermal pain thresholds and surgical/non-surgical hindlimb weight-bearing ratios. The augmented neurobehavioral symptoms observed in the Atg5flox/flox LysMCre + mice were associated with more severe paw inflammation, higher pro-inflammatory mediator mRNA expression, and more monocytes/macrophages at the surgical site. CONCLUSION: The lack of macrophage autophagy augmented postoperative pain and inflammation, which were accompanied by enhanced pro-inflammatory cytokine secretion and surgical-site monocyte/macrophage infiltration. Macrophage autophagy plays a protective role in postoperative pain and inflammation and can be a novel therapeutic target.

Propofol ameliorates ischemic brain injury by blocking TLR4 pathway in mice.

Ischemic brain injury is one of the most serious perioperative complications. However, effective preventative methods have not yet been established. This study aimed to investigate whether propofol has neuroprotective effects against ischemic brain injury, with a specific focus on Toll-like receptor 4 (TLR4). Focal brain ischemia was induced via a combination of left common carotid artery occlusion and distal left middle cerebral artery coagulation in mice. Either propofol (10 mg/kg) or vehicle was intravenously injected 10 min prior to the induction of brain ischemia in wild-type and TLR4 knockout mice. Infarct volume, pro-inflammatory cytokine expression, inflammatory cell infiltration, and neurobehavioral function were assessed. Propofol administration significantly reduced infarct volume in wild-type mice (26.9 ± 2.7 vs 15.7 ± 2.0 mm3 at day 7), but not in TLR4 knockout mice. Compared with the control mice, the propofol-treated wild-type mice exhibited lower levels of IL-6 (0.57 ± 0.23 vs 1.00 ± 0.39 at 24 h), and smaller numbers of TLR4-expressing microglia in the penumbra (11.7 ± 3.1 vs 25.1 ± 4.7 cells/0.1 mm2). In conclusion, propofol administration prior to ischemic brain insult attenuated brain injury by blocking the TLR4-dependent pathway and suppressing pro-inflammatory cytokine production.

Short Sleep Duration on the Night Before Surgery Is Associated With Postoperative Cognitive Decline in Elderly Patients: A Prospective Cohort Study.

As the world is rapidly aging, and the number of elderly patients who undergo surgery is rising, postoperative cognitive decline among those patients has become an increasing healthcare problem. Although understanding the risk factors and mechanisms underlying the pathogenesis of postoperative cognitive decline is critically important from a preventative viewpoint, such knowledge and evidence are lacking. A growing body of evidence suggest an association between cognitive function and sleep duration. The purpose of this study was to investigate the association between postoperative cognitive function and sleep duration on the night before surgery using a wearable sleep tracker. In this 6-month prospective cohort study, we analyzed data from 194 patients aged ≥ 65 years who underwent elective non-cardiac and non-cranial surgery under general anesthesia. According to the sleep duration on the night before surgery, patients were categorized into following four groups: <5, 5-7, 7-9, and >9 h. Perioperative cognitive function and domains were assessed using a neuropsychological test battery, and the incidence and prevalence of cognitive decline over 6 months after surgery were analyzed using the multiple logistic regression analysis. During the 6-month follow-up period, 41 patients (21%) developed cognitive decline. The incidence of cognitive decline was significantly elevated for the patients with sleep duration < 5 h (vs. 7-9 h; surgical duration-adjusted odds ratio, 3.50; 95% confidence interval, 1.20-10.2; P < 0.05). The association between sleep duration and prevalence of cognitive decline was limited to the early postoperative period (at 1 week and 1 month). Among the cognitive domains assessed, attentional function was significantly impaired in patients with a sleep duration < 5 h [vs. 7-9 h at 1 week; 4/37 (10.8%) vs. 0/73 (0%); P < 0.05]. In conclusion, sleep duration < 5 h on the night before surgery was significantly associated with worse attentional function after surgery and higher incidence of cognitive decline. The present results indicate that sleep deprivation on the night before surgery may have a temporary but significantly negative influence on the patient's postoperative cognitive function and is a potential target for preventing cognitive decline.

Effects of ß1-adrenergic receptor blockade on the cerebral microcirculation in the normal state and during global brain ischemia/reperfusion injury in rabbits.

BACKGROUND: Although recent studies using experimental models of ischemic brain injury indicate that systemically-administered ß1-blockers have potential protective effects on the cerebrovascular system, the precise mechanisms remain unclear. In addition to their cardiovascular effects, water-soluble ß1-blockers can pass the blood-brain barrier and may exert their vascular action on cerebral microvessels. The aim of this study was to investigate the direct effects of ß1-blockade on the cerebral microvasculature both in the normal state and ischemia/reperfusion state using the cranial window method. METHODS: The closed cranial window method was used to visualize the cerebral microcirculation and changes in the pial arteriole diameter in adult male rabbits. In the first experiment, various concentrations of the selective ß1-blocker landiolol were administered into the cranial window to evaluate the dose-response. In the second experiment, the effect of ß1-blockade on the brain during ischemic/reperfusion injury was investigated. Global brain ischemia/reperfusion was induced by clamping the brachiocephalic, left common carotid, and left subclavian arteries for 15 min. Either landiolol or artificial cerebrospinal fluid was infused 5 min after initiation of ischemia through 120 min after reperfusion. Pial arteriole diameter and hemodynamic and physiological parameters were recorded before ischemia, during ischemia, and 5, 10, 20, 40, 60, 80, 100, and 120 min after reperfusion. RESULTS: In the first experiment, topical administration of landiolol at higher concentrations produced slight pial arteriole dilation (10- 8 mol/L: 4.3 ± 3.4%, 10- 6 mol/L: 8.0 ± 5.8%, 10- 4 mol/L: 7.3 ± 4.0%). In the second experiment, the topical administration of landiolol significantly dilated the pial arteriole diameters during ischemia/reperfusion injury (ischemia: 30.6 ± 38.6%, 5 min: 47.3 ± 42.2%, 10 min: 47.8 ± 34.2%, 20 min: 38.0 ± 39.0%). There were no statistical differences in hemodynamic and physiological parameters between the landiolol and control groups. CONCLUSIONS: The blockade of ß1-adrenergic receptors induced significant vasodilation of pial arterioles during ischemia/reperfusion injury. By contrast, only a slight dilation of the arterioles was observed in the normal state, indicating that ischemic cerebral microvessels are more susceptible to the vasodilatory effect induced by selective blockade of ß1-adrenergic receptors than normal microvessels.

Amiodarone exacerbates brain injuries after hypoxic-ischemic insult in mice.

BACKGROUND: Sodium ion transportation plays a crucial role in the pathogenesis of hypoxic-ischemic brain injury. Amiodarone, a Vaughan-Williams class III antiarrhythmic drug, has been widely used to treat life-threatening arrhythmia and cardiac arrest worldwide. In addition to its inhibitory effects on the potassium channel, amiodarone also blocks various sodium ion transporters, including the voltage-gated sodium channel, sodium pump, and Na+/Ca+ exchanger. Considering these pharmacological profile, amiodarone may affect the influx-efflux balance of sodium ion in the hypoxic-ischemic brain. Previous studies suggest that the blockade of the voltage-gated sodium channel during hypoxic-ischemic brain injury exerts neuroprotection. On the contrary, the blockade of sodium pump or Na+/Ca+ exchanger during hypoxia-ischemia may cause further intracellular sodium accumulation and consequent osmotic cell death. From these perspectives, the effects of amiodarone on sodium ion balance on the hypoxic-ischemic brain can be both protective and detrimental depending on the clinical and pathophysiological conditions. In this study, we therefore investigated the effect of amiodarone on hypoxic-ischemic brain injury using a murine experimental model. RESULTS: Compared with the control group mice, mice that received amiodarone after induction of 40-min hypoxic-ischemic brain injury exhibited lower survival rates over 7 days and worse neurological function. After 25-min hypoxic-ischemic brain injury, amiodarone treated mice exhibited larger infarct volumes (16.0 ± 6.9 vs. 24.2 ± 6.8 mm3, P < 0.05) and worse neurological function. In addition, the brains harvested from the amiodarone-treated mice contained larger amounts of sodium (194.7 ± 45.1 vs. 253.5 ± 50.9 mEq/kg dry weight, P < 0.01) and water (259.3 ± 8.9 vs. 277.2 ± 12.5 mg, P < 0.01). There were no significant differences in hemodynamic parameters between groups. CONCLUSIONS: Amiodarone exacerbated brain injuries and neurological outcomes after hypoxic-ischemic insults. Severe brain sodium accumulation and brain edema were associated with the detrimental effects of amiodarone. Amiodarone at the clinical dose can exacerbate brain injury after hypoxic-ischemic insult by affecting sodium ion transportation and facilitate intracellular sodium accumulation in the brain.

Neuroprotective effects of neurotropin in a mouse model of hypoxic-ischemic brain injury.

PURPOSE: Ischemic-hypoxic insult leads to detrimental effects on multiple organs. The brain is especially vulnerable, and it is hard to regenerate once damaged. Currently, therapeutic options are very limited. Previous studies have reported neuroprotective effects of neurotropin, a non-protein extract derived from the inflamed skin of rabbits inoculated with vaccinia virus, using a murine model of peripheral nerve injury and cultured cell lines. However, whether neurotropin might have protective effects against brain injuries remains unclear. We, therefore, investigated the neuroprotective effect of neurotropin and possible underlying mechanisms, using a mouse model of hypoxic-ischemic brain injury. METHODS: Hypoxic-ischemic brain injury was induced via a combination of the left common carotid artery occlusion and exposure to hypoxic environment (8% oxygen) in adult male C57BL/6 mice. Immediately following induction of hypoxia-ischemia, mice received either saline or 2.4 units of neurotropin. The survival rate, neurological function, infarct volume, and expression of inflammatory cytokines were evaluated. RESULTS: Compared to the control group, the neurotropin group exhibited a significantly higher survival rate (100% vs. 62.5%, p < 0.05) and lower neurological deficit scores (1; 0-2 vs. 3; 0-5, median; range, p < 0.05) after the hypoxic-ischemic insult. The administration of neurotropin also reduced infarct volume (18.3 ± 5.1% vs. 38.3 ± 7.2%, p < 0.05) and mRNA expression of pro-inflammatory cytokines. CONCLUSIONS: The post-treatment with neurotropin improved survival and neurological outcomes after hypoxic-ischemic insult. Our results indicate that neurotropin has neuroprotective effects against hypoxic-ischemic brain injury by suppressing pro-inflammatory cytokines.

Nicorandil increased the cerebral blood flow via nitric oxide pathway and ATP-sensitive potassium channel opening in mice.

PURPOSE: Nicorandil has dual properties and acts as a nitric oxide donor and an ATP-sensitive potassium (KATP) channel opener. Considering its pharmacological profile, nicorandil might exert protective effects on the brain as well as on the heart. The purpose of this study was to directly evaluate the effect of nicorandil on cerebral blood flow (CBF) in mice using a transcranial Doppler method. METHODS: Under general anesthesia, the nicorandil groups received a single-bolus intraperitoneal injection of the respective doses of nicorandil (1, 5, or 10 mg/kg), while the control group received vehicle only. CBF was measured using a transcranial Doppler flowmeter. NG-nitro-L-arginine methyl ester and glibenclamide were used to elucidate the underlying mechanisms. RESULTS: A single-bolus injection of 1 mg/kg of nicorandil increased the CBF (11.6 ± 3.6 vs. 0.5 ± 0.7%, p < 0.001) without affecting the heart rate and blood pressure. On the contrary, 5 and 10 mg/kg of nicorandil significantly decreased the cerebral blood flow by decreasing the mean blood pressure below the cerebral autoregulation range. The positive effect of 1 mg/kg of nicorandil on the cerebral blood flow was inhibited by co-administration of either NG-nitro-L-arginine methyl ester or glibenclamide. CONCLUSIONS: A clinical dose of nicorandil increases CBF without affecting systemic hemodynamics. The positive effect of nicorandil on CBF is most likely caused via both the nitric oxide pathway and KATP channel opening.

Neuroprotective effects of amiodarone in a mouse model of ischemic stroke.

BACKGROUND: Ion channels play a crucial role in the development of ischemic brain injury. Recent studies have reported that the blockade of various types of ion channels improves outcomes in experimental stroke models. Amiodarone, one of the most effective drugs for life-threatening arrhythmia, works as a multiple channel blocker and its characteristics cover all four Vaughan-Williams classes. Although it is known that amiodarone indirectly contributes to preventing ischemic stroke by maintaining sinus rhythm in patients with atrial fibrillation, the direct neuroprotective effect of amiodarone has not been clarified. The purpose of this study was to investigate the direct effect of amiodarone on ischemic stroke in mice. METHODS: Focal cerebral ischemia was induced via distal permanent middle cerebral artery occlusion (MCAO) in adult male mice. The amiodarone pre-treatment group received 50 mg/kg of amiodarone 1 h before MCAO; the amiodarone post-treatment groups received 50 mg/kg of amiodarone immediately after MCAO; the control group received vehicle only. In addition, the sodium channel opener veratrine and selective beta-adrenergic agonist isoprotelenol were used to elucidate the targeted pathway. Heart rate and blood pressure were monitored perioperatively. Infarct volume analysis was conducted 48 h after MCAO. The body asymmetry test and the corner test were used for neurological evaluation. RESULTS: Amiodarone pre-treatment and post-treatment reduced the heart rate but did not affect the blood pressure. No mice showed arrhythmia. Compared with the control group, the amiodarone pre-treatment group had smaller infarct volumes (8.9 ± 2.1% hemisphere [mean ± SD] vs. 11.2 ± 1.4%; P < 0.05) and improved functional outcomes: lower asymmetric body swing rates (52 ± 17% vs. 65 ± 18%; P < 0.05) and fewer left turns (7.1 ± 1.2 vs. 8.3 ± 1.2; P < 0.05). In contrast, amiodarone post-treatment did not improve the outcomes after MCAO. The neuroprotective effect of amiodarone pre-treatment was abolished by co-administration of veratrine but not by isoproterenol. CONCLUSIONS: Amiodarone pre-treatment attenuated ischemic brain injury and improved functional outcomes without affecting heart rhythm and blood pressure. The present results showed that amiodarone pre-treatment has neuroprotective effects, at least in part, via blocking the sodium channels.

[Femoral nerve block for total knee arthroplasty].

BACKGROUND: Femoral nerve block and sciatic nerve block are used to provide intraoperative and postoperative analgesia for total knee arthroplasty. Sciatic nerve block is contraindicated in our hospital, because orthopedists want to assess peroneal nerve function after the surgery. We retrospectively assessed postoperative analgesic effect and complications of the continuous femoral nerve block for total knee arthroplasty. METHODS: We included 19 cases in 17 patients scheduled to undergo total knee arthroplasty under femoral nerve block combined with general anesthesia. Ultrasound-guided femoral nerve block was performed before the surgery. The ultrasound linear probe was used to visualize the femoral nerve. A 22 gauge needle attached to a nerve stimulator, was inserted with in-plane method. Five percent glucose solution was injected through the needle to encircle the femoral nerve. Then, the 22 gauge needle was withdrawn and an 18 gauge needle was inserted with out-of-plane method. Five percent glucose solution was injected through the needle to confirm the needle tip and perineural catheter was inserted through the needle. To achieve femoral nerve block, 0.375% ropivacaine 20 ml was injected through the needle. Perineural infusion with 0.15% ropivacaine at 4 ml x hr(-1) was initiated at the end of the surgery. Intravenous patient-controlled analgesia (IV-PCA) was also conducted postoperatively. We assessed pain at rest with a verbal numeric pain rating score (0-10) including pain on moving, and nausea as well as vomiting. RESULTS: Patients with numeric pain scores at 3 or less were 14 out of 19. Two patients complained of severe pain. There were 4 cases suffering pain on moving. CONCLUSIONS: Femoral nerve separation with 5% glucose solution using in-palne method and catheter placement with out-of-plane method could be useful for perineural catheter placement. Perineural infusion of 0.15% ropivacaine at 4 ml x hr(-1) combined with IV-PCA provided a good postoperative analgesia in patients receiving total knee arthroplasty.

Danaparoid sodium inhibits systemic inflammation and prevents endotoxin-induced acute lung injury in rats.

INTRODUCTION: Systemic inflammatory mediators, including high mobility group box 1 (HMGB1), play an important role in the development of sepsis. Anticoagulants, such as danaparoid sodium (DA), may be able to inhibit sepsis-induced inflammation, but the mechanism of action is not well understood. We hypothesised that DA would act as an inhibitor of systemic inflammation and prevent endotoxin-induced acute lung injury in a rat model. METHODS: We used male Wistar rats. Animals in the intervention arm received a bolus of 50 U/kg of DA or saline injected into the tail vein after lipopolysaccharide (LPS) administration. We measured cytokine (tumour necrosis factor (TNF)alpha, interleukin (IL)-6 and IL-10) and HMGB1 levels in serum and lung tissue at regular intervals for 12 h following LPS injection. The mouse macrophage cell line RAW 264.7 was assessed following stimulation with LPS alone or concurrently with DA with identification of HMGB1 and other cytokines in the supernatant. RESULTS: Survival was significantly higher and lung histopathology significantly improved among the DA (50 U/kg) animals compared to the control rats. The serum and lung HMGB1 levels were lower over time among DA-treated animals. In the in vitro study, administration of DA was associated with decreased production of HMGB1. In the cell signalling studies, DA administration inhibited the phosphorylation of IkappaB. CONCLUSION: DA decreases cytokine and HMGB1 levels during LPS-induced inflammation. As a result, DA ameliorated lung pathology and reduces mortality in endotoxin-induced systemic inflammation in a rat model. This effect may be mediated through the inhibition of cytokines and HMGB1.

Transplantation of GABAergic neurons into adult mouse neocortex.

GABAergic neurons in the neocortex contribute to various brain functions by regulating cortical pyramidal neurons. A deficiency of GABAergic neurons in the neocortex leads to the dysregulation of cortical neuronal circuits, but this can be overcome by cell transplantation, which provides a practical approach to repair damaged neuronal circuits. Here, we focused on the transplantation of committed neuronal progenitor cells. Because neuronal differentiation is considerably suppressed in the adult neocortex, we transfected proneural bHLH transcription factors into neural precursor cells to commit them to a neuronal lineage prior to the cell transplantation. We show that ventral neural stem cells transfected with Ngn1 are integrated as GABAergic neurons within a few days of transplantation into the adult mouse neocortex. These results demonstrate that the transplantation of committed neuronal progenitor cells is an effective method for brain repair.

Decrease in expression of alpha 5 beta 1 integrin during neuronal differentiation of cortical progenitor cells.

Neuronal differentiation of embryonic neural progenitor cells is regulated by both intrinsic and extrinsic signals. Since dynamic changes in cell shape typify neuronal differentiation, cell adhesion molecules could be relevant to this process. Although it has been reported that fibronectin-integrin interactions are important for the proliferation of neural progenitor cells, little is known about the contribution of integrins to neuronal differentiation. In order to address this shortfall, we examined integrin expression on cortical progenitor cells by using immunohistochemistry and FACS analysis of cells in which GFP expression was driven by regulatory (promoter) regions of the nestin gene (nestin-GFP(+)). We here report that high levels of nestin promoter activity correlated with high expression levels of alpha(5)beta(1) integrin (alpha(5)beta(1)(high) cells). FACS analysis of nestin-GFP(+) cortical cells revealed an additional subpopulation with reduced expression of alpha(5)beta(1) integrin (alpha(5)beta(1)(low) cells). The size of the alpha(5)beta(1)(low) subpopulation increased during cortical development. To investigate the correlation between integrin and neuronal differentiation, nestin-GFP(+) cortical progenitor cells were sorted into alpha(5)beta(1)(high) or alpha(5)beta(1)(low) populations, and each potential to differentiate was analyzed. We show that the nestin-GFP(+) alpha(5)beta(1)(high) population corresponded to broadly multipotential neural progenitor cells, whereas nestin-GFP(+) alpha(5)beta(1)(low) cells appeared to be committed to a neuronal fate. These findings suggest that alpha(5)beta(1) expression on cortical progenitor cells is developmentally regulated and its downregulation is involved in the process of neuronal differentiation.