Synergistic Antitumor Effect of Dichloroacetate in Combination with 5-Fluorouracil in Colorectal Cancer

Dichloroacetate (DCA), an inhibitor of pyruvate dehydrogenase kinase (PDK), has been recently demonstrated as a promising nontoxic antineoplastic agent that promotes apoptosis of cancer cells. In the present study, we aimed to investigate the antitumor effect of DCA combined with 5-Fluorouracil (5-FU) on colorectal cancer (CRC) cells. Four human CRC cell lines were treated with DCA or 5-FU, or a combination of DCA and 5-FU. The cell viability was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The interaction between DCA and 5-FU was evaluated by the median effect principle. Immunocytochemistry with bromodeoxyuridine (BrdU) was carried out to determine the proliferation of CRC cells. Cell cycle and apoptosis were measured by flow cytometry, and the expression of apoptosis-related molecules was assessed by western blot. Our results demonstrated that DCA inhibited the viability of CRC cells and had synergistic antiproliferation in combination with 5-FU. Moreover, compared with 5-FU alone, the apoptosis of CRC cells treated with DCA and 5-FU was enhanced and demonstrated with the changes of Bcl-2, Bax, and caspase-3 proteins. Our results suggest that DCA has a synergistic antitumor effect with 5-FU on CRC cell lines in vitro

Sr-Nd isotopic geochemistry of Holocene sediments from the South Yellow Sea: Implications for provenance and monsoon variability

Elemental geochemical and Sr-Nd isotopic signatures are used to decipher terrigenous sediments provenances and transport mechanisms in the South Yellow Sea during the Holocene. Sr-87/Sr-86 ratios in the Chinese and Korean riverine sediments overlap each other, whereas epsilon Nd values of Korean riverine sediments are generally less radiogenic in comparison to the Changjiang and Huanghe. Moreover, eNd values of these two large rivers appear unaffected by mineral sorting and are relative stable during the Holocene. We propose a three end-members (i.e., the Changjiang, the Huanghe, and Korean rivers) mixing model to explain sediment provenances in the Central Yellow Sea Mud (CYSM). Mixing calculations show that the Huanghe is the major sediment contributor to the CYSM before similar to 8 ka (thousand years before 1950 CE), whereas the Changjiang has become the predominant sediment source after similar to 8 ka. Holocene changes in riverine sediment supplies to the CYSM are closely related to the oceanic circulation, monsoon climate, and drainage changes. After examining several hypotheses to explain the variations in Sr-87/Sr-86 ratios of Core YSC-1 during the past similar to 8 kyr, we tentatively attribute that to changes in the erosion patterns of the Changjiang Basin. This in turn is associated with the asynchronous evolution of monsoon precipitation in the upper (Indian Summer Monsoon) and middle-lower Changjiang (East Asian Summer Monsoon). Therefore, our results highlight significant influences of monsoon climate on erosion patterns within the Changjiang catchment at millennial timescales

Molecular characterization of cathepsin B from Clonorchis sinensis excretory/secretory products and assessment of its potential for serodiagnosis of clonorchiasis

<p>Abstract</p> <p>Background</p> <p>Cathepsin cysteine proteases play multiple roles in the life cycle of parasites such as food uptake, immune invasion and pathogenesis, making them valuable targets for diagnostic assays, vaccines and drugs. The purpose of this study was to identify a cathepsin B of <it>Clonorchis sinensis </it>(<it>Cs</it>CB) and to investigate its diagnostic value for human helminthiases.</p> <p>Results</p> <p>The predicted amino acid sequence of the cathepsin B of <it>C. sinensis </it>shared 63%, 52%, 50% identity with that of <it>Schistosoma japonicum</it>, <it>Homo sapiens </it>and <it>Fasciola hepatica</it>, respectively. Sequence encoding proenzyme of <it>Cs</it>CB was overexpressed in <it>Escherichia coli</it>. Reverse transcription PCR experiments revealed that <it>Cs</it>CB transcribed in both adult worm and metacercaria of <it>C. sinensis</it>. <it>Cs</it>CB was identified as a <it>C. sinensis </it>excretory/secretory product by immunoblot assay, which was consistent with immunohistochemical localization showing that <it>Cs</it>CB was especially expressed in the intestine of <it>C. sinensis </it>adults. Both ELISA and western blotting analysis showed recombinant <it>Cs</it>CB could react with human sera from clonorchiasis and other helminthiases.</p> <p>Conclusions</p> <p>Our findings revealed that secreted CsCB may play an important role in the biology of C. sinensis and could be a diagnostic candidate for helminthiases.</p

The secreted FolAsp aspartic protease facilitates the virulence of Fusarium oxysporum f. sp. lycopersici

Pathogens utilize secretory effectors to manipulate plant defense. Fusarium oxysporum f. sp. lycopersici (Fol) is the causal agent of Fusarium wilt disease in tomatoes. We previously identified 32 secreted effector candidates by LC-MS analysis. In this study, we functionally identified one of the secreted proteins, FolAsp, which belongs to the aspartic proteases (Asp) family. The FolAsp was upregulated with host root specifically induction. Its N-terminal 1–19 amino acids performed the secretion activity in the yeast system, which supported its secretion in Fol. Phenotypically, the growth and conidia production of the FolAsp deletion mutants were not changed; however, the mutants displayed significantly reduced virulence to the host tomato. Further study revealed the FolAsp was localized at the apoplast and inhibited INF1-induced cell death in planta. Meanwhile, FolAsp could inhibit flg22-mediated ROS burst. Furthermore, FolAsp displayed protease activity on host protein, and overexpression of FolAsp in Fol enhanced pathogen virulence. These results considerably extend our understanding of pathogens utilizing secreted protease to inhibit plant defense and promote its virulence, which provides potential applications for tomato improvement against disease as the new drug target

Identification and Characterization of Paramyosin from Cyst Wall of Metacercariae Implicated Protective Efficacy against Clonorchis sinensis Infection

Human clonorchiasis has been increasingly prevalent in recent years and results in a threat to the public health in epidemic regions, motivating current strategies of vaccines to combat Clonorchis sinensis (C. sinensis). In this study, we identified C. sinensis paramyosin (CsPmy) from the cyst wall proteins of metacercariae by proteomic approaches and characterized the expressed recombinant pET-26b-CsPmy protein (101 kDa). Bioinformatics analysis indicated that full-length sequences of paramyosin are conserved in helminthes and numerous B-cell/T-cell epitopes were predicted in amino acid sequence of CsPmy. Western blot analysis showed that CsPmy was expressed at four life stages of C. sinensis, both cyst wall proteins and soluble tegumental components could be probed by anti-CsPmy serum. Moreover, immunolocalization results revealed that CsPmy was specifically localized at cyst wall and excretory bladder of metacercaria, as well as the tegument, oral sucker and vitellarium of adult worm. Both immunoblot and immunolocalization results demonstrated that CsPmy was highly expressed at the stage of adult worm, metacercariae and cercaria, which could be supported by real-time PCR analysis. Both recombinant protein and nucleic acid of CsPmy showed strong immunogenicity in rats and induced combined Th1/Th2 immune responses, which were reflected by continuous high level of antibody titers and increased level of IgG1/IgG2a subtypes in serum. In vaccine trials, comparing with control groups, both CsPmy protein and DNA vaccine exhibited protective effect with significant worm reduction rate of 54.3% (p<0.05) and 36.1% (p<0.05), respectively. In consistence with immune responses in sera, elevated level of cytokines IFN-γ and IL-4 in splenocytes suggested that CsPmy could induce combined cellular immunity and humoral immunity in host. Taken together, CsPmy could be a promising vaccine candidate in the prevention of C. sinensis regarding its high immunogenicity and surface localization

Transcriptome Analysis of Female and Male Xiphophorus maculatus Jp 163 A

Background: Xiphophorus models are important for melanoma, sex determination and differentiation, ovoviviparity and evolution. To gain a global view of the molecular mechanism(s) whereby gene expression may influence sexual dimorphism in Xiphophorus and to develop a database for future studies, we performed a large-scale transcriptome study. Methodology/Principal Findings: The 454-FLX massively parallel DNA sequencing platform was employed to obtain 742,771 and 721,543 reads from 2 normalized cDNA libraries generated from whole adult female and male X. maculatus Jp 163 A, respectively. The reads assembled into 45,538 contigs (here, a "contig" is a set of contiguous sequences), of which, 11,918 shared homology to existing protein sequences. These numbers estimate that the contigs may cover 53% of the total number of Xiphophorus transcriptome. Putative translations were obtained for 11,918 cDNA contigs, of which, 3,049 amino acid sequences contain Pfam domains and 11,064 contigs encode secretory proteins. A total of 3,898 contigs were associated with 2,781 InterPro (IPR) entries and 5,411 contigs with 132 KEGG (Kyoto Encyclopedia of Genes and Genomes) pathways. There were 10,446 contigs annotated with 69,778 gene ontology (GO) terms and the three corresponding organizing principles. Fifty-four potential sex differentially expressed genes have been identified from these contigs. Eight and nine of these contigs were confirmed by real-time PCR as female and male predominantly expressed genes respectively. Based on annotation results, 34 contigs were predicted to be differentially expressed in male and female and 17 of them were also confirmed by real-time PCR. Conclusions/Significance: This is the first report of an annotated overview of the transcriptome of X. maculatus and identification of sex differentially expressed genes. These data will be of interest to researchers using the Xiphophorus model. This work also provides an archive for future studies in molecular mechanisms of sexual dimorphism and evolution, and can be used in comparative studies of other fish