脓毒症患者血清HMGB1、IGF-1水平变化及与T淋巴细胞亚群、预后的关系研究

目的:探讨脓毒症患者血清高迁移率族蛋白1(HMGB1)、胰岛素样生长因子-1(IGF-1)水平变化及与T淋巴细胞亚群、预后的关系。方法:选取2016年2月～2018年12月期间我院收治的脓毒症患者139例,根据Sepsis 3.0定义,将脓毒症患者分成一般脓毒症组(n=73)及脓毒症休克组(n=66),根据患者进入重症监护室28d后的转归资料,将其分为存活组和死亡组。比较不同预后、不同病情严重程度的脓毒症患者血清IGF-1、HMGB1水平、急性病生理与慢性健康评价系统Ⅱ(APACHEⅡ)评分以及T淋巴细胞亚群;采用Pearson相关分析血清HMGB1、IGF-1水平与T淋巴细胞亚群、APACHEⅡ评分的关系。结果:一般脓毒症组CD3~+、CD4~+、CD4~+/CD8~+高于脓毒症休克组,CD8~+低于脓毒症休克组(P0.05)。脓毒症休克组血清HMGB1水平、APACHEⅡ评分均高于一般脓毒症组,血清IGF-1水平则低于一般脓毒症组(P0.05)。存活组CD8~+低于死亡组,CD3~+、CD4~+、CD4~+/CD8~+高于死亡组(P0.05)。存活组血清HMGB1水平、APACHEⅡ评分低于死亡组,血清IGF-1水平高于死亡组(P0.05)。Pearson相关分析显示,脓毒症患者血清HMGB1水平与CD8~+、APACHEⅡ评分呈正相关,与CD3~+、CD4~+、CD4~+/CD8~+呈负相关(P0.05);血清IGF-1水平与CD8~+、APACHEⅡ评分呈负相关,与CD3~+、CD4~+、CD4~+/CD8~+呈正相关(P0.05)。结论:脓毒症血清HMGB1、T淋巴细胞亚群、IGF-1均存在异常变化,可用于评估脓毒症患者的病情和预后。     

Characterization and expression of cry4Cb1 and cry30Ga1 from Bacillus thuringiensis strain HS18-1

We characterized a novel Bacillus thuringiensis isolate native to China (HS18-1) that shows a spherical crystal harboring two major proteins of about 70 and 130 kDa, and contains three novel cry genes (cry4Cb1, cry30Ga1, cry54-type). Furthermore, the cry4Cb1 and cry30Ga1 genes were expressed in Escherichia coli BL21 (DE3): pLysS. Insecticidal activity tests showed that the cry4Cb1 protein exhibited larvicidal activity against Aedes aegypti (Diptera) and the cry30Ga1 protein was toxic to both A. aegypti and P. xylostella (Lepidoptera).     

The extracellular release of Schistosoma mansoni HMGB1 nuclear protein is mediated by acetylation

Schistosoma mansoni HMGB1 (SmHMGB1) was revealed to be a substrate for the parasite histone acetyltransferases SmGCN5 and SmCBP1. We found that full-length SmHMGB1, as well as its HMG-box B (but not HMG-box A) were acetylated in vitro by SmGCN5 and SmCBP1. However, SmCBP1 was able to acetylate both substrates more efficiently than SmGCN5. Interestingly, the removal of the C-terminal acidic tail of SmHMGB1 (SmHMGB1ΔC) resulted in increased acetylation of the protein. We showed by mammalian cell transfection assays that SmHMGB1 and SmHMGB1ΔC were transported from the nucleus to the cytoplasm after sodium butyrate (NaB) treatment. Importantly, after NaB treatment, SmHMGB1 was also present outside the cell. Together, our data suggest that acetylation of SmHMGB1 plays a role in cellular trafficking, culminating with its secretion to the extracellular milieu. The possible role of SmHMGB1 acetylation in the pathogenesis of schistosomiasis is discussed.     

A new species of ferret-badger, Genus Melogale, from Vietnam

Ferret-badgers, genus Melogale, are distributed in the Indochinese region, Java, Bali and NE-Borneo. There are currently four species described each having very similar phenotypes. In March 2005, a living ferret-badger of a different phenotype was confiscated by rangers from Cuc Phuong National Park, Vietnam. This individual died and the carcass was not preserved. In January 2006, a newly deceased individual of the same phenotype was found at the Endangered Primate Rescue Center, Cuc Phuong National Park. Due to several different characteristics these individuals vary greatly from the current species. Thus, we describe an additional species, M. cucphuongensis sp. nov. from northern Vietnam, which occurs sympatrically with M. moschata and M. personata, but differs from both species clearly in skull morphology and other features.Based on a 423 bp-long fragment of the mitochondrial cytochrome b gene, M. cucphuongensis sp. nov. is a member of the genus Melogale and represents a sister lineage to a clade consisting of M. personata and M. moschata.     

A recombinant immunosuppressive protein from Pimpla hypochondriaca (rVPr1) increases the susceptibility of Lacanobia oleracea and Mamestra brassicae larvae to Bacillus thuringiensis

The precise mechanisms underlying Bacillus thuringiensis-mediated killing of pest insects are not clear. In some cases, death may be due to septicaemia caused by Bt and/or gut bacteria gaining access to the insect haemocoel. Since insects protect themselves from microbes using an array of cellular and humoral immune defences, we aimed to determine if a recombinant immunosuppressive wasp venom protein (rVPr1) could increase the susceptibility of two pest Lepidoptera (Lacanobia oleracea and Mamestra brassicae) to Bt. Bio-assays indicated that injection of 6 μl of rVPr1 into the haemocoel of both larvae caused similar levels of mortality (less than 38%). On the other hand, the LD_30-40 of Bt for M. brassicae larvae was approximately 20 times higher than that for L. oleracea larvae. Furthermore, in bio-assays where larvae were injected with rVPr1, then fed Bt, a significant reduction in survival of larvae for both species occurred compared to each treatment on its own (P < 0.001); and for L. oleracea larvae, this effect was more than additive. The results are discussed within the context of insect immunity and protection against Bt.     

miR-142通过靶向HMGB1对宫颈癌细胞生物学行为影响的实验研究

摘要 目的：通过实验探究miR-142靶向高迁移率族蛋白1（high-mobility group box 1 protein,HMGB1）对宫颈癌（cervical cancer,CC）细胞生物学行为的影响及其潜在的作用机制。方法：采用实时荧光定量PCR（RT-PCR）和蛋白质免疫印迹法（Western Blot）检测CC组织和正常组织中miR-142和HMGB1 mRNA及蛋白表达水平,采用荧光素酶报告实验分析miR-142与HMGB1的靶向关系,CCK-8法检测CC细胞生存能力,克隆形成实验检测CC细胞增殖能力,划痕修复实验检测CC细胞迁移能力,基质胶侵袭实验检测CC细胞侵袭能力。结果：CC组miR-142 mRNA和蛋白表达水平显著低于正常组（P<0.05）,HMGB1 mRNA和蛋白表达水平显著高于正常组（P<0.05）,且CC癌组织中miR-142和HMGB1 mRNA和蛋白表达水平均呈显著负相关（r=-0.399,P=0.002;r=-0.429,P=0.001）;miR-142与HMGB1存在靶向关系;CCK-8法实验、克隆形成实验、划痕修复实验和基质胶侵袭实验结果显示,miR-142 mimic组细胞生存、增殖、迁移和侵袭能力显著低于miR-NC组（P<0.05）,miR-142 inhibitor组细胞生存、增殖、迁移和侵袭能力显著高于miR-NC组;Western Blot实验结果显示,HMGB1过表达时miR-142 mimic+plasmid组HMGB1蛋白表达水平显著高于miR-142 mimic+control plasmid组（P<0.05）,显著低于miR-NC+plasmid组（P<0.05）;CCK-8法实验、克隆形成实验、划痕修复实验和基质胶侵袭实验结果显示,HMGB1过表达时miR-142 mimic+plasmid组细胞生存、增殖、迁移和侵袭能力显著高于miR-142 mimic+control plasmid组（P<0.05）,显著低于miR-NC+plasmid组（P<0.05）。结论：miR-142可通过靶向负调控HMGB1表达,进而抑制CC细胞生存、增殖、迁移和侵袭。     

Characterization of resistance to Bacillus thuringiensis toxin Cry1Ac in Plutella xylostella from China

A field population (SZ) of Plutella xylostella, collected from the cabbage field in Shenzhen, Guangdong Province of China in 2002, showed 2.3-fold resistance to Cry1Aa, 110-fold to Cry1Ab, 30-fold to Cry1Ac, 2.1-fold to Cry1F, 5.3-fold to Cry2Aa and 6-fold resistance to Bacillus thuringiensis var. kurstaki (Btk) compared with a susceptible strain (ROTH). The SZBT strain was derived from the SZ population through 20 generations of selection with activated Cry1Ac in the laboratory. While the SZBT strain developed 1200-fold resistance to Cry1Ac after selection, resistance to Cry1Aa, Cry1Ab, Cry1F, and Btk increased to 31-, 1900-,>33- and 17-fold compared with the ROTH strain. However, little or no cross-resistance was detected to Cry1B, Cry1C and Cry2Aa in the SZBT strain. Genetic cross analyses between the SZBT and ROTH strains revealed that Cry1Ac-resistance in the SZBT strain was controlled by a single, autosomal, incompletely recessive gene. Binding studies with ¹²⁵I-labeled Cry1Ac showed that the brush border membrane vesicles (BBMVs) of midguts from the resistant SZBT insects had lost binding to Cry1Ac. Allelic complementation tests demonstrated that the major Bt resistance locus in the SZBT strain was same as that in the Cry1Ac-R strain which has “mode 1” resistance to Bt. An F₁ screen of 120 single-pair families between the SZBT strain and three field populations collected in 2008 was carried out. Based on this approach, the estimated frequencies of Cry1Ac-resistance alleles were 0.156 in the Yuxi population from Yunnan province, and 0.375 and 0.472 respectively in the Guangzhou and Huizhou populations from Guangdong province.     

br regulates the expression of the ecdysone biosynthesis gene npc1

The growth and metamorphosis of insects are regulated by ecdysteroid hormones produced in the ring gland. Ecdysone biosynthesis-related genes are both highly and specifically expressed in the ring gland. However, the intrinsic regulation of ecdysone biosynthesis has received little attention. Here we used the Drosophila npc1 gene to study the mechanism of ring gland-specific gene expression. npc1 is important for sterol trafficking in the ring gland during ecdysone biosynthesis. We have identified a conserved ring gland-specific cis-regulatory element (RSE) in the npc1 promoter using promoter fusion reporter analysis. Furthermore, genetic loss-of-function analysis and in vitro electrophoretic mobility shift assays revealed that the ecdysone early response gene broad complex (br) is a vital factor in the positive regulation of npc1 ring gland expression. Moreover, br also affects the ring gland expression of many other ecdysone biosynthetic genes as well as torso and InR, two key factors in the regulation of ecdysone biosynthesis. These results imply that ecdysone could potentially act through its early response gene br to achieve positive feedback regulation of ecdysone biosynthesis during development.     

The ascl1a and dlx genes have a regulatory role in the development of GABAergic interneurons in the zebrafish diencephalon

During development of the mouse forebrain interneurons, the Dlx genes play a key role in a gene regulatory network (GRN) that leads to the GABAergic phenotype. Here, we have examined the regulatory relationships between the ascl1a, dlx, and gad1b genes in the zebrafish forebrain. Expression of ascl1a overlaps with dlx1a in the telencephalon and diencephalon during early forebrain development. The loss of Ascl1a function results in a loss of dlx expression, and subsequent losses of dlx5a and gad1b expression in the diencephalic prethalamus and hypothalamus. Loss of Dlx1a and Dlx2a function, and, to a lesser extent, of Dlx5a and Dlx6a, impairs gad1b expression in the prethalamus and hypothalamus. We conclude that dlx1a/2a act downstream of ascl1a but upstream of dlx5a/dlx6a and gad1b to activate GABAergic specification. This pathway is conserved in the diencephalon, but has diverged between mammals and teleosts in the telencephalon.     

Metabolic characterization of green pods from Vanilla planifolia accessions grown in La Réunion

Large phenotypic variation has been observed between the cultivated vanillas since a single genetic source of Vanilla planifolia was spread to the Indian Ocean and the Indonesia in the 19th century. In order to differentiate the cultivated vanilla plants, genetic studies have been conducted in the past on the plants grown in various regions such as the French island, La Réunion. However, the genetic difference was not big enough to differentiate diverse accessions of V. planifolia. In this study, metabolomics, in which genetic variation could be amplified, was employed to delve into the variation between the cultivated vanilla plants. To obtain a broad view of the metabolome, nuclear magnetic resonance (NMR) spectroscopy was applied to the analysis of V. planifolia green pods. Principal component analysis (PCA) and partial least square-discriminant analysis (PLS-DA) of the data showed that the accessions could be differentiated according to their glucovanillin and glucosides A and B contents. Furthermore, a correlation between the glucovanillin content and the pod length, number of flower and growth capacity of the accessions has been observed from the multivariate data analysis.     

Isolation and Characterization of OsGSTL1 Promoter from Rice

OsGSTL1 gene was isolated from the rice genomic library. Semi-quantitative RT-PCR analysis demonstrated that the expression of the OsGSTL1 in rice was not induced by chlorsulfuron, ethylene, abscisic acid, salicylic acid, and methyl jasmonate. In order to investigate the cis-elements of OsGSTL1 promoter, the promoter regions with different lengths were fused to the β-glucuronidase (GUS) reporter gene. All constructs were transformed into onion epidermal cells or A. thaliana plants to detect the expression patterns. In onion epidermal cells, the 160 bp fragment and longer ones were functional for directing GUS expression. In transgenic A. thaliana, the 2?155 bp upstream region of OsGSTL1 gene directed the GUS expression only in cotyledon after germination, but not in the root of young seedlings. In the later seedling, the 2?155 bp upstream region of OsGSTL1 gene directed GUS expression in roots, stems, and leaves. However, the GUS gene directed by a 1?224 bp upstream fragment is expressed in all the checked tissues. These results suggest that the spatiotemporal expression response elements of OsGSTL1 existed in the 5′-upstream region between −2?155 and −1?224 bp.     

The anamorphic genus Monotosporella (Ascomycota) from Eocene amber and from modern Agathis resin

The anamorphic fungal genus Monotosporella (Ascomycota, Sordariomycetes) has been reco-vered from a piece of Early Eocene Indian amber, as well as from the surface of extant resin flows in New Caledonia. The fossil fungus was obtained from the Tarkeshwar Lignite Mine of Gujarat State, western India, and was part of the biota of an early tropical angiosperm rainforest. The amber inclusion represents the second fossil record of Sordariomycetes, as well as the first fossil of its particular order (either Savoryellales or Chaetosphaeriales). The fossil fungus is distinguished from extant representatives by possessing both short conidiophores and small two-septate pyriform conidia, and is described as Monotosporella doerfeltii sp. nov. Inside the amber, the anamorph is attached to its substrate, which is likely the degraded thallus of a cladoniform lichen. The extant New Caledonian species is assigned to Monotosporella setosa. It was found growing on semi-solidified resin flows of Agathis ovata (Araucariaceae), and is the first record of Monotosporella from modern resin substrates.     

An Eimeria vaccine candidate based on Eimeria tenella immune mapped protein 1 and the TLR-5 agonist Salmonella typhimurium FliC flagellin

Immune mapped protein-1 (IMP1) is a new protective protein in apicomplexan parasites, and exits in Eimeria tenella. But its structure and immunogenicity in E. tenella are still unknown. In this study, IMPI in E. tenella was predicted to be a membrane protein. To evaluate immunogenicity of IMPI in E. tenella, a chimeric subunit vaccine consisting of E. tenella IMP1 (EtIMP1) and a molecular adjuvant (a truncated flagellin, FliC) was constructed and over-expressed in Escherichia coli and its efficacy against E. tenella infection was evaluated. Three-week-old AA broiler chickens were vaccinated with the recombinant EtIMP1-truncated FliC without adjuvant or EtIMP1 with Freund’s Complete Adjuvant. Immunization of chickens with the recombinant EtIMP1-truncated FliC fusion protein resulted in stronger cellular immune responses than immunization with only recombinant EtIMP1 with adjuvant. The clinical effect of the EtIMP1-truncated FliC without adjuvant was also greater than that of the EtIMP1 with adjuvant, which was evidenced by the differences between the two groups in body weight gain, oocyst output and caecal lesions of E. tenella-challenged chickens. The results suggested that the EtIMP1-flagellin fusion protein can be used as an effective immunogen in the development of subunit vaccines against Eimeria infection. This is the first demonstration of antigen-specific protective immunity against avian coccidiosis using a recombinant flagellin as an apicomplexan parasite vaccine adjuvant in chickens.