Effects of L-3-n-butylphthalide on caspase-3 and nuclear factor kappa-B expression in primary basal forebrain and hippocampal cultures after beta-amyloid peptide 1-42 treatment

BACKGROUND： L-3-n-butylphthalide （L-NBP） can inhibit phosphorylation of tau protein and reduce the neurotoxicity of beta-amyloid peptide 1-42 （Aβ1-42）. OBJECTIVE： To observe the neuroprotective effects of L-NBP on caspase-3 and nuclear factor kappa-B （NF- K B） expression in a rat model of Alzheimer＇s disease. DESIGN, TIME AND SETTING： A cell experiment was performed at the Central Laboratory of Provincial Hospital affiliated to Shandong University between January 2008 and August 2008. MATERIALS： L-NBP （purity 〉 98%） was provided by Shijiazhuang Pharma Group NBP Pharmaceutical Company Limited. Aβ1-42, 3-[4,5-dimethylthiazolo-2]-2,5 iphenyltetrazolium bromide （MTT）, and rabbit anti-Caspase-3 polyclonal antibody were provided by Cell Signaling, USA; goat anti-choactase and rabbit anti-NF- kB antibodies were provided by Santa Cruz, USA. METHODS： Primary cultures were generated from rat basal forebrain and hippocampal neurons at 17 or 19 days of gestation. The cells were assigned into five groups： the control group, the Aβ1-42 group （2 μmol/L）, the Aβ1-42 ＋ 0.1 μmol/L L-NBP group, the Aβ1-42 ＋ 1 μ mol/L L-NBP group, and the Aβ1-42 ＋ 10μmol/L L-NBP group. The neurons were treated with Aβ1-42 （2 μmol/L） alone or in combination with L-NBP （0.1, 1, 10 μmol/L） for 48 hours. Cells in the control group were incubated in PBS. MAIN OUTCOME MEASURES： Morphologic changes were evaluated using inverted microscopy, viability using the M-I-I- method, and the changes in caspase-3 and NF- k B expression using Western blot. RESULTS： Induction with Aβ1-42 for 48 hours caused cell death and soma atrophy, and increased caspase-3 and NF- K B expression （P 〈 0.05）. L-NBP blocked these changes in cell morphology, decreased caspase-3 and NF- k B expression （P 〈 0.05）, and improved cell viability, especially at the high dose （P 〈 0.05）. CONCLUSION： AI3^-42 is toxic to basal forebrain and hippocampal primary neurons; L-NBP protects against this toxicity and inhibits the induction of caspase-3 and NF- K B expression.

Improved extracellular endo-1,4-β-mannosidase activity of recombinant Pichia pastoris by optimizing signal peptide

In order to improve the extracellular endo-1,4-β-mannosidase(MAN) activity of recombinant Pichia pastoris, optimization of signal peptides was investigated. At first, five potential signal peptides(W1, MF4 I, INU1 A, αpre, HFBI) were chosen to be analyzed by Signal P 4.0, among which W1 was designed. Then, the widely used signal peptide α-factor in expression vector p GAPZαA was replaced by those five signal peptides to reconstruct five new expression vectors. MAN activity was assayed after expression vectors were transformed into Pichia pastoris. The data show that the relative efficiencies of W1, MF4 I, INU1 A, αpre, and HFBI signal peptides are 23.5%, 203.5%, 0, 79.7%, and 120.3% compared with α-factor, respectively. The further gene copy number determination by the quantitative real-time PCR reveals that the MAN activities mediated by α-factor from 1 to 6 gene copy number levels are 12.95, 43.33, 126.63, 173.53, 103.23 and 88.63 U/m L, while those mediated by MF4 I are 79.22, 133.89, 260.14, 347.5, 206.15 and 181.89 U/m L, respectively. The maximum MAN activity reached 347.5 U/m L with 4 gene copies mediated by MF4 I. These results indicate that replacing the signal peptide α-factor with MF4 I and increasing MAN gene copies to a proper number can greatly improve the secretory expression of MAN.

Neuroprotective Effect of Peptides Extracted from Walnut(Juglans Sigilata Dode) Proteins on Aβ25-35-induced Memory Impairment in Mice

Alzheimer＇s disease（AD） is one of the major neurodegenerative disorders of the elderly, which is characterized by the accumulation and deposition of amyloid-beta（Aβ） peptide in human brains. Oxidative stress and neuroinflammation induced by Aβ in brain are increasingly considered to be responsible for the pathogenesis of AD. The present study aimed to determine the protective effects of walnut peptides against the neurotoxicity induced by Aβ25-35 in vivo. Briefly, the AD model was induced by injecting Aβ25-35 into bilateral hippocampi of mice. The animals were treated with distilled water or walnut peptides（200, 400 and 800 mg/kg, p.o.） for five consecutive weeks. Spatial learning and memory abilities of mice were investigated by Morris water maze test and step-down avoidance test. To further explore the underlying mechanisms of the neuroprotectivity of walnut peptides, the activities of superoxide dismutase（SOD）, glutathione（GSH）, acetylcholine esterase（ACh E）, and the content of malondialdehyde（MDA） as well as the level of nitric oxide（NO） in the hippocampus of mice were measured by spectrophotometric method. In addition, the levels of 8-hydroxy-2＇-deoxyguanosine（8-OHd G）, tumor necrosis factor-α（TNF-α）, interleukin 1β（IL-1β） and IL-6 in the samples were determined using ELISA. The hippocampal expressions of inducible nitric oxide synthase（i NOS） and nuclear factor κB（NF-κB） were evaluated by Western blot analysis. The results showed that walnut peptides supplementation effectively ameliorated the cognitive deficits and memory impairment of mice. Meanwhile, our study also revealed effective restoration of levels of antioxidant enzymes as well as inflammatory mediators with supplementation of walnut peptides（400 or 800 mg/kg）. All the above findings suggested that walnut peptides may have a protective effect on AD by reducing inflammatory responses and modulating antioxidant system.

Protective effects of proanthocyanidins on beta-amyloid peptide (25-35)-induced PC12 cell apoptosis by blocking S-phase and increasing p53 gene expression

BACKGROUND： Current studies related to the effects of proanthocyanidins on Alzheimer＇s disease have focused primarily on the signal transduction pathway of cellular apoptosis. However, the influence of p53 gene expression on cell cycle regulation, with regard to the protective mechanisms of proanthocyanidins, has not been reported. OBJECTIVE： To observe the effect of proanthocyanidins on cell cycle distribution, cellular apoptosis and p53 gene expression in β-amyloid peptide （25-35） （Aβ25-35）-induced PC12 cells cultured in serum-free media, and to investigate the molecular neuroprotective mechanisms of proanthocyanidins with regard to cell cycle regulation. DESIGN, TIME AND SETTING： A parallel, controlled, at the Institute of Biochemistry and Molecular Biology cellular, and molecular study was performed Guangdong Medical College from July 2006 to July 2008. MATERIALS： Proanthocyanidins were provided by Nanjing Xuezi Medical and Chemical Research Center, China; Aβ25-35 was provided by Sigma, USA; PC12 cells were provided by the Institute of Basic Medical Science, Academy of Military Medical Sciences; and rabbit anti-p53 polyclonal antibody was provided by Santa Cruz Biotechnology, USA. METHODS： PC12 cells were cultured in serum-free media for 24 hours. Cells from the model group were treated with 25 μmol/L Aβ25-35 for 24 hours. Cells in the drug protection group were pre-treated with 30 mg/L proanthocyanidins for 1 hour and then treated with 25 μmol/LAβ2^-35 for 24 hours. The control group was not treated. MAIN OUTCOME MEASURES： Flow cytometry was used to detect cell cycle distribution and rate of apoptosis; reverse-transcriptase polymerase chain reaction was used to detect p53 mRNA expression; and Western blot was used to detect p53 protein expression. RESULTS： After treating with 25 μmol/LAβ25-35 for 24 hours, the rate of apoptosis and the percentage of cells in S phase were significantly increased （P 〈 0.01 ）, and p53 mRNA and protein expressions were decreased. Pretreatment with proanthocyanidins for 1 hour blocked the increase in apoptosis and the percentage of cells in S phase in Aβ25-35-induced PC12 cells （P 〈 0.01 ） and increased p53 mRNA and protein expressions. CONCLUSION： Proanthocyanidins blocked apoptosis and S-phase arrest in Aβ25-35-induced PC12 cells cultured in serum-free media. The protective mechanism could be related to increased p53 mRNA and protein expressions.

STUDY ON THE THERAPEUTIC EFFECTS OF GINSENOSIDE Rg-1 AND GASTRODINE ON AD MODEL RATS INDUCED BY β-AMYLOID PEPTIDE (25-35)

Objective To study the therapeutic effects of Ginsenoside Rg-1 and Gastrodine on rats model of Alzheimer's disease(AD). Methods Aggregated β-Amyloid peptide (25-35) was injected into the lateral ventricle of rats to establish AD models. Ginsenoside Rg-1, Gastrodine and Ginsenoside Rg-1+Gastrodine were intraperitoneally injected into rats of each test group(Ginsenoside Rg-1∶10mg/kg·day; Gastrodine 100mg/kg·day) for 4 weeks, the rats of control group received equal volume of saline. Passive avoidance task and Morris maze test were done to assess the ability of learning and memory. The content of superoxide dismutase (SOD), malondiadehyde (MDA), total-antioxidative capability (T-AOC), Choline acetyltransferase (ChAT) and acetylcholinesterase (AchE) in brain tissue were measured. Results Ginsenoside Rg-1 and Gastrodine significantly improved learning and memory deficits in the rats with AD induced by β-Amyloid peptide (25-35) (P<0.05). Ginsenoside Rg-1+Gastrodine group were better than Ginsenoside Rg-1 group and Gastrodine group (P<0.05). Ginsenoside Rg-1 reduced the increase of SOD, MDA, but inhibited the decrease of T-AOC, AchE and ChAT; Gastrodine reduced the increase of SOD, MDA, while inhibited the decrease of T-AOC. Gastrodine could also prevent the activity of ChAT and AchE decline in AD rats. Conclusion Both Ginsenoside Rg-1 and Gastrodine have therapeutic effects on rats with AD; Ginsenoside Rg-1 and Gastrodine injection at the same time were better than only using one of them. Their mechanisms might different. Ginsenoside Rg-1 can not only inhibit peroxidation but also increase the activity of AchE and ChAT in brain tissue, while Gastrodine can inhibit peroxidation only, but it can't prevent the decline of ChAT and AchE activity in AD rats.

Inhibitory effects of scorpion venom heat-resistant protein on neurotoxicity of exogenous amyloid beta peptide 1-40

BACKGROUND： Studies have shown that scorpion venom heat-resistant protein （SVHRP） exhibits protective effects on primary cultured hippocampal neurons. OBJECTIVE： To determine the effects of SVHRP on astrocyte activity and synaptic density in the hippocampus induced by amyloid β peptide 1-40 （Aβ1-40） neurotoxicity. DESIGN, TIME AND SETTING： The randomized, controlled, animal experiment was performed at the Central Laboratory, the Laboratory of Human Anatomy, and the Laboratory of Physiology, in Dalian Medical University between March 2006 and June 2008. MATERIALS： Aβ1-40 was provided by Biosource, USA; SVHRP was a patented biological product of Dalian Medical University （No. ZL01 1 06166.9）. METHODS： A total of 27 healthy, 2-month-old, male SD rats were randomly assigned to 3 groups： control, Aβ, and SVHRP, with 9 rats in each group. Alzheimer＇s disease was simulated with 10 μg Aβ1-40 bilaterally injected into the hippocampus of the Aβ and SVHRP groups. The control group was injected with 2 μL 0.05% trifluoroacetic acid. One day following model establishment, the SVHRP group received an intraperitoneal injection of 2 μg/100 g SVHRP, while the control group and Aβ group received 0.5 mL/100 g tri-distilled water, once per day, for 10 consecutive days. MAIN OUTCOME MEASURES： At 16 days following model establishment, synaptophysin （p38） expression in CA1-CA4 regions of the rat hippocampus was determined by immunohistochemistry. Glial fibrillary acidic protein （GFAP） expression surrounding the hippocampal Aβ1-40 injected area was also detected. At 11 days following model establishment, escape latency, swimming time, and distance to target quadrant were measured using the Morris water maze. RESULTS： Compared with the control group, the Aβ group exhibited notably reduced p38 expression （P 〈 0.05） and notably increased GFAP expression in the rat hippocampus （P 〈 0.05）. Water maze results demonstrated that escape latency was prolonged （P 〈 0.05）, and swimming time and distance to the target quadrant were shortened in the Aβ group. Compared with the Aβ group, the SVHRP group exhibited notably increased p38 expression （P 〈 0.05） and notably decreased GFAP expression in the rat hippocampus （P 〈 0.05）. Water maze results demonstrated that escape latency was significantly reduced （P 〈 0.05）, and swimming time and distance to the target quadrant were significantly prolonged. CONCLUSION： SVHRP inhibited exogenous Aβ1-40-induced astrocyte activation and synaptic density decline in the rat hippocampus. Place navigation and spatial searching results showed that SVHRP blocked Aβ1-40-induced impaired learning and memory.

Regulation of adenosine triphosphate-sensitive potassium channels suppresses the toxic effects of amyloid-beta peptide(25-35)

In this study, we treated PC12 cells with 0-20 μM amyloid-β peptide （25-35） for 24 hours to induce cytotoxicity, and found that 5-20 μM amyloid-β peptide （25-35） decreased PC12 cell viability, but adenosine triphosphate-sensitive potassium channel activator diazoxide suppressed the decrease in PC12 cell viability induced by amyloid-β peptide （25-35）. Diazoxide protected PC12 cells against amyloid-β peptide （25-35）-induced increases in mitochondrial membrane potential and intracellular reactive oxygen species levels. These protective effects were reversed by the selective mitochondrial adenosine triphosphate-sensitive potassium channel blocker 5-hydroxydecanoate. An inducible nitric oxide synthase inhibitor, Nw-nitro-L-arginine, also protected PC12 cells from amyloid-β peptide （25-35）-induced increases in both mitochondrial membrane potential and intracellular reactive oxygen species levels. However, the H202-degrading enzyme catalase could not reverse the amyloid-β peptide （25-35）-induced increase in intracellular reactive oxygen species. A 24-hour exposure to amyloid-13 peptide （25-35） did not result in apoptosis or necrosis, suggesting that the increases in both mitochondrial membrane potential and reactive oxygen species levels preceded cell death. The data suggest that amyloid-β peptide （25-35） cytotoxicity is associated with adenosine triphosphate-sensitive potassium channels and nitric oxide. Regulation of adenosine triphosphate-sensitive potassium channels suppresses PC12 cell cytotoxicity induced by amyloid-β peptide （25-35）.

Prognostic Potential of B - type Natriuetic Peptide and Quality of Life in Patients with Chronic Heart Failure

Objectives To study wheth- er change of BNP levels reflect the change of cardiac function and to investigate the short - term prognostic potential of BNP and QOL in patients with CHF. Methods 96 consecutive patients admittedwith CHF between September 2002 and January 2003 were stud- ied , upon entry the study, BNP levels were measured, Patients administered the disease - specificquality of life questionnaire Minnesota living with heart failure questionnaire (LiHFe) within 1 day. BNP levels and administering LiHFe were repeated three months later. Results BNP levels were increased proportional to the severity of cardiac function. Physical domain and total score of LiHFe were significantly correlated to the severity of CHF ( p < 0. 05 ). BNP levels were de- creased in improving group(p =0. 032) . In deteriora- ting group BNP levels increased (P = 0. 043 ) . Kaplan - meier analysis according to BNP level cutoff point 150 ng/1, the life curve of higher BNP level group was significantly lower than the lower group ( p = 0. 001 ) . In univariate logistic regression, NYHA class, BNP, LVEF, LVEDD, heart size, total score of LiHFe, phys- ical domain of LiHFe and the emotional domain of LiH- Fe were all significant prognostic factors of CHF ( p < 0. 05 for all). While in multiple regression, only BNP level( p = 0. 036) and the emotional domain of LiHFe ( p = 0. 025 ) were independent prognostic factors. Conclusions Change of BNP reflects the treatment efficacy of CHF. BNP and QOL are the two major short - time prognostic factors of the chronic heart failure patients.

Antibacterial and antibiofi lm activity of peptide PvGBP2 against pathogenic bacteria that contaminate Auricularia auricular culture bags

Bacteria contamination in Auricularia auricular culture bags reduces yield and increases the risk of food safety.In this study,5 species of bacteria,mainly gram-positive bacteria including three species of Bacillus spp.,Arthrobacter arilaitensis and Staphylococcus warneri,were isolated and identifi ed from bacteria-contaminated A.auricular culture bags.An in silico predicted antimicrobial peptide from theβ-1,3-glucan-binding protein sequence of Penaeus vannamei,designated Pv GBP2(FLKLGRKSRYGMLKL),was screened and its antibacterial effect and mechanism of action on the isolated Bacillus spp.explored.The minimal inhibitory concentrations(MIC)of Pv GBP2 on Bacillus spp.were 15.6–31.25μg/m L.Peptide Pv GBP2 could inhibit Bacillus subtilis in A.auricular culture bags to maintain growth and yield of A.auricular.Transmission electron microscopy(TEM)revealed that Pv GBP2 kills bacteria by perforating the cell wall,destroying membrane integrity and resulting in the leakage of intracellular solutes.In addition,Pv GBP2 inhibits biofi lm formation by B.subtilis by 90.6%at 1×MIC.Thus,peptide Pv GBP2 could be potentially applied as an antibacterial agent to control bacterial infection of A.auricular cultivation and the spread of foodborne pathogens.

β-淀粉样蛋白在Alzheimer病中的神经毒性

文综述了β淀粉样蛋白多肽(βamyloid peptide,Aβ)在老年性痴呆症(Alzheimer's disease,AD)中的毒性作用。Aβ是老年斑形成的始动因子,也是老年斑核中的核心成分;Aβ沉积所形成的神经炎性斑,即老年斑是AD脑内特征性病理变化之一;Aβ本身也可形成自由基损伤神经元;Aβ引起的细胞凋亡在AD病神经元丢失中起着重要作用;Aβ可作用于胆碱能神经元末梢,通过抑制胆碱的摄取而减少乙酰胆碱合成,减少海马和皮质脑片释放乙酰胆碱。总之,Aβ是通过多个环节引发神经毒性作用的。

Peptide RL-QN15 promotes wound healing of diabetic foot ulcers through p38 mitogen-activated protein kinase and smad3/miR-4482-3p/vascular endothelial growth factor B axis

Background:Wound management of diabetic foot ulcers(DFUs)is a complex and challenging task,and existing strategies fail to meet clinical needs.Therefore,it is important to develop novel drug candidates and discover new therapeutic targets.However,reports on peptides as molecular probes for resolving issues related to DFUs remain rare.This study utilized peptide RL-QN15 as an exogenous molecular probe to investigate the underlying mechanism of endogenous non-coding RNA in DFU wound healing.The aim was to generate novel insights for the clinical management of DFUs and identify potential drug targets.Methods:We investigated the wound-healing efficiency of peptide RL-QN15 under diabetic con-ditions using in vitro and in vivo experimental models.RNA sequencing,in vitro transfection,quantitative real-time polymerase chain reaction,western blotting,dual luciferase reporter gene detection,in vitro cell scratches,and cell proliferation and migration assays were performed to explore the potential mechanism underlying the promoting effects of RL-QN15 on DFU repair.Results:Peptide RL-QN15 enhanced the migration and proliferation of human immortalized keratinocytes(HaCaT cells)in a high-glucose environment and accelerated wound healing in a DFU rat model.Based on results from RNA sequencing,we defined a new microRNA(miR-4482-3p)related to the promotion of wound healing.The bioactivity of miR-4482-3p was verified by inhibiting and overexpressing miR-4482-3p.Inhibition of miR-4482-3p enhanced the migration and proliferation ability of HaCaT cells as well as the expression of vascular endothelial growth factor B(VEGFB).RLQN15 also promoted the migration and proliferation ability of HaCaT cells,and VEGFB expression was mediated via inhibition of miR-4482-3p expression by the p38 mitogen-activated protein kinase(p38MAPK)and smad3 signaling pathways.Conclusions:RL-QN15 is an effective molecule for the treatment of DFUs,with the underlying mechanism related to the inhibition of miR-4482-3p expression via the p38MAPK and smad3 signaling pathways,ultimately promoting re-epithelialization,angiogenesis and wound healing.This study provides a theoretical basis for the clinical application of RL-QN15 as a molecular probe in promoting DFU wound healing.

血浆NT-proBNP水平与不稳定型心绞痛近期预后的关系

目的:探讨血浆N末端-脑钠肽前体(NT-proBNP)水平与不稳定型心绞痛(UAP)患者近期预后的关系。方法:因胸痛住院并经冠状动脉造影检查确诊为UAP的患者52例,分别检测症状发作后18h和症状缓解96h血浆NT-proBNP水平,并计算差值比。对UAP患者随访30天,记录患者主要心血管事件(MACE)的发生情况,并应用受试者工作曲线(ROC)分析差值比对UAP患者发生近期心血管事件的预测价值。结果:发生心血管事件患者的NT-proBNP差值比高于未发生者,差异具有统计学意义(P<0.05)。ROC结果显示,NT-proBNP差值比为0.99时,其预测UAP患者近期发生心血管事件的灵敏度和特异度分别为64.3%和81.8%。结论:NTproBNP差值比可用来预测UAP患者近期心血管事件的发生情况,可作为筛选高危病例的一个指标。

Protective Effect of Ecdysterone on PC12Cells Cytotoxicity Induced by Beta-amyloid_(25-35)

Objective： To examine the protective effect of ecdysterone （ECR） against beta-amyloid peptide fragment25-35 （Aβ25-35）-induced PC12 cells cytotoxicity, and to further explore its mechanism. Methods： Experimental PC12 cells were divided into the Aβ group （treated by Aβ25-35 100μmol/L）, the blank group （untreated）, the positive control group （treated by Vit E 100 μmol/L after induction） and the ECR treated groups （treated by ECR with different concentrations of 1, 50 and 100 μmol/L）. The damaged and survival condition of PC12 cells in various groups was monitored by lactate dehydrogenase （LDH） release and MTT assay. The content of malondialdehyde （MDA） was measured by fluorometric assay to indicate the lipid peroxidation. And the antioxidant enzymes activities in PC12 cells, including superoxide dismutases（SOD）, catalase （CAT） and glutathione peroxidase（GSH-Px）, were detected respectively. Results： After PC12 cells were treated with Aβ25-35 （100 μmol/L） for 24 hrs, they revealed a great decrease in MTT absorbance and activity of antioxidant enzymes, including SOD, CAT and GSH-Px as well as a significant increase of LDH activity and MDA content in PC12 cells （P〈0.01）. When the cells was pretreated with 1-100 μmol/L ECR for 24 hrs before Aβ25-35 treatment, the above-mentioned cytotoxic effect of Aβ25-35 could be significantly attenuated dose-dependently, for ECR 50 μmol/L, P〈0.05 and for ECR 100 μmol/L, P〈0.01. Moreover, ECR also showed significant inhibition on the Aβ25-35 induced decrease of SOD and GSH-Px activity, but not on that of CAT. Conclusion： ECR could protect PC12 cells from cytotoxicity of Aβ25-35, and the protective mechanism might be related to the increase of SOD and GSH-Px activities and the decrease of MDA resulting from the ECR-pretreatment.

S4B-4 A Novel Allosteric Phosphodiesterase 4D Inhibitor BPN14770 Reverses Cognitive Impairment in Humanized PDE4D Mice

Classical inhibitors of PDE4 lack subtype selectivity due to exact amino acid sequence conservation of the catalytic site,and consequently,development of these drugs has stalled due to dose-limiting side effects of nausea and emesis.While use of subtype-selective inhibitors(i.e.,for PDE4A,B,or D)could overcome this issue,conservation of the catalytic region,to which classical inhibitors bind,limits this approach.The present study examined the effects of BPN14770,an allosteric inhibitor of PDE4D,which binds to a primate-specific,N-terminal region,conferring greater than 260-fold selectivity for PDE4D.BPN14770 was 100-fold more potent for improving memory and cognition in humanized PDE4D(hPDE4D)mice,which expressed the primate-specific binding sequence,compared to wild-type mice;meanwhile,it exhibited low potency in a mouse surrogate model for emesis.The behavioral and matching neurochemical data presented established a relationship between PDE4D target engagement and effects on cognition for BPN14770.Furthermore,BPN14770 reversed memory and cognitive deficits induced byβ-amyloid peptide 1-42(Aβ42)in Morris water maze,Y maze and novel object recognition tests in the humanized PDE4D mice.The morphological analyses suggested that the number of dendrites and the dendritic length in the CA1 of hippocampus were significantly increased after the Aβ42-treated hPDE4D mice were administered of BPN14770 for two weeks.The neurochemical and molecular biological assays suggested that neuroplasticity-related proteins and neurotrophic factor BDNF in the hippocampus of hPDE4D mice were significantly increased after the hPDE4D mice were treated with BPN14770.These findings suggest clinical potential for PDE4D selective inhibitors in disorders with cognitive deficits such as Alzheimer’s disease,which affects approximately 20 million people worldwide and nearly 5 million people in the United States.

Biodistribution and preparation of technetium-99m-labeled D-D3 monoclonal antibody against pro-gastrin-releasing peptide ~31-98） in mice

Background We previously reported that iodine-131（131^I）-Iabeled anti-pro-gastrin-releasing peptide （ProGRP（31-98）） monoclonal antibody D-D3 could selectively accumulate in the tumor sites of nude mice bearing small cell lung cancer （SCLC） xenografts. However, 1311-D-D3 was cleared slowly from the body, and the best radioimmunoimaging time for SCLC was 72-96 hours after injection. The aims of this study were to radiolabel anti-ProGRP（31-98） D-D3 monoclonal antibody with technetium-99m（99m^Tc） and to investigate the biodistribution of this antibody in healthy ICR mice. Methods D-D3was labeled with 99m^Tc via the 2-mercaptoethanol reduction method. 99m^Tc-D-g3 was purified by the gel column separation method. The labeling efficiency and radiochemical purity were measured by thin-layer chromatography. The immunological activity of 99m^Tc-D-O3 was determined with cell conjugation assays. 99m^Tc-D-D3 was injected into healthy ICR mice via a tail vein, and all the healthy ICR mice were sacrificed by cervical dislocation at a designated time. Then, the blood and major organs were removed and weighed, and counted in a gamma scintillation counter to determine the percentage of the injected dose per gram （%ID/g）. Results The labeling rate and the radiochemical purity of 99m^TC-D-D3 were （73.87±2.89）% and （94.13±4.49）%, respectively. The immunobinding rates of 99m^Tc-O-D3 to the human small cell lung cancer NCI-H446 cell line and lung adenocarcinoma A549 cell line were （81.2±2.37）% and （24.3±1.46）%, respectively. The distribution data of normal ICR mice demonstrated that 99m^TC-D-D3was mainly distributed in the liver, kidney and lung, and less in the brain tissue and muscle. Conclusions 99m^Tc-D-D3 antibody not only had high radiochemical purity, but also had good stability both in vitro and in vivo, and maintained good immunological activity. 99m^Tc-D-D3 was metabolized mainly in the kidney and liver, and the blood radioactivity decreased rapidly. Thus, 99m^Tc-O-D3 is conducive to the radioimmunoimaging of SCLC.

Evaluating Anti-SmDl-amino-acid 83-119 Peptide Reactivity in Children with Systemic Lupus Erythematosus and Other Immunological Diseases

Background： SmD1-amino-acid 83-119 peptide （SmD183-119） is the major epitope of Smith （Sm） antigen, which is specific for adult systemic lupus erythematosus （SLE）. The anti-SmD183-119 antibody has exhibited higher sensitivity and specificity than anti-Sm antibody in diagnosing adult SLE. However, the utility of anti-SmD183-119 antibodies remains unclear in children with SLE （cSLE）. This study aimed to assess the characteristics of anti-StuD 183-119 antibody in the diagnosis of cSLE. Methods： Samples from 242 children with different rheumatological and immunological disorders, including autoimmune diseases （SLE [n = 46] and ankylosing spondylitis [AS, n = 11]）, nonautoimmune diseases （Henoch-Schonlein purpura [HSP, n = 60], idiopathic thrombocytopenia purpura [n = 27], hematuria [n = 59], and arthralgia [n = 39]） were collected from Shanghai Children＇s Medical Center from March 6, 2012 to February 27, 2014. Seventy age- and sex-matched patients were enrolled in this study as the negative controls. All the patients＇ sera were analyzed for the anti-SmD 183-119, anti-Sm, anti-U 1-nRNP, anti-double-stranded DNA （dsDNA）, anti-nucleosome, anti-SSA/Ro60, anti-SSA/Ro52, anti-SSB, anti-Scl-70, and anti-histone antibodies using the immunoblotting assay. The differences in sensitivity and specificity between anti-SmD183-119 and anti-Sm antibodies were compared by Chi-square test. The correlations between anti-SmD183-119 and other auto-antibodies were analyzed using the Spearman＇s correlation analysis. A value of P 〈 0.05 was considered statistically significant. Results： Thirty-six out of 46 patients with cSLE were found to be positive for anti-SmD183-119, while 12 patients from the cSLE cohort were found to be positive for anti-Sm. Compared to cSLE, it has been shown that anti-SmD183-119 was only detected in 27.3% of patients with AS and 16.7% of patients with HSP. In comparison with anti-Sm, it has been demonstrated that anti-StuD 183-119 had a higher sensitivity （78.3% vs. 26.1%, 2＇2 = 25.1, P 〈 0.05） and a lower specificity （90.8% vs. 100%, x^2 = 13.6, P 〈 0.05） in the diagnosis of cSLE. Further analysis revealed that anti-StuD 183-119 antibodies were positively correlated with anti-dsDNA, anti-nucleosome, and anti-histone antibodies in cSLE. Moreover, it has been clearly shown that anti-SmD183-119was more sensitive than anti-Sm in discriminating autoimmune diseases from nonautoimmune disorders in patients with arthralgia or hematuria. Conclusions： Measurement of anti-SmD183-119 in patients with cSLE has a higher sensitivity and a marginally lower specificity than anti-Sin. It has been suggested that inclusion of anti-SmD183-119 testing in the integrated laboratory diagnosis ofcSLE may significantly improve the overall sensitivity in child populations.

[Gly14]-Humanin Protects Against Amyloid b Peptide-Induced Impairment of Spatial Learning and Memory in Rats

Alzheimer disease （AD）, a progressive neurodegenerative disorder, is characterized by cognitive decline and the accumulation of senile plaques in the brain. Amyloid β protein （Aβ） in the plaques is thought to be responsible for the memory loss in AD patients. [Glyl4]- humanin （HNG）, a derivative of humanin （HN）, has much stronger neuroprotective effects than natural HN in vitro. However, clarification of the Aβ active center and the neuroprotective mechanism of HN still need in vivo evidence. The present study first compared the in vivo bio- logical effects of three Aβ fragments （1-42, 31-35, and 35-31） on spatial memory in rats, and investigated the neuroprotective effects and molecular mechanisms of HNG. The results showed that intrahippocampal injection of Aβ1-2 and Aβ1-35 almost equally impaired spatial learning and memory, but the reversed sequence Aβ1-31 did not have any effect; a high dose of Aβ1-35 （20 nmol） produced a more detrimental response than a low dose （2 nmol）; Aβ1-35 injection also disrupted gene and protein expression in the hippocampus, with up-regulation of caspase3 and down-regulation of STAT3; pretreatment with HNG not only protected spatial memory but also rescued STAT3 from Aβ-induced disruption; and the neuropro- tective effects of HNG were effectively counteracted by genistein, a specific tyrosine kinase inhibitor. These results clearly show that sequence 31-35 in AI5 is the shortest active center responsible for the neurotoxicity of A[3 from molecule to behavior; and HNG protects spatial learning and memory in rats against Aβ-indnced insults; and prob- ably involves the activation of tyrosine kinases and sub- sequent beneficial modulation of STAT3 and caspase3.