Construction and delivery of gene therapy vector containing soluble TNFalpha receptor-IgGFc fusion gene for the treatment of allergic rhinitis

Tumor necrosis factor alpha plays primary role in the pathogenesis of inflammatory diseases. TNFalpha is essential for antigen-specific IgE production and for the induction of Th2-type cytokines. The lack of TNFalpha inhibited the development of allergic rhinitis. In this study, the chimeric gene of soluble TNF receptor and IgGFc fragment (sTNFR-IgGFc) was cloned into the EBV-based plasmid pGEG. When the plasmid pGEG.sTNFR-IgGFc was transferred to endothelium cell, a considerable expression of the sTNFR-IgGFc fusion protein was detected. Moreover, the expression product in the supernatant could antagonize the cytolytic activity of TNFalpha on L929 cells. Then the plasmid was delivered into nasal mucosa of allergic rhinitis mice to determine its effect on this animal model. Results showed that symptoms in treated group were improved. Pathological examination showed the numbers of eosinophil, mast cell and IL-5(+) cells in treated groups were reduced compared with placebo group. These data showed that pGEG.sTNFR-IgGFc expression plasmid is potential for the treatment of allergic rhinitis, and suggest that the antagonist of TNFalpha may provide a new approach for the treatment of allergic rhinitis.     

99mTc-labeled cyclic RGDfK dimer: initial evaluation for SPECT imaging of glioma integrin alphavbeta3 expression

This report describes the evaluation of biodistribution properties of three radiotracers, [(99m)Tc(SQ168)(EDDA)], [(99m)Tc(SQ168)(tricine)(PDA)], and [(99m)Tc(SQ168)(tricine)(TPPTS)] (SQ168 = [2-[[[5-[carboonyl]-2-pyridinyl]hydrazono]methyl]benzenesulfonic acid]-Glu(cyclo{Lys-Arg-Gly-Asp-d-Phe})-cyclo{Lys-Arg-Gly-Asp-d-Phe}; EDDA = ethylenediamine-N,N'-diacetic acid; PDA = 2,5-pyridinedicarboxylic acid; TPPTS = trisodium triphenylphosphine-3,3',3' '-trisulfonate), and their potential to image the glioma integrin alpha(v)beta(3) expression in BALB/c nude mice bearing the U87MG human glioma xenografts. It was found that all three radiotracers were able to localize in glioma tumors with a relatively high tumor uptake and long tumor retention time by binding to the integrin alpha(v)beta(3) expressed on both tumor cells and endothelial cells of tumor neovasculature. It seems that the coligand has minimal effect on integrin alpha(v)beta(3) targeting capability of the (99m)Tc-labeled RGDfK dimer, but it has a significant impact on their biodistribution properties. For example, the complex [(99m)Tc(SQ168)(tricine)(TPPTS)] has the lowest liver uptake and the highest metabolic stability in normal BALB/c nude mice. Results from SPECT imaging studies show that the glioma tumors can be clearly visualized with all three radiotracers at 4 h postinjection. Among the three radiotracers evaluated in this study, [(99m)Tc(SQ168)(tricine)(TPPTS)] has the best imaging quality and is a promising candidate for more preclinical evaluations in the future.     

New method for HPLC separation and fluorescence detection of malonaldehyde in normal human plasma

A new method for the detection of free and total malonaldehyde (MDA) in human plasma samples based on the derivatization of MDA with 9-fluorenylmethoxycarbonyl hydrazine (FMOC-hydrazine) in an acidic medium was developed. Derivatization was achieved after 4 h at 50 degrees C. The derivatized samples were analyzed by HPLC using a reversed-phase C18 column with fluorescence detection (Ex=270 nm, Em=310 nm). The benefit of this direct injection of deproteinized plasma is to avoid the use of an internal standard. The detection limit was 0.1 pmol (4.0 nmol/L). The recovery of MDA spiked in different human plasma samples was 95.3% (n=25; R.S.D. 5.1%) for the hydrolysation procedure. The total and free MDA in plasma of 15 healthy male volunteers are 426+/-29.8 nmol/L and 153+/-9.6 nmol/L, respectively.     

Determination of omeprazole in rat plasma by high-performance liquid chromatography without solvent extraction

A HPLC method without solvent extraction and using ultraviolet detection at 302 nm for the determination of omeprazole in rat plasma has been validated. Plasma samples after pretreatment with acetonitrile to effect deproteinization were dried under N(2) at 40 degrees C and reconstituted with mobile phase. The standard calibration curve for omeprazole was linear (r(2)=0.9999) over the concentration range of 0.02-3 microgml(-1). The intra- and inter-day assay variability range was 4.8-9.2% and 5.2-10.3% individually. This method has been successfully applied to a pharmacokinetic study of omeprazole in rats.     

Plasma phospholipid metabolic profiling and biomarkers of mouse IgA nephropathy

IgA nephropathy is the most common form of glomerulonephritis (GN) and it could progress to end-stage renal failure within 10 years. Participating in biological processes in various pathways, phospholipids as a class of important constituents in the biomembranes have been paid increasing attention in many fields. However, phospholipids metabolism in glomerular disease was not clear, especially in IgA nephropathy. In this paper, the plasma phospholipid metabolic profile in mouse IgA nephropathy was investigated to discover the potential biomarkers on the progression of this disease by using high performance liquid chromatography/mass spectrometry (HPLC/MS) and the principal components analysis (PCA) as well as partial least squares-discriminant analysis (PLS-DA). The experimental mouse models of IgA nephropathy were established by oral immune and BSA injection. It was found that expression of intercellular adhesion molecule-1 (ICAM-1) in the glomeruli had a significant correlation with proteinuria in mouse IgA nephropathy. The association between plasma phospholipids and expression of ICAM-1 in the glomeruli of IgA nephropathy suggested C18:0/C18:0 PS (phosphatidylserine), C18:0/C22:5 PS (phosphatidylserine) and C18:0/C20:4 PI (phosphatidylinositol) were possible biomarkers of IgA nephropathy. The results show that the plasma phospholipid metabolic profiles from HPLC/MS combining with PCA and PLS-DA can be used not only to differentiate the IgA nephropathy from the controls, but also to discover and identify the potential biomarkers.     

New bisabolane sesquiterpenes and coumarin from Leontopodium longifolium

From the roots of Leontopotium longifolium, three new bisabolane sesquiterpenes, rel-(1S,4R,5S,6R)-4,5-diacetoxy-6-[(R)-1,5-dimethylhexa-3,5-dienyl]-3-methylcyclohex-2-enyl (Z)-2-methylbut-2-enoate (1), rel-(1S,4R,5S,6R)-4,5-diacetoxy-6-[(R)-5-hydroxy-1,5-dimethylhex-3-enyl]-3-methylcyclohex-2-enyl (Z)-2-methylbut-2-enoate (2), rel-(1R,2S,4R,5S)-4-acetoxy-2-[(R)-5-hydroxy-1,5-dimethylhex-3-enyl]-5-methylcyclohexyl (Z)-2-methylbut-2-enoate (3), and a new coumarin, 2,3-dihydro-5-hydroxy-2-(1-methylethenyl)-7H-pyrano[2,3-g][1,4]benzodioxin-7-one (4) together with nine known compounds have been isolated. The structures of these compounds were established by spectroscopic methods. Compounds 1 and 2 exhibited moderate cytotoxic activities against human promyelocytic leukemia (HL-60) cells.     

Proteome analysis of up-regulated proteins in the rat spinal cord induced by transection injury

The inability of the CNS to regenerate in adult mammals propels us to reveal associated proteins involved in the injured CNS. In this paper, either thoracic laminectomy (as sham control) or thoracic spinal cord transection was performed on male adult rats. Five days after surgery, the whole spinal cord tissue was dissected and fractionated into water-soluble (dissolved in Tris buffer) and water-insoluble (dissolved in a solution containing chaotropes and surfactants) portions for 2-DE. Protein identification was performed by MS and further confirmed by Western blot. As a result, over 30 protein spots in the injured spinal cord were shown to be up-regulated no less than 1.5-fold. These identified proteins possibly play various roles during the injury and repair process and may be functionally categorized as several different groups, such as stress-responsive and metabolic changes, lipid and protein degeneration, neural survival and regeneration. In particular, over-expression of 11-zinc finger protein and glypican may be responsible for the inhibition of axonal growth and regeneration. Moreover, three unknown proteins with novel sequences were found to be up-regulated by spinal cord injury. Further characterization of these molecules may help us come closer to understanding the mechanisms that underlie the inability of the adult CNS to regenerate.     

华山松胚性愈伤组织诱导与幼胚离体培养

探索华山松(Pinus armandii)体细胞胚胎发生技术对其实施规模化无性繁殖和开展遗传转化具有重要意义。本文以1/2LM为基本培养基, 通过激素调节等措施对华山松的胚性愈伤组织诱导和幼胚的离体培养技术进行了初步研究。研究结果: 胚性愈伤组织诱导率最高可达52.71%, 但愈伤组织继代培养后没有体细胞胚胎的分化; 首次从其子叶期的幼胚中直接诱导出具有根和茎的完整植株, 诱导率达92%以上。文章确认了采集的幼胚发育状态对胚性愈伤组织的诱导有重要影响, 并对诱导的培养条件等进行了探讨。     

促炎基因ALOX5AP基因多态性与脑卒中的相关性研究

白三烯是作用较强的促炎症因子,在动脉粥样硬化的发生发展中发挥着重要作用。5-脂氧合酶激活蛋白是白三烯合成的关键调控因素,通过全基因组扫描的连锁分析和关联分析发现编码5-脂氧合酶激活蛋白的基因ALOX5AP在白种人中与心肌梗塞和脑卒中的患病风险相关。然而,目前尚无关于该基因与亚洲人脑卒中患病风险的遗传学资料。本研究探讨了ALOX5AP基因多态与脑卒中及其亚型易感性的关系。采用PCR—RFLP方法,对来自7个临床中心的1713名对照和1773名脑卒中患者检测了ALOX5AP基因的4个SNPs：SG13S25、SG13S114、SG13S89和SG13S32。多元logistic回归方法校正传统危险因素后分析基因多态与脑卒中患病风险的独立相关性。结果表明,人群未发现SG13S25和SG13S32具有多态性;ALOX5AP基因多态SG13S114A等位基因频率在男性脑梗塞组显著高于对照组（33．6％VS29．2％;P=0．014）,SG13S114AA基因型增加男性脑梗塞1．62倍的发病风险（95％CI：1．1-2．35：P=0．012）。多态SG13S89G／A与脑梗塞的易感性不相关。单体型分析表明单体型频率在脑卒中患者和对照组间无显著的统计学差异。因此,本研究结果提示ALOX5AP基因多态的等位基因和基因型频率在东、西方人群存在种族差异,SG13S114AA基因型增加中国人群男性脑梗塞的易感性。     

假苍耳的生活史进程中几种生理生化指标的变化

本文试图从生理生化的角度对假苍耳(Iva xanthifolia)生活史进程中可溶性糖类、赤霉素、单宁以及黄酮的变化进行探讨。通过对假苍耳在生长发育期间几种生理生化指标的测定, 结果表明, 在假苍耳生活史进程的不同阶段, 其体内各种代谢产物的含量基本都在种子或芽阶段具有最高含量。此外, 不同发育阶段可溶性还原糖含量的变化规律相似: 芽>花序>苗>成株>种子。除在花序和苗阶段没有测到海藻糖, 其他各阶段海藻糖的含量变化如下: 芽>成株>种子。另外, 只有在种子阶段检测到棉子糖, 其含量为15.43 mg.g-1。赤霉素含量的变化规律如下: 种子＞芽≈苗≈花序＞成株。单宁含量的变化趋势: 种子＞成株＞苗＞芽＞花序。黄酮含量的变化趋势: 种子＞芽＞成株≈花序＞苗。值得注意的是, 当单宁/黄酮的比值接近1时, 植物体内需要的单宁和黄酮的含量则相对较低; 相反, 当单宁/黄酮的比值接近0时, 植物体内需要的单宁和黄酮的含量则较高。