Deep sequencing of non-ribosomal peptide synthetases and polyketide synthases from the microbiomes of Australian marine sponges

The biosynthesis of non-ribosomal peptide and polyketide natural products is facilitated by multimodular enzymes that contain domains responsible for the sequential condensation of amino and carboxylic subunits. These conserved domains provide molecular targets for the discovery of natural products from microbial metagenomes. This study demonstrates the application of tag-encoded FLX amplicon pyrosequencing (TEFAP) targeting non-ribosomal peptide synthetase (NRPS) and polyketide synthase (PKS) genes as a method for determining the identity and diversity of natural product biosynthesis genes. To validate this approach, we assessed the diversity of NRPS and PKS genes within the microbiomes of six Australian marine sponge species using both TEFAP and metagenomic whole-genome shotgun sequencing approaches. The TEFAP approach identified 100 novel ketosynthase (KS) domain sequences and 400 novel condensation domain sequences within the microbiomes of the six sponges. The diversity of KS domains within the microbiome of a single sponge species Scopalina sp. exceeded that of any previously surveyed marine sponge. Furthermore, this study represented the first to target the condensation domain from NRPS biosynthesis and resulted in the identification of a novel condensation domain lineage. This study highlights the untapped potential of Australian marine sponges for the isolation of novel bioactive natural products. Furthermore, this study demonstrates that TEFAP approaches can be applied to functional genes, involved in natural product biosynthesis, as a tool to aid natural product discovery. It is envisaged that this approach will be used across multiple environments, offering an insight into the biological processes that influence the production of secondary metabolites.     

An Evaluation Model of Supply Chain Performances Using 5DBSC and LMBP Neural Network Algorithm

A high efficient Supply Chain (SC) would bring great benefits to an enterprise such as integrated resources, reduced logistics costs, improved logistics efficiency and high quality of overall level of services. So it is important to research various methods, performance indicator systems and technology for evaluating, monitoring, predicting and optimizing the performance of a SC. In this paper, the existing performance indicator systems and methods are discussed and evaluated. Various nature-inspired algorithms are reviewed and their applications for SC Performance Evaluation (PE) are discussed. Then, a model is proposed and developed using 5 Dimensional Balanced Scorecard (5DBSC) and LMBP (Levenberg-Marquardt Back Propagation) neural network for SC PE. A program is written using Matlab tool box to implement the model based on the practical values of the 14 indicators of 5DBSC of a given previous period. This model can be used to evaluate, predict and optimize the performance of a SC. The analysis results of a case study of a company show that the proposed model is valid, reliable and effective. The convergence speed is faster than that in the previous work.     

对比不同类型干细胞对于缺血性脑卒中治疗的研究进展

成人中枢神经系统存在着一定量的神经干细胞,其具有两大关键能力;自我更新和多向分化潜能。缺血性脑卒中是一种由于由脑血流的缺失或减少引起的脑动脉闭塞,进而导致脑组织梗死的脑血管疾病。虽然对于脑损伤的药物治疗已经取得了一定的成果,但目前以干细胞为基础的治疗方法仍成为了研究热点。无论是内源性神经干细胞还是外源性神经干细胞移植均可在脑损伤后向远端损伤区迁移并分化成新的神经细胞,从而在中枢神经系统疾病尤其是脑梗死后进行组织修复和功能恢复。因此在这篇综述中,我们主要探讨不同类型的干细胞对脑梗死介导的脑损伤的应用潜能,对比不同类型干细胞对缺血性脑卒中的治疗优缺点。     

恒磁场增强心肌梗死大鼠骨髓间充质干细胞移植后心脏功能改善

目的：研究恒磁场对心肌梗死大鼠骨髓间充质干细胞（bone marrow-derived mesenchymal stem cells,BMSCs）移植后心脏功能的影响。方法：取180g,9-12周龄雄性SD大鼠骨髓,以密度梯度离心分离出单个核细胞（MNCs）,于体外培养并传代培养出骨髓间充质干细胞（MSCs）。制作大鼠心肌梗死模型,将1.5×106BMSCs注射入梗死梗死区周围,分为磁场照射＋BMSCs植入组、BMSCs植入组及空白对照组。4周后处死动物,每组5只大鼠。磁场照射组用0.4 T恒磁场置于心前区30 min,每日1次,共7天。用颈动脉插管法测定心脏功能,Masson三色染色测定梗死面积,VWF VIII染色计算血管密度。结果：与对照组相比BMSCs组以及磁场组均可以显著提高左心室收缩压（LVSP）,dp/dtmax以及-dp/dtmax,减少LVEDP（P〈0.05）。但是,磁场组与BMSCs组相比LVSP,左心室内压最大上升速率（dp/dtmax）以及左心室内压最大下降速率（-dp/dtmax）增高,左心室舒张末压（LVEDP）减少（P〈0.05）。与对照组相比BMSCs组以及磁场组均可以显著提高减少心梗面积（P〈0.05）。磁场组与BMSCs组相比心梗面积减少（P〈0.05）。与对照组相比BMSCs组以及磁场组均可以显著提高增加血管密度（P〈0.05）。磁场组与BMSCs组相比血管密度增加（P〈0.05）。结论：恒磁场具有加强移植BMSCs改善心脏功能的作用。     

咪喹莫特对慢性哮喘模型小鼠肺组织中基质金属蛋白酶-9-表达的影响

目的：观察Toll样受体7配体咪喹莫特对慢性哮喘小鼠模型气道重塑及肺组织中基质金属蛋白酶MMP-9表达的影响。方法：36只BALB／c小鼠按随机原则分成正常对照组、哮喘模型组、咪喹莫特组,每组12只。通过卵蛋白致敏,气道激发8周,末次激发24h后,检测各组小鼠气道反应性,HE染色观察气道炎症变化;Masson三色染色观察气道纤维化的改变;real-timePCR和western—blot分别检测肺组织中MMP-9的mRNA和蛋白表达。结果：慢性哮喘组小鼠气道炎症、气道高反应性和气道重塑较正常对照小鼠明显加重,而咪喹莫特组小鼠模型的气道炎症和气道反应性及气道重塑均较哮喘模型组小鼠减少或降低。慢性哮喘组小鼠肺组织MMP-9的mRNA和蛋白水平均较正常对照小鼠明显增加（P〈0．05）,而咪喹莫特治疗可显著降低哮喘小鼠肺组织MMP-9的mRNA和蛋白水平（P〈0．05）。结论：咪喹莫特能够显著抑制慢性哮喘小鼠模型的气道炎症、降低气道高反应性并减轻气道重塑,这可能与其抑制MMP-9的表达有关。     

异丙酚和氯胺酮对大鼠离体缺血再灌注损伤心肌脂质过氧化的影响

目的：比较异丙酚和氯胺酮对大鼠离体缺血再灌注损伤心肌脂质过氧化的影响。方法：成年Wistar大鼠18只,雌雄不拘。体重240-300g,随机分为3组（T1=6）：心肌缺血再灌注损伤组（I／R组）,异丙酚组（P组）,氯胺酮组（K组）。采用Langendorff灌装置建立离体心脏缺血再灌注模型,将心脏连接至Langendorff逆灌装置,3组均以K-H液平衡灌注10min后,再分别以K．H液、含30μmol/L。异丙酚的K-H液、含10μmol-L-1氯胺酮的K-H液灌注10min,然后全心停灌25min,再分别以停灌前相同的灌注液恢复灌注30min。留取冠脉流出液测定总LDH活性;灌注末取左室心肌组织置于2．5％的戊二醛固定,观察心肌的超微结构;心尖部心肌组织留待检测8-异前列腺素和SOD活性。结果：与I／R组比较,P组8-异前列腺素含量降低,SOD活性升高,LDH活性降低（P〈0．05）;K组8-异前列腺素含量,SOD及LDH活性均无统计学意义（P〉0．05）;与P组比较,K组8-异前列腺素含量升高,SOD及LDH活性降低（P〈0．05）;P组心肌超微结构损伤较m组和K组也明显改善。结论：异丙酚可显著减轻心肌缺血再灌注损伤大鼠的脂质过氧化和心肌缺血再灌注损伤,而氯胺酮没有抗心肌缺血再灌注损伤心肌脂质过氧化的作用。     

Secretory/releasing proteome-based identification of plasma biomarkers in HBV-associated hepatocellular carcinoma

For successful therapy, hepatocellular carcinoma (HCC) must be detected at an early stage. Herein, we used a proteomic approach to analyze the secretory/releasing proteome of HCC tissues to identify plasma biomarkers. Serum-free conditioned media (CM) were collected from primary cultures of cancerous tissues and surrounding noncancerous tissues. Proteomic analysis of the CM proteins permitted the identification of 1365 proteins. The enriched molecular functions and biological processes of the CM proteins, such as hydrolase activity and catabolic processes, were consistent with the liver being the most important metabolic organ. Moreover, 19% of the proteins were characterized as extracellular or membrane-bound. For validation, secretory proteins involved in transforming growth factor-β signaling pathways were validated in plasma samples. Alphafetoprotein (AFP), metalloproteinase (MMP)1, osteopontin (OPN), and pregnancy-specific beta-1-glycoprotein (PSG)9 were significantly increased in HCC patients. The overall performance of MMP1 and OPN in the diagnosis of HCC remained greater than that of AFP. In addition, this study represents the first report of MMP1 as a biomarker with a higher sensitivity and specificity than AFP. Thus, this study provides a valuable resource of the HCC secretome with the potential to investigate serological biomarkers. MMP1 and OPN could be used as novel biomarkers for the early detection of HCC and to improve the sensitivity of biomarkers compared with AFP.     

Knockdown of OsHox33, a member of the class III homeodomain-leucine zipper gene family, accelerates leaf senescence in rice

The class III homeodomain-leucine zipper (HD-Zip III) gene family plays important roles in plant growth and development, including regulation of apical embryo patterning, embryonic shoot meristem formation, leaf polarity, vascular development, and meristem function, with a particularly crucial function in leaf development. Although HD-Zip III members are highly conserved in land plants, previous studies, such as genetic analyses based on multiple mutants in Arabidopsis and other plants, suggest that various HD-Zip III family genes have evolved with distinct functions and pleiotropic effects on plant growth and development. In this study, we analyzed a HD-Zip III member, OsHox33, and demonstrated that it plays an important role in age-dependent leaf senescence in rice. We constructed two specific RNAi vectors derived from the 5′-end region and 3′-UTR of OsHox33 to knockdown its expression. Transgenic plants harboring either RNAi construct displayed similar phenotypes of precocious leaf senescence symptoms, suggesting that knockdown of OsHox33 accelerates leaf senescence in rice. pOsHox33::GUS fusion expression and RT-PCR revealed that OsHox33 is highly expressed in young organs, especially in young meristems such as shoot apical meristems, intercalary meristems, and young callus. In addition, real-time PCR indicated that OsHox33 was more highly expressed in young leaves than in old leaves. To further investigate OsHox33 function, we analyzed chloroplast ultrastructure in different-aged leaves of RNAi plants, and found that OsHox33 knockdown accelerated chloroplast degradation, which is consistent with RNAi phenotypes. Finally, real-time PCR studies showed that OsHox33 can regulate the expression of GS1 and GS2, two senescence-associated genes. Taken together, the work presented here provides new insights into the function of HD-Zip III members in plants.     

Insecticidal activity of the chitinase from the Spodoptera litura nucleopolyhedrovirus

Baculovirus chitinase gene (chiA) is a late gene essential for liquefying the host insect at a late stage of infection for its hydrolyzing chitin function. In a previous report, baculovirus ChiA has been shown to offer many interesting new opportunities for pest control. Recently, a putative chiA gene was identified in the Korean isolate of the Spodoptera litura nucleopolyhedorvirus (SpliMNPV‐K1) genome. The open reading frame (ORF) contains 1692 nucelotides and encodes a protein of 563 amino acids with a predicted molecular weight of about 62.6 kDa. To study the insecticidal activity of ChiA from SpliMNPV‐K1, we constructed a recombinant AcMNPV, Ap‐SlChiA, which is designed to express the ChiA under the control of a polyhedrin promoter. Western blot analysis indicated that ChiA was successfully expressed by this recombinant virus. Chitinase assay revealed that the chitobiosidase and endochitinase activity of the recombinant virus was 2.5‐ and 3.9‐flods higher than those of wild‐type AcMNPV, respectively. In addition, the recombinant virus showed higher evident insecticidal activity against 3rd instar larvae of Spodotera exigua than that of the AcMNPV. These results suggest that the chiA gene from SpliMNPV‐K1 could be successfully applied to improve pathogenicity of baculoviruses.