Histone H2A as a transfection agent in crayfish hematopoietic tissue cells

We report a novel and highly efficient dsRNA transfection system based on one of the nuclear proteins, namely, histone H2A. RT-PCR semi-quantitative analysis of silencing target gene shows that the transfection efficiency of histone H2A is higher than Effectene or liposome-based transfection systems. Importantly, the high efficiency of histone H2A was associated with very low toxicity to the transfected crayfish hematopoietic tissue (Hpt) cells. The non-toxicity, effectiveness and specificity of histone H2A as a transfection agent provides a cheap, simple, highly efficient and reproducible gene delivery system, particularly for the sensitive cell cultures of crustacean animals such as crayfish and shrimp. (c) 2006 Elsevier Ltd. All rights reserved

不同品种、盐度及月份菊芋叶中绿原酸的含量测定

建立HPLC 法测定菊芋叶中绿原酸含量的方法,对菊芋植株中不同部位绿原酸的分布规律,以及不同品系、盐度、采摘时间等因素对菊芋叶中绿原酸含量的影响情况进行研究。方法:色谱柱为Hypersil BDS C-18(4.6×250 mm,5 μm),流速为1 mL min-1 ,检测波长为327 nm,进样量为10 μL,柱温为30℃,流动相为0.1%三氟乙酸水(A)-乙腈(B)。结果:菊芋叶中绿原酸的含量(3.40%)显著高于其他部位绿原酸的含量;10 月份菊芋叶中绿原酸的平均含量达最大值(2.74%)。南芋(N)受盐度影响较小,菊芋叶中绿原酸的平均含量(3.15%)明显高于莱芋(L)叶中绿原酸的平均含量(2.32%)。结论:菊芋植株中的绿原酸主要存在于菊芋叶中;不同采摘时间对菊芋叶中绿原酸的积累影响很大,10 月中旬可以作为菊芋叶的最佳采收时间;不同菊芋品种对盐度高低的适应能力不同,其中,南芋可能更适 合在盐碱地土壤类型中生长

Effects of the Xitongxiao prescription on apoptosis and proliferation of chondrocytes in knee

目的：探讨膝痛消方对促进软骨细胞增殖和抑制IL-1β诱导的软骨细胞凋亡的作用。方法：体外培养膝关节软骨细胞，用阿利新蓝染色方法检测软骨细胞蛋白多糖，用Ⅱ型胶原免疫荧光染色检测软骨细胞Ⅱ型胶原的表达，以鉴定软骨细胞；用MTS检测软骨细胞的增殖；用IL-1β诱导软骨细胞凋亡，AnnexinV/PI双染检测细胞凋亡。结果：膝关节剥离得到的细胞形态为多角形或梭形，阿利新蓝与Ⅱ型胶原免疫荧光染色均呈阳性，证明分离培养的细胞为软骨细胞。MTS实验结果发现膝痛消方提取物处理软骨细胞后检测到的OD值随浓度增加而增加且显著高于对照组。此外，IL-1β处理诱导软骨细胞凋亡，而膝痛消方提取物能够有效地抑制IL-1β引起的细胞凋亡。结论：膝痛消方能促进软骨细胞的增殖，抑制IL-1β诱导的软骨细胞的凋亡。以上发现为膝痛消方临床上治疗膝关节骨性关节炎提供药理作用依据。Objective:To investigate the effects of the Xitongxiao prescription on the cell proliferation and the inhibition of IL-1β-induced apoptosis of chondrocytes.Methods:To identify chondrocytes,the articular chondrocytes were cultured in vitro;staining with alcian blue was used to detect the protein polysaccharide,and staining with collagen immunofluorescence was used to examine the expression of type II collagen in chondrocytes.The proliferation of chondrocytes was determined by MTS.Apoptosis of chondrocytes was induced by IL-1β,and the effect of apoptosis was tested by Annexin V/PI double staining.Results:The morphology of the cells obtained from the knee joint was polygonal or fusiform.The tested results of Alcian blue and collagen immunofluorescence staining were positive.These together certified that the isolated cells were knee chondrocytes.MTS assay results showed that the detected OD values were dosedependently increased and they were significantly higher than those of the control group.In addition,the apoptosis of chondrocytes was induced by IL-1βtreatment.While the extract of the Xitongxiao prescription could effectively inhibit the IL-1β-induced apoptosis of chondrocytes.Conclusion:The Xitongxiao prescription could promote the proliferation of chondrocytes and inhibit the apoptosis of chondrocytes induced by IL-1β.The above findings may provide one of pharmacological mechanisms of the Xitongxiao for clinical treatment of knee osteoarthritis.厦门市科技计划项目(3502z20144031);福建省自然科学基金计划资助项目(2015J01065)

Hemocyte-lineage marker proteins in a crustacean, the freshwater crayfish, Pacifastacus leniusculus

To identify proteins associated with development of different hemocyte types in the freshwater crayfish Pacifastacus leniusculus, 2-DE followed by MS analysis was carried out with hematopoietic tissue (Hpt) cells, semigranular cells (SGC) and granular cells (GC). Within the hemocyte lineages one two-domain Kazal proteinase inhibitor (KPI) was found to be specific,for SGC, while a superoxide dismutase (SOD) was specific for GC at protein as well as at mRNA level. The proliferation cell nuclear antigen (PCNA) was detected at the mRNA level in Hpt cells only. We also provide evidence that SGC and GC most likely differentiate to maturation as separate lineages. We found that after laminarin or lipopolysaccharide (LPS) injection into crayfish, the transcript levels of PCNA and SOD increased in the Hpt cells, whereas the KPI transcript never was present in Hpt regardless of any challenge. RNA interference of PCNA in the Hpt cells led to that most of the cells did not spread or attach to the tissue culture dish. These results suggest that PCNA, KPI and SOD can be used as markers for Hpt cells, SGC and GC, respectively, and in conjunction with these results, a model is proposed how the Hpt responds to a microbial challenge by proliferation and release of Hpt cells

Effect of pretreatment temperature on volatile fatty acids production by activated sludge

考察了预处理温度对剩余污泥水解酸化生产挥发性脂肪酸(VFAs)的影响。分别在70、80、90、100℃下处理剩余活性污泥2 h,考察其水解酸化情况。结果表明:热处理可提高溶液中的SCOD及碳水化合物含量,提高效果随温度升高而增大;预处理温度对污泥产酸的促进效果为:90℃>80℃>100℃>70℃。经加90℃处理的活性污泥在缺氧条件下水解酸化30 h后,VFAs的产酸量达到最大,为946 mg/L。VFAs包括乙酸、丙酸、丁酸和戊酸,其中乙酸和丙酸为主要产物,热预处理污泥可提高VFAs中乙酸的含量。The effect of thermal pretreatment temperature on solubilization an acidification of waste sludge(WAS) was investigated.Pretreatment temperatures of WAS ranged from 70℃ to 100℃ and holding time was 2 h.The results showed that COD solubilization was found to increase with treatment temperature.Whereas,total volatile fatty acid(TVFAs)concentration was found to be in the following order:90℃>80℃>100℃>70℃.The optimal pretreatment temperature was 90℃.The VFAs concentration of WAS pretreated under 90℃ was 946 mg/L after 30 h of fermentation.The VFAs consisted of acetic,propionic,butyric and valeric acids,but acetic and propionic acids were the two main products.The proportion of acetic acid was increased by thermal pretreatment.福建省科技重点项目(2008Y0060);; 厦门市科技计划项目(3502Z20073005);; 新加坡厦大校友会研发与学术交流基金项

Isolation, gene cloning and expression profile of a pathogen recognition protein: A serine proteinase homolog (Sp-SPH) involved in the antibacterial response in the crab Scylla paramamosain

To identify the frontline defense molecules against microbial infection in the crab Scylla paramamosain, a live crab pathogenic microbe. Vibrio parahaemolyticus, was recruited as an affinity matrix to isolate innate immune factors from crab hemocytes lysate. Interestingly, a serine proteinase homolog (Sp-SPH) was obtained together with an antimicrobial peptide-antilipopolysaccharide factor (Sp-ALF). We then determined the full-length cDNA sequence of Sp-SPH, which contained 1298 bp with an open reading frame of 1107 bp encoding 369 amino acid residues. Multiple alignment analysis showed that the deduced amino acid sequences of Sp-SPH shared overall identity (83.8%) with those of SPH-containing proteins from other crab species. Tissue distribution analysis indicated that the Sp-SPH transcripts were present in various tissues including eye stalk, subcuticular epidermis, gill, hemocyte, stomach, thorax ganglion, brain and muscle of S. paramamosain. The Sp-SPH was highly expressed in selected different development stages including embryo (I, II, III and V), zoea (I), megalopa, and juvenile. Importantly, the prophenoloxidase was also present in the embryos, zoea, juvenile and adult crabs, but relatively lower in megalopa compared to those of other stages. Furthermore. the Sp-SPH mRNA expression showed a statistically significant increase (P < 0.05) in both hemocyte and subcuticular epidermis at 24 h, and in gill at 96 h after challenge of V. parahaemolyticus determined by quantitative real-time PCR. Taken together, the live-bacterial-binding activity and the acute-phase response against bacterial infection of Sp-SPH suggested that it might function as an innate immune recognition molecule and play a key role in host defense against microbe invasion in the crab S. paramamosain. (C) 2010 Elsevier Ltd. All rights reserved.Natural Science Foundation of Fujian Province China (NSFC-FPC) [2009J05084]; Minjiang Scholars Program [2009

Antioxidant enzymes from the crab Scylla paramamosain: Gene cloning and gene/protein expression profiles against LPS challenge

Recent studies revealed that antioxidant enzymes play important roles in antioxidant responses caused by metabolic process or pathogen invasion. Catalase is one of these key enzymes which has been characterized and highly conserved from invertebrates to vertebrates. In the present study, a full-length cDNA sequence of catalase was cloned from the hemocyte suppression subtractive hybridization library of the crab Scylla paramamosain. The Sp-catalase (Sp-CAT) cDNA sequence contained 2551 bp with an open reading frame of 1551 bp encoding 517 amino acid residues. The conserved catalytic active residues His-71, Asn-144 and Tyr-354 were predicted in the amino acid sequence of Sp-CAT. The deduced Sp-CAT protein had a calculated molecular mass of 59 kDa with an estimated isoelectric point of 6.4. Multiple alignment analysis revealed that the deduced amino acid sequence of Sp-CAT shared high identity (75.4%) with those of other species. The Sp-CAT mRNA transcripts were demonstrated in multiple tissues of normal S. paramamosain. After LPS challenge, the expression level of Sp-CAT gene was increased significantly in hemocyte at 3 and 6 h, and in hepatopancreas at 6 h, respectively, determined by quantitative real-time PCR. Furthermore, the activities of CAT and SOD were also measured in different tissues and serum after LPS challenge. The CAT activity was significantly increased at 3, 6, 24 and 48 h in hemocyte lysate, at 3 h in serum, and at 24 and 48 h in hepatopancreas after LPS challenge. In addition, the SOD activity was significantly induced at 3 and 6 h in hemocyte lysate, 3 and 12 h in serum, 12 and 48 h in hepatopancreas post LPS stimulation, indicating a tissue and time-dependent antioxidant response in the crab. Taken together, these data demonstrated that a strong antioxidant response occurred in the LPS-challenged crab, which might be involved in the protection of host against microbial infections. (C) 2010 Elsevier Ltd. All rights reserved.National Natural Science Foundation of China (NSFC) [40676083]; National High Technology Research and Development Program of China [2007AA091406]; Minjiang Scholars Program [2009]; Natural Science Foundation of Fujian Province China (NSFC-FPC) [2009J05084

“建筑适应性”主题沙龙

"地域建筑"是当下建筑学界共同尊重的准则,但这一术语的过度使用,也使得其内涵在当代建筑创作中逐渐模糊。本期沙龙主题"建筑适应性",是对"地域建造"这一命题的具体化。"适度"的"适应"是当下应当倡导的地域建筑设计创作原则。"建筑适应性"中的"适应",强调对历史文脉与所处环境的积极回应,研究建筑与环境之间的复杂应答关联;除此,"建筑适应性"还强调对经济、技术、材料的"适度

Identification of genes differentially expressed in hemocytes of Scylla paramamosain in response to lipopolysaccharide

Although the crab Scylla paramamosain has been cultured in China for a long time, little knowledge is available on how crabs respond to infection by bacteria. A forward suppression subtractive hybridization (SSH) cDNA library was constructed from their hemocytes and the up-regulated genes were identified in order to isolate differentially expressed genes in S. paramamosain in response to bacterial lipopolysaccharide (LPS). A total of 721 clones on the middle scale in the SSH library were sequenced. Among these genes, 271 potentially functional genes were recognized based on the BLAST searches in NCBI and were categorized into seven groups in association with different biological processes using AmiGO against the Gene Ontology database. Of the 271 genes, 269 translatable DNA sequences were predicted to be proteins, and the putative amino acid sequences were searched for conserved domains and proteins using the CD-Search service and BLASTp. Among 271 genes, 179 (66.1%) were annotated to be involved in different biological processes, while 92 genes (33.9%) were classified as an unknown-function gene group. It was noted that only 18 of the 271 genes (6.6%) had previously been reported in other crustaceans and most of the screened genes showed less similarity to known sequences based on BLASTn results, suggesting that 253 genes were found for the first time in S. paramamosain. Furthermore, two upregulated genes screened from the SSH library were selected for full-length cDNA sequence cloning and in vivo expression study, including Sp-superoxide dismutase (Sp-Cu-ZnSOD) gene and Sp-serpin gene. The differential expression pattern of the two genes during the time course of LPS challenge was analyzed using real-time PCR. We found that both genes were significantly expressed in LPS-challenged crabs in comparison with control. Taken together, the study primarily provides the data of the upregulated genes associated with different biological processes in S. paramamosain in response to LPS, by which the interesting genes or proteins potentially involved in the innate immune defense of S. paramamosain will be investigated in future. (C) 2009 Elsevier Ltd. All rights reserved.National Natural Science Foundation of China (NSFC) [40676083]; National High Technology Research and Development Program of China [2007AA091406]; Program for Minjiang Scholars of Fujian Provinc

1978～2008年中国湿地类型变化

分别基于美国陆地卫星(Landsat MSS/TM/ETM+)和中巴资源卫星(CBERS-02B)影像数据,以人工目视解译为主,完成了中国1978～2008年4期(基准年分别为1978,1990,2000和2008年)湿地遥感制图,并进行了大量的室内外验证.在此基础上,对我国湿地现状及近30年来湿地变化进行了初步分析,得到以下主要结论:(ⅰ)截止2008年,中国湿地面积约为324097km2,其中以内陆沼泽(35%)和湖泊湿地(26%)为主.(ⅱ)1978～2008年,中国湿地面积减少了约33%,而人工湿地增加了约122%.过去30年里湿地减少的速度大幅降低,由最初5523km2/a(1978～1990年)降为831km2/a(2000～2008年).(ⅲ)减少的自然湿地(包括滨海湿地和内陆湿地),其类型变化由湿地向非湿地转化的比例逐渐降低.初期(1978～1990年)几乎全部(98%)转换为非湿地;在1990～2000年间减少的自然湿地约有86%转化为非湿地,而在2000～2008年,这一比例下降为77%.(ⅳ)气候变化和农业活动是中国湿地变化的主要驱动因素,湿地变化在中国分为三大不同特征区域,即西部三省/自治区(西藏、新疆和青海)、北部两省/自治区(黑龙江和内蒙古)和其他省市区.其中西部区域尤其是青藏高原,湿地变化的驱动因子以气候增温为主;新疆湿地由于气候增温和农业活动共同作用造成变化不大.北部省/自治区的湿地变化则主要由农业活动引起;而其他省市区的湿地变化几乎完全受控于人类的农业经济活动