Knockdown of BMP7 induced oligodendrocyte apoptosis, demyelination and motor function loss.

BACKGROUND: Demyelinating diseases, including multiple sclerosis (MS) and spinal cord injury (SCI), lead to significant neurological deficits primarily due to the loss of oligodendrocytes (OLs). Bone Morphogenetic Protein 7 (BMP7) is expressed abundantly in the central nervous system and previous studies showed its protective effect in reducing OL loss. In this study, we aim to explore BMP7's potential as a biomarker and therapeutic target for demyelinating diseases by investigating its expression and effects on OLs and myelin sheath integrity. METHOD: We analyzed multiple Gene Expression Omnibus datasets for BMP7 expression profiles in demyelinating conditions such as MS and SCI. Experimentally, we employed a BMP7 knockdown model in rat spinal cords using adeno-associated virus8 vectors to specifically reduce BMP7 expression. Western blotting, immunofluorescence, and Nissl staining were used to assess the effect on OL and other types of cells. The structure of myelin sheath and locomotor function were evaluated using transmission electron microscopy and BBB scores, and statistical analysis included ROC curves and ANOVA to evaluate BMP7's diagnostic and therapeutic potential. RESULTS: BMP7 expression consistently decreased across various demyelinating models, and BMP7 knockdown led to increased OL apoptosis through the Smad1/5/9 pathway, with no apparent effect on other cell types. This reduction in OLs was associated with myelin degeneration, axonal damage, and impaired motor function. CONCLUSION: The study confirms BMP7's significant involvement in the pathophysiology of demyelinating diseases and supports its potential as a therapeutic target or biomarker. Future research should focus on therapeutic strategies to enhance BMP7 function and further investigate the mechanisms by which BMP7 supports myelin integrity.

Technical guideline for intra-prepontine cisternal drug delivery via spinal puncture through subarachnoid catheterization. / ç»è„ŠæŸ±æ¤Žé—´éš™ç©¿åˆºè››ç½‘è†œä¸‹è ”è„‘æ¡¥å‰æ± ç½®ç®¡è¯ç‰©è¾“æ³¨æŠ€æœ¯åº”ç”¨æŒ‡å—.

OBJECTIVES: The distribution characteristics of intrathecal drugs and the limitation of current catheterization techniques make traditional intrathecal analgesic treatment nearly useless for refractory craniofacial pain, such as trigemina neuralgia. This technical guideline aims to promote the widespread and standardize the application of intra-prepontine cisternal drug delivery via spinal puncture and catheterization. METHODS: A modified Delphi approach was used to work for this guideline. On the issues related to the intra-prepontine cisternal targeted drug delivery technique, the working group consulted 10 experts from the field with 3 rounds of email feedback and 3 rounds of conference discussion. RESULTS: For the efficacy and safety of the intra-prepontine cisternal targeted drug delivery technique, a consensus was formed on 7 topics (with an agreement rate of more than 80%), including the principles of the technique, indications and contraindications, patient preparation, surgical specifications for intra-prepontine cisternal catheter placement, analgesic dosage coordination, analgesic management, and prevention and treatment of complications. CONCLUSIONS: Utilizing the intra-prepontine cisternal drug infusion system to manage refractory craniofacial pain could provide advantages in terms of minimally invasive, secure, and effective treatment. This application can not only alleviate the suffering of individuals experiencing the prolonged pain but also support the maintenance of quality of life and dignity in their final moments, justifiing its widespread dissemination and standardized adoption in domestic and international professional fields.

Cerebrospinal Fluid Proteomic Profiles in Patients with Postherpetic Neuralgia.

The intrinsic mechanism of postherpetic neuralgia (PHN) remains unclear. Herein, we aimed to seek the hub proteins in the cerebrospinal fluid (CSF), which display significant changes between the PHN and nonpainful patients (Control). First, the proteomic results showed that compared with the Control-CSF, there were 100 upregulated and 50 downregulated differentially expressed proteins (DEPs) in the PHN-CSF. Besides, functional analyses including gene ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and gene set enrichment analysis (GSEA) revealed that biological processes and pathways including complement activation, infection, coagulation, and lipid metabolism were activated, while synaptic organization was suppressed. Next, the protein-protein interaction (PPI) analysis indicated that increased PLG, F2, APOA1, APOA2, SERPINC1, and KNG1 and reduced APOE, which were all enriched in the top pathways according to the KEGG analysis, were defined as hub proteins. Finally, three of the hub proteins, such as PLG, APOA1, and APOE, were reconfirmed in a larger cohort using both enzyme-linked immunosorbent assay (ELISA) and Western blotting methods. Above all, the results indicated that PLG, APOA1, and APOE and their involved processes such as infection, inflammation, cholesterol metabolism, and coagulation shall be potential therapeutic approaches. (The raw mass spectrometry proteome data and search results have been deposited to the iProx-integrated Proteome Resources (http://www.iprox.cn) with the data set identifier IPX0007372000.).

BMP7 alleviates trigeminal neuralgia by reducing oligodendrocyte apoptosis and demyelination.

BACKGROUND: BMP7 has been shown to have neuroprotective effects and to alleviate demyelination. However, its role in trigeminal neuralgia (TN) has not been well investigated. The current study aims to determine whether BMP7 plays a role in demyelination, its effects on pain behaviors and mechanism of action in rats with TN. METHODS: We used an infraorbital-nerve chronic-constriction injury (ION-CCI) to establish a rat model of TN. Adeno-associated viruses (AAVs) were injected into the rats to upregulate or downregulate BMP7. The mechanical withdrawal thresholds (MWT) of the injured rats were detected using Von Frey filaments. The changes in expression levels of BMP7 and oligodendrocyte (OL) markers were examined by western blotting, quantitative real-time PCR, immunofluorescence, and transmission electron microscopy. RESULTS: The ION-CCI induced mechanical allodynia, demyelination, and loss of OLs with a reduction of BMP7. Short-hairpin RNA (shRNA)-BMP7 that inhibited BMP7 expression also caused mechanical allodynia, demyelination, and loss of OLs, and its mechanism may be OL apoptosis. Overexpressing BMP7 in the trigeminal spinal subnucleus caudalis(VC) with AAV-BMP7 relieved all three phenotypes induced by the CCI, and its mechanism may be alleviating OLs apoptosis. Two signal pathways associated with apoptosis, STAT3 and p65, were significantly downregulated in the VC after CCI and rescued by BMP7 overexpression. CONCLUSION: BMP7 can alleviate TN by reducing OLs apoptosis and subsequent demyelination. The mechanism behind this protection could be BMP7-mediated activation of the STAT3 and NF-κB/p65 signaling pathway and subsequent decrease in OL apoptosis. Importantly, our study presents clear evidence in support of BMP7 as a possible therapeutic target for the treatment of TN.

Thalidomide Reduces Hemorrhage of Brain Arteriovenous Malformations in a Mouse Model.

BACKGROUND AND PURPOSE: Brain arteriovenous malformation (bAVM) is an important risk factor for intracranial hemorrhage. Current treatments for bAVM are all associated with considerable risks. There is no safe method to prevent bAVM hemorrhage. Thalidomide reduces nose bleeding in patients with hereditary hemorrhagic telangiectasia, an inherited disorder characterized by vascular malformations. In this study, we tested whether thalidomide and its less toxic analog, lenalidomide, reduce bAVM hemorrhage using a mouse model. METHODS: bAVMs were induced through induction of brain focal activin-like kinase 1 (Alk1, an AVM causative gene) gene deletion and angiogenesis in adult Alk1-floxed mice. Thalidomide was injected intraperitoneally twice per week for 6 weeks, starting either 2 or 8 weeks after AVM induction. Lenalidomide was injected intraperitoneally daily starting 8 weeks after AVM induction for 6 weeks. Brain samples were collected at the end of the treatments for morphology, mRNA, and protein analyses. The influence of Alk1 downregulation on PDGFB (platelet-derived growth factor B) expression was also studied on cultured human brain microvascular endothelial cells. The effect of PDGFB in mural cell recruitment in bAVM was explored by injection of a PDGFB overexpressing lentiviral vector to the mouse brain. RESULTS: Thalidomide or lenalidomide treatment reduced the number of dysplastic vessels and hemorrhage and increased mural cell (vascular smooth muscle cells and pericytes) coverage in the bAVM lesion. Thalidomide reduced the burden of CD68+ cells and the expression of inflammatory cytokines in the bAVM lesions. PDGFB expression was reduced in ALK1-knockdown human brain microvascular endothelial cells and in mouse bAVM lesion. Thalidomide increased Pdgfb expression in bAVM lesion. Overexpression of PDGFB mimicked the effect of thalidomide. CONCLUSIONS: Thalidomide and lenalidomide improve mural cell coverage of bAVM vessels and reduce bAVM hemorrhage, which is likely through upregulation of Pdgfb expression.

Soluble FLT1 Gene Therapy Alleviates Brain Arteriovenous Malformation Severity.

BACKGROUND AND PURPOSE: Brain arteriovenous malformation (bAVM) is an important risk factor for intracranial hemorrhage. Current therapies are associated with high morbidities. Excessive vascular endothelial growth factor has been implicated in bAVM pathophysiology. Because soluble FLT1 binds to vascular endothelial growth factor with high affinity, we tested intravenous delivery of an adeno-associated viral vector serotype-9 expressing soluble FLT1 (AAV9-sFLT1) to alleviate the bAVM phenotype. METHODS: Two mouse models were used. In model 1, bAVM was induced in R26CreER;Eng2f/2f mice through global Eng gene deletion and brain focal angiogenic stimulation; AAV2-sFLT02 (an AAV expressing a shorter form of sFLT1) was injected into the brain at the time of model induction, and AAV9-sFLT1, intravenously injected 8 weeks after. In model 2, SM22αCre;Eng2f/2f mice had a 90% occurrence of spontaneous bAVM at 5 weeks of age and 50% mortality at 6 weeks; AAV9-sFLT1 was intravenously delivered into 4- to 5-week-old mice. Tissue samples were collected 4 weeks after AAV9-sFLT1 delivery. RESULTS: AAV2-sFLT02 inhibited bAVM formation, and AAV9-sFLT1 reduced abnormal vessels in model 1 (GFP versus sFLT1: 3.66±1.58/200 vessels versus 1.98±1.29, P<0.05). AAV9-sFLT1 reduced the occurrence of bAVM (GFP versus sFLT1: 100% versus 36%) and mortality (GFP versus sFLT1: 57% [12/22 mice] versus 24% [4/19 mice], P<0.05) in model 2. Kidney and liver function did not change significantly. Minor liver inflammation was found in 56% of AAV9-sFLT1-treated model 1 mice. CONCLUSIONS: By applying a regulated mechanism to restrict sFLT1 expression to bAVM, AAV9-sFLT1 can potentially be developed into a safer therapy to reduce the bAVM severity.

[Role of the autophagy in the treatment of neuropathic pain with pulsed radiofrequency].

OBJECTIVE: To detect the effect of pulse radiofrequency (PRF) treatment on the neuropathic pain established by L5-spinal nerve ligation (SNL) on rats, and to investigate if PRF treatment would affect the expression of autophagy related protein LC3 and autophagy related receptor P62 at the dorsal horn.
 Methods: A total of 36 male Sprague-Dawley rats were randomly divided into 3 groups: a Sham group, a SNL group, and a SNL+PRF group. The 50% paw withdrawal mechanical threshold (PWMT) was detected at 1 day before and 1, 3, 7, 14 and 28 days post-operation by using Von-Frey filaments. The autophagy related protein LC3 and autophagy related receptor P62 were investigated by Western blot.
 Results: Compared with the Sham group, the PWMT significantly decreased in the SNL group at each time points (P<0.05); in SNL+PRF group, PRF treatment could elevate the PWMT at the 1st day post-operation and lasted for 28 days (P<0.05). What's more, SNL could elevate the LC3-II and P62 levels at the 7th day post-operation (P<0.05), which were decreased by the PRF treatment (P<0.05).
 Conclusion: PRF treatment could improve SNL-induced the neuropathic pain, which might be partly due to the regulatory effects on the autophagy levels at the spinal dorsal horn.

Efficacy Analysis of Temporary Spinal Cord Stimulation in the Treatment of Refractory Postherpetic Neuralgia.

BACKGROUND: Spinal cord stimulation can be considered in PHN patients if conservative treatment is not effective. However, the long-term pain outcomes of temporary (7-14 days) spinal cord stimulation (tSCS) in refractory PHN patients with a course of more than 3 months have not been  documented. OBJECTIVES: To investigate the efficacy of tSCS as a treatment for refractory PHN. STUDY DESIGN: Retrospective study. SETTING: Pain Department in a university hospital. METHODS: A total of 52 patients with refractory PHN were treated with tSCS between March 2018 and February 2021. Their medical records were collected, and the patients were divided into 3 groups according to the course of their disease into the medium-term group, long-term group and ultra-long-term group. The changes in the numeric rating scale (NRS) scores, Pittsburgh sleep quality index (PSQI) responses, pain relief rate, postoperative efficiency and patients' use of analgesics were recorded before the operation, 3 days, 10 days, one month, 3 months, 6 months and 12 months after the operation. RESULTS: The average NRS scores, the maximum NRS scores and the PSQI scores at 3 days, 10 days, one month, 3 months, 6 months and 12 months after the operation were significantly lower than those before the operation (P < 0.05). The average NRS scores and the maximum NRS scores of all groups increased significantly from one month to 6 months compared to those at 10 days after the tSCS treatment, and they decreased significantly at 12 months compared with 6 months post-operation. The average NRS scores of the medium-term and long-term group were significantly lower than that of the ultra-long-term group at 1-3 months after the operation, and the maximum NRS scores at one month, 3 months and 12 months after the operation were also significantly lower in the medium-term and long-term group compared to the ultra-long-term group. The average PSQI scores at 1-12 months after the operation were not significantly higher than that at 10 days after the operation, but it decreased significantly at 12 months compared with 6 months after the operation. Among the 3 groups, the PSQI scores of the medium-term and long-term group were significantly lower than those of the ultra-long-term group at 6 months after the operation. The postoperative pain relief rate ranged from 41.51%-59.81%, and the total effective rate was 42.31%-69.23%, and there was no significant difference among the 3 groups. Some patients still needed analgesics at 12 months after the operation, but the number of patients who were taking medications post-operation was significantly lower than that before the operation. LIMITATIONS: This is a single-center retrospective study with the inability to completely control for variables. Additionally, the number of cases is small and the follow-up duration is short. CONCLUSION: tSCS can be used as a safe and effective method to relieve refractory PHN, and the curative effect is substantially higher in patients with a disease course of 3-12 months compared to that in patients with a course of more than 12 months.

Gadolinium-based MR cisternography with prepontine cisternal routine for evaluating distribution pattern of intrathecal targeted drug delivery in pain management.

Gadolinium-based MR cisternography has been mainly applied in clinical evaluation of cerebrospinal fluid leaking, that is conducted by intrathecal administration of contrast media. Recently, we have reported one novel technique of intrathecal targeted drug delivery with prepontine cisternal routine to treat orofacial cancer pain. The aim of this study was to examine the distribution pattern of this intrathecal drug delivery strategy. Here, we introduce one case who suffered severe orofacial pain caused by sublingual gland tumor, and successfully attenuated by prepontine cisternal administration of analgesic agents. To assess the distribution of intrathecal drugs, postoperative MR images of brain, cervical, thoracic, and lumbar segments in axial, coronal, and sagittal planes were obtained after application of gadolinium. The perfusion rate of contrast medium was set at 0.01 mmol per hour for 24 hours prior to MR scanning. In the T1-weighted images, we can identify contrast spread not only locating around the site of the intrathecal catheter tip, but also concentrated to the lateral sides. None obvious side effect was found after intrathecal injection of contrast media. Thus, our finding demonstrated the local distribution phenomenon of intrathecal drugs through prepontine cisternal access, and the bilateral perfusion pattern may provide insights underlying the analgesic mechanism of trigeminal pain provided by this novel intrathecal therapy. Gadolinium-based MR cisternography may serve as a potential tool to confirm the therapeutic effect of intrathecal targeted drug delivery via prepontine cisternal routine in orofacial pain management.

[Limb ischemic preconditioning reduces rabbit hepatic ischemia-reperfusion injury through inhibition the phosphorylation of proteins in the MAPK signal pathway in the late phase].

OBJECTIVE: To investigate the liver protection mechanisms of MAPK signaling pathway of limb ischemia preconditioning in the late phase. METHODS: Thirty-six adult male New Zealand white rabbits, weighing 1.8-2.0 kg, were randomly divided equally into 3 groups: group C (sham operation), group L (liver ischemia-reperfusion 24 h after limb ischemia preconditioning), group IR (liver ischemia-reperfusion without limb ischemia preconditioning). Serum alanine transaminase (ALT) was measured during ischemia reperfusion. The tissue and cell injury of liver were examined by optical and electron microscopy. Activation of P38MAPK, P44/P42MAPK, and JNK in hepatic tissue was assessed by western blot after 30 min of reperfusion. RESULTS: Serum ALT and cell injury in the liver as examined by optical and electron microscopy was decreased in group L as compared with the group IR. Phosphorylation of P38MAPK, P44/ P42MAPK, and JNK were all increased significantly after 30 min of reperfusion. Phosphorylation of P38MAPK and JNK was reduced by limb ischemia pre-treatment. CONCLUSION: Limb ischemia pre-treatment can induce the late phase of preconditioning in rabbit liver through the inhibition of the phosphorylation of P38MAPK and JNK.

Prepontine cisternal routine for intrathecal targeted drug delivery in craniofacial cancer pain treatment: technical note.

Intrathecal targeted drug delivery provides effective relief for cancer-related pain. However, its validation in management of craniofacial pain remains much less widely practiced, mainly due to the localized diffusion of analgesic agent with current approach. Here, we report our experience of prepontine cisternal routine for placement and implantation of intrathecal targeted drug delivery in two cases of cancer-related craniofacial pain. Lumbar cannulation was applied and the intrathecal catheter tip was positioned at the prepontine cistern under fluoroscopic guidance during the surgical implantation. Postoperative imaging confirmed that the catheter tip was successfully placed in the prepontine cisternal space. Satisfactory control of pain was achieved after intrathecal therapy, with significant reduction of background and breakthrough cancer pain. None obvious complications were observed in this study. Thus, our novel intrathecal routine may provide an alternative option for craniofacial pain caused by tumor, who were insufficiently treated by oral analgesic agents.

[Proteomics analysis of adenosine A1 receptor agonist-induced delayed myocardial protection in rabbits].

OBJECTIVE: To investigate the changes of myocardial protein expression profiles in 2-chloro-N6-cyclopentyladenosine (CCPA), an adenosine A1 receptor agonist-induced delayed myocardial protection in New Zealand rabbits . METHODS: A total of 8 rabbits were randomly divided into a CCPA group (CCPA group) and a normal saline group (NS group). CCPA and NS were infused into rabbits in the CCPA group and the NS group respectively. Twenty-four hours later, the rabbits were subjected to 30 min left anterior descending coronary artery occlusion and were reperfused for 2 hours, then the ischemic zone tissues of left ventricle were sampled for proteomic analysis.A total of 12 other New Zeland rabbits were divided into a sham group (Sham group), a normal saline group (NS group) and a CCPA group (CCPA group). The expression of αB-crystalline, one of the differential proteins, was confirmed by Western blot. RESULTS: Analysis of two dimensional gel electrophoresis showed that the expression of 55 protein spots were different between the two groups, 17 protein spots were preliminarily identified with the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and Mascot and Expasy bioinformatics software. These proteins included stress proteins, metabolism-associated proteins, signal transduction pathway-related proteins, ionophorous proteins, immunity-associated proteins, and so on. Western blot showed that the expression of αB-crystalline was significantly up-regulated in the CCPA group. CONCLUSION: The myocardial protein expression profiles are changed markedly in the preconditioning late phase of CCPA .The differential proteins might be involved in the delayed cardioprotection induced by CCPA.

Expression of bone morphogenetic proteins and their receptors in the normal adult rat spinal cord.

OBJECTIVE: To observe the expression distribution of bone morphogenetic proteins (BMP) in the spinal cord of normal adult rats. METHODS: Expression of BMP2, BMP4, and BMP7, and their receptors BMPR Ia, BMPR Ib, and BMP II were detected by immunochemistry analysis in the spinal cord of normal adult rats. RESULTS: Expression of BMPR Ia or BMPR Ib was observed in the motor neurons of the anterior horn, sensory neurons of the dorsal horn, oligodentrocytes, some microglia, and some astrocytes. Expression of receptor BMPR II was found in the oligodentrocytes and motor neurons in the gray matter of anterior horn. It was also expressed in some glial fibrillary acidic protein (GFAP)-positive astrocytes in the white matter but not in the gray matter. BMP2 and BMP4 were not expressed in the spinal cord of normal adult rats by immunohistochemistry. BMP7 was expressed in all the APC-positive oligodentrocytes, all the NeuN-positive motor neurons in the anterior horn, and some astrocytes in the normal spinal cord. Phosphated pSmad 1/5/8 protein was expressed in all the oligodentrocytes, all the neurons, and some astrocytes, especially in the GFAP-positive astrocytes which were RC2-positive radial glia in the subventricular zone. CONCLUSION: BMP7, BMP receptors, and phosphated pSmad 1/5/8 are expressed in many types of cells whereas BMP2 and BMP4 are not expressed in the spinal cord of normal adult rats, which suggests an important function of BMP signal pathway in the neuron and glia of spinal cord.

[Proteomic analysis of morphine rabbit myocardium with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry].

OBJECTIVE: To analyze the morphine rabbit myocardium with matrix-assisted laser desorption/ ionization time-of-flight mass spectrometry (MALDI-TOF-MS). METHODS: Six New Zealand white rabbits were randomly assigned to a control group (Group C) and a morphine group (Group M). Group C were pretreated with bolus injection of saline 1 mL/kg. Group M were pretreated with bolus injection of morphine 3 mg/kg. The myocardium tissue proteins of the rabbits 24 hours after the injection of morphine or saline preconditioned were extracted and separated by two dimensional gel electrophoresis(2-DE), and the images were analyzed and different proteins were found. Some of the different proteins were determined with MALDI-TOF-MS. RESULTS: There were 51 protein spots that displayed quantitative changes in expression (P < 0.05), 15 protein spots were chosen for MS analysis, and 8 proteins were preliminarily identified.They were aldose reductase, zinc finger protein 312, src related tyrosine kinase, carbonic anhydrase 12 precursor, electron transfer flavoprotein beta-subunit, glyceraldehyde-3-phosphate dehydrogenase, tumor necrosis factor ligand superfamily member 11 and transmembrane emp24 domain-containing protein. CONCLUSION: These proteins may be involved in the cardioprotection of morphine preconditioning.

[Delayed protective effect of morphine preconditioning on rabbit myocardium].

OBJECTIVE: To investigate whether morphine preconditioning has the delayed protective effect on rabbit myocardium. METHODS: Thirty New Zealand white rabbits were randomly divided into a NS group, a Mor-12 group and a Mor-24 group (n=10). In the Mor-12 group and Mor-24 group, morphine (3 mg/kg) was infused into rabbits, while the same volume of normal saline (NS) was given to rabbits in the NS group. Twelve hours after morphine infusion in the Mor-12 group, 24 h after NS or morphine infusion in the NS group and Mor-24 group, rabbits were subjected to 30 min left anterior descending coronary artery occlusions and were reperfused for 120 min. In 8 of the 10 rabbits in each group, arterial blood samples were taken before the ischemia (T1), 30 min after the ischemia (T2) and 120 min after the reperfusion (T3) to determine the concentration of cardiac troponin I (cTnI), and the myocardial infarct area was determined at the end of reperfusion. In the other 2 of the 10 rabbits in each group,the cell ultramicro-structure injury of myocardium was examined by electron microscope at the end of reperfusion. RESULTS: The concentration of cTnI at T2 and T3 in the Mor-24 group was lower than that in the NS group and Mor-12 group.The myocardial infarct size, and cell ultramicrostructure injury of myocardium in the Mor-24 group were decreased compared with the NS group and Mor-12 group. CONCLUSION: Morphine preconditioning has delayed protective effect on rabbit myocardium.

[Effect of isoflurane delayed preconditioning on myocardial ischemia reperfusion injury in rabbits].

OBJECTIVE: To investigate the protective effect of isoflurane delayed preconditioning on myocardial ischemia reperfusion injury and the potential mechanism in rabbits. METHODS: Thirty New Zealand male white rabbits were randomly assigned to 3 groups: Control group; I/R group; and 2.0% isoflurane group. Isoflurane group was exposed to 2.0% isoflurane-100% oxygen for 2 hours. Control group and I/R group were exposed to 100% oxygen for 2 hours and served as untreated controls. Twenty-four hours later I/R group and isoflurane group underwent 40 minutes of coronary occlusion followed by 2 hours of reperfusion. Blood samples were taken from the arterial line at 20 minutes before the occlusion(T1), 20 minutes after the occlusion(T2), 40 minutes after the occlusion(T3), 1 hours after the reperfusion(T4), and 2 hours after the reperfusion(T5) to determine the plasma level of TNF-alpha. At the end of the reperfusion, infarct size and area at risk were defined by Evans and TTC staining. The heart was harvested and levels of the p38MAPK activity were determined by Western blot, and ultrastructures were observed under the electron microscope. RESULTS: The p38MAPK activity of isoflurane group was significantly lower than that of I/R group (P<0.05). Isoflurane significantly (P<0.05) reduced the infarct size(19.7%+/-2.8% in isoflurane group) of the left ventricular area at risk as compared with the controls (37.8%+/-1.7% in I/R group).The injury of I/R group was worse than that of isoflurane group under the light microscope. Isoflurane group had a lower level of TNF-alpha than I/R group. CONCLUSION: Isoflurane can inhibit p38MAPK activity during myocardial ischemia reperfusion and modulate the cytokine expression, which may be one of the molecular mechanisms of isoflurane delayed preconditioning on cardioprotection.

[Protective effect of heat-shock protein 27 on myocardium with isoflurane preconditioning in myocardial ischemia/reperfusion injury of myocardium in rabbit].

OBJECTIVE: To investigate the mechanism and the protective effect of heart-shock protein 27 (HSP27) on rabbit myocardium with isoflurane preconditioning in myocardial ischemia/reperfusion (I/R) injury. METHODS: Thirty New Zealand white rabbits were randomly assigned to three groups (each n = 10):(1)Sham operation group (C group); (2)I/R group; (3)Two percent in volume is of isoflurane group (S group). S group was exposed to 2.0% isoflurane-pure oxygen for 2 hours. C group and I/R group were exposed 2 hours to pure oxygen to serve as untreated controls. Twenty-four hours later the rats in I/R group and S group underwent 40 minutes of coronary occlusion followed by 120 minutes of reperfusion. At the end of the reperfusion, infarct size (IS) was defined by Evan's blue and triphenyltetrazolium chloride (TTC) staining. Blood samples were taken from arterial line for determination of malondialdehyde (MDA) levels. Western Blotting was used to determine the expression of HSP27 and nuclear factor-KappaB (NF-KappaB) in myocardium. RESULTS: Isoflurane preconditioning could decrease I/R induced myocardial infarct size [(19.7 +/- 2.8)% vs.(37.8 +/- 1.7)%]. This was accompanied by an increase in the expression in HSP27 [(84.5 +/- 4.3) gray scale value vs. (53.1 +/- 3.8) gray scale value] and a decrease in NF-KappaB [(58.6 +/- 4.2) gray scale value vs. (119.3+/-5.6) gray scale value] and MDA [(5.24 +/- 0.45)kU/L vs. (9.42 +/- 0.83)kU/L]. CONCLUSION: The expression of HSP27 induced by isoflurane preconditioning plays an important role in protecting myocardium against ischemia/reperfusion injury.

Activation of Alpha-7 Nicotinic Acetylcholine Receptor Reduces Brain Edema in Mice with Ischemic Stroke and Bone Fracture.

Stroke is an important risk factor for bone fracture. We showed previously that bone fracture at the acute stage of ischemic stroke worsens, and activation of α-7 nicotinic acetylcholine receptor (α-7 nAchR) improves, stroke recovery by attenuating inflammation. We hypothesized that activation of α-7 nAchR also improves the blood-brain barrier (BBB) integrity. Permanent distal middle cerebral artery occlusion (pMCAO) was performed on C57BL/6J mice followed by tibia fracture 1 day later. Mice were treated with 0.8 mg/kg PHA 568487 (PHA, α-7 nAchR-specific agonist), 6 mg/kg methyllycaconitine (MLA, α-7 nAchR antagonist), or saline 1 and 2 days after pMCAO. Brain water content, the expression of monoamine oxidase B (MAO-B), and tight junction protein (claudin-5) were assessed. We found that tibia fracture increased water content in the ischemic stroke brain (p = 0.006) and MAO-B-positive astrocytes (p < 0.001). PHA treatment reduced water content and MAO-B-positive astrocytes and increased claudin-5 expression in stroke and stroke + tibia fracture mice (p < 0.05), while MLA had the opposite effect. Our findings suggest that in addition to inhibiting inflammation, activation of α-7 nAchR also reduces brain edema, possibly through diminished astrocyte oxidative stress and improved BBB integrity. Thus, the α-7 nAchR-specific agonist could be developed into a new therapy for improving recovery of patients with stroke or stroke + bone fracture.

Correction: Bone Fracture Pre-Ischemic Stroke Exacerbates Ischemic Cerebral Injury in Mice.

[This corrects the article DOI: 10.1371/journal.pone.0153835.].