Shaoyang Chen

Pretreatment With Electroacupuncture Induces Rapid Tolerance to Focal Cerebral Ischemia Through Regulation of Endocannabinoid System

Background and Purpose— Our previous study demonstrated that pretreatment with electroacupuncture (EA) induces rapid tolerance to focal cerebral ischemia. The present study was aimed to investigate the involvement of the endocannabinoid system in the early neuroprotection conferred by EA pretreatment in the animal model of focal cerebral ischemia. Methods— Two hours after the end of EA pretreatment, focal cerebral ischemia was induced by middle cerebral artery occlusion for 120 minutes in male Sprague-Dawley rats or male C57BL/6 mice. The neurobehavioral scores, infarction volumes, and neuronal apoptosis were evaluated at 24 hours or 7 days after reperfusion in the presence or absence of AM251 (a selective cannabinoid receptor type 1 [CB1] receptor antagonist) or CB1 short interfering RNA. The expression of CB1 receptor and the content of endocannabinoids in the brains were also investigated. Results— EA pretreatment reduced infarct size, improved neurological outcome, and inhibited neuronal apoptosis at 24 hours or 7 days after reperfusion. The beneficial effects were abolished by AM251. CB1 knockdown by CB1 short interfering RNA attenuated EA pretreatment-induced neuroprotection. EA pretreatment upregulated the neuronal expression of CB1 receptor in the rat brains and elevated the brain tissue content of the endocannabinoid 2-arachidonylglycerol and N-arach-idonoylethanolamine-anandamide. Pretreatment with 2-arachidonylglycerol and N-arach-idonoylethanolamine-anandamide also reduced infarct size and improved neurological outcome. Conclusion— We conclude that pretreatment with EA increases the production of endocannabinoid 2-arachidonylglycerol and N-arach-idonoylethanolamine-anandamide, which elicits protective effects against transient cerebral ischemia through CB1 receptors. These results suggest a novel mechanism of EA pretreatment-induced rapid tolerance to focal cerebral ischemia.

Preconditioning with hyperbaric oxygen and hyperoxia induces tolerance against spinal cord ischemia in rabbits.

Background The aim of this study was to determine if the ischemic tolerance could be induced in the spinal cord by pretreatment with hyperbaric oxygen (HBO) and what components of HBO (hyperoxia, hyperbaricity, and combination of these two) were critical in the induction of tolerance against ischemic injury. Methods In experiment 1, 21 rabbits were randomly assigned to one of three groups (n = 7 each): animals in the control group received no HBO before spinal cord ischemia; animals in the HBO-1 and HBO-2 groups received HBO (2.5 atmosphere absolute [ATA], 100% O2) pretreatment 1 h/day for 3 and 5 days before ischemia, respectively. In experiment 2, 48 rabbits were randomly assigned to one of four groups (n = 12 each): the control group received no HBO (21% O2, 1 ATA, 1 h/day, 5 days) before spinal cord ischemia; the HB group received 1-h treatment in 21% O2 at 2.5 ATA each day for 5 days; the HO group received 1-h treatment in 100% oxygen at 1 ATA each day for 5 days; and the HBO group received HBO (2.5 ATA, 100% O2) treatment 1 h/day for 5 days. Twenty-four hours after the last treatment, spinal cord ischemia was induced by an infrarenal aorta clamping for 20 min. Forty-eight hours after reperfusion, hind-limb motor function and histopathology of the spinal cord were examined in a blinded fashion. Results In experiment 1, the neurologic outcome in the HBO-2 group was better than that of the control group (P = 0.004). The number of normal neurons in the anterior spinal cord in the HBO-2 group was more than that of the control group (P = 0.021). In experiment 2, the neurologic and histopathologic outcomes in the HBO group were better than that of the control group (P < 0.01). The histopathologic outcome in the HO group was better than that in the control group (P < 0.05). Conclusions Serial exposure to high oxygen tension induced ischemic tolerance in spinal cord of rabbits. Simple hyperbaricity (2.5 ATA, 21% O2) did not induce ischemic tolerance.

PROTECTIVE EFFECTS OF HYDROGEN GAS ON MURINE POLYMICROBIAL SEPSIS VIA REDUCING OXIDATIVE STRESS AND HMGB1 RELEASE

Despite recent advances in antibiotic therapy and intensive care, sepsis is still considered to be the most common cause of death in intensive care units. Excessive production of reactive oxygen species plays an important role in the pathogenesis of sepsis. Recently, it has been suggested that molecular hydrogen (H2) exerts a therapeutic antioxidant activity by selectively reducing hydroxyl radicals (•OH, the most cytotoxic reactive oxygen species) and effectively protects against organ damage induced by I/R. Therefore, we hypothesized that H2 treatment had a beneficial effect on sepsis. In the present study, we found that H2 inhalation starting at 1 and 6 h after cecal ligation and puncture (CLP) or sham operation significantly improved the survival rate of septic mice with moderate or severe CLP in a concentration- and time-dependent manner. Furthermore, moderate or severe CLP mice showed significant multiple organ damage characterized by the increases of lung myeloperoxidase activity, wet-to-dry weight ratio, protein concentration in bronchoalveolar lavage, serum biochemical parameters, and organ histopathologic scores at 24 h after CLP operation, which was significantly attenuated by 2% H2 treatment. In addition, we found that the beneficial effects of H2 treatment on sepsis and sepsis-associated organ damage were associated with the decreased levels of oxidative product, increased activities of antioxidant enzymes, and reduced levels of high-mobility group box 1 in serum and tissue. Thus, H2 inhalation may be an effective therapeutic strategy for patients with sepsis.ABBREVIATIONS-ALI-acute lung injury; ALT-alanine aminotransferase; AST-aspartate aminotransferase; BAL-bronchoalveolar lavage; BUN-blood urea nitrogen; CAT-catalase; CLP-cecal ligation and puncture; Cr-creatinine; H2-hydrogen; H2O2-hydrogen peroxide; HMGB1-high-mobility group box 1; 8-iso-PGF2&agr;-8-iso-prostaglandin F2&agr;; MPO-myeloperoxidase; •OH-hydroxyl radicals; ROS-reactive oxygen species; SOD-superoxide dismutase; W/D-wet-to-dry

Noninvasive limb remote ischemic preconditioning contributes neuroprotective effects via activation of adenosine A1 receptor and redox status after transient focal cerebral ischemia in rats

PURPOSES To investigate whether activation of adenosine A1 receptor (A1R) through limb remote ischemic preconditioning (RIPC) by a noninvasive tourniquet contribute neuroprotective effects against rat focal cerebral ischemic injury induced by transient middle cerebral artery occlusion (MCAO). METHODS One hundred twenty-eight Sprague-Dawley (SD) rats were randomly assigned into eight groups (n=16 each): MCAO, Control, 8-cyclopentyl-1,3-dipropulxanthine (DPCPX, Adenosine A1 receptor antagonist), RIPC, DPCPX+RIPC, Vehicle+RIPC, 2-chloro-N(6)-cyclopentyladenosine (CCPA, Adenosine A1 receptor agonist) and CCPA+DPCPX groups. All animals underwent right middle cerebral artery occlusion (MCAO) for 2 h. Limb RIPC consisted of three cycles of 5-minute ischemia followed by 5-minute reperfusion in right hind-limb by tourniquet application. Neurological deficit scores were evaluated 24 h after reperfusion, and then the infarct volume was assessed with diffusion weighted imaging (DWI) and 2, 3, 5-triphenyltetrazolium chloride (TTC) staining. Inflammation was assessed by serum tumor necrosis factor α (TNF(α)) and NO; oxidative stress was estimated by malondialdehyde (MDA) and 4-hydroxyalkenals (4-HAD), superoxide dismutase (SOD) activity and GSH. RESULTS Animals in the RIPC, Vehicle+RIPC and CCPA groups developed lower neurological deficit scores and smaller brain infarct volumes than the Control group (P<0.01). Animals in the DPCPX, DPCPX+RIPC and CCPA+DPCPX groups developed higher neurological deficit scores and larger brain infarct volumes than the RIPC, Vehicle+RIPC and CCPA groups (P<0.01). DPCPX abolished the protective effects of RIPC and CCPA. RIPC or CCPA induced a significant increase in brain MnSOD (manganese SOD) activity and NO generation, and this activity was abolished by DPCPX pretreatment. RIPC or CCPA induced a significant reduction (P<0.05) in the GSH and MDA+4HDA concentration and an accumulation in the GSSG concentration in both compartments (serum and tissue) as compared with the MCAO group. CONCLUSIONS The present study demonstrates that limb RIPC induced by noninvasive tourniquet reduces cerebral ischemic injury in rats, and the effect of neuroprotection may depend on the activation of adenosine A1 receptors. CCPA pretreatment can induce delayed ischemic tolerance against cerebral ischemia/reperfusion injury. These protective effects are associated with a reduction in oxidative stress, inflammation and endogenous antioxidant preservation.

Rapid tolerance to focal cerebral ischemia in rats is induced by preconditioning with electroacupuncture: window of protection and the role of adenosine.

The aim of the present study was to investigate the first protective window of preconditioning with electroacupuncture (EA) against focal cerebral ischemia, and to explore whether adenosine is involved in the rapid tolerance phenomenon. Sixty-four male Sprague-Dawley rats were randomly assigned to eight groups (n=8 in each). Animals in the control group received no treatment, and animals in EA1-EA4 groups received EA at 0.5, 1, 2 and 3 h before induction of focal cerebral ischemia, respectively. Rats in DPCPX group were intraperitoneally injected with 1 mg kg-1 8-cyclopentyl-1,3-dipropylxanthine (DPCPX), 3 h before induction of focal cerebral ischemia. Animals in vehicle group and EA+DPCPX group were pretreated with 1 ml kg-1 dimethyl sulfoxide (DMSO, the solvent of DPCPX) and 1 mg kg-1 DPCPX 30 min before preconditioning with EA, respectively. All rats were anesthetized with 40 mg kg-1 pentobarbital sodium intraperitoneally. Animals that required EA preconditioning, received EA with intensity of 1 mA and frequency of 15 Hz at the Baihui acupoint (GV 20) for 30 min. The focal cerebral ischemia was produced by the right middle cerebral artery occlusion (MCAO) for 120 min. The neurologic deficit scores (NDS) and brain infarct volumes were evaluated at 24 h after reperfusion. All rats survived until 24 h after reperfusion. Preconditioning with EA at 2 h before induction of focal cerebral ischemia improved neurologic outcome (P<0.05 versus control) and reduced the infarct volume (P<0.01 versus control) at 24 h after reperfusion. These beneficial effects were reversed by pretreatment with 1 mg kg-1 DPCPX, whereas this agent itself did not affect the NDS and volume in drug-ischemic controls after ischemia. The results show that preconditioning with single EA session induces rapid tolerance to focal cerebral ischemia. The rapid ischemic tolerance appears at 2 h (but not at 0.5, 1, or 3 h) after preconditioning, and is possibly mediated through an adenosine A1 receptor-related mechanism.

Activation of Epsilon Protein Kinase C-Mediated Anti-Apoptosis Is Involved in Rapid Tolerance Induced by Electroacupuncture Pretreatment Through Cannabinoid Receptor Type 1

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–&psgr;ϵRACK). Results— EA pretreatment enhanced ϵPKC activation. Systemic delivery of TAT–&psgr;ϵRACK 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.

Ethyl Pyruvate Attenuates Spinal Cord Ischemic Injury with a Wide Therapeutic Window through Inhibiting High-mobility Group Box 1 Release in Rabbits

Background:Ethyl pyruvate (EP) has been reported to offer a protective effect against ischemic injury through its antiinflammatory action. The nuclear protein high-mobility group box 1 (HMGB1) can activate inflammatory pathways when released from ischemic cells. This study was designed to investigate the neuroprotective effect of EP against spinal cord ischemic injury and the potential role of HMGB1 in this process. Methods:EP was administered at various time points before or after 20 min of spinal cord ischemia in male New Zealand rabbits. All animals were sacrificed at 72 h after reperfusion with modified Tarlov criteria, and the spinal cord segment (L4) was harvested for histopathological examination and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling staining. The HMGB1 levels in serum and spinal cord tissue were analyzed by enzyme-linked immunosorbent assay. Results:The treatment of EP at 30 min before ischemia or at 6 h after reperfusion significantly improved the hind-limb motor function scores and increased the numbers of normal motor neurons, which was accompanied with reduction of the number of apoptotic neurons and levels of HMGB1 in serum and spinal cord tissue. The HMGB1 contents of spinal cord tissue correlated well with the numbers of apoptotic motor neurons in the anterior spinal cord at 72 h after reperfusion. Conclusion:These results suggest that EP affords a strong protection against the transient spinal cord ischemic injury with a wide therapeutic window through inhibition of HMGB1 release.

Isoflurane tolerance against focal cerebral ischemia is attenuated by adenosine A1 receptor antagonists.

PurposeTo investigate the role of the adenosine A1 receptor in the rapid tolerance to cerebral ischemia induced by isoflurane preconditioning.MethodsSeventy-five rats were randomly assigned into five groups (n = 15 each): Control, 8-cyclopentyl-1,3-dipropulxanthine (DPCPX), Isoflurane, DPCPX+Isoflurane and Vehicle+Isoflurane groups. All animals underwent right middle cerebral artery occlusion (MCAO) for two hours. Isoflurane preconditioning was conducted one hour before MCAO in Isoflurane, DPCPX+Isoflurane and Vehicle+Isoflurane groups by exposing the animals to 1.5% isoflurane in 98% oxygen for one hour. In the Control and DPCPX groups, animals were exposed to 98% oxygen one hour before MCAO for one hour. A selective adenosine A1 receptor antagonist, DPCPX, was administered (0.1 mg·kg-1) 15 min before isoflurane/oxygen exposure in the DPCPX and DPCPX+Isofluranegroups to evaluate the effect of adenosine A1 receptor antagonist on isoflurane preconditioning. Dimethyl sulfoxide, the solvent of DPCPX, was administered (1 mL·kg-1) 15 min before isoflurane exposure in the Vehicle+Isoflurane group. Neurological deficit scores and brain infarct volumes were evaluated 24 hr after reperfusion.ResultsAnimals in the Isoflurane and Vehicle+Isoflurane groups developed lower neurological deficit scores and smaller brain infarct volumes than the Control group (P < 0.01). Animals in the DPCPX+Isoflurane group developed higher neurological deficit scores and larger brain infarct volumes than the Isoflurane and Vehicle+Isoflurane groups (P < 0.01).ConclusionThe present study demonstrates that preconditioning with isoflurane reduces focal cerebral ischemic injury in rats, and the adenosine A1 receptor antagonist (DPCPX) attenuates the neuroprotection induced by isoflurane preconditioning.RésuméObjectifExplorer le rôle des récepteurs de ľadénosine A1 dans la tolérance rapide à ľischémie cérébrale induite par le préconditionnement à ľisoflurane.MéthodeNous avons réparti 75 rats en cinq groupes de 15: Témoin, 8-cyclopentyl-1,3-dipropulxanthine (DPCPX), Isoflurane, DPCPX+Isoflurane et Véhicule+Isoflurane. Tous les rats ont subi une occlusion de deux heures de ľartère cérébrale moyenne droite (OACM). Le préconditionnement avec ľisoflurane a été fait une heure avant ľOACM dans les groupes Isoflurane, DPCPX+Isoflurane et Véhicule+Isoflurane par une exposition à un mélange de 1,5 % ďisoflurane et de 98 % ďoxygène pendant une heure. Dans les groupes Témoin et DPCPX, les rats ont été exposés à 98 % ďoxygène une heure avant ľOACM qui a duré une heure. Un antagoniste sélectif des récepteurs de ľadénosine A1, DPCPX, a été administré (0,1 mg·kg-1) 15 min avant ľexposition à ľisoflurane/oxygène dans les groupes DPCPX et DPCPX+Isoflurane pour évaluer ľeffet de ľantagoniste des récepteurs de ľadénosine A1 sur le préconditionnement à ľisoflurane. Le sulfoxyde de diméthyle, le solvant de DPCPX, a été administré (1 mL·kg-1) 15 min avant ľexposition à ľisoflurane dans le groupe Véhicule+Isoflurane. Les scores de déficit neurologique et les volumes ďinfarctus cérébral ont été évalués 24 h après la reperfusion.RésultatsDans les groupes Isoflurane et Véhicule+Isoflurane, les scores de déficit neurologique étaient plus bas et les infarctus cérébraux de moindre volume que dans le groupe Témoin (P < 0,01). Dans le groupe DPCPX+Isoflurane, les scores de déficit neurologique étaient plus élevés et les infarctus cérébraux de plus grand volume que dans les groupes Isoflurane et Véhicule+Isoflurane (P < 0,01).ConclusionLe préconditionnement avec de ľisoflurane réduit la lésion ischémique cérébrale focale chez les rats et ľantagoniste (DPCPX) des récepteurs de ľadénosine A1 atténue la neuroprotection induite par le préconditionnement à ľisoflurane.Objectif Explorer le role des recepteurs de ľadenosine A1 dans la tolerance rapide a ľischemie cerebrale induite par le preconditionnement a ľisoflurane.Purpose: To investigate the role of the adenosine A 1 receptor in the rapid tolerance to cerebral ischemia induced by isoflurane preconditioning. Methods: Seventy-five rats were randomly assigned into five groups (n = 15 each): Control, 8-cyclopentyl-1,3-dipropulxanthine (DPCPX), Isoflurane, DPCPX+Isoflurane and Vehicle+Isoflurane groups. All animals underwent right middle cerebral artery occlusion (MCAO) for two hours. Isoflurane preconditioning was conducted one hour before MCAO in Isoflurane, DPCPX+Isoflurane and Vehicle+Isoflurane groups by exposing the animals to 1.5% isoflurane in 98% oxygen for one hour. In the Control and DPCPX groups, animals were exposed to 98% oxygen one hour before MCAO for one hour. A selective adenosine A 1 receptor antagonist, DPCPX, was administered (0.1 mg·kg –1 ) 15 min before isoflurane/oxygen

Cardioprotective Effects of Electroacupuncture Pretreatment on Patients Undergoing Heart Valve Replacement Surgery: A Randomized Controlled Trial

BACKGROUND Cardiac ischemia-reperfusion injury after cardiopulmonary bypass contributes to postoperative morbidity and mortality in patients with open-heart surgery. This randomized controlled trial was designed to address the protective effects of electroacupuncture (EA) pretreatment on myocardial injury in patients undergoing heart valve replacement surgery. METHODS Sixty patients with acquired heart valve disease were randomly allocated to the EA pretreatment group or the control group. Patients in the EA group received EA stimulus at bilateral Neiguan (PC 6), Lieque (LU 7), and Yunmen (LU 2) for 30 minutes each day for five consecutive days before surgery. Hemodynamic data, mechanical ventilation time, inotropic drug use in the intensive care unit, serum cardiac troponin I concentrations, morbidities, and mortalities were compared between the two groups. This trial is registered with ClinicalTrials.gov, number NCT00732459. RESULTS At 6 hours, 12 hours, and 24 hours after reperfusion, levels of serum cardiac troponin I were significantly decreased in the EA group (5.74 +/- 0.67, 6.22 +/- 0.66, and 5.21 +/- 0.58) compared with that in the control group (7.89 +/- 0.74, 8.34 +/- 1.08, and 7.57 +/- 0.89, p < 0.05). The EA pretreatment significantly reduced overall serum troponin I release at 6 hours, 12 hours, and 24 hours after aortic cross-clamp removal. Meanwhile, EA pretreatment also reduced the inotrope score at 12 hours, 24 hours, and 48 hours after the intensive care unit arrival and shortened intensive care unit stay time (p < 0.05). CONCLUSIONS The present study demonstrated that EA pretreatment may alleviate cardiac ischemia-reperfusion injury in adult patients undergoing heart valve replacements. This simple and convenient treatment has the potential to be used in the clinic for reducing myocardial injury in patients with heart valve replacement surgery.

Trans-activator of Transcription-mediated Delivery of NEP1-40 Protein into Brain Has a Neuroprotective Effect against Focal Cerebral Ischemic Injury via Inhibition of Neuronal Apoptosis

Background:The Nogo-66 antagonistic peptide (NEP1-40) is a potential candidate for therapeutic intervention of neuronal injury. However, delivery of the proteins across the blood–brain barrier is severely limited by its size and biochemical properties. The current study was designed to evaluate the transducible effects of the trans-activator of transcription (TAT) transduction system for NEP1-40 to cross the blood–brain barrier and to clarify whether intraperitoneal administration of TAT-NEP1-40 could protect cerebral neurons from ischemic injury. Methods:Adult male Sprague-Dawley rats were submitted to a 120-min focal ischemia and received an intraperitoneal injection of No-TAT-NEP1-40, TAT-NEP1-40, TAT-&bgr;-galactosidase, or vehicle. The existence of the proteins in the brain was analyzed with immunofluorescence and Western blot techniques at 6 h after injection. Brain ischemic injury was evaluated by neurologic deficit scores, infarction volumes, terminal deoxynucleotidyl transferase-mediated dUDP-biotin nick end labeling staining, and assay of caspase-3 activity. Results:Western blot analysis and immunofluorescence staining confirmed the presence of TAT-NEP1-40 protein in the brains 6 h after injection. Intraperitoneal injection of TAT-NEP1-40 could attenuate the numbers of terminal deoxynucleotidyl transferase-mediated dUDP-biotin nick end labeling–positive cells and activated caspase-3 positive cells, and increase the viability of the cells in the ischemic bounder zone, compared with that treated with No-TAT-NEP1-40, TAT-&bgr;-Gal, or vehicle. Furthermore, treatment with TAT-NEP1-40 significantly improved the neurologic outcomes and reduced the size of the infarction in rats. Conclusions:The results demonstrate that the TAT-NEP1-40 could be efficiently delivered into the rat brains and improve ischemia-induced neurologic outcomes through attenuating cell apoptosis in ischemic brains.