Environmental stress sensitivity of an ascorbic acid-deficient Arabidopsis mutant.

L-ascorbic acid (vitamin C) is a powerful reducing agent found in millimolar concentrations in plants, and is proposed to play an important role in scavenging free radicals in plants and animals. However, surprisingly little is known about the role of this antioxidant in plant environmental stress adaptation or ascorbate biosynthesis. We report the isolation of soz1, a semi-dominant ozone-sensitive mutant that accumulates only 30% of the normal ascorbate concentration. The results of genetic approaches and feeding studies show that the ascorbate concentration affects foliar resistance to the oxidizing gas ozone. Consistent with the proposed role for ascorbate in reactive oxygen species detoxification, lipid peroxides are elevated in soz1, but not in wild type following ozone fumigation. We show that the soz1 mutant is hypersensitive to both sulfur dioxide and ultraviolet B irradiation, thus implicating ascorbate in defense against varied environmental stresses. In addition to defining the first ascorbate deficient mutant in plants, these results indicate that screening for ozone-sensitive mutants is a powerful method for identifying physiologically important antioxidant mechanisms and signal transduction pathways. Analysis of soz1 should lead to more information about the physiological roles and metabolism of ascorbate.     

下坡运动对大鼠骨骼肌肌浆网功能的影响

目的：观测研究下坡(离心)运动对大鼠骨骼肌肌浆网Ca2+-ATP酶活性,Ca2+摄取与释放在量与时程上的影响。此外,测定离子载体的刺激作用,即测定在含与不含(Ca2+离子载体)A23187时Ca2+-ATP酶活性的比值,用以评定囊泡的完整性。方法：成年雄性SD大鼠随机分为对照与离心运动组, 离心运动的大鼠分别于运动后即刻, 4, 24, 48, 72 和144h后取样 (n=7). 离心运动方式采用90min持续跑台下坡运动(-16°;15m/min)。取大鼠红股肌制备组织匀浆, 测定肌浆网Ca2+-ATP酶活性,Ca2+摄取与释放。结果：与对照组[19.25±1.38 nmol ·min-1·(mg protein)-1]相比, 肌浆网Ca2+摄取分别于运动后即刻和4h下降了29% and 36% (P<0.05), 24h依然降低(P<0.05). 肌浆网Ca2+释放与对照组[31.06±2.36 nmol·min-1·(mg protein)-1] 相比,也分别于运动后即刻和4h下降了37% and 39% (P<0.05), 24h持续降低(P<0.05). 用含离子载体测定的肌浆网Ca2+-ATP酶活性运动后4h降低了31%(P<0.05), 并于运动后24h仍然降低 (P<0.05)。运动后, 含与不含A23187时测定的Ca2+-ATP酶活性的比值未见显著性改变, 表明该运动没有明显改变肌浆网膜的完整性。结论：一次性低强度，长时间下坡运动导致肌浆网功能长时间降低, 运动后恢复期两天尚未完全恢复, 亦可构成离心运动诱导的骨骼肌某些功能降低的基础。提示这些变化可能产生于离心收缩时肌节长度不匀一性所造成的张力应激。     

体外制成复合软质再生颅骨修复材料填充犬颅骨缺损

目的:目前颅骨修补材料有很多种,但都为异源性无机骨替代物,并且应用该方法又要给患者再次行开颅手术,实验拟开展新型颅骨再生材料的研究。方法:实验于2006-05/11在解放军第一五七医院动物中心及中山大学附属第三医院动物实验室完成。①实验动物:30只犬随机分为实验组20只,对照组10只。②实验方法:应用纳米级羟基磷灰石为支架和成骨细胞培养,加入脱矿的犬类骨基质为载体的重组人类骨形成蛋白2,制成复合软质再生颅骨。实验组犬在右侧颅骨缺损中填补藻酸钙凝胶、成骨细胞、纳米级骨粉的复合材料,左侧颅骨缺损中填补藻酸钙凝胶、成骨细胞、纳米级骨粉和重组人类骨形成蛋白2的复合材料。对照组犬在右侧为单纯颅骨缺损,左侧颅骨缺损中填补藻酸钙凝胶、成骨细胞、纳米级骨粉和重组人类骨形成蛋白2复合材料。实验过程中对动物处置符合动物伦理学标准。③实验评估:手术后1,2,3,6个月X射线片检查颅骨缺损修复情况,对再生的颅骨组织标本进行茜素红S染色,观察成骨能力及再生材料骨膜组织细胞体外培养情况。结果:实验动物均进入结果分析。术后1个月,成骨活跃,骨端新生骨小梁基本覆盖骨断端,缺损区可见较多新生骨小梁形成,骨端新生骨小梁向缺损区长入;术后2个月可见较多散在骨岛形成;术后3个月可见成熟骨,并有髓腔形成,缺损区大量新骨形成。而各对照组骨断端处有散在骨岛,或被增生的纤维结缔组织占据,可见大量纤维组织及毛细血管长入,植入的基质材料基本被吸收,无新骨生成。结论:应用纳米级羟基磷灰石为支架和成骨细胞培养,加入脱矿骨基质为载体的重组人类骨形成蛋白2,制成的复合软质再生颅骨能自身代谢并逐渐骨化,形成新的颅骨。     

Antitumor activity of antisense clusterin oligonucleotides is improved in vitro and in vivo by incorporation of 2'-O-(2-methoxy)ethyl chemistry

Phosphorothioate (P=S) antisense oligonucleotides (ASO) targeting the cell survival gene clusterin synergistically enhance castration- and chemotherapy-induced apoptosis in prostate cancer xenografts. This study compares efficacy, tissue half-lives, and toxicity of P=S clusterin ASO to third-generation backbone 2'-O-(2-methoxy)ethyl (2'MOE) ribose-modified clusterin ASO. Northern analysis quantified changes in clusterin mRNA levels in human PC-3 cells and tumors. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay measured effects of combined clusterin ASO plus paclitaxel on PC-3 cell growth. Athymic mice bearing PC-3 tumors were treated with paclitaxel plus either P=S clusterin ASO, 2'-MOE clusterin ASO, or mismatch control oligonucleotides for 28 days. Weekly body weights and serum parameters were measured to assess toxicity. Tissue half-life of P=S and 2'-MOE ASO in PC-3 tumors was assessed using capillary gel electrophoresis (CGE). Both 2'-MOE and P=S ASO decreased clusterin mRNA levels in a dose-dependent and sequence-specific manner. 2'-MOE ASO more potently suppressed clusterin mRNA (80 versus 40% at 500 nM) compared with P=S ASO. IC(50) of paclitaxel was equally reduced (50--75%) by both compounds. In vivo tissue half-life was significantly longer for 2'-MOE-modified ASO than for P=S ASO (5 versus 0.5 days). Using CGE, >90% of detected 2'-MOE ASO in tumor tissue was full length. Weekly administration of 2'-MOE clusterin ASO was equivalent to daily P=S clusterin ASO in enhancing paclitaxel efficacy in vivo. 2'-MOE-modified ASO potently suppressed clusterin expression and prolonged tissue half-lives with no additional side effects. These results support the use of 2'-MOE-modified ASO over conventional P=S ASO by potentially increasing potency and allowing longer dosing intervals in clinical trials.     

New serological biomarkers of inflammatory bowel disease

Serological biomarkers in inflammatory bowel disease （IBD） are a rapidly expanding list of non-invasive tests for objective assessments of disease activity, early diagnosis, prognosis evaluation and surveillance. This review summarizes both old and new biomarkers in IBD, but focuses on the development and characterization of new serological biomarkers （identifi ed since 2007）. These include fi ve new anti-glycan antibodies, anti-chitobioside IgA （ACCA）, anti-laminaribioside IgG （ALCA）, anti-manobioside IgG （AMCA）, and antibodies against chemically synthesized （∑） two major oligomannose epitopes, Man α-1,3 Man α-1,2 Man （∑Man3） and Man α-1,3 Man α-1,2 Man α-1,2 Man （∑Man4）. These new biomarkers serve as valuable complementary tools to existing biomarkers not only in differentiating Crohn＇s disease （CD）, ulcerative colitis （UC）, normal and other non-IBD gut diseases, but also in predicting disease involvement （ileum vs colon）, IBD risk （as subclinical biomarkers）, and disease course （risk of complication and surgery）. Interestingly, the prevalence of the antiglycan antibodies, including anti-Saccharomyces cerevisiae antibodies （ASCA）, ALCA and AMCA, was found to be associated with single nucleotide polymorphisms （SNPs） of IBD susceptible genes such as NOD2/CARD15, NOD1/CARD4, toll-likereceptors （TLR） 2 and 4, and β-defensin-1. Further-more, a gene dosage effect was observed： anti-glycan positivity became more frequent as the number of NOD2/CARD15 SNPS increased. Other new serum/ plasma IBD biomarkers reviewed include ubiquitination factor E4A （UBE4A）, CXCL16 （a chemokine）, resistin, and apolipoprotein A-IV. This review also discusses the most recent studies in IBD biomarker discovery by the application of new technologies such as proteomics, fourier transform near-infrared spectroscopy, and multiplex enzyme-linked immunosorbent assay （ELISA）＇s （with an emphasis on cytokine/chemokine profiling）. Finally, the prospects o     

Long-term Functional and Quality of Life Outcomes of Patients After Repair of Large Perianal Skin Defects for Paget’s and Bowen’s Disease

Introduction  

The assessment of long- term functional and quality of life outcomes of these patients following repair of large defects after surgical excision has not been reported.     

Use of risk stratification to target therapies in patients with recent onset arthritis; design of a prospective randomized multicenter controlled trial

Background  

Early and intensive treatment is important to inducing remission and preventing joint damage in patients with rheumatoid arthritis. While intensive combination therapy (Disease Modifying Anti-rheumatic Drugs and/or biologicals) is the most effective, rheumatologists in daily clinical practice prefer to start with monotherapy methotrexate and bridging corticosteroids. Intensive treatment should be started as soon as the first symptoms manifest, but at this early stage, ACR criteria may not be fulfilled, and there is a danger of over-treatment. We will therefore determine which induction therapy is most effective in the very early stage of persistent arthritis. To overcome over-treatment and under-treatment, the intensity of induction therapy will be based on a prediction model that predicts patients' propensity for persistent arthritis.     

Acrolein activates matrix metalloproteinases by increasing reactive oxygen species in macrophages

Acrolein is a ubiquitous component of environmental pollutants such as automobile exhaust, cigarette, wood, and coal smoke. It is also a natural constituent of several foods and is generated endogenously during inflammation or oxidation of unsaturated lipids. Because increased inflammation and episodic exposure to acrolein-rich pollutants such as traffic emissions or cigarette smoke have been linked to acute myocardial infarction, we examined the effects of acrolein on matrix metalloproteinases (MMPs), which destabilize atherosclerotic plaques. Our studies show that exposure to acrolein resulted in the secretion of MMP-9 from differentiated THP-1 macrophages. Acrolein-treatment of macrophages also led to an increase in reactive oxygen species (ROS), free intracellular calcium ([Ca²⁺]_i), and xanthine oxidase (XO) activity. ROS production was prevented by allopurinol, but not by rotenone or apocynin and by buffering changes in [Ca²⁺]_I with BAPTA-AM. The increase in MMP production was abolished by pre-treatment with the antioxidants Tiron and N-acetyl cysteine (NAC) or with the xanthine oxidase inhibitors allopurinol or oxypurinol. Finally, MMP activity was significantly stimulated in aortic sections from apoE-null mice containing advanced atherosclerotic lesions after exposure to acrolein ex vivo. These observations suggest that acrolein exposure results in MMP secretion from macrophages via a mechanism that involves an increase in [Ca²⁺]_I, leading to xanthine oxidase activation and an increase in ROS production. ROS-dependent activation of MMPs by acrolein could destabilize atherosclerotic lesions during brief episodes of inflammation or pollutant exposure.