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Find video protocols related to scientific articles indexed in Pubmed.
Bone Morphogenetic Protein (BMP) signaling in development and human diseases.
Genes Dis
PUBLISHED: 11-18-2014
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Bone Morphogenetic Proteins (BMPs) are a group of signaling molecules that belongs to the Transforming Growth Factor-? (TGF-?) superfamily of proteins. Initially discovered for their ability to induce bone formation, BMPs are now known to play crucial roles in all organ systems. BMPs are important in embryogenesis and development, and also in maintenance of adult tissue homeostasis. Mouse knockout models of various components of the BMP signaling pathway result in embryonic lethality or marked defects, highlighting the essential functions of BMPs. In this review, we first outline the basic aspects of BMP signaling and then focus on genetically manipulated mouse knockout models that have helped elucidate the role of BMPs in development. A significant portion of this review is devoted to the prominent human pathologies associated with dysregulated BMP signaling.
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A pedigree analysis of two homozygous mutant Gitelman Syndrome cases.
Endocr. J.
PUBLISHED: 10-03-2014
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Gitelman Syndrome (GS) is a salt-wasting tubulointerstitial disease of autosomal recessive inheritance (OMIM613395) caused by genic mutation of SLC12A3, which codes thiazide-sensitive Na-Cl cotransporter (NCCT) gene. The gene mutation of the majority of GS patients is compound heterozygous. This study analyzes two cases of GS gene mutation and the clinical phenotype. Twenty patients of two GS pedigrees underwent direct sequence alignment of 26 exons of SLC12A3 to spot and locate mutant site. Proband A of Pedigree I had three mutant sites: Arg928Cys, a homozygote, missense mutation, and two homozygous silent mutations, Ala122Ala and Thr465Thr, and 8 members of Pedigree I carried Arg928Cy heterozygous mutation. Proband B of Pedigree II had a homozygote, Ser710X, and a termination codon was spotted, which would inevitably be translated into abridged and defective protein, and 7 members had Ser710X heterozygous mutation. The heterozygous mutation carriers of the two pedigrees often have stimulus-controlled hypokalemia after strenuous exercise. The parents of Proband A are cousins, a case of intermarriage. Both probands show hypokalemia, hypochloraemia, hypocalcinuria, hyperreninemia, and hyperaldosteronemia; Proband A has normal serum magnesium and increased urinary sodium excretion, while Proband B has hypomagnesemia and increased urinary magnesium ion excretion. Both probands have normal or lower blood pressure, weakness and numbness of lower extremities, muscular soreness, and occasional palpitations and chest discomfort. Proband A wearies easily and Proband B has occasional joint numbness and pain. These two homozygous mutations are responsible for the morbidity of two GS families and they show heterogenicity of clinical phenotype.
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[Clinical efficacy of unipedicular versus bipedicular percutaneous vertebroplasty for Kummell's disease].
Nan Fang Yi Ke Da Xue Xue Bao
PUBLISHED: 09-30-2014
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To compare the clinical efficacy and safety of percutaneous vertebroplasty by unipedicular and bipedicular approach for treatment of Kummell's disease.
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Psychrophilins E-H and Versicotide C, Cyclic Peptides from the Marine-Derived Fungus Aspergillus versicolor ZLN-60.
J. Nat. Prod.
PUBLISHED: 09-23-2014
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Four new cyclic peptides, psychrophilins E-H (1-4), possessing a rare amide linkage between the carboxylic acid in anthranilic acid (ATA) and the nitrogen from an indole moiety, along with a new ATA-containing hexapeptide, versicotide C (5), were obtained from the culture of the marine-derived fungus Aspergillus versicolor ZLN-60. The structures, including absolute configurations, were elucidated by a combination of HRESIMS, NMR, X-ray crystallography, TDDFT ECD calculations, and Marfey's method. Versicotide C (5) is the first natural cyclic hexapeptide containing two anthranilic acids. Compounds 1-5 were not cytotoxic, and compound 3 showed potent lipid-lowering effects.
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The complete mitochondrial genome of Ophicephalus argus kimurai (Perciformes: Channidae).
Mitochondrial DNA
PUBLISHED: 09-04-2014
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Abstract The complete sequence of the mitochondrial genome of Ophicephalus argus kimurai was determined in this study. The genome was 16,558?bp in length. As in other vertebrates, it consisted of 22 transfer RNA (tRNA) genes, 13 protein-coding genes, 2 ribosomal RNA (rRNA) genes and 1 noncoding control region. The overall base composition was estimated to be A, 27.2%; T, 24.2%; C, 31.6% and G, 17% with AT bias of 51.4%. The genomic composition, organization, and gene order of O. argus kimurai was similar to that obtained in most vertebrates. These results may provide the basis for the study of genetic structure as well as resource conservation and protection of O. argus kimurai.
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Collagen type-I leads to in vivo matrix mineralization and secondary stabilization of Mg-Zr-Ca alloy implants.
Colloids Surf B Biointerfaces
PUBLISHED: 08-19-2014
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Biodegradable magnesium-zirconia-calcium (Mg-Zr-Ca) alloy implants were coated with Collagen type-I (Coll-I) and assessed for their rate and efficacy of bone mineralization and implant stabilization. The phases, microstructure and mechanical properties of these alloys were analyzed using X-ray diffraction (XRD), optical microscopy and compression test, respectively, and the corrosion behavior was established by their hydrogen production rate in simulated body fluid (SBF). Coll-I extracted from rat tail, and characterized using fourier transform infrared (FT-IR) spectroscopy, was used for dip-coating the Mg-based alloys. The coated alloys were implanted into the femur bones of male New Zealand white rabbits. In vivo bone formation around the implants was quantified by measuring the bone mineral content/density (BMC/BMD) using dual-energy X-ray absorptiometry (DXA). Osseointegration of the implant and new bone mineralization was visualized by histological and immunohistochemical analysis. Upon surface coating with Coll-I, these alloys demonstrated high surface energy showing enhanced performance as an implant material that is suitable for rapid and efficient new bone tissue induction with optimal mineral content and cellular properties. The results demonstrate that Coll-I coated Mg-Zr-Ca alloys have a tendency to form superior trabecular bone structure with better osteoinduction around the implants and higher implant secondary stabilization, through the phenomenon of contact osteogenesis, compared to the control and uncoated ones in shorter periods of implantation. Hence, Coll-I surface coating of Mg-Zr-Ca alloys is a promising method for expediting new bone formation in vivo and enhancing osseointegration in load bearing implant applications.
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Prenylated indole diketopiperazines from the marine-derived fungus Aspergillus versicolor.
J. Org. Chem.
PUBLISHED: 08-11-2014
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Seven new prenylated indole diketopiperazines, versicamides A-G (1-7) and a novel chemical derivative from 7, versicamide H (8), along with three known analogic diketopiperazines (9-11) were obtained from the marine-derived fungus Aspergillus versicolor HDN08-60. Their structures were determined by spectroscopic techniques, including 2D NMR, ECD calculations, and single-crystal X-ray diffraction analysis, together with the assistance of further chemical conversions. The cytotoxicities of 1-8 were tested against the HeLa, HCT-116, HL-60, and K562 cell lines, but only 8 exhibited moderate activity against HL-60 cells, with an IC50 value of 8.7 ?M. Further investigation with target screening showed that 8 exhibited selective PTK inhibitory activities.
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A hop count based heuristic routing protocol for mobile delay tolerant networks.
ScientificWorldJournal
PUBLISHED: 06-23-2014
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Routing in delay tolerant networks (DTNs) is a challenge since it must handle network partitioning, long delays, and dynamic topology. Meanwhile, routing protocols of the traditional mobile ad hoc networks (MANETs) cannot work well due to the failure of its assumption that most network connections are available. In this paper, we propose a hop count based heuristic routing protocol by utilizing the information carried by the peripatetic packets in the network. A heuristic function is defined to help in making the routing decision. We formally define a custom operation for square matrices so as to transform the heuristic value calculation into matrix manipulation. Finally, the performance of our proposed algorithm is evaluated by the simulation results, which show the advantage of such self-adaptive routing protocol in the diverse circumstance of DTNs.
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Associations between sleep duration, daytime nap duration, and osteoporosis vary by sex, menopause, and sleep quality.
J. Clin. Endocrinol. Metab.
PUBLISHED: 05-21-2014
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Associations between sleep, daytime nap duration, and osteoporosis remain uncertain, and far less is even known about the influence of sex, menopause, and sleep quality on them.
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Differential expression of Nrat1 is responsible for Al-tolerance QTL on chromosome 2 in rice.
J. Exp. Bot.
PUBLISHED: 05-12-2014
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Although rice (Oryza sativa) is the most Al-tolerant species among small-grain cereal crops, there is wide genotypic variation in its tolerance to Al toxicity. A number of quantitative trait loci (QTLs) for Al tolerance have been detected, but the responsible genes have not been identified. By using chromosome segment substitution lines, this work found that Nrat1, a gene encoding an Al transporter, is responsible for a QTL previously detected on chromosome 2. Substitution of the chromosome segment containing Nrat1 from Koshihikari (Al-tolerant variety) by that from Kasalath (Al-sensitive variety) decreased Nrat1 expression and Al uptake and tolerance, but increased binding of Al to the cell wall. Nrat1 in Kasalath showed tissue localization similar to Koshihikari in the roots. Although Koshihikari and Kasalath differed in four amino acids in Nrat1 protein, Nrat1 from Kasalath also showed transport activity for Al. Analysis with site-directed mutagenesis revealed that these differences did not affect the Al-transport activity much. Furthermore, there was no correlation between Al tolerance and the open-reading-frame sequence differences in other rice varieties. On the other hand, there was good correlation between Nrat1 expression and Al tolerance; however, sequence comparison of the promoter region up to 2.1kb did not give a clear difference between the Al-tolerant and -sensitive varieties. Taken together, these results indicate that differential expression of Nrat1 is responsible for the QTL for Al tolerance on chromosome 2, although the mechanism controlling Nrat1 expression remains to be examined.
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Reversibly Immortalized Mouse Articular Chondrocytes Acquire Long-Term Proliferative Capability while Retaining Chondrogenic Phenotype.
Cell Transplant
PUBLISHED: 05-08-2014
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Cartilage tissue engineering holds great promise for treating cartilaginous pathologies including degenerative disorders and traumatic injuries. Effective cartilage regeneration requires an optimal combination of biomaterial scaffolds, chondrogenic seed cells and biofactors. Obtaining sufficient chondrocytes remains a major challenge due to the limited proliferative capability of primary chondrocytes. Here, we investigate if reversibly immortalized mouse articular chondrocytes (iMACs) acquire long-term proliferative capability while retaining the chondrogenic phenotype. Primary mouse articular chondrocytes (MACs) can be efficiently immortalized with a retroviral vector expressing SV40 large T antigen flanked with Cre/loxP sites. iMACs exhibit long-term proliferation in culture, although the immortalization phenotype can be reversed by Cre recombinase. iMACs express the chondrocyte markers Col2a1 and aggrecan and produce chondroid matrix in micromass culture. iMACs form subcutaneous cartilaginous masses in athymic mice. Histologic analysis and chondroid matrix staining demonstrate that iMACs can survive, proliferate and produce chondroid matrix. The chondrogenic growth factor BMP2 promotes iMACs to produce more mature chondroid matrix resembling mature articular cartilage. Taken together, our results demonstrate that iMACs acquire long-term proliferative capability without losing the intrinsic chondrogenic features of MACs. Thus, iMACs provide a valuable cellular platform to optimize biomaterial scaffolds for cartilage regeneration, to identify biofactors that promote the proliferation and differentiation of chondrogenic progenitors and to elucidate the molecular mechanisms underlying chondrogenesis.
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Bone morphogenetic protein 2 inhibits the proliferation and growth of human colorectal cancer cells.
Oncol. Rep.
PUBLISHED: 04-15-2014
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Colorectal cancer (CRC) is one of the most deadly cancers worldwide. Significant progress has been made in understanding the molecular pathogenesis of CRC, which has led to successful early diagnosis, surgical intervention and combination chemotherapy. However, limited therapeutic options are available for metastatic and/or drug-resistant CRC. While the aberrantly activated Wnt/?-catenin pathway plays a critical initiating role in CRC development, disruption of the bone morphogenetic protein (BMP) pathway causes juvenile polyposis syndrome, suggesting that BMP signaling may play a role in CRC development. However, conflicting results have been reported concerning the possible roles of BMP signaling in sporadic colon cancer. Here, we investigated the effect of BMP2 on the proliferation, migration, invasiveness and tumor growth capability of human CRC cells. Using an adenovirus vector that overexpresses BMP2 and the piggyBac transposon-mediated stable BMP2 overexpression CRC line, we found that exogenous BMP2 effectively inhibited HCT116 cell proliferation and colony formation. BMP2 was shown to suppress colon cancer cell migration and invasiveness. Under a low serum culture condition, forced expression of BMP2 induced a significantly increased level of apoptosis in HCT116 cells. Using a xenograft tumor model, we found that forced expression of BMP2 in HCT116 cells suppressed tumor growth, accompanied by decreased cell proliferation activity. Taken together, our results strongly suggest that BMP2 plays an important inhibitory role in governing the proliferation and aggressive features of human CRC cells.
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Population genetic diversity of the northern snakehead (Channa argus) in China based on the mitochondrial DNA control region and adjacent regions sequences.
Mitochondrial DNA
PUBLISHED: 04-15-2014
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Abstract Genetic variation and population structure of northern snakehead (Channa argus) from eight locations in China were investigated using mitochondrial DNA control region and adjacent regions sequences. Sequence analysis showed that there were 105 haplotypes in 260 individuals, 48 unique haplotypes and 57 shared haplotypes, but no common haplotype shared by all populations. As a whole, the haplotype diversity was high (h?=?0.989), while the nucleotide diversity was low (??=?0.00482). AMOVA analysis detected significant genetic differentiation among all eight populations (FST?=?0.328, p?
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Regio- and diastereoselective construction of ?-hydroxy-?-amino ester derivatives via 1,4-conjugate addition of ?,?-unsaturated N-sulfonylimines.
J. Org. Chem.
PUBLISHED: 04-15-2014
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A first example of 1,4-conjugate addition of ?,?-unsaturated N-sulfonylimines via the oxonium ylides trapping process was developed. This method afforded a novel and efficient access for the high regio- and diastereoselective construction of ?-hydroxyl-?-amino esters derivatives, which exhibit inhibitory activity on PTP1B and SIRT1 enzymes in vitro. The synthetic potentials and the biological activity of the resulting products were well demonstrated to be promising for drug discovery.
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An appropriate concentration of arginine is required for normal root growth in rice.
Plant Signal Behav
PUBLISHED: 04-04-2014
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Plant roots play an important role in uptake of water and nutrients, support of above-ground part and environmental sensing, but the molecular mechanisms underlying the root development are poorly understood in rice. We found that a gene (OsASL1) encoding argininosuccinate lyase is involved in normal root development of rice. OsASL1 cleaves argininosuccinate to arginine and fumarate reversibly, the last step in the arginine biosynthetic pathway. Here, we further characterized OsASL1 in terms of expression pattern, subcellular localization, and arginine effect on the root growth. A detailed expression analysis revealed that 2 transcripts of OsASL1, OsASL1.1 and OsASL1.2, showed different expression patterns; OsASL1.1 was expressed in most organs throughout the whole growth period, whereas OsASL1.2 was mainly expressed in the roots. In contrast to plastid-localized OsASL1.1, OsASL1.2 was localized to the cytosol and nucleus. The short-root phenotype of the mutant was not rescued by exogenous addition of the sodium nitroprusside, a nitric oxide donor, but rescued by an appropriate concentration of Arg. Our results indicate that the subcellular localization was determined by the N terminus of OsASL1 and that appropriate concentration of Arg is required for normal root elongation in rice.
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Bone morphogenetic protein-9 effectively induces osteo/odontoblastic differentiation of the reversibly immortalized stem cells of dental apical papilla.
Stem Cells Dev.
PUBLISHED: 03-21-2014
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Dental pulp/dentin regeneration using dental stem cells combined with odontogenic factors may offer great promise to treat and/or prevent premature tooth loss. We previously demonstrated that bone morphogenetic protein 9 (BMP9) is one of the most potent factors in inducing bone formation. Here, we investigate whether BMP9 can effectively induce odontogenic differentiation of the stem cells from mouse apical papilla (SCAPs). Using a reversible immortalization system expressing SV40 T flanked with Cre/loxP sites, we demonstrate that the SCAPs can be immortalized, resulting in immortalized SCAPs (iSCAPs) that express mesenchymal stem cell markers. BMP9 upregulates Runx2, Sox9, and PPAR?2 and odontoblastic markers, and induces alkaline phosphatase activity and matrix mineralization in the iSCAPs. Cre-mediated removal of SV40 T antigen decreases iSCAP proliferation. The in vivo stem cell implantation studies indicate that iSCAPs can differentiate into bone, cartilage, and, to lesser extent, adipocytes upon BMP9 stimulation. Our results demonstrate that the conditionally iSCAPs not only maintain long-term cell proliferation but also retain the ability to differentiate into multiple lineages, including osteo/odontoblastic differentiation. Thus, the reversibly iSCAPs may serve as an important tool to study SCAP biology and SCAP translational use in tooth engineering. Further, BMP9 may be explored as a novel and efficacious factor for odontogenic regeneration.
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Cloning and characterization of cytoplasmic dynein intermediate chain in Fenneropenaeus chinensis and its essential role in white spot syndrome virus infection.
Fish Shellfish Immunol.
PUBLISHED: 03-20-2014
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To investigate the role of cytoplasmic dynein in white spot syndrome virus (WSSV) infection, the full-length cDNA of cytoplasmic dynein intermediate chain (FcDYNCI) was cloned in Fenneropenaeus chinensis, which consists of 2582 bp and encodes a polypeptide of 660 amino acids. Sequence analysis and multiple sequence alignment displayed that FcDYNCI was a member of cytoplasmic dynein 1 family. The FcDYNCI mRNA was most highly expressed in hemocytes, which was significantly up-regulated post WSSV infection. At 12 h post infection (hpi), confocal microscopic observation showed that WSSV could be co-localized with cytoplasmic dynein in hemocytes. After silencing by specific FcDYNCI dsRNA, the FcDYNCI mRNA level and the protein amount of FcDYNCI in hemocytes both exhibited a significant reduction, and the expression levels of three WSSV genes ie1, wsv477 and vp28 all exhibited the greatest decreases at 24 hpi. These results suggested that cytoplasmic dynein was involved in WSSV infection.
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Complete sequencing and phylogenetic analysis of porcine kobuvirus in domestic pigs in Northwest China.
Arch. Virol.
PUBLISHED: 03-05-2014
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Porcine kobuvirus, a member of the genus Kobuvirus that is associated with diarrhea, has been reported in many countries. We determined the complete genome sequence and investigated the genetic evolution of the kobuvirus strain swKoV CH441, which was detected in the highland of Gansu province in Northwest China. The viral genome is 8149 nucleotides (nt) long, including a 29-nt poly(A) tail of the 3' end, and is 90 nt shorter in the 2B coding region than those of other kobuvirus strains whose sequences are available in the GenBank database. Phylogenetic analysis showed that swKoV CH441 was most closely related to porcine kobuvirus CH/HNXX-4 but more distantly related to other strains, including the strains GS-1/2012/CH and GS-2/2012/CH, which were detected in Gansu province, indicating that porcine kobuvirus may have geographic and host differences in evolution.
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Complete sequence and phylogenetic analysis of a porcine sapovirus strain isolated from western China.
Virus Genes
PUBLISHED: 03-03-2014
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Sapovirus (SaV) is a type of calicivirus that can cause acute viral gastroenteritis in humans and animals. SaVs have been found in several mammalian species, including humans, pigs, minks, dogs, and bats. Porcine sapovirus (PoSaV) was first identified in 1980 in the United States and has been found to be circulating throughout China in recent years. In this study, the complete genomic characterization of PoSaV CH430, first found in west China, was reported and analyzed. The genome was 7,342 bp excluding the 30 nt poly(A) tail at the 3' terminus and comprised two major open reading frames. Comprehensive evolutionary and phylogenetic analyses indicated that the CH430 strain belongs to genotype III SaVs. However, this particular isolate and DG24 strain occupied an independent branch of the phylogenetic tree we generated, indicating that they could form a separate subgenotype in the near future. We predicted the cleavage sites for the ORF1 polyprotein located at Q56/G57, Q310/A311, E649/A650, E934/A935, E1047/G1048, and E1712/A1713, separately. This is the first PoSaV strain isolated from western China to be fully sequenced and characterized. It provided a reliable experimental basis for studying the genetic nature of emerging PoSaVs.
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Is alanine aminotransferase associated with osteopenia in middle-aged and elderly chinese?
Endocr Pract
PUBLISHED: 02-13-2014
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To investigate the association between alanine aminotransferase (ALT) levels and risk of osteopenia in middle-aged and elderly Chinese with ALT within the normal range.
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Sorbicatechols A and B, antiviral sorbicillinoids from the marine-derived fungus Penicillium chrysogenum PJX-17.
J. Nat. Prod.
PUBLISHED: 02-04-2014
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Two novel sorbicillinoids combining a bicyclo[2.2.2]octane with a 2-methoxyphenol moiety, named sorbicatechols A (1) and B (2), were isolated from the culture of the marine sediment-derived fungus Penicillium chrysogenum PJX-17, together with the known protocatechuic acid methyl ester and caffeic acid methyl ester (3). Their structures, including absolute configurations, were assigned by analysis of NMR, MS data, and TDDFT ECD calculations. Compounds 1 and 2 exhibited activities against influenza virus A (H1N1), with IC50 values of 85 and 113 ? M, respectively.
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Targeting BMP9-promoted human osteosarcoma growth by inactivation of notch signaling.
Curr Cancer Drug Targets
PUBLISHED: 02-02-2014
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Osteosarcoma (OS) is the most common primary malignancy of bone and is usually associated with poor prognosis due to its high incidence of metastasis and chemoresistance. Molecular pathogenesis of OS is poorly understood. We previously showed that OS cells are refractory to BMP9-induced osteogenesis and respond favorably to proliferation and tumor growth. Here we investigate if Notch signaling mediates the BMP9-promoted cell proliferation and tumor growth of human osteosarcoma (OS). We find that the expression of Notch1, Notch2, Notch3, DLL1, JAG1 and JAG2 is readily detected in most of the tested OS cell lines. BMP9-promoted OS cell proliferation, migration, and cell cycle S/G2 progression are effectively inhibited by a dominant-negative mutant of Notch1 (dnNotch1) or the ?-secretase inhibitor Compound E (ComE). Furthermore, BMP9-promoted tumor growth and osteolytic lesions in vivo are significantly inhibited by dnNotch1. BMP9 up-regulates the expression of Notch1, Notch3, DLL1, and JAG1 in OS cells. Accordingly, BMP9 stimulation induces a nuclear accumulation of NICD, which is blocked by ComE. Our results demonstrate that BMP9-promoted OS proliferation and tumor growth is at least in part mediated by Notch signaling, suggesting that osteogenic BMPs may function as upstream regulators of Notch signaling in OS tumorigenesis. Thus, pharmacologic intervention of Notch signaling may be explored as a new therapeutic strategy for human OS tumors.
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Complete sequence and phylogenetic analysis of a porcine bocavirus strain swBoV CH437.
Virus Genes
PUBLISHED: 01-28-2014
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Porcine bocavirus (PBoV), a member of genus Bocavirus, family Parvoviridae, was first identified in 2009 in Swedish swine herds suffering from postweaning multisystemic wasting syndrome. Up to date, the different species of PBoVs have been reported in different countries. Especially, the virus isolated in China was complicated. In this study, we detected a novel PBoV strain swBoV CH437 from clinical samples collected in Gansu Province, Northwest China. The complete genome of swBoV CH437 was 5,275 nucleotides (nt) in length and contains three ORFs: ORF1 encodes NS1 (2,004 nt, 667 aa), ORF3 encodes NP1 (681 nt, 226 aa), and ORF2 encodes VP1 (2,049 nt, 682 aa) and VP2 (1,641 nt, 546 aa). Sequence analysis demonstrated that the NS1 gene shared 24.2-88.6 % nucleotide sequence identity, the NP1 shared 21.3-89.9 %, less than 95 % nucleotide sequence identity with other PBoV strains. Therefore, we propose that swBoV CH437 should be classified as a novel PBoV species.
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Normal root elongation requires arginine produced by argininosuccinate lyase in rice.
Plant J.
PUBLISHED: 01-24-2014
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Plant roots play an important role in the uptake of water and nutrients, structural support and environmental sensing, but the molecular mechanisms involved in root development are poorly understood in rice (Oryza sativa), which is characterized by a dense fibrous root system. Here we report a rice mutant (red1 for root elongation defect 1) with short roots. Morphological and physiological analyses showed that the mutant had a shorter length from the quiescent center (QC) to the starting point of the elongation zone but a similar cell size and number of lateral and crown roots compared with the wild type. Furthermore, the mutant had similar radial structure and nutrient uptake patterns to the wild type. Map-based cloning revealed that the mutant phenotype was caused by a point mutation of a gene encoding an argininosuccinate lyase (ASL), catalyzing the last step of arginine biosynthesis. The OsASL1 gene has two distinct transcripts, OsASL1.1 and OsASL1.2, which result from different transcription start sites, but only OsASL1.1 was able to complement the mutant phenotype. OsASL1.1 was expressed in both the roots and shoots. The protein encoded by OsASL1.1 showed ASL activity in yeast. OsALS1.1 was localized to the plastid. The short root of the mutant was rescued by exogenous addition of arginine, but not by other amino acids. These results indicate that arginine produced by ASL is required for normal root elongation in rice.
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Complete mitochondrial genomes of domesticated and wild common carp (Cyprinus carpio L.).
Mitochondrial DNA
PUBLISHED: 01-21-2014
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Abstract Glass red common carp (Cyprinus carpio var. wananensis) were occasionally found among wild common carp (Cyprinus carpio) 50 years ago. In this paper, we determined the complete sequences of Glass red common carp and wild common carp mitogenomes. Both mitogenomes exhibited the same length of 16,581?bp, order in 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes and 1 control region. The sequence similarity between them reached 99.62% and 100% in 21 tRNA genes (excluding tRNA(Ser)) and 2 protein-coding genes (ATP8 and ND4L), respectively. The two mitogenomes will be useful in elucidating the evolutional relationship of the common carp.
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A Simplified and Versatile System for the Simultaneous Expression of Multiple siRNAs in Mammalian Cells Using Gibson DNA Assembly.
PLoS ONE
PUBLISHED: 01-01-2014
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RNA interference (RNAi) denotes sequence-specific mRNA degradation induced by short interfering double-stranded RNA (siRNA) and has become a revolutionary tool for functional annotation of mammalian genes, as well as for development of novel therapeutics. The practical applications of RNAi are usually achieved by expressing short hairpin RNAs (shRNAs) or siRNAs in cells. However, a major technical challenge is to simultaneously express multiple siRNAs to silence one or more genes. We previously developed pSOS system, in which siRNA duplexes are made from oligo templates driven by opposing U6 and H1 promoters. While effective, it is not equipped to express multiple siRNAs in a single vector. Gibson DNA Assembly (GDA) is an in vitro recombination system that has the capacity to assemble multiple overlapping DNA molecules in a single isothermal step. Here, we developed a GDA-based pSOK assembly system for constructing single vectors that express multiple siRNA sites. The assembly fragments were generated by PCR amplifications from the U6-H1 template vector pB2B. GDA assembly specificity was conferred by the overlapping unique siRNA sequences of insert fragments. To prove the technical feasibility, we constructed pSOK vectors that contain four siRNA sites and three siRNA sites targeting human and mouse ?-catenin, respectively. The assembly reactions were efficient, and candidate clones were readily identified by PCR screening. Multiple ?-catenin siRNAs effectively silenced endogenous ?-catenin expression, inhibited Wnt3A-induced ?-catenin/Tcf4 reporter activity and expression of Wnt/?-catenin downstream genes. Silencing ?-catenin in mesenchymal stem cells inhibited Wnt3A-induced early osteogenic differentiation and significantly diminished synergistic osteogenic activity between BMP9 and Wnt3A in vitro and in vivo. These findings demonstrate that the GDA-based pSOK system has been proven simplistic, effective and versatile for simultaneous expression of multiple siRNAs. Thus, the reported pSOK system should be a valuable tool for gene function studies and development of novel therapeutics.
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The piggyBac transposon-mediated expression of SV40 T antigen efficiently immortalizes mouse embryonic fibroblasts (MEFs).
PLoS ONE
PUBLISHED: 01-01-2014
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Mouse embryonic fibroblasts (MEFs) are mesenchymal stem cell (MSC)-like multipotent progenitor cells and can undergo self-renewal and differentiate into to multiple lineages, including bone, cartilage and adipose. Primary MEFs have limited life span in culture, which thus hampers MEFs' basic research and translational applications. To overcome this challenge, we investigate if piggyBac transposon-mediated expression of SV40 T antigen can effectively immortalize mouse MEFs and that the immortalized MEFs can maintain long-term cell proliferation without compromising their multipotency. Using the piggyBac vector MPH86 which expresses SV40 T antigen flanked with flippase (FLP) recognition target (FRT) sites, we demonstrate that mouse embryonic fibroblasts (MEFs) can be efficiently immortalized. The immortalized MEFs (piMEFs) exhibit an enhanced proliferative activity and maintain long-term cell proliferation, which can be reversed by FLP recombinase. The piMEFs express most MEF markers and retain multipotency as they can differentiate into osteogenic, chondrogenic and adipogenic lineages upon BMP9 stimulation in vitro. Stem cell implantation studies indicate that piMEFs can form bone, cartilage and adipose tissues upon BMP9 stimulation, whereas FLP-mediated removal of SV40 T antigen diminishes the ability of piMEFs to differentiate into these lineages, possibly due to the reduced expansion of progenitor populations. Our results demonstrate that piggyBac transposon-mediated expression of SV40 T can effectively immortalize MEFs and that the reversibly immortalized piMEFs not only maintain long-term cell proliferation but also retain their multipotency. Thus, the high transposition efficiency and the potential footprint-free natures may render piggyBac transposition an effective and safe strategy to immortalize progenitor cells isolated from limited tissue supplies, which is essential for basic and translational studies.
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Characterization of constitutive promoters for piggyBac transposon-mediated stable transgene expression in mesenchymal stem cells (MSCs).
PLoS ONE
PUBLISHED: 01-01-2014
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Multipotent mesenchymal stem cells (MSCs) can undergo self-renewal and give rise to multi-lineages under given differentiation cues. It is frequently desirable to achieve a stable and high level of transgene expression in MSCs in order to elucidate possible molecular mechanisms through which MSC self-renewal and lineage commitment are regulated. Retroviral or lentiviral vector-mediated gene expression in MSCs usually decreases over time. Here, we choose to use the piggyBac transposon system and conduct a systematic comparison of six commonly-used constitutive promoters for their abilities to drive RFP or firefly luciferase expression in somatic HEK-293 cells and MSC iMEF cells. The analyzed promoters include three viral promoters (CMV, CMV-IVS, and SV40), one housekeeping gene promoter (UbC), and two composite promoters of viral and housekeeping gene promoters (hEFH and CAG-hEFH). CMV-derived promoters are shown to drive the highest transgene expression in HEK-293 cells, which is however significantly reduced in MSCs. Conversely, the composite promoter hEFH exhibits the highest transgene expression in MSCs whereas its promoter activity is modest in HEK-293 cells. The reduced transgene expression driven by CMV promoters in MSCs may be at least in part caused by DNA methylation, or to a lesser extent histone deacetlyation. However, the hEFH promoter is not significantly affected by these epigenetic modifications. Taken together, our results demonstrate that the hEFH composite promoter may be an ideal promoter to drive long-term and high level transgene expression using the piggyBac transposon vector in progenitor cells such as MSCs.
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Adenovirus-mediated efficient gene transfer into cultured three-dimensional organoids.
PLoS ONE
PUBLISHED: 01-01-2014
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Three-dimensional organoids have been recently established from various tissue-specific progenitors (such as intestinal stem cells), induced pluripotent stem cells, or embryonic stem cells. These cultured self-sustaining stem cell-based organoids may become valuable systems to study the roles of tissue-specific stem cells in tissue genesis and disease development. It is thus conceivable that effective genetic manipulations in such organoids may allow us to reconstruct disease processes and/or develop novel therapeutics. Recombinant adenoviruses are one of the most commonly used viral vectors for in vitro and in vivo gene deliveries. In this study, we investigate if adenoviruses can be used to effectively deliver transgenes into the cultured "mini-gut" organoids derived from intestinal stem cells. Using adenoviral vectors that express fluorescent proteins, we demonstrate that adenoviruses can effectively deliver transgenes into the cultured 3-D "mini-gut" organoids. The transgene expression can last at least 10 days in the cultured organoids. As a proof-of-principle experiment, we demonstrate that adenovirus-mediated noggin expression effectively support the survival and self-renewal of mini-gut organoids, while adenovirus-mediated expression of BMP4 inhibits the self-sustainability and proliferation of the organoids. Thus, our results strongly suggest that adenovirus vectors can be explored as effective gene delivery vehicles to introduce genetic manipulations in 3-D organoids.
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Adenovirus-mediated gene transfer in mesenchymal stem cells can be significantly enhanced by the cationic polymer polybrene.
PLoS ONE
PUBLISHED: 01-01-2014
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Mesenchymal stem cells (MSCs) are multipotent progenitors, which can undergo self-renewal and give rise to multi-lineages. A great deal of attentions have been paid to their potential use in regenerative medicine as potential therapeutic genes can be introduced into MSCs. Genetic manipulations in MSCs requires effective gene deliveries. Recombinant adenoviruses are widely used gene transfer vectors. We have found that although MSCs can be infected in vitro by adenoviruses, high virus titers are needed to achieve high efficiency. Here, we investigate if the commonly-used cationic polymer Polybrene can potentiate adenovirus-mediated transgene delivery into MSCs, such as C2C12 cells and iMEFs. Using the AdRFP adenovirus, we find that AdRFP transduction efficiency is significantly increased by Polybrene in a dose-dependent fashion peaking at 8 ?g/ml in C2C12 and iMEFs cells. Quantitative luciferase assay reveals that Polybrene significantly enhances AdFLuc-mediated luciferase activity in C2C12 and iMEFs at as low as 4 ?g/ml and 2 ?g/ml, respectively. FACS analysis indicates that Polybrene (at 4 ?g/ml) increases the percentage of RFP-positive cells by approximately 430 folds in AdRFP-transduced iMEFs, suggesting Polybrene may increase adenovirus infection efficiency. Furthermore, Polybrene can enhance AdBMP9-induced osteogenic differentiation of MSCs as early osteogenic marker alkaline phosphatase activity can be increased more than 73 folds by Polybrene (4 ?g/ml) in AdBMP9-transduced iMEFs. No cytotoxicity was observed in C2C12 and iMEFs at Polybrene up to 40 ?g/ml, which is about 10-fold higher than the effective concentration required to enhance adenovirus transduction in MSCs. Taken together, our results demonstrate that Polybrene should be routinely used as a safe, effective and inexpensive augmenting agent for adenovirus-mediated gene transfer in MSCs, as well as other types of mammalian cells.
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Sorbicillamines A-E, Nitrogen-Containing Sorbicillinoids from the Deep-Sea-Derived Fungus Penicillium sp. F23-2.
J. Nat. Prod.
PUBLISHED: 11-11-2013
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Five new nitrogen-containing sorbicillinoids named sorbicillamines A-E (1-5) were isolated from an agitated culture of the deep-sea-derived fungus Penicillium sp. F23-2, which has previously produced indole alkaloids and terpenoids when cultured under static conditions. The structures of 1 to 5, including absolute configurations, were determined based on MS, NMR, and circular dichroism (CD) data. The cytotoxicities of the five new sorbicillin alkaloids against the HeLa, BEL-7402, HEK-293, HCT-116, and P388 cell lines were evaluated.
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The complete mitochondrial genome of the sea carp, Cyprinus acutidorsalis (Cypriniformes: Cyprinidae).
Mitochondrial DNA
PUBLISHED: 10-15-2013
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Abstract The complete mitochondrial genome of sea carp Cyprinus acutidorsalis was determined in this study. The genome was 16,579?bp in length. As in other vertebrates, it consisted of 13 protein-coding genes, 22 transfer RNA (tRNA) genes, 2 ribosomal RNA (rRNA) genes, and 1 non-coding control region. The overall base composition was estimated to be A, 31.9%; T, 24.8%; C, 27.6% and G, 15.7% with AT bias of 56.7%. The genomic composition, organization, and gene order of C. acutidorsalis was similar to most of vertebrates. This result may provide the basis for the study of genetic structure as well as resource conservation and protection of C. acutidorsalis.
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Generation of a monoclonal antibody to S1 protein of porcine epidemic diarrhea virus.
Monoclon Antib Immunodiagn Immunother
PUBLISHED: 10-12-2013
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Gene encoding the N-terminal half of spike protein (S1) of porcine epidemic diarrhea virus (PEDV) was cloned and expressed as a recombinant protein in Escherichia coli BL21 (DE3). Then, female BALB/c mice were immunized with the purified recombinant S1 protein (rS1), and a monoclonal antibody (MAb designated as 5E12) against the rS1 protein was achieved by hybridoma technique. MAb 5E12 not only reacted with rS1 protein indirect ELISA and Western blot, but also recognized PEDV transiently expressed in Vero E6 cells in indirect immunofluorescence examinations. This work suggests that 5E12 would be a useful tool as a specific diagnostic reagent for detecting PEDV S protein.
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A member of the heavy metal P-type ATPase OsHMA5 is involved in xylem loading of copper in rice.
Plant Physiol.
PUBLISHED: 09-24-2013
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Heavy metal-transporting P-type ATPase (HMA) has been implicated in the transport of heavy metals in plants. Here, we report the function and role of an uncharacterized member of HMA, OsHMA5 in rice (Oryza sativa). Knockout of OsHMA5 resulted in a decreased copper (Cu) concentration in the shoots but an increased Cu concentration in the roots at the vegetative stage. At the reproductive stage, the concentration of Cu in the brown rice was significantly lower in the mutants than in the wild-type rice; however, there was no difference in the concentrations of iron, manganese, and zinc between two independent mutants and the wild type. The Cu concentration of xylem sap was lower in the mutants than in the wild-type rice. OsHMA5 was mainly expressed in the roots at the vegetative stage but also in nodes, peduncle, rachis, and husk at the reproductive stage. The expression was up-regulated by excess Cu but not by the deficiency of Cu and other metals, including zinc, iron, and manganese, at the vegetative stage. Analysis of the transgenic rice carrying the OsHMA5 promoter fused with green fluorescent protein revealed that it was localized at the root pericycle cells and xylem region of diffuse vascular bundles in node I, vascular tissues of peduncle, rachis, and husk. Furthermore, immunostaining with an antibody against OsHMA5 revealed that it was localized to the plasma membrane. Expression of OsHMA5 in a Cu transport-defective mutant yeast (Saccharomyces cerevisiae) strain restored the growth. Taken together, OsHMA5 is involved in loading Cu to the xylem of the roots and other organs.
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Development of a reverse genetics system for the aG strain of rabies virus in China.
Arch. Virol.
PUBLISHED: 07-16-2013
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The aG rabies virus strain has been attenuated through multiple passages in cells and is now used as a vaccine strain in China. We attempted to develop a reverse genetics system using the aG strain. Recombinant full-length genomic cDNA was flanked by a hammerhead ribozyme and the hepatitis delta virus ribozyme. Three helper plasmids encoding the nucleoprotein, the phosphoprotein, and the large protein were produced and introduced together with a plasmid containing the full-length aG viral genome into BHK-21 cells by transfection. Recombinant virus was successfully recovered from the cloned cDNA under the control of a CMV promoter driven by RNA polymerase II. The recombinant virus was confirmed by RT-PCR, and the titer of the recombinant virus was 6.2 log LD50.
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Antiviral alkaloids produced by the mangrove-derived fungus Cladosporium sp. PJX-41.
J. Nat. Prod.
PUBLISHED: 06-12-2013
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Six new indole alkaloids including five new glyantrypine derivatives (1, 2a, 2b, 3, 4) and a new pyrazinoquinazoline derivative (5), together with eight known alkaloids (6-13), were isolated from the culture of the mangrove-derived fungus Cladosporium sp. PJX-41. Their structures were elucidated primarily by spectroscopic and physical data. The absolute configurations of compounds 1-9 were established on the basis of CD, NOESY data, and single-crystal X-ray diffraction analysis. Compounds 2b, 5, 7-9, and 11 exhibited significant activities against influenza virus A (H1N1), with IC50 values of 82-89 ?M.
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Phylogenetic analysis reveals the coexistence of interfamily and interspecies horizontal gene transfer in Streptococcus thermophilus strains isolated from the same yoghurt.
Mol. Phylogenet. Evol.
PUBLISHED: 05-29-2013
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Horizontal gene transfer (HGT) is an important evolutionary mechanism that has shaped prokaryotic genomes. For Streptococcus thermophilus, there is no direct evidence that the bacteria might acquire a second paralog from a different origin in the same niche. In this study, we found that four isolates of S. thermophilus (B, C, E and F) from the same yoghurt contained two putative homologs of the eno genes (eno-1 and eno-2) and two putative homologs of the guaB genes (guaB-1 and guaB-2). Both eno-1 and guaB-1 shared 100% nucleotide identity among the four isolates, and with isolate A and S. thermophilus ND03. Phylogenetic and nucleotide divergence analyses indicated that guaB-2 of these isolates may have been acquired from species in the genus Streptococcus, while eno-2 of isolates B and C may have been acquired from a donor in the genus Streptococcus. The eno-2 genes of isolates E and F may have been acquired from a donor in the Enterococcus genera. Relative synonymous codon usage analysis confirmed the eno-2 genes of isolates E and F as being acquired from a donor in genus Enterococcus. This study provides evidence that interfamily and interspecies HGT occur in S. thermophilus strains isolated from the same niche.
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Characterization and genetic mapping of a Photoperiod-sensitive dwarf 1 locus in rice (Oryza sativa L.).
Theor. Appl. Genet.
PUBLISHED: 04-30-2013
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Plant height is an important agronomic trait for crop architecture and yield. Most known factors determining plant height function in gibberellin or brassinosteroid biosynthesis or signal transduction. Here, we report a japonica rice (Oryza sativa ssp. japonica) dominant dwarf mutant, Photoperiod-sensitive dwarf 1 (Psd1). The Psd1 mutant showed impaired cell division and elongation, and a severe dwarf phenotype under long-day conditions, but nearly normal growth in short-day. The plant height of Psd1 mutant could not be rescued by gibberellin or brassinosteroid treatment. Genetic analysis with R1 and F2 populations determined that Psd1 phenotype was controlled by a single dominant locus. Linkage analysis with 101 tall F2 plants grown in a long-day season, which were derived from a cross between Psd1 and an indica cultivar, located Psd1 locus on chromosome 1. Further fine-mapping with 1017 tall F2 plants determined this locus on an 11.5-kb region. Sequencing analysis of this region detected a mutation site in a gene encoding a putative lipid transfer protein; the mutation produces a truncated C-terminus of the protein. This study establishes the genetic foundation for understanding the molecular mechanisms regulating plant cell division and elongation mediated by interaction between genetic and environmental factors.
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Preferential delivery of zinc to developing tissues in rice is mediated by P-type heavy metal ATPase OsHMA2.
Plant Physiol.
PUBLISHED: 04-10-2013
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Developing tissues such as meristems and reproductive organs require high zinc, but the molecular mechanisms of how zinc taken up by the roots is preferentially delivered to these tissues with low transpiration are unknown. Here, we report that rice (Oryza sativa) heavy metal ATPase2 (OsHMA2), a member of P-type ATPases, is involved in preferential delivery of zinc to the developing tissues in rice. OsHMA2 was mainly expressed in the mature zone of the roots at the vegetative stage, but higher expression was also found in the nodes at the reproductive stage. The expression was unaffected by either zinc deficiency or zinc excess. OsHMA2 was localized at the pericycle of the roots and at the phloem of enlarged and diffuse vascular bundles in the nodes. Heterologous expression of OsHMA2 in yeast (Saccharomyces cerevisiae) showed influx transport activity for zinc as well as cadmium. Two independent Tos17 insertion lines showed decreased zinc concentration in the crown root tips, decreased concentration of zinc and cadmium in the upper nodes and reproductive organs compared with wild-type rice. Furthermore, a short-term labeling experiment with (67)Zn showed that the distribution of zinc to the panicle and uppermost node I was decreased, but that, to the lower nodes, was increased in the two mutants. Taken together, OsHMA2 in the nodes plays an important role in preferential distribution of zinc as well as cadmium through the phloem to the developing tissues.
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A plasma membrane-localized small peptide is involved in rice aluminum tolerance.
Plant J.
PUBLISHED: 03-12-2013
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A transcription factor for Al tolerance, ART1, regulates the expression of at least 30 genes in rice. Here we functionally characterized one of the downstream genes, OsCDT3, which encodes a predicted peptide of only 53 amino acid residues rich in cysteine. Knockdown of this gene resulted in decreased tolerance to Al, but did not affect the tolerance to Cd. The aluminum (Al) content in the root residues including cell wall and the plasma membrane of knockdown lines decreased, but the Al concentration in the root cell sap increased compared with those of the wild-type rice. OsCDT3 was mainly expressed in the roots and its expression was specifically induced by Al exposure, not by low pH and other metals. There was a small genotypic variation in OsCDT3 expression level, but no correlation between Al tolerance and the OsCDT3 variation was found among 17 rice cultivars. Analysis of pOsCDT3::GFP transgenic rice showed that OsCDT3 was expressed at all cells in the root tips. Transient expression of OsCDT3 fused with GFP at both N- and C-termini showed that OsCDT3 was anchored to the plasma membrane. Expression of OsCDT3 in yeast conferred tolerance to Al, but not to Cd. Furthermore, OsCDT3 did not show transport activity for Al in yeast, but was able to directly bind Al in vitro. Taken together, our results indicate that OsCDT3 anchoring to the plasma membrane may play a role in stopping entry of Al into the root cells by binding Al, therefore, contributing to high Al tolerance in rice.
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Associations between age at menarche and menopause with cardiovascular disease, diabetes, and osteoporosis in Chinese women.
J. Clin. Endocrinol. Metab.
PUBLISHED: 03-07-2013
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Ages at menarche and menopause are associated with cardiovascular disease (CVD), diabetes, and osteoporosis in Caucasian women, but associations remain unexplored in Chinese women.
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Wnt10b promotes differentiation of mouse hair follicle melanocytes.
Int J Med Sci
PUBLISHED: 02-28-2013
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Previous research has revealed that Wnt10b activates canonical Wnt signaling, which is integral to melanocyte differentiation in hair follicles (HFs). However, the function of Wnt10b in HF melanocytes remains poorly understood. We determined using Dct-LacZ transgenic mice that Wnt10b is mainly expressed near and within melanocytes of the hair bulbs during the anagen stage of the hair cycle. We also found that Wnt10b promotes an increase in melanocyte maturation and pigmentation in the hair bulbs of the mouse HF. To further explore the potential functions of Wnt10b in mouse HF melanocytes, we infected iMC23 cells with Ad-Wnt10b to overexpress Wnt10b. We demonstrated that Wnt10b promotes the differentiation of melanocytes by activating canonical Wnt signaling in melanocytes.
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Complete genome sequence of rabies virus CVS-24 from China.
Arch. Virol.
PUBLISHED: 01-30-2013
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The entire genome of the mouse-adapted rabies virus strain CVS-24 (challenge virus standard 24), was sequenced. The overall length of the genome was 11,927 nucleotide (nt), comprising a leader sequence of 58 nt, a nucleoprotein (N) gene of 1353 nt, phosphoprotein (P) gene of 894 nt, a matrix protein (M) gene of 609 nt, a glycoprotein (G) gene of 1575 nt, an RNA-dependent RNA polymerase (RdRp, L) gene of 6384 nt and a trailer region of 70 nt. There was a TGAAAAAAA (TG7) consensus sequence at the end of each gene, except the G gene which had an AGAAAAAAA sequence at the end, and the L/trailer region had the sequence CGAAAAAAA. Three were AACAYYYCT consensus start signals close to TG7. The five cistrons were separated by intergenic regions (IGRS) of 2, 5, 5, 24 nt, respectively. Residue 333 of the mature G protein, which is considered to be associated with pathogenicity, was Ala in CVS-24. The topology of the phylogenetic trees generated using N protein sequences suggested that CVS-11 and CVS-N2C have a close relationship to CVS-24.
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Associations of green tea and rock tea consumption with risk of impaired fasting glucose and impaired glucose tolerance in chinese men and women.
PLoS ONE
PUBLISHED: 01-01-2013
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To explore the associations of green tea and rock tea consumption with risk of impaired fasting glucose (IFG) and impaired glucose tolerance (IGT).
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Serum level of endogenous secretory receptor for advanced glycation end products and other factors in type 2 diabetic patients with mild cognitive impairment.
Diabetes Care
PUBLISHED: 10-19-2011
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Determine the serum levels of endogenous secretory receptor for advanced glycation end products (esRAGEs) in patients with type 2 diabetes and mild cognitive impairment (MCI) and in control patients with type 2 diabetes but no MCI, and examine the relationship of esRAGE and MCI with other clinical factors.
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OsYSL6 is involved in the detoxification of excess manganese in rice.
Plant Physiol.
PUBLISHED: 10-10-2011
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Yellow Stripe-Like (YSL) proteins belong to the oligopeptide transporter family and have been implicated in metal transport and homeostasis in different plant species. Here, we functionally characterized a rice (Oryza sativa) YSL member, OsYSL6. Knockout of OsYSL6 resulted in decreased growth of both roots and shoots only in the high-manganese (Mn) condition. There was no difference in the concentration of total Mn and other essential metals between the wild-type rice and the knockout line, but the knockout line showed a higher Mn concentration in the leaf apoplastic solution and a lower Mn concentration in the symplastic solution than wild-type rice. OsYSL6 was constitutively expressed in both the shoots and roots, and the expression level was not affected by either deficiency or toxicity of various metals. Furthermore, the expression level increased with leaf age. Analysis with OsYSL6 promoter-green fluorescent protein transgenic rice revealed that OsYSL6 was expressed in all cells of both the roots and shoots. Heterogolous expression of OsYSL6 in yeast showed transport activity for the Mn-nicotianamine complex but not for the Mn-mugineic acid complex. Taken together, our results suggest that OsYSL6 is a Mn-nicotianamine transporter that is required for the detoxification of excess Mn in rice.
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Molecular characterization of China rabies virus vaccine strain.
Virol. J.
PUBLISHED: 09-20-2011
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Rabies virus (RV), the agent of rabies, can cause a severe encephalomyelitis in several species of mammals, including humans. As a human rabies vaccine strain employed in China, the genetic knowledge of the aG strain has not been fully studied. The main goal of the present study is to amplify the whole genome of aG strain, and genetic relationships between other vaccine strains and wild strains were analyzed.
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Molecular detection of a mixed infection of Goatpox virus, Orf virus, and Mycoplasma capricolum subsp. capripneumoniae in goats.
J. Vet. Diagn. Invest.
PUBLISHED: 06-13-2011
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The current study investigated an outbreak of mixed infection with Goatpox virus (GTPV), Orf virus (ORFV), and Mycoplasma capricolum subsp. capripneumoniae (MCCP) that occurred on a Chinese goat farm, with a case fatality rate of 60.2%. The observed clinical signs were ecthyma and accelerated respiration with frequent coughing. Specific fragments of the p32 gene of GTPV, B2L gene of ORFV, and 16S ribosomal RNA gene of MCCP were synchronously amplified by polymerase chain reaction (PCR) from the tissues of 12 dead goats. The PCR products were cloned, sequenced, and aligned with related reference sequences in GenBank for further identification of the pathogens. The present study reports a mixed infection with GTPV, ORFV, and MCCP in goats.
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Expression and antigenicity characterization for truncated capsid protein of porcine circovirus type 2.
Can. J. Vet. Res.
PUBLISHED: 04-05-2011
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Three pairs of specific primers were designed to amplify F2-1, F2-2, and XF2-2 truncated capsid protein genes of porcine circovirus type 2 (PCV-2). Amplified sequences were subcloned to pET-32a(+) vectors and expressed in Rosetta (DE3) Escherichia coli by induction of isopropy-?-D-thiogalactoside (IPTG). All of the fusion proteins had positive reactions to PCV-2 antiserum and His-XF2-2 showed the best reactivity. Proteins were used to immunize BALB/c mice to produce monoclonal antibodies (mAbs), and 7 mAbs were selected. Capsid protein N-terminal parts 55 to 96 amino acid (aa), 97 to 141 aa, and 143 to 211 aa were confirmed as binding regions of the 7 mAbs. Reactivity between His-XF2-2 and the 7 mAbs was detected, FmAb-8 showed the best reactivity. The dominant B-cell epitope was located at 97 to 141 aa. The PEPSCAN indicated that the P122-136 peptide contained the dominant B-cell epitope.
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Cardiovascular disease (CVD) risk, insulin resistance and ?-cell function in prehypertension population of China.
Atherosclerosis
PUBLISHED: 03-19-2011
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To explore the cardiovascular disease (CVD) risk in prehypertensive subjects and evaluate whether high blood pressure (BP) is associated with insulin resistance (IR) and ?-cell dysfunction.
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Further characterization of an aluminum influx transporter in rice.
Plant Signal Behav
PUBLISHED: 01-01-2011
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Nrat1 is a plasma membrane-localized aluminum transporter recently identified in rice, which is a member of Nramp family. Here, we further characterized this transporter in terms of transport substrate specificity. Heterologous assay in yeast showed that Al transport activity by Nrat1 was unaffected by the presence of high concentration of Ca, but significantly inhibited by trivalent ions including Yb and Ga, analogs of Al. Knockout of Nrat1 did not affect the uptake of Cd and Mn in rice. On the other hand, over-expression of Nrat1 led to enhanced Al uptake by rice roots compared with wild-type rice, but did not affect Cd uptake. These results provide further evidence that unlike other Nramp members, Nrat1 is an influx transporter for trivalent Al ion.
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Plasma membrane-localized transporter for aluminum in rice.
Proc. Natl. Acad. Sci. U.S.A.
PUBLISHED: 10-11-2010
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Aluminum (Al) is the most abundant metal in the Earths crust, but its trivalent ionic form is highly toxic to all organisms at low concentrations. How Al enters cells has not been elucidated in any organisms. Herein, we report a transporter, Nrat1 (Nramp aluminum transporter 1), specific for trivalent Al ion in rice. Nrat1 belongs to the Nramp (natural resistance-associated macrophage protein) family, but shares a low similarity with other Nramp members. When expressed in yeast, Nrat1 transports trivalent Al ion, but not other divalent ions, such as manganese, iron, and cadmium, or the Al-citrate complex. Nrat1 is localized at the plasma membranes of all cells of root tips except epidermal cells. Knockout of Nrat1 resulted in decreased Al uptake, increased Al binding to cell wall, and enhanced Al sensitivity, but did not affect the tolerance to other metals. Expression of Nrat1 is up-regulated by Al in the roots and regulated by a C2H2 zinc finger transcription factor (ART1). We therefore concluded that Nrat1 is a plasma membrane-localized transporter for trivalent Al, which is required for a prior step of final Al detoxification through sequestration of Al into vacuoles.
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FMD subunit vaccine produced using a silkworm-baculovirus expression system: protective efficacy against two type Asia1 isolates in cattle.
Vet. Microbiol.
PUBLISHED: 10-04-2010
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Cattle vaccinated with a single dose of subunit vaccine containing the capsid and 3C proteinase coding regions of foot-and-mouth disease virus (FMDV) Asia I/HNK/CHA/05 strain were protected when challenged 28 days later with a homologous virus. Here, the 50% bovine protective dose (PD(50)) test was performed to assess the potency of the subunit vaccine. When challenged with two Chinese isolates, the subunit vaccine could achieve 6.5 PD(50) (challenged with Asia I/HNK/CHA/05 strain) and 5.2 PD(50) (challenged with Asia I/JSL/05 strain) per dose.
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Co-infection with influenza A/H1N1 and A/H3N2 viruses in a patient with influenza-like illness during the winter/spring of 2008 in Shanghai, China.
J. Med. Virol.
PUBLISHED: 06-30-2010
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Co-infection with different influenza viruses occurs naturally and plays an important role in epidemiology and pathogenicity. To monitor the prevalence of influenza viruses in humans during seasonal influenza epidemics in Shanghai, China, and to analyze the genetic characteristics of the viruses, 365 nasopharyngeal swabs collected from patients with influenza-like illness between January and April 2008, were tested by a colloidal gold assay, viral isolation in Madin-Darby canine kidney (MDCK) cells, direct immunofluorescence assay and multiplex RT-PCR. The genetic characteristics of the viruses were analyzed by full-length PCR amplification of the HA segment. One case of co-infection with influenza A/H1N1 and A/H3N2 viruses was detected among the 7 cases of A/H1N1, 84 cases of A/H3N2 and 48 cases of influenza B virus during the winter/spring of 2008. All influenza A/H1N1 and A/H3N2 isolates were similar, including the co-infecting isolates. The present study demonstrates the possibility of natural co-infection with different types of influenza viruses in humans, which could provide the opportunity for the occurrence of viral genetic reassortment within the human respiratory tract.
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Overweight, obesity, and their associations with insulin resistance and ?-cell function among Chinese: a cross-sectional study in China.
Metab. Clin. Exp.
PUBLISHED: 03-11-2010
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The aim of this study was to evaluate the associations of body mass index (BMI) with insulin resistance and ?-cell function in subjects with normal glucose tolerance. A cross-sectional study was carried out in Fujian province by multistratified sampling from July 2007 to May 2008. The sample consisted of 2931 subjects aged from 20 to 79 years. The questionnaires, physical examinations, and laboratory tests were obtained from all the participants. The homeostasis model assessment of insulin resistance (HOMA-IR) index was used to estimate insulin sensitivity, insulin secretion was assessed using the HOMA-? index, and ?-cell function was quantified as the ratio of the incremental insulin to glucose responses over the first 30 minutes during the oral glucose tolerance test (?I30/?G30). Another measure was adjusted for insulin sensitivity as it modulates ?-cell function ([?I30/?G30]/HOMA-IR). Associations of BMI with morbidities were estimated using multiple logistic regression analysis. Relationships of BMI to insulin resistance and ?-cell function were assessed using multiple linear regression analysis and analysis of covariance. The age- and sex-adjusted prevalence of overweight and obesity was 23.04% (27.44% in men and 18.40% in women) and 2.65% (2.75% in men and 2.55% in women), respectively. After adjustment for covariables, BMI was independently associated with morbidity conditions; and there were increasing trend for odds ratios of morbidities across the BMI categories. There were independent differences for HOMA-IR, HOMA-?, and ?I30/?G30 between the normal-weight, overweight, and obese groups except for (?I30/?G30)/HOMA-IR. Body mass index was significantly and independently associated with HOMA-IR, HOMA-?, and ?I30/?G30 in the multiple linear regression analysis. Body mass index was an independent risk factor for hypertension, type 2 diabetes mellitus, dyslipidemia, metabolic syndrome, as well as the indexes of insulin resistance and ?-cell function. It is imperative that the whole society pay more attention to the identification and intervention of overweight and obesity to prevent obesity-related diseases at the very early stage.
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Clinical analysis of 249 cases of adrenal tumors in a Chinese hospital.
Urol. Int.
PUBLISHED: 01-21-2010
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This study was designed to evaluate the clinical and pathological characteristics of adrenal masses.
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Age-related changes in the orientation of lumbar facet joints.
Spine
PUBLISHED: 08-01-2009
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A measurement and analysis of the orientation of the lumbar facet joints at the L4-L5 level of different age groups.
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[Status and application prospect in repair of spinal cord injury by stem cells].
Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi
PUBLISHED: 04-16-2009
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To review the status and application prospect in repair of spinal cord injury by stem cells.
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Preparation and characterization of bisphenol A-cationized bovine serum albumin.
J. Immunol. Methods
PUBLISHED: 03-14-2009
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To detect BPA residue in the environment quickly by immune method, preparation of high quality antibody against BPA was the most critical step of project. Based on Mannich-type reaction and EDC method, we produced BPA-cBSA and BHPVA-BSA against BPA and the conjugates were characterized by UV-Vis and FT-IR spectroscopy. Compared with BSA, cBSA as carrier protein could improve the efficiency of coupling and significantly enhance the immune response against BPA. The mole coupling ratio of BPA-cBSA (8:1) was greater than that of BHPVA-BSA (5:1). We also observed that the similar sensitivity of antisera against BPA between the BPA-cBSA group and BHPVA-BSA group.
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Use of tuf as a target for sequence-based identification of Gram-positive cocci of the genus Enterococcus, Streptococcus, coagulase-negative Staphylococcus, and Lactococcus.
Ann. Clin. Microbiol. Antimicrob.
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Accurate identification of isolates belonging to genus Enterococcus, Streptococcus, coagulase-negative Staphylococcus, and Lactococcus at the species level is necessary to provide a better understanding of their pathogenic potential, to aid in making clinical decisions, and to conduct epidemiologic investigations,especially when large blind samples must be analyzed. It is useful to simultaneously identify species in different genera using a single primer pair.
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Rabies virus nucleoprotein expressed in silkworm pupae at high-levels and evaluation of immune responses in mice.
J. Biotechnol.
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Rabies is one of the most fatal zoonotic diseases in developing countries, where a safe, cheap and effective vaccine against the disease remains unaffordable. In this paper, we describe a new silkworm-baculovirus expression system to express the nucleoprotein (N) gene of rabies virus and evaluation of the immune response in BALB/c mice. A recombinant baculovirus -rBmNPV(RV-N) carrying the N gene of rabies virus Evelyn Rokitniki Abelseth (ERA) strain was constructed and the N protein expression was evaluated in Bombyx mori (BmN) cells and silkworm pupae by immunofluorescence staining, Western blots and enzyme-linked immunosorbent assay (ELISA). The immune response to vaccines was evaluated based on serum IgG antibody titers and challenge experiments. The study revealed that N protein of rabies virus can be highly expressed in silkworm baculovirus expression system and the vaccine of N antigen presents a promising approach for the prevention of rabies virus.
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Growth factors from tumor microenvironment possibly promote the proliferation of glioblastoma-derived stem-like cells in vitro.
Pathol. Oncol. Res.
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Glioblastoma multiform is a lethal brain glial tumor characterized by low survival and high recurrence, partially attributed to the glioblastoma stem cells according to recent researches. Microenvironment or niche in tumor tissue is believed to provide essential support for the aberrant growth of tumor stem cells. In order to explore the effect of growth factors in tumor microenvironment on glioblastoma stem cells behavior, glioblastoma-derived stem-like cells (GDSCs) were isolated from adult human glioblastoma specimen with antibody against surface marker CD133 and were co-cultured with various tumor cells including U87MG cells, unsorted glioblastoma tumor cells, CD133(-) cells and normal rat primary astrocytes. Results suggested that tumor cells could promote GDSCs proliferation while non-tumor cells could not, and several growth factors were exclusively detected in the co-culture system with tumor cells. It was concluded that growth factors derived from tumor microenvironment possibly contributed to the uncontrolled proliferation of GDSCs.
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Development of a foot-and-mouth disease virus serotype A empty capsid subunit vaccine using silkworm (Bombyx mori) pupae.
PLoS ONE
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Foot-and-mouth disease (FMD) is a highly contagious disease of cloven-hoofed animals that inflicts severe economic losses in the livestock industry. In 2009, FMDV serotype A caused outbreaks of FMD in cattle in China. Although an inactivated virus vaccine has proven effective to control FMD, its use may lead to new disease outbreaks due to a possible incomplete inactivation of the virus during the manufacturing process. Here, we expressed the P1-2A and the 3C coding regions of a serotype A FMDV field isolate in silkworm pupae (Bombyx mori) and evaluated the immunogenicity of the expression products. Four of five cattle vaccinated with these proteins developed high titers of FMDV-specific antibody and were completely protected against virulent homologous virus challenge with 10,000 50% bovine infectious doses (BID(50)). Furthermore, the 50% bovine protective dose (PD(50)) test was performed to assess the bovine potency of the empty capsid subunit vaccine and was shown to achieve 4.33 PD(50) per dose. These data provide evidence that silkworm pupae can be used to express immunogenic FMDV proteins. This strategy might be used to develop a new generation of empty capsid subunit vaccines against a variety of diseases.
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The complete mitochondrial genome of black grunt Hapalogenys nigripinnis.
Mitochondrial DNA
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The complete sequence of the mitochondrial genome of Hapalogenys nigripinnis was determined in this study. The genome was 16,478 bp in size with a base composition of 28.6% A, 15.8% G, 27.4% T, and 28.2% C, containing a typical structure of 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNA genes, and 1 noncoding control region. The genomic composition, organization, and gene order of H. nigripinnis was similar to that obtained in most vertebrates. These results may provide molecular information on the future phylogenetic relationships of H. nigripinnis and its position within the suborder Percoidei.
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Association between serum endogenous secretory receptor for advanced glycation end products and risk of type 2 diabetes mellitus with combined depression in the Chinese population.
Diabetes Technol. Ther.
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The role of the endogenous secretory receptor for advanced glycation end products (esRAGE) in depression of diabetes patients and its clinical significance are unclear. This study investigated the role of serum esRAGE in patients with type 2 diabetes mellitus with depression in the Chinese population.
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Molecular phylogenetic relationships of family Haemulidae (Perciformes: Percoidei) and the related species based on mitochondrial and nuclear genes.
Mitochondrial DNA
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Haemulidae species are morphologically diversified fishes with wondrous and changeable coloration. These species are prevalent in global tropical, subtropical, and temperate inshore reef areas. Previous morphological classification within Haemulidae and some related families was problematic, and no comprehensive molecular evaluation was conducted on these groups. In this study, we revealed the first molecular phylogenetic analysis involving representative species within Haemulidae and the relevant families (Nemipteridae and Teraponidae). The analysis was performed using both mitochondrial DNA (16S rRNA) and nuclear DNA (TMO-4c4) genes to construct maximum parsimony, maximum likelihood, and Bayesian analysis tree topologies. The molecular phylogeny recovered well-resolved relationships within Haemulidae species and the problematic taxa. In the trees, the Haemulidae species (except for genus Hapalogenys) were divided into two distinct sister lineages, including grunts (Haemulinae) and sweetlips (Plectorhynchinae). Hapalogenys was positioned outside the major Haemulidae tribe and formed an independent group, which challenged the traditional taxonomy that Hapalogenys was classified into Haemulidae. The results did agree with most current studies that Hapalogenys was only distantly related to Haemulidae and could potentially be removed from this family. Additionally, the genus Diagramma was observed to be tightly grouped inside the clade Plectorhinchus, indicating its highly close affinity within Plectorhinchus. Regarding interfamily relationships, the phylogenetic constructions suggested distant relationships within two pairs: between Scolopsis and Haemulidae and between Teraponidae and Haemulidae. Scolopsis was revealed to be highly close with Nemipteridae, and Teraponidae was clustered in an independent group, which was in accordance with most current taxonomic studies.
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[Salidroside biosynthesis pathway: the initial reaction and glycosylation of tyrosol].
Sheng Wu Gong Cheng Xue Bao
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Salidroside, the 8-O-beta-D-glucoside of tyrosol, is a novel adaptogenic drug extracted from the medicinal plant Rhodiola sachalinensis A. Bor. Due to the scarcity of R. sachalinensis and its low yield of salidroside, there is great interest in enhancing the production of salidroside by biotechnological process. Glucosylation of tyrosol is thought to be the final step in salidroside biosynthesis. In our related works, three UGT clones were isolated from the roots and the cultured cells. Our intention was to combine the catalytic specificity of these UGTs in vitro in order to change the level of salidroside in vivo by over-expression of the above UGTs. However, as the aglycone substrate of salidroside, the biosynthetic pathway of tyrosol and its regulation are less well understood. The results of related studies revealed that there are two different possibilities for the tyrosol biosynthetic pathway. One possibility is that tyrosol is produced from a p-coumaric acid precursor, which is derived mainly from phenylalanine. The second possibility is that the precursor of tyrosol might be tyramine, which is synthesized from tyrosine. Our previous work demonstrated that over-expression of the endogenous phenylalanine ammonia-lyase gene (PALrs1) and accumulation of p-coumaric acid did not facilitate tyrosol biosynthesis. In contrast, the data presented in our recent work provide in vitro and in vivo evidence that the tyrosine decarboxylase (RsTyrDC) is most likely to have an important function in the initial reaction of the salidroside biosynthesis pathway in R. Sachalinensis.
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Pyronepolyene C-glucosides with NF-?B inhibitory and anti-influenza A viral (H1N1) activities from the sponge-associated fungus Epicoccum sp. JJY40.
Bioorg. Med. Chem. Lett.
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A new pyronepolyene C-glucoside, named iso-D8646-2-6 (1) together with the known related compound D8646-2-6 (2), was isolated from the sponge-associated fungus Epicoccum sp. JJY40. They showed NF-?B inhibitory and anti-influenza A viral (H1N1) activities.
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JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

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In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.