Objective To evaluate the effects of different ratios of medicine dosage for isoflurane and propofol on postoperative cognitive function of rats with mild cognitive impairment (MCI).Methods Healthy male Wistar rats,aged 16-18 months,were used in the study.MCI model was established by occlusion of bilateral common carotid arteries.One hundred and fifty rats with MCI were divided into 5 groups (n=30 each) using a random number table:sham operation group (group S),isoflurane group (group I),propofol group (group P) and different ratios of medicine dosage for isoflurane and propofol groups (IP1,2 groups).The rats inhaled 1.9％ isoflurane for 3 h in group I.Propofol 40 mg · kg-1 · h-1 was infused intravenously for 3 h in group P.The rats inhaled 1.0％ isoflurane,and propofol 20 mg · kg 1 · h-1 was infused intravenously for 3 h in group IP1.The rats inhaled 1.4％ isoflurane,and propofol 10 mg· kg-1 · h-1 was infused intravenously for 3 h in group IP2.After disappearance of eyelash reflex,open reduction and internal fixation was performed after tibial fracture was induced.On 7 days after operation,contextual fear conditioning test and Y maze test were used to assess the cognitive function,and hippocampal tissues were obtained to count the viable neurons (using Nissl's staining) and CCAAT/enhancer-binding protein homologous protein (CHOP) and caspase-12 positive neurons (by immunofluorescence) in hippocampal CA1 region.Results Compared with group S,the rate of time spent in N arm,the rate of time spent freezing,and the number of viable neurons were significantly decreased,and the number of CHOP and caspase-12 positive neurons were significantly increased in IP2,I and P groups (P＜0.05),and no significant change was found in the parameters mentioned above in group IP1 (P＞0.05).Compared with group IP1,the rate of time spent in N arm,the rate of time spent freezing,and the number of viable neurons were significantly decreased,and the number of CHOP and caspase-12 positive neurons was significantly increased in IP2,I and P groups (P＜0.05).Compared with group IP2,the rate of time spent in N arm,the rate of time spent freezing,and the number of viable neurons were significantly decreased,and the nunber of CHOP and caspase-12 positive neurons was significantly increased in I and P groups (P＜0.05).Conclusion Combination of 1.0％ isoflurane and propofol 20 mg · kg-1 · h-1 does not aggravate the postoperative cognitive dysfunction of the rats with MCI.

Objective To investigate the specific anti-tumor immune response induced by dendritic cells (DCs) transfected with total RNA of human pancreatic cancer Capan-2 cells.Methods DCs were isolated and cultured from peripheral blood mononuclear cells (PBMCs) derived from six patients with pancreatic cancer.Total RNA of Capan-2 cells and MUC4 mRNA were transfected into DCs by electroporation.The survival rate of transfected DCs was determined by MTT method and the expression of MUC4 mRNA in DCs was detected by Western blotting.The activity of cytotoxic T lymphocyte cells (CTLs) induced by DCs transfected with total RNA of Capan-2 cells were evaluated by IFN-γ ELISA and the induction of specific CTL response to the killing effect on pancreatic cancer cell in vitro were measured by 51 Cr standard cytotoxicity test.Results The survival rate of DCs transfected with total RNA of Capan-2 cells (DC-Capan-2-total RNA) showed a decrease in a time dependent manner and the survival rate was reduced to 60.81％ after transfection for 96 h.The survival rate of MUC4 mRNA transfection DCs (DC-MUC4 mRNA) was stable at around 80％.The difference of DCs surviral rate between the two groups was statistically significant (P ＜ 0.05).The amount of MUC4 protein expression of DC-Capan-2-total RNA was significantly lower than that of DC-MUC4 mRNA (P ＜0.05).The quantity of CTL IFN-γ release induced by DC-Capan-2-total RNA was (89.34 ± 3.85)U/mL and the quantity of DC-MUC4 mRNA induced CTL IFN-γ release was (21.77 ± 21.77)U/ml There was statistically significant between the two groups (P ＜0.05).In addition,the specific CTLs induced by DC-Capan-2-total RNA could effectively identify and kill the HLA-A2+/MUC4+ Capan-2 and the HLA-A2+/MUC4-PANC 1 cells,and could not effectively identify and kill the HLA-A2 /MUC4-MiaPaCa-2 cells and the HLA-A2-/MUC4 + AsPC-1 cells.Conclusions A more pronounced CTL anti-tumor immune response can be induced by DCs transfected with total RNA of Capan-2 cells compared with a single tumor associated antigen,but it is limited by MHC class Ⅰ antigen presented.

Objective To investigate the effect of neutral amino acid on preventing Parkinson disease.Methods Mice were injected with L-Valine,L-Pheylalanine,D-Valine or L-Lysine before or after paraquat administration,by which prakinsonian mouse model was constructed.The paraquat immunoreactivity was observed within nigral cell bodies.Then neurodegeneration and ?-synuclein aggregation were observed by immunohistochemistry and Western blot.Results Paraquat immunoreactivity was abolished by the administration of L-Valine,L-Pheylalanine before paraquat exposure.Pre-treatment with these two amino acids also protected the paraquat-induced loss of nigrostriatal dopaminergic cells and formation of thioflavine S-positive aggregates.In contrast, paraquat-induced toxicity was unaffected if animals were injected with these two amino acids after paraquat exposure or pre-treated with D-Valine or L-Lysine.Conclusions L-type neutral amino acids such as L Valine and L-Pheylalanine can prevent paraquat-induced neurodegeneration and a synuclein pathology through a competitive inhibition mechanism with stereospecificity in the central nervous system (CNS).Neutral amino acid could protect the dopaminergic neuron in substantia nigra and may prevent Parkinson disease.

AIM To establish the HPLC-ELSD fingerprints of oligosaccharide sites from mycelia of Hericium erinaceum solid cultures and Weilening Tablets.METHODS The analysis of aqueous extract from samples was performed on a 80 ℃ thermostatic Waters XBridge TM Amide column (4.6 mm × 150 mm,3.5 μm),with the mobile phase comprising of acetonitrile-0.2％ ammonium acetate flowing at 1 mL/min in a gradient elution manner.RESULTS There were eight and nine common peaks in two HPLC-ELSD fingerprints with the similarties of 0.994-0.966 and 0.990-0.997,respectively.Three of them were mannitol,lactose and trehalose,which showed good linear relationships within their own ranges (r ≥ 0.999 0),the average recoveries were 95.08％-104.82％ with the RSDs of 1.12％-2.90％.CONCLUSION This simple and accurate method can be used for the rapid quality control of mycelia of Hericium erinaceum solid cultures and Weilening Tablets.

Objective To investigate the appropriate compatibility of appropriate compatibility of sevoflurane and propofol for patients with mild cognitive impairment (MCI) undergoing posterior lumbar interbody fusion in order to protect their cognitive function.Methods Eighty patients, 41 males, 39 females, aged 65-75 years, BMI 17-26 kg/m2, ASA physical status Ⅰ or Ⅱ, scheduled to undergo elective posterior lumbar interbody fusion, were to be scored according to Montreal cognitive assessment (MoCA), mini mental state examination (MMSE), dementia scale (CDR) and daily living ability scale (ADL) to identify patients with MCI before the surgery.They were randomly assigned to 4 groups (n=20 each) using a random number table: TCI propofol 2.0-2.5 μg/ml group (group P), TCI propofol 1.2 μg/ml+sevoflurane 0.6 MAC group (group PS1), TCI propofol 0.6 μg/ml+sevoflurane 0.9 MAC group (group PS2), 1.0-1.5 MAC sevoflurane group (group S).MoCA and MMSE were used to evaluate the cognitive function of patients 1 d before the operation (T0), after patients become wide-awake (T1), 3 d and 7 d after operation (T2 and T3).Apolipoprotein J (ApoJ) concentration related to cognitive function in blood samples, which were drawn at T0-T3 would be measured with ELISA method.Results Compared with T0, the scores of MMSE and MoCA in four groups decreased significantly (P＜0.05) at T1, the scores of MMSE and MoCA in group S decreased significantly (P＜0.05) at T2;compared with T1, the score of MMSE in the four groups increased significantly at T2, T3 (P＜0.05).The scores of MMSE at T1, T3 in group S decreased significantly compared with groups P, PS1 and PS2 (P＜0.05).The scores of MoCA at T2, T3 in group S decreased significantly compared with groups P, PS1 and PS2 (P＜0.05).Compared with T0, the concentration of plasma ApoJ in the four groups increased significantly at T1 (P＜0.05).Compared with T1, the concentration of plasma ApoJ in the four groups decreased significantly at T2 and T3 (P＜0.05).Compared with group PS1, the concentration of plasma ApoJ at T1, T3 increased significantly in groups S and group PS2 (P＜0.05).Conclusion TCI propofol 1.2 μg/ml combined with 0.6 MAC sevoflurane group is the appropriate compatibility of sevoflurane and propofol for patients with MCI undergoing posterior lumbar interbody fusion,because it has less negative influence on cognitive function and lower concentration of plasma ApoJ.

Objective To explore the relationship between the genesis of dyskinesia and the degree of substantia nigra lesion in Parkinson disease(PD).Methods The hemi-parkinsonian rat model was established by injecting 6-OHDA stereotaxically to right medial forebrain bundle(MFB). Then the hemi-parkinsonian rat was injected intraperitoneally with levodopa methylester(25 mg?kg~(-1)?d~(-1),twice a day)for 21 days,the abnormal involuntary movements were estimated.After being sacrificed,the midbrain was removed,and the injured degree of dopaminergic neurons at substantia nigra was observed by tyrosine hydroxylase immunohistochemistry staining.The relationship between the abnormal involuntary movement scores and dopaminergic neurons loss at substantia nigra was evaluated by sigmoid equation analysis by using Excel software.Results The apomorphine-induced rotation rate above 7 r/min was found in 10 of 25 rats,those rats were regarded as successful hemi-parkinsonian model rats.After the treatment with levodopa methylester,8 of 10 rats displayed abnormal involuntary movements,including stereotype and contralateral rotation,the types of movements varied.Abnormal involuntary movements were appeared in the rats with dopaminergic neurons loss above 90%.The positive relationship was observed between the degree of lesion in substantia nigra and the severity of abnormal involuntary movements.Conclusions The severe loss of dopaminergic neurons at substantia nigra probably plays a role in the development of levodopa-induced dyskinesia in patient with Parkinson disease.

Objective To investigate cellular and behavioural effects of 5-HT1A receptor agonist 8- OH-DPAT in a rat model of levodopa-induced motor complications.Methods The hemi-parkinsonian rat model was produced by stereotaxically injecting 6-OHDA to right medial forebrain bundle(MFB).Two sets of experiments were performed.First,rats were intrapefitoneally treated with levodopa 50 mg/kg plus benserazide 12.5 mg/kg twice a day for 22 days.On day 23,rats intraperitoneally received either 8-OH- DPAT(1 mg/kg)or 8-OH-DPAT plus WAY-100635(0.1 mg/kg)or dissolvent with each levodopa dose as controls.In the second set,rats were intraperitoneally treated either with levodopa(50 mg/kg)plus 8-OH- DPAT(1 mg/kg)or levodopa 50 mg/kg plus dissolvent,administered twice daily for 22 consecutive days. Rotational duration and frequency of off period were estimated.After sacrificed,subcellualr distribution of GluR1 and GluR1Ser845 phosphorylation was observed by Western blot.Results 8-OH-DPAT,reversing the shortened rotational duration induced by levodopa,prolonged the rotational duration by 27.8%?6.1% and reduced the frequency of failures to levodopa by 7.2%?1.7%.Co-administration of WAY-100635,a 5-HT1A receptor antagonist,with 8-OH-DPAT eliminated the effect of 8-OH-DPAT on motor complications, indicating that the observed 8-OH-DPAT responses were probably mediated via the 5-HT1A autoreceptor. Moreover,8-OH-DPAT could regulate subcellular distribution of GluR1 and reduce hyperphosphorylation of GluR1 Ser845 by 22.1%?3.5%,which was closely associated with levodopa-induced motor complications. Conclusions These results suggest that pharmaceuticals stimulating 5-HT1A receptors could be useful in the treatment and prevention of the motor complications in parkinsonian patients.

Dengue virus (DENV) envelope [E] protein is the major surface protein of the virions that indued neutralizing antibodies. The domain III of envelope protein (EDIII) is an immunogenic region that holds potential for the development of vaccines; however, the epitopes of DENV EDIII, especially neutralizing B-cell linear epitopes, have not been comprehensively mapped. We mapped neutralizing B-cell linear epitopes on DENV-1 EDIII using 27 monoclonal antibodies against DENV-1 EDIII proteins from mice immunized with the DENV-1 EDIII. Epitope recognition analysis was performed using two set of sequential overlapping peptides (16m and 12m) that spanned the entire EDIII protein from DENV-1, respectively. This strategy identified a DENV-1 type- specific and a group-specific neutralizing epitope, which were highly conserved among isolates of DENV-1 and the four DENV serotypes and located at two regions from DENV-1 E, namely amino acid residues 309-320 and 381-392(aa 309-320 and 381-392), respectively. aa310 -319(310KEVAETQHGT319)was similar among the four DENV serotypes and contact residues on aa 309 -320 from E protein were defined and found that substitution of residues E309 , V312, A313 and V320 in DENV-2, -3, -4 isolates were antigenically silent. We also identified a DENV-1 type-specific strain-restricted neutralizing epitope, which was located at the region from DENV-1 E, namely amino acid residues 329-348 . These novel type- and group-specific B-cell epitopes of DENV EDIII may aid help us elucidate the dengue pathogenesis and accelerate vaccine design.
Amino Acid Sequence ; Animals ; Antibodies, Neutralizing ; immunology ; Dengue ; virology ; Dengue Virus ; chemistry ; genetics ; immunology ; Epitope Mapping ; Epitopes, B-Lymphocyte ; chemistry ; genetics ; immunology ; Humans ; Mice ; Molecular Sequence Data ; Viral Envelope Proteins ; chemistry ; genetics ; immunology

BACKGROUND: The content of type Ⅰ and Ⅲ collagen and the ratio of both are crucial factors to promote heart geometric morphology change,and ventricular systolic and diastolic myocardial performance.Matrix metalloproteinase 2 (MMP-2) and tissue inhibitor of metalloproteinase 1 (TIMP1) are primary regulatory substances of collagen metabolism.After myocardial infarction,chronic heart failure rats were subjected to bone marrow meseuchymal stem cell transplantation.What changes in MMP-2 and TIMPI would occur?OBJECTIVE: To observe the content of type Ⅰ and Ⅲcollagen and the ratio between both,as well as expression levels of MMP-2 and TIMP1 in the left ventricular tissue of rats with myocardial infarction-caused chronic heart failure following bone marrow mesenchymal stem cell (MSC) transplantation.DESIGN: A randomized controlled experiment.SETTING: Department of Senile Angiocardiopathy,General Hospital of Chinese PLA & Department of Biochemistry,Peking University Health Science Center.MATERIALS: This study was performed at the laboratory of Department of Biochemistry,Peking University Health Science Center between July 2004 and December 2005.Male Sprage Dawley rats of clean grade,aged 4 weeks old,were provided by the Laboratory Animal Center,Beijing Medical University and used for preparation of MSCs.Fourteen female rats,weighing 200-250g,were developed into models of heart failure-caused by myocardial infarction.METHODS: MSCs were isolated and,purified by gradient centrifugation and adherent cells were allowed to proliferate.Female rats underwent coronary artery ligation to induce chronic ischemic heart failure.Four weeks later,the rats were randomly divided into 2 groups: (1) experimental group (n=7),rats received transplantation of MSCs harvested from male rats [5×106 in 50 μL phosphate buffered saline(PBS)]by injection into the ischemic myocardium; (2) control group (n=7),rats received the same volume of PBS.MAIN OUTCOME MEASURES: Twenty-one days after therapy,(1) left ventricular fusion was tested by hematoxylin-eosinstaining and Masson staining; (2) Expression of MMP-2 and 1TMPI as well as contents of type I and llI collagen was analyzed by immunohistochemistry; (3) MMP-2 and TIMP1 expression levels were examined by Western blot.RESULTS: Fourteen rats were included in the final analysis.Type Ⅰ collagen expression in the scar area was much higher in theexperimental group than in the control group,while type Ⅲ collagen expression was much lower in the experimental group.MMP-2 expression was reduced and TIMPI expression was increased in the experimental group compared with the control group.Together,ventricular wall was thickened,ventricular chamber was reduced,and heart function was strengthened in the experimental group compared with the control group.CONCLUSION: MSC transplantation alleviated left ventricular remodeling in chronic ischemie heart failure,which results from dynamic regulation of MMP-2/TIMP1.

BACKGROUND: There are still few effective methods to repair injured myocardium after myocardial failure and pathologically rebuild reverral myocardium. As a new therapy, normal myocytes and therapeutic gene to interfere injured myocardium have advantageous effects in improving heart function.OBJECTIVE: To observe the efficiency and stability of adenovirus-medicated gene transferred into different passages of bone marrow mesenchymal stem cell (MSC) and investigate the effect of MSC-based sarcoplasmic reticulum Ca2+ ATPase gene (SERCA2a) gene therapy for rats with chronic heart failure. To compare the effects of gene therapy, cell transplantation and MSC-based SERCA2a gene therapy for chronic heart failure. DESIGN: Randomized controlled study.SETTING: Department of Senile Angiocardiopathy, General Hospital of Chinese PLA; Department of Biochemistry, Beijing Medical University. MATERIALS: Male Sprague-Dawley (SD) rats with 4 weeks old, clean grade and weighing 45-50 g provided by the Animal Experimental Center, Peking Medical University were used as donators of bone marrow. Other female SD rats of 12 weeks old, clean grade and weighing 200-250 g were used as receptors of cell transplantation and gene therapy. Sry gene of Y chromosome in male rats was used to evaluate whether transplanted cells of donators lived in myocardium of receptor rats. Ad-SERCa2a and Ad-EGFP were constructed by Doctor Lu Xiao-chun; MSC in the 3rd and 8th generations was isolating cultured on its own. METHODS: The experiment was carried out in the Zhou CY Laboratory (BSL-2), Department of Biochemistry, Beijing Medical University from July 2004 to December 2005. Thirty female SD rats received ligation at the left coronary artery to make models with chronic cardiac failure following acute myocardial infarction. And then, 29 rats were randomly divided into four groups, including gene therapy group (n=7), MSC group (n=7), gene-modified MSC group (n=8) and control group (n=7). Rats in the four groups were given the interventions of SERCA2a gene, MSC transplantation, MSC+Ad/SERCa2a and empty adenoviral vector, respectively. MSCs were separated and cultured, and then Ad-SERCA2a-GFP was used to transfer MSC in the 3rd and 8th generations.MAIN OUTCOME MEASURES: Ad-SERCA2a-GFP transfection rate of MSC was measured by using flow cytometer. Before and at 14 and 21 days after treatment, cardiac function was evaluated by ultrasonic echocardiogram. Expression of cytokine Ⅷ was tested by immunohistochemical staining. SERCA2a gene and protein expression were evaluated by RT-PCR and Western blot respectively, as well as SERCA2a enzyme activity. RESULTS: ① Transfection rate: The infection efficiency of adenovirus-medicated gene into different passages of MSC was over 80%, and there was no difference between passage three (P3) MSC and P8 MSC (P > 0.05). ② Heart function: Left ventricle wall was thickened obviously in group MSC and group MSC+Ad/SERCa2a on the 21st day after treatment, while volume was shortened and gradually rounded. Compared to control group, ejection fraction (EF) and shortening fraction (FS) of group Ad-SERCa2a, group MSC and group MSC+Ad/SERCa2a were elevated significantly on the 14th day after therapy (P < 0.01). While the elevation values of EF and FS began to reduce in group Ad-SERCa2a on 14th day after therapy, it continued to increase in both group MSC and group MSC+Ad/SERCa2a (P < 0.01). Improvement rate of EF at 21 days after therapy (EF D21) increased in group MSC and group MSC+Ad/SERCa2a respectively, but decreased in group Ad-SERCa2a. Compared to group Ad-SERCa2a, peak systolic flow velocity of anterior wall and interventricular septum in group MSC+Ad/SERCa2a increased significantly on the 21st day after therapy, and peak diastolic flow velocity of anterior wall and interventricular septum elevated in group MSC+Ad/SERCa2a, too (P < 0.01). ③ SERCA2a gene, protein expression and enzyme activity in group MSC+Ad/SERCa2a were significantly stronger in group MSC and control group. Parts of MSC transplanted into scar zone expressed Ⅷ.CONCLUSION: ① MSC is an effective platform for the targeted delivery of therapeutic gene. It suggests that different passages of MSC from P3 MSC to P8 MSC are regarded as high-effectively gene vehicles. MSC-based SERCA2a gene therapy showed much strong and lasting beneficial effect on exhausted myocardium. ② Effect of MSC transplantation on improving heart function may be related to promoting vascular neogenesis.