Objective:To identify the differentially expressed long-chain non-coding RNA(lncRNA) and mRNA using ribonucleic acid sequencing(RNA-seq), and construct a competing endogenous RNA(ceRNA) regulatory network in mice with sepsis-associated encephalopathy.Methods:Ten clean-grade healthy male C57BL/6 mice, aged 6-8 weeks, weighing 20-25 g, were divided into 2 groups( n=5 each) using a random number table method: sham operation group(group Sham) and sepsis group(group Sepsis). Sepsis was induced by cecal ligation and puncture(CLP) in group Sepsis, while group Sham only underwent laparotomy without CLP. Morris water maze test and contextual fear conditioning test were performed to detect the cognitive function on 1 day before CLP and 3 days after CLP. Three mice were randomly sacrificed in group Sham, and 3 mice with the worst results in the cognitive function test were sacrificed in group Sepsis. The hippocampal tissues were obtained for RNA-seq via the BGISEQ-500 platform, and the differentially expressed mRNA and lncRNA were identified. The differentially expressed mRNAs and lncRNAs were visualized and analyzed by Dr. Tom platform provided by Shenzhen BGI Technology Service Co., Ltd., and the ceRNA regulatory network was constructed using the online visualization tool Cytoscape software. Results:Compared with group Sham, the escape latency was significantly prolonged, and the percentage of time of staying at the target quadrants and percentage of time spent freezing were decreased in group Sepsis( P<0.05). A total of 62 differentially expressed lncRNAs were obtained from RNA-seq, of which the expression of 45 lncRNAs was up-regulated and the expression of 17 lncRNAs was down-regulated.There were 282 differentially expressed mRNAs identified from RNA-seq, of which the expression of 173 mRNAs was up-regulated, and the expression of 109 mRNAs was down-regulated.Gene Ontology enrichment analysis revealed that the differentially expressed mRNAs were involved in biological processes such as memory, learning or memory, inflammatory responses, regulation of aging-related behavioral decline, and regulation of synaptic plasticity. Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis showed that differentially expressed mRNAs were enriched in IL-17 signaling pathway, TNF signaling pathway, NF-κB signaling pathway and etc. KDA analysis was performed on the differentially expressed mRNAs to identify the key driver genes, and the results showed that Ch25h, Il6ra, Lcn2, Sgk1, Nr4a3, Osm, Saa3, Ccl7, Sqle, Dhcr24 were the key SAE genes.A competing endogenous RNA regulatory network was successfully constructed based on 9 lncRNAs, 28 mRNAs and 134 miRNAs in the hippocampus of mice with SAE. Conclusions:The results of RNA-seq find that 10 mRNAs including Ch25h, Il6ra, Lcn2, Sgk1, Nr4a3, Osm, Saa3, Ccl7, Sqle, Dhcr24 and lncRNAs such as Rian, Gm35874 and Gm34347 are key genes regulating SAE in mice. Meanwhile, a ceRNA regulatory network based on lncRNA-miRNA-mRNA is successfully constructed in the hippocampus of mice with SAE.

Abstract To study the antiviral effect ofZedoary TurmericOil Injection on novel coronavirus, SARS-CoV-2 viroid cell lines were preparedin vitroand treated with different concentrations of Zedoary Oil. The cell number and relative fluorescence value(RLU) were observed and measured, and the 50% effective inhibitory concentration(IC 50) was calculated. Four patients with Coronavirus Disease 2019 were clinically included, including 2 in the control group and 2 in the experimental group. The control group received conventional treatment, and the experimental group receivedZedoary TurmericOil Injection in addition to conventional treatment. The nucleic acid conversion rate, conversion time, pulmonary imaging changes, fever reduction time, clinical improvement time and adverse events of the patients were observed.In vitroexperiment, the relative fluorescence value decreased with increasing concentration ofZedoary TurmericOil, which was significantly different from that of the control group(P<0.05). The IC50 was 0.26 μg/ml.In vivostudy, the novel coronavirus nucleic acid in stool of case 1 in the test group turned negative in 3 days, the cough symptom of case 2 was significantly relieved, and there was obvious absorption in pulmonary imaging. The negative conversion time of novel coronavirus nucleic acid in the control group was 5 and 7 days respectively. No adverse events occurred in the experimental group.Zedoary TurmericOil had strong inhibitory effect on SARS-COV-2 virusin vitrowhich was dose-dependent.In vivotreatment of COVID-19,Zedoary TurmericOil Injection combined with conventional treatment can improve the cough caused by SARS-COV-2 infection, promote SARS-COV-2 to turn negative, promote absorption of lung lesions, and reduce lung injury, with no obvious adverse events.

Objective:To investigate the neuroprotective effect of long-term prophylactic use of buphthalein on mice with permanent distal middle cerebral artery occlusion and its relationship with the nuclear factor erysid 2 related factor 2 (Nrf2) pathway.Methods:Nrf2 + /+ wild-type and Nrf2 -/- knockout mice were randomly divided into control group (equal volume vegetable oil), low-dose butylphthalide group (20 mg/kg) and high-dose butylphthalide group (60 mg/kg), with 6 mice in each group. The drug was administered once a day by gavage for 1 month, and then a permanent middle cerebral artery occlusion model was induced by electrocoagulation. After the model was made, the drug was continued and the mice were sacrificed on the 10 th day. The modified Longa grading scale and the rotating rod test were used to evaluate neurological deficits on the 3 rd and 10 th day after the model was made. After the mice were sacrificed, the cerebral infarct volume was measured by triphenyltetrazolium chloride staining. The brain water content was measured by dry and wet weight method. The expression of Nrf2 pathway related factors, including Nrf2, heme oxygenase 1 (HO-1) and NAD(P)H quinone oxidoreductase 1 (NQO1) were measured by quantitative real-time PCR and Western blotting. Results:On the 10 th day after modeling, compared with the Nrf2 -/- control group, the neurological deficit was significantly milder, the volume of cerebral infarction and brain water content were significantly smaller, and the mRNA and protein levels of Nrf2, HO-1 and NQO1 were significantly higher in the Nrf2 + /+ control group, and the differences were statistically significant ( P<0.05). For Nrf2 + /+ mice, compared with the control group, the cerebral infarct volume was significantly reduced ( P<0.05), the brain water content was significantly reduced ( P<0.05), and the neurological function recovery was significantly better ( P<0.05), and the levels of Nrf2, HO-1, and NQO1 mRNA and protein were significantly higher in the high-dose butylphthalide group (all P<0.05). For Nrf2 -/- mice, there were no significant differences in neurological function, cerebral infarction group volume, brain water content, Nrf2, HO-1, NQO1 mRNA and protein levels among the groups. Conclusion:Long-term butylphthalide pretreatment can significantly improve the neurological function, reduce cerebral infarction volume, reduce brain water content, and increase Nrf2, HO-1, NQO1 mRNA and protein expression levels in mice with permanent distal middle cerebral artery occlusion, suggesting butylphthalide may play a neuroprotective effect by up-regulating the expression of Nrf2 gene and its downstream antioxidant stress factors HO-1 and NQO1.

Objective:To investigate the relationship between autophagy and nuclear factor E2 related factor 2(Nrf2) signaling pathway during high glucose-induced damage to Schwann cells.Methods:RSC96 were cells cultured in vitro and seeded in 96-well plates (1×10 4 cells/ml, 200 μl/well) or in 6-well plates (1×10 6 cells/ml, 2 ml/well) for 48 h. The cells were divided into 3 groups ( n=25 each) using a random number table method: control group (group C), high glucose group (group H) and high glucose+ autophagy agonist rapamycin group ( group H+ RAP). The cells were cultured in the common culture medium in group C. In group H, 50 mmol/L of glucose was added to the culture medium.In group H+ RAP, 50 mmol/L of glucose and 5 μmol/L rapamycin were added to the culture medium.At 48 h of incubation, the growth of cells was observed with inverted phase contrast microscope, the cell viability was measured using MTT method, apoptosis rate was determined by flow cytometry, malondialdehyde (MDA) content was determined by thiobarbituric acid method, superoxide dismutase (SOD) activity was detected using xanthine oxidase method, and the expression of Nrf2, P62 and microtubule-associated protein 1 light chain 3 Ⅱ (LC3 Ⅱ) was determined by Western blot. Results:Compared with group C, the cell viability and SOD activity were significantly decreased, apoptotic rate and MDA content were increased, and expression of Nrf2, P62 and LC3Ⅱ was up-regulated in group H and group H+ RAP ( P<0.05). Compared with group H, the cell viability and SOD activity were significantly increased, apoptosis rate and MDA content were decreased, the expression of Nrf2 and LCII was up-regulated and P62 expression was down-regulated in group H+ RAP ( P<0.05). Conclusion:Enhanced autophagy can activate Nrf2 signaling pathway, which is the endogenous protective mechanism of Schwann cell injury induced by high glucose.

Objective:To evaluate the effect of hydrogen on lipopolysaccharide (LPS)-caused inflammatory responses in BV-2 microglia and the role of autophagy.Methods:The BV-2 microglial cells cultured in vitro were seeded in 6- or 96-well plates and were divided into 4 groups ( n=24 each) using a random number table method: control group (group C), group LPS, hydrogen-rich medium group (group H) and autophagy inhibitor 3-methylpurine group (group 3-MA). In group C, cells were cultured in MEM culture medium supplemented with 15% fetal bovine serum for 24 h. In group LPS, LPS was added at a final concentration of 1 μg/ml, and cells were incubated for 24 h. In group H, LPS was added at a final concentration of 1 μg/ml, the culture medium was replaced with a hydrogen-rich medium at a final concentration of 0.6 mmol/L, and cells were incubated for 24 h. In group 3-MA, 3-methylpurine was added at a final concentration of 2 mmol/L, and the subsequent treatment was similar to those previously described in group H. The cell survival rate was detected by CCK-8 assay.The concentrations of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), IL-10 and transforming growth factor-β (TGF-β) in supernatant were detected by enzyme-linked immunosorbent assay.The percentage of ionized calcium binding adaptor molecule-1 (Iba-1) +, Iba-1 + CD86 + and Iba-1 + CD206 + cells was detected by flow cytometry.The expression of microtubule-associated protein 1 light chain 3 Ⅰ (LC3 Ⅰ), LC3Ⅱ, Beclin-1 and p62 was detected by Western blot, and the ratio of LC3Ⅱ/LC3Ⅰ was calculated. Results:There was no significant difference in the cell survival rate among the four groups ( P>0.05). Compared with group C, the concentrations of TNF-α, IL-6, IL-10 and TGF-β and percentage of Iba-1 +, Iba-1 + CD86 + and Iba-1 + CD206 + cells were significantly increased in LPS, H and 3-MA groups, the LC3Ⅱ/LC3Ⅰ ratio and Beclin-1 expression was significantly down-regulated, and p62 expression was up-regulated in LPS and 3-MA groups, and the ratio of LC3LC3Ⅱ/LC3Ⅰ and Beclin-1 expression was significantly up-regulated, and p62 expression was down-regulated in group H ( P<0.05). Compared with group LPS, the concentrations of TNF-α and IL-6 were significantly decreased, the concentrations of IL-10 and TGF-β were increased, the percentage of Iba-1 + and Iba-1 + CD86 + cells were decreased, the percentage of Iba-1 + CD206 + cells was increased, the LC3Ⅱ/LC3Ⅰ ratio and Beclin-1 expression was up-regulated, and p62 expression was down-regulated in group H ( P<0.05), and no significant change was found in the above indexes in group 3-MA ( P>0.05). Compared with group H, the concentrations of TNF-α and IL-6 were significantly increased, the concentrations of IL-10 and TGF-β were decreased, the percentage of Iba-1 + and Iba-1 + CD86 + cells was increased, the percentage of Iba-1 + CD206 + cells was decreased, the LC3Ⅱ/LC3Ⅰ ratio and Beclin-1 expression was down-regulated, and p62 expression was up-regulated in group 3-MA ( P<0.05). Conclusion:The mechanism by which hydrogen reduces LPS-caused inflammatory responses in BV-2 microglia is related to enhancing autophagy and inhibiting microglial activation.

Objective: To evaluate the role of DNA methylation in sepsis-associated encephalopathy in mice. Methods: A total of 144 clean-grade healthy male C57BL/6 mice, aged 6-8 weeks, weighing 20-25 g, were divided into 4 groups (n=36 each) using a random number table method: sham operation group (group Sham), sepsis group (group Sepsis), sham operation plus S-adenosyl methionine (SAM) group (group Sham+ SAM) and sepsis plus SAM group (group Sepsis+ SAM). Sepsis was produced by cecum ligation and puncture (CLP). In Sham+ SAM and Sepsis+ SAM groups, DNA methylated methyl donor SAM 100 mg/kg was intraperitoneally injected at 1 h before operation and 12 h after operation, while the equal volume of normal saline was given instead in Sham and Sepsis groups.The cognitive function was assessed using Y-maze and contextual fear conditioning test at 1, 3 and 7 days after CLP.Mice were sacrificed at 1, 3 and 7 days after CLP, and the hippocampal tissues were taken for determination of genome-wide DNA methylation (by colorimetric assay) and expression of DNA methyltransferase enzymes (DNMT1, DNMT3a, and DNMT3b), ten-eleven translocation (TET) enzymes (TET1, TET2 and TET3) and thymine-DNA glycosylase (TDG) mRNA (by fluorescent quantitative real-time). Results: Compared with group Sham, the time of staying at the novel arm was significantly shortened, the percentage of time spent freezing and the total number of entries into each arm were reduced, genome-wide DNA methylation in hippocampal tissues was decreased at 1, 3 and 7 days after CLP, the expression of DNMT1, DNMT3a, TET1, TET2, TET3 and TDG was up-regulated, and the expression of DNMT3b was down-regulated in group Sepsis (P<0.05). Compared with group Sepsis, the time of staying at the novel arm was significantly prolonged, the percentage of time spent freezing and the total number of entries into each arm were increased, the expression of DNMT1, DNMT3a, TET1, TET2, TET3 and TDG mRNA was down-regulated, and the expression of DNMT3b was up-regulated in group Sepsis+ SAM(P<0.05). Conclusion DNA methylation is involved in the development of sepsis-associated encephalopathy in mice.

Objective Septal reduction therapies were recommended for drug-refractory patients with hypertrophic ob-structive cardiomyopathy(HOCM).To explore and compare the effectiveness and safety in patients with hypertrophic obstruc-tive cardiomyopathy(HOCM) treated with surgery myectomy(SM) and alcohol septal ablation(SA).Methods The clinical data of 260 patients performed SA(n=184) or SM(n=76)between September 2002 and September 2014 in our institute were retrospectively reviewed.The t-test, rank sum test and chi-square test were used to compare the differences between the two groups, and the Cox regression model was used for multivariate survival analysis.All-cause mortality, cardiac cause death(peri-operative death were included ) , heart function improvement , procedure-related complications and permanent pacemaker de-pendence( PPM) constituted the main contents of this study .Results Compared with patients treated with SM , patients un-dergone SA were poor heart function status(2.97 ±0.29 vs 2.50 ±0.56, P =0.01), more prevalence of atrial fibrillation( 15.14％ vs 6.80％, P=0.046) and longer follow-up period[(5.4 ±3.8) years vs(2.5 ±2.2) years, P =0.01)].All-cause mortality for SA and SM were 3.3％ and 14.5％ respectively(P=0.001).The fatal cardiac events of SA and SM were 1.63％ and 13.16％ respectively(P<0.001).Sudden cardiac arrest were the main cardiac cause death for both patients trea-ted with SA and SM.The cardiac death of left ventricular systolic dysfunction was main found in patients treated with SM . Heart function improvement(NYHA) after SA and SM were 1.23 ±0.61 and 0.88 ±0.64 respectively(all P <0.01).And SA had a lower procedure-related PPM implantation(1.63％ vs 4.20％, P<0.05).Conclusion Our results shown that SA have survival advantage, lower PPM and similar heart function improvement compared with SM for refractory patients with HOCM.

Objective To investigate the effect of high frequency (10 Hz),low frequency (1 Hz) and theta burst stimulation (TBS) mode of repetitive transcranial magnetic stimulation (rTMS) on the recovery of motor function in hemiplegic patients following acute ischemic stroke.Methods Seventy-two patients with hemiplegia after acute ischemic stroke were randomly grouped with the random number table.They were treated with low frequency (n=18),high frequency (n=18),and TBS (n=18) rTMS or sham stimulation (control group,n=18),once a day,for 2 weeks.Fugl-Meyer Assessment (FMA) and National Institutes of Health Stroke Scale (NIHSS) were used to evaluate neurological function in all patients before rTMS treatment (on the day before the first treatment) and after treatment (on the day after the last treatment).Results After treatment,the FMA and NIHSS scores in the 4 groups were significantly improved compared with before treatment (all P<0.05).After rTMS treatment,the FMA and NIHSS scores were improved significantly in the high frequency group,low frequency group and TBS group compare with the control group (all P<0.05).There were no significant differences among all the treatment groups.Conclusion sHigh frequency,low frequency and TBS rTMS can improve the recovery of motor function in hemiplegic patients following acute ischemic stroke.There were no significant differences among all the treatment modes.

Objective: To evaluate the clinical effect of pereutaneous vertbroplasty (PVP) combined with implantation of iodine-125 (125 I)radioactive particle in the treatment of vertebral metastasis,and to provide basis for the treatment of vertebral metastasis.Methods:A total of 69 patients with vertebral metastasis were divided into test group (n=32)and control group (n=37);the patients in test group were treated with PVP comined with implantation 125 I radioactive particle and the patients in control group were treated with PVP only.The heights of anterior and posterior vertebral bodies of the patients before and after treatment were detected by X-ray.The numerical rating scale (NRS)scores,pain relief rate and the incidence of surgical complications of the patients were recorded before operation and 1 d,1 week,1 month,3 months,and 6 months after operation.Results:The operation was successfully performed in all the patients without local bleeding;there were no movement dysfunction and nerve compression phenomenon.There was no leakage of bone cement.All the 125 I radioactive particles located well and there was no particle obscission.The heights of vertebral bodies of the patients in two groups after operation were increased compared with before operation (P <0.05).The NRS scores of the patients in two groups s at 1 d,1 week,1 month,3 months,6 months after operation were significantly decreased compared with before operation (P <0.05);compared with control group,the NRS scores of the patients in test group at 1 d,1 week, 1 month,3 months,6 months after operation were decreased (P <0.05).The incidence of pulmonary embolism or radiation myelitis complications was about 4.3% in 69 patients.Compared with control group,the difference in the incidence of complications of the patients in test group was not significant (P < 0.05 ).Conclusion:PVP combined with 125 I radioactive particle implantation is a safe and effective method in the treatment of vertebral metastasis,which can relieve the pain of the patients obviously compared with PVP.

Objective To determine collateral circulation in patients with acute ischemic stroke using capillary index score (CIS)in order to evaluate the prognosis of endovascular treatment. Methods From January 2013 to December 2015,46 consecutive patients with acute ischemic stroke treated with endovascular treatment at the Department of Neurology,Central Hospital of Baotou were enrolled retrospectively. Angiography was performed before endovascular treatment in order to complete CIS score. The patients were divided into a good prognosis group (n = 21)and a poor prognosis group (n = 25)according to the modified Rankin scale (mRS)scores. Univariate analysis was used to compare the baseline data and the clinical data of the two groups,including age,sex,history of diabetes,pretreatment systolic blood pressure,conducting intravenous thrombolysis or not,time from ictus to intravenous thrombolysis,National Institutes of Health Stroke Scale (NIHSS)score,Alberta stroke program early CT score (ASPECTS),vascular filling,time from onset to revascularization,and postoperative vascular recanalization (the modified Thrombolysis in Cerebral Infarction [mTICI]). Multivariate analysis was used to analyze the effect of CIS score on good prognosis. Results There were no significant differences in age,sex,history of diabetes,pretreatment systolic blood pressure,conducting intravenous thrombolysis or not,time from ictus to thrombolysis,and number of mechanical thrombectomy between the good prognosis group and the poor prognosis group (all P > 0. 05). There were significant differences in the NIHSS score (15 ± 3 vs. 19 ± 4),ASPECTS score (8 [7,10]vs. 6 [5,8]),filling well 85. 7% (18 / 21)vs. 44. 0% [11 / 25]),time from ictus to recanalization (363 ± 42 min vs. 398 ± 53 min),and postoperative vascular recanalization (mTICI≥Ⅱb)(100. 0% [21 / 21]vs. 68. 0%[17 / 25];all P < 0. 05). CIS (OR,8. 600,95% CI 2. 670 -33. 800)and mTICI grade (OR,5. 720, 95%CI 12. 170-22. 300)were significantly associated with the prognosis. Conclusion The CIS score can be used to evaluate brain perfusion. fCIS is closely associated with the good clinical prognosis. When screening the suitable patients for endovascular therapy,increasing the CIS score to evaluate the salvageable brain tissue is effective and feasible.