流体剪应力对成骨细胞OPG和ODF表达的影响

通过平板流动腔装置对大鼠成骨细胞施以流体剪应力,研究剪应力作用对成骨细胞中骨保护因子(osteoprotegerin,OPG),破骨细胞分化因子(osteoclast differentiation factor,ODF)基因表达的影响.分别考察了剪应力大小和作用时间,以及单一水平和梯度变化的剪切力加载方式的影响.运用RT-PCR和蛋白质印迹技术检测OPG、ODFmRNA和蛋白质表达的变化,结果显示,剪切力作用下OPG的表达得到促进,ODF的表达受到抑制,mRNA与蛋白质表达的变化一致,这种影响与剪切力的大小和作用时间有关.1.0和1.5N/m2的剪应力作用效果比0.5N/m2明显,梯度变化的作用方式在作用效果上与最后一个梯度水平相当的恒定剪应力单独作用没有显著差异,在加载的24h内剪应力对OPG、ODF表达的影响始终存在.这种影响使得OPG/ODF的平衡向着OPG占优的方向发展,这种变化意味着骨吸收会受到抑制,提示力学刺激可能通过OPG/ODF调控系统对骨代谢平衡进行调控.     

重组胰高血糖素样肽-1与人血清白蛋白融合蛋白的纯化及活性研究

利用构建的重组菌株Pichia pastoris GS115/GLP-1/HSA,在10L发酵罐中表达了胰高血糖素样肽-1与人血清白蛋白融合蛋白(GLP-1/HSA),表达量为63.6mg/L。发酵液经中空纤维柱浓缩、疏水层析、阴离子交换层析和凝胶过滤分离纯化,获得了较高纯度的GLP-1/HSA,经HPLC分析纯度达95.8%。进一步的体内活性分析结果表明,GLP-1/HSA不仅具有天然GLP-1的生物活性,而且在给药后4h仍能发挥显著性降血糖作用。以上结果表明,利用Pichia pastoris分泌型表达系统和建立的分离纯化方法,能获得大量较高纯度的GLP-1/HSA,为进一步研究和开发能够用于糖尿病临床治疗的长效GLP-1类似物奠定了基础。     

Carnosol protects against spinal cord injury through Nrf-2 upregulation

Background: Carnosol is an ortho-diphenolic diterpene with excellent antioxidant potential. The present study was designed to identify the protective role of carnosol against spinal cord injury (SCI)-induced oxidative stress and inflammation in Wistar rats. Methods: In the present study, oxidative stress status was determined through estimating total antioxidant capacity, total oxidant status, lipid peroxide content, protein carbonyl and sulfhydryl levels, reactive oxygen species (ROS), antioxidant status (superoxide-dismutase, catalase, glutathione, glutathione peroxidase, glutathione-S-transferase). Inflammatory effects were determined by analyzing the expression of NF-κB and COX-2 through Western blot analysis. Further, carnosol-mediated redox homeostasis was analyzed by determining p-AKT and Nrf-2 levels. Results: SCI resulted in a significant increase in oxidative stress status through increased ROS generation, total oxidant levels, lipid peroxide content, protein carbonyl and sulfhydryl levels. The antioxidant status in SCI rats was significantly reduced, indicating imbalance in redox status. In addition, the expression of NF-κB and COX-2 was significantly upregulated, while p-AKT and Nrf-2 levels were downregulated in SCI rats. However, treatment with carnosol showed a significant enhancement in the antioxidant status with concomitant decline in oxidative stress parameters. Further, carnosol treatment regulated the key proteins in inflammation and redox status through significant downregulation of NF-κB and COX-2 levels and upregulation of p-AKT and Nrf-2 expression. Conclusion: Thus, the present study shows for the first time on the protective role of carnosol against SCI-induced oxidative stress and inflammation through modulating NF-κB, COX-2 and Nrf-2 levels in Wistar rats.     

Phosphoenolpyruvate Carboxykinase Is Involved in the Decarboxylation of Aspartate in the Bundle Sheath of Maize

We recently showed that maize (Zea mays L.) leaves contain appreciable amounts of phosphoenolpyruvate carboxykinase (PEPCK; R.P. Walker, R.M. Acheson, L.I. Técsi, R.C. Leegood [1997] Aust J Plant Physiol 24: 459–468). In the present study, we investigated the role of PEPCK in C₄ photosynthesis in maize. PEPCK activity and protein were enriched in extracts from bundle-sheath (BS) strands compared with whole-leaf extracts. Decarboxylation of [4-¹⁴C]aspartate (Asp) by BS strands was dependent on the presence of 2-oxoglutarate and Mn²⁺, was stimulated by ATP, was inhibited by the PEPCK-specific inhibitor 3-mercaptopicolinic acid, and was independent of illumination. The principal product of Asp metabolism was phosphoenolpyruvate, whereas pyruvate was a minor product. Decarboxylation of [4-¹⁴C]malate was stimulated severalfold by Asp and 3-phosphoglycerate, was only slightly reduced in the absence of Mn²⁺ or in the presence of 3-mercaptopicolinic acid, and was light dependent. Our data show that decarboxylation of Asp and malate in BS cells of maize occurs via two different pathways: Whereas malate is mainly decarboxylated by NADP-malic enzyme, decarboxylation of Asp is dependent on the activity of PEPCK.     

不同EGF和胰岛素水平对体外培养的大熊猫皮肤成纤维细胞的影响

用DME∶Ham sF12(1∶1)培养液,添加3个水平的EGF和2个水平的胰岛素,组合成6种培养体系(CS)分别培养大熊猫皮肤成纤维细胞。通过对细胞生长速度和染色体数目变异率进行测定,测得在添加10μg/ml的胰岛素和40 ng/ml的表皮生长因子的培养体系中,以1.673±0.185×105/ml密度接种细胞,经3.5天,密度达到6.890×105/ml,其生长速度最快;染色体数目为二倍体的百分率为75.77%。综合衡量,CS-5在本研究中更适合大熊猫皮肤成纤维细胞的培养。     

氮、磷、钾肥配施对桂花品种'金玉台阁'开花性状及叶片中叶绿素和营养元素含量的影响

以桂花品种'金玉台阁'(Osmanthus fragrans'Jinyu Taige')盆栽苗为供试材料,采用"3414"肥料效应试验设计方案,研究了不同单株施用量CO(NH2)2(0.0、1.0、2.0和3.0 g)、NH4H2PO4(0.0、1.5、3.0和4.5 g)和KCl(0.0、0.5、1.0、1.5 g)对秋冬生长期(2014年10月至2015年2月)'金玉台阁'开花性状及叶片中叶绿素和营养元素的影响进行了比较和分析,并筛选出适宜的氮、磷、钾肥施用量.结果表明:与CK〔CO(NH2)2、NH4H2PO4和KCl的单株施用量均为0.0 g〕组相比,各氮、磷、钾肥配施处理组'金玉台阁'的单株产花量、花瓣相对含水量、花径以及叶片中叶绿素、全氮、全磷和全钾的含量总体上显著升高.CO(NH2)2单株施用量对'金玉台阁'单株产花量、叶片中叶绿素含量和全氮含量的影响最大,NH4H2PO4单株施用量对这3个指标的影响次之;NH4H2PO4单株施用量对'金玉台阁'叶片中全磷含量的影响最大;KCl单株施用量对'金玉台阁'花瓣相对含水量、花径及叶片中全钾含量的影响最大.在秋冬生长期内叶片中叶绿素含量呈先降低后升高的趋势,均在12月份降至最低值.通过建立'金玉台阁'单株产花量(y)与CO(NH2)2、NH4 H2 PO4和KCl的单株施用量(分别为xN、xP和xK)间的肥料效应模型,得到的三元二次肥料效应模型为y=1.2027+0.7561×xN+0.2632×xP+0.4590×xK-0.2761×xN2-0.1201×xP2-0.5007×xK2+0.1901×xN×xP-0.0960×xN×xK+0.1185×xP×xK.以单株产花量为目标,'金玉台阁'的CO(NH2)2、NH4 H2 PO4和KCl的最佳单株施用量分别为2.58、3.56和0.86 g.     

喀斯特山区粗裂地钱风兜亚种冬季芽胞杯的形态多样性特征及芽胞传播观察

芽胞杯是地钱属特有的无性繁殖器官,关于其冬季形态特征及繁殖传播的行为研究较少。现以贵州喀斯特山区常见的粗裂地钱风兜亚种（Marchantia paleacea subsp.diptera）为代表,在最冷的冬季1月份,对其芽胞杯、杯内芽胞产量及传播方式进行野外定点观察和采样分析。结果显示：（1）冬季芽胞杯形态多样。根据其颜色和杯内芽胞特点将其划分为4个生长时期：未成熟期（透明）、成熟期（绿色）、衰退期（紫色）和衰亡期（紫黑色）,反映了冬季芽胞杯生长发育的不同阶段。（2）各生长时期的芽胞杯数量不同,表现出有序的凋亡特征。在统计的708个芽胞杯中,4个时期芽胞杯数量分别为62、209、254和183个,且不同时期的芽胞杯内芽胞的平均产量明显不同,不同时期单杯芽胞的平均产量分别为42、131、87和0 个;（3）冬季芽胞杯及芽胞在配子体上的密度较高,每平方米分别达到10 139和754 889个;（4）除春夏季常见的被雨滴敲打传播外,通过重力作用传播是冬季芽胞的一种重要传播方式。冬季粗裂地钱风兜亚种配子体上的芽胞杯处在不同的生长时期,形成的芽胞仍十分丰富,这对该物种适应喀斯特山区最冷月严苛环境条件具有积极的意义。     

氨氮和亚硝酸盐对红螯螯虾幼虾和亚成虾的急性毒力

【目的】近年来,红螯螯虾养殖面积越来越广泛,明确不同规格的红螯螯虾对氨氮和亚硝酸盐的耐受力,有利于提高其养成率,促进其养殖业的健康发展。【方法】在水温24~25℃、p H 7.9~8.0的条件下,研究了氨氮和亚硝酸盐对红螯螯虾幼虾和亚成虾的急性毒性,分析半致死浓度(LC50)和安全浓度(SC)。【结果】总氨氮对红螯螯虾幼虾的24、48、72和96 h LC50分别为188.0、136.15、104.67和88.00 mg·L~(-1),SC为8.80 mg·L~(-1);总氨氮对亚成虾的24、48、72和96 h LC50分别为344.01、270.46、205.15和167.68 mg·L~(-1),SC为16.77 mg·L~(-1);非离子氨对幼虾的24、48、72和96 h LC50分别为10.16、7.35、5.65和4.75 mg·L~(-1),SC为0.48 mg·L~(-1);非离子氨对亚成虾的24、48、72和96 h LC50分别为18.58、14.60、11.08和9.05 mg·L~(-1),SC为0.91 mg·L~(-1);亚硝酸盐对幼虾的24、48、72和96 h LC50分别为46.76、33.88、27.97和22.81 mg·L~(-1),SC为2.28 mg·L~(-1);亚硝酸盐对亚成虾的24、48、72和96 h LC50分别为77.56、59.33、45.41和37.48 mg·L~(-1),SC为3.75 mg·L~(-1)。【结论】红螯螯虾对氨氮的耐受力高于亚硝酸盐,亚成虾对氨氮和亚硝酸盐的耐受力高于幼虾。