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Find video protocols related to scientific articles indexed in Pubmed.
Validity of 18F-fluorodeoxyglucose Positron Emission Tomography/Computed Tomography for Pretreatment Evaluation of Patients With Cervical Carcinoma: A Retrospective Pathology-Matched Study.
Int. J. Gynecol. Cancer
PUBLISHED: 10-01-2014
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The aim of this study was to evaluate the validity of F-fluorodeoxyglucose (F-FDG) positron emission tomography (PET)/computed tomography (CT) for pretreatment evaluation of patients with cervical carcinoma.
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Expression profiles of glyceraldehyde-3-phosphate dehydrogenase from Clonorchis sinensis: a glycolytic enzyme with plasminogen binding capacity.
Parasitol. Res.
PUBLISHED: 08-28-2014
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Globally, 15-20 million people are infected with Clonorchis sinensis (C. sinensis) which results in clonorchiasis. In China, clonorchiasis is considered to be one of the fastest-growing food-borne parasitic diseases. That more key molecules of C. sinensis are characterized will be helpful to understand biology and pathogenesis of the carcinogenic liver fluke. Glyceraldehyde-3-phosphate dehydrogenases (GAPDHs) from many species have functions other than their catalytic role in glycolysis. In the present study, we analyzed the sequence and structure of GAPDH from C. sinensis (CsGAPDH) by using bioinformatics tools and obtained its recombinant protein by prokaryotic expression system, to learn its expression profiles and molecular property. CsGAPDH could bind to human intrahepatic biliary epithelial cell in vivo and in vitro by the method of immunofluorescence assays. CsGAPDH also disturbed in lumen of biliary tract near to the parasite in the liver of infected rat. Western blotting analysis together with immunofluorescence assay indicated that CsGAPDH was a component of excretory/secretory proteins (CsESPs) and a surface-localized protein of C. sinensis. Quantitative real-time PCR (Q-PCR) and Western blotting demonstrated that CsGAPDHs are expressed at the life stages of adult worm, metacercaria, and egg, but the expression levels were different from each other. Recombinant CsGAPDH (rCsGAPDH) was confirmed to have the capacity to catalyze the conversion of glyceraldehyde 3-phosphate to D-glycerate 1,3-bisphosphate which was inhibited by AMP in a dose-dependent manner. In addition, rCsGAPDH was able to interact with human plasminogen in a dose-dependent manner by ELISA. The interaction could be inhibited by lysine. The plasminogen binding capacity of rCsGAPDH along with the distribution of CsGAPDH in vivo and in the liver of C. sinensis-infected rat hinted that surface-localized CsGAPDH might play an important role in host invasion of the worm besides its glycolytic activity. Our work will be a cornerstone for getting more messages about CsGAPDH and its role in biology and parasitism of C. sinensis.
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Fluvastatin attenuated the effect of expression of ?1 integrin in PAN-treated podocytes by inhibiting reactive oxygen species.
Mol. Cell. Biochem.
PUBLISHED: 06-12-2014
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It is well accepted that ?1 integrin plays a key role in maintaining normal podocytes form and functions; however, its mechanism of the potential protective effect remains unclear. Furthermore, the investigation and understanding of the non-lipid-dependent renal protection of Statins in addition to well-known lipid-lowering effect may provide the therapeutic utility and ultimately improve clinical outcome for patients with renal diseases. In the present study, we investigated the effect and mechanism of fluvastatin (FLV) on the expression of ?1 integrin in puromycin aminonucleoside (PAN)-treated podocytes in vitro. Cultured human podocytes were treated with PAN, and/or different concentrations of FLV (1 × 10(-8)-1 × 10(-5 )mol/l), superoxide dismutase (SOD), or H2O2, respectively. The expression of ?1 integrin and reactive oxygen species (ROS) in human podocytes under each experimental condition was evaluated by western blot, RT-PCR, and 2'7'-dichlorofluorescein 3'6'-diacetate, respectively. The viability of podocytes was also assessed by MTT colorimetry in the present study. The expression of ?1 integrin was significantly decreased, and the synthesis of ROS was significantly increased in podocytes following either PAN or H2O2 treatment (p < 0.05). The up-regulation of ?1 integrin and down-regulation of ROS were also observed in PAN-treated podocytes following lower concentrations of FLV or SOD treatment (p < 0.05, respectively). The cytotoxicity data derived from MTT assay revealed that lower podocyte viability was found in the presence of higher concentrations of FLV, PAN, or H2O2. Lower concentration of FLV or SOD can protect podocytes from being impaired by PAN treatment. FLV attenuated the podocyte injury induced by PAN and increased the production of ?1 integrin in human podocytes in vitro. This underlying mechanism of FLV may be through inhibiting the activity of ROS in human podocytes.
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[Triptolide inhibits the inflammatory response of monocytes from rheumatoid arthritis patients by regulating miR-155].
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi
PUBLISHED: 06-10-2014
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To explore the anti-inflammatory effect of triptolide (TPT) by regulating miR-155 in monocytes pre-stimulated by lipopolysaccharide (LPS) from rheumatoid arthritis (RA) patients.
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Farnesol induces apoptosis-like cell death in the pathogenic fungus Aspergillus flavus.
Mycologia
PUBLISHED: 06-03-2014
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Farnesol (FOH) is known to induce apoptosis in some fungi and mammalian cells. We treated Aspergillus flavus, one of the leading causes of human invasive aspergillosis and a key producer of the most potent naturally occurring hepatocarcinogenic compounds, with FOH to assess its effect on the viability of the fungus. FOH strongly inhibited germination and growth of A. flavus and induced markers for apoptosis including nuclear condensation, phosphatidylserine (PS) externalization, DNA fragmentation and intracellular reactive oxygen species (ROS) generation, metacaspase activation and abnormal cellular ultrastructure. Moreover, FOH-induced apoptosis in A. flavus was inhibited by the broad-spectrum caspase inhibitor Z-VAD-fmk and partially inhibited by the ROS scavenger l-proline, which suggests that FOH induces apoptosis in A. flavus via a mechanism involving metacaspase activation and ROS production.
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Characterization of the secreted cathepsin B cysteine proteases family of the carcinogenic liver fluke Clonorchis sinensis.
Parasitol. Res.
PUBLISHED: 05-02-2014
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Clonorchis sinensis excretory/secretory products (ESP) have gained high attentions because of their potential to be vaccine candidates and drug targets in C. sinensis prevention. In this study, we extensively profiled the characteristics of four C. sinensis cathepsin B cysteine proteases (CsCB1, CsCB2, CsCB3, and CsCB4). Bioinformatics analysis showed all CsCBs contained signal peptides at the N-terminal. Functional domains and residues were found in CsCB sequences. We expressed four CsCBs and profiled immune responses followed by vaccine trials. Recombinant CsCBs could induce high IgG titers, indicating high immunogenicity of CsCB family. Additionally, ELISA results showed that both IgG1 and IgG2a levels apparently increased post-immunization with all four CsCBs, showing that combined Th1/Th2 immune responses were triggered by CsCB family. Both Real-time polymerase chain reaction (RT-PCR) and Western blotting confirmed that four CsCBs have distinct expression patterns in C. sinensis life stages. More importantly, we validated our hypothesis that CsCBs were C. sinensis excretory/secretory products. CsCBs could be recognized by C. sinensis-infected sera throughout the infection period, indicating that secreted CsCBs are immune triggers during C. sinensis infection. The protective effect was assessed by comparing the worm burden and egg per gram (EPG) between CsCB group and control group, showing that worm burden (P?
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Vitamin C mitigates oxidative stress and tumor necrosis factor-alpha in severe community-acquired pneumonia and LPS-induced macrophages.
Mediators Inflamm.
PUBLISHED: 04-24-2014
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Oxidative stress is an important part of host innate immune response to foreign pathogens. However, the impact of vitamin C on oxidative stress and inflammation remains unclear in community-acquired pneumonia (CAP). We aimed to determine the effect of vitamin C on oxidative stress and inflammation. CAP patients were enrolled. Reactive oxygen species (ROS), DNA damage, superoxide dismutases (SOD) activity, tumor necrosis factor-alpha (TNF-?), and IL-6 were analyzed in CAP patients and LPS-stimulated macrophages cells. MH-S cells were transfected with RFP-LC3 plasmids. Autophagy was measured in LPS-stimulated macrophages cells. Severe CAP patients showed significantly increased ROS, DNA damage, TNF-?, and IL-6. SOD was significantly decreased in severe CAP. Vitamin C significantly decreased ROS, DNA damage, TNF-?, and IL-6. Vitamin C inhibited LPS-induced ROS, DNA damage, TNF-?, IL-6, and p38 in macrophages cells. Vitamin C inhibited autophagy in LPS-induced macrophages cells. These findings indicated that severe CAP exhibited significantly increased oxidative stress, DNA damage, and proinflammatory mediator. Vitamin C mitigated oxidative stress and proinflammatory mediator suggesting a possible mechanism for vitamin C in severe CAP.
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Molecular and biochemical characterizations of three fructose-1,6-bisphosphate aldolases from Clonorchis sinensis.
Mol. Biochem. Parasitol.
PUBLISHED: 04-13-2014
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Fructose-1,6-bisphosphate aldolase (FbA) is a ubiquitous enzyme in glycolysis. In the present study, we screened out three distinct genes encoding FbA isozymes (CsFbAs, CsFbA-1/2/3) from Clonorchis sinensis (C. sinensis) and characterized their sequences and structures profiles as well as biochemical properties. The amino acid sequences of CsFbAs shared homology with those of Class I FbAs from other species. The putative quaternary structures revealed that CsFbA-2 and CsFbA-3 were tetramers, while CsFbA-1 was dimer. Recombinant CsFbA-2 and CsFbA-3 (rCsFbA-2/3) were confirmed to be Class I FbAs for their stable enzymatic activities in the presence of EDTA or metal ions. However, recombinant CsFbA-1 (rCsFbA-1) did not show the catalytic activity, which might be due to the inappropriate fold and interaction between its subunits. Both rCsFbA-2 and rCsFbA-3 showed similar enzymatic properties such as optimal temperatures and broad pH ranges that similar to human FbA isozymes. They showed relatively higher affinities for fructose-1,6-bisphosphate (FBP) than fructose-1-phosphate (F-1-P). Their kcat ratios of FBP to F-1-P were in accordance with those of human FbA-A or C. In addition, CsFbAs were differentially transcribed in the developmental stages of C. sinensis, suggesting their essential roles throughout the life stages. Extensive distribution of CsFbAs in adult worms indicated that ubiquitous activities of CsFbAs took place in these organs. Collectively, these results suggested that long-term parasitic environment might adapt these isozymes similar to host FbAs for metabolic requirement. Our study will provide new insight into CsFbAs in the glycometabolism of C. sinensis and relationship between the host and the parasite.
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Proteomic identification of potential Clonorchis sinensis excretory/secretory products capable of binding and activating human hepatic stellate cells.
Parasitol. Res.
PUBLISHED: 03-28-2014
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Epidemiological and experimental evidence demonstrated that Clonorchis sinensis is an important risk factor of hepatic fibrosis and cholangiocarcinoma. C. sinensis excretory/secretory products (CsESPs) are protein complex including proteases, antioxidant enzymes, and metabolic enzymes, which may contribute to pathogenesis of liver fluke-associated hepatobiliary diseases. However, potential CsESP candidates involved into hepatic fibrosis and cholangiocarcinoma still remain to be elucidated. In the present study, we performed proteomic identification of CsESP candidates capable of binding and activating human hepatic stellate cell line LX-2. Immunofluorescence analysis confirmed the interaction of CsESPs with LX-2 cell membrane. LX-2 cells could be stimulated by CsESPs from 24 h post incubation (p?
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Highly enantioselective Michael addition of diethyl malonate to chalcones catalyzed by cinchona alkaloids-derivatived bifunctional tertiary amine-thioureas bearing multiple hydrogen-bonding donors.
Org. Biomol. Chem.
PUBLISHED: 03-28-2014
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Chalcones are still challenge substrates in Michael reactions, and only limited success has been achieved. This work describes a highly enantioselective Michael addition of diethyl malonate with chalcones catalyzed by cinchona alkaloids-derivatived bifunctional tertiary amine-thioureas bearing multiple hydrogen-bonding donors.
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Biochemical and immunological characterization of annexin B30 from Clonorchis sinensis excretory/secretory products.
Parasitol. Res.
PUBLISHED: 03-19-2014
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Clonorchis sinensis has been classified as group I biological carcinogen for cholangiocarcinoma by the World Health Organization. Biological studies on excretory/secretory products (ESPs) enabled us to understand the pathogenesis mechanism of C. sinensis and develop new strategies for the prevention of clonorchiasis. In this study, sequence analysis showed that annexin B30 from C. sinensis (CsANXB30) is composed of four annexin repeats which were characterized by type II and III Ca(2+)-binding sites or KGD motif with the capability of Ca(2+)-binding. In addition, immunoblot assay revealed that recombinant CsANXB30 (rCsANXB30) could be recognized by the sera from rats infected with C. sinensis and the sera from rats immunized by CsESPs. Real-time PCR showed that its transcriptional level was the highest at the stage of metacercaria. Immunofluorescence assay was employed to confirm that CsANXB30 was distributed in the tegument, intestine, and egg of adult worms, as well as the tegument and vitellarium of metacercaria. rCsANXB30 was able to bind phospholipid in a Ca(2+)-dependent manner and human plasminogen in a dose-dependent manner. Moreover, cytokine and antibody measurements indicated that rats subcutaneously immunized with rCsANXB30 developed a strong IL-10 production in spleen cells and a high level of IgG1 isotype, indicating that rCsANXB30 could trigger specific humoral and cellular immune response in rats. The present results implied that CsANXB30 might be involved in a host-parasite interaction and affected the immune response of the host during C. sinensis infection.
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Abnormal platelet kinetics are detected before the occurrence of thrombocytopaenia in HBV-related liver disease.
Liver Int.
PUBLISHED: 03-12-2014
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Thrombocytopaenia is a frequent feature in patients with HBV-related liver disease. Its underlying mechanism is not fully understood. Multiple factors might contribute to the development of thrombocytopaenia. In this study, we investigated the reticulated platelets (RP), glycocalicin (GC), serum thrombopoietin (TPO) and platelet glycoprotein (GP) in different stages of the disease.
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Molecular characterization of Clonorchis sinensis secretory myoglobin: delineating its role in anti-oxidative survival.
Parasit Vectors
PUBLISHED: 02-22-2014
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Clonorchiasis is a globally important, neglected food-borne disease caused by Clonorchis sinensis (C. sinensis), and it is highly related to cholangiocarcinoma and hepatocellular carcinoma. Increased molecular evidence has strongly suggested that the adult worm of C. sinensis continuously releases excretory-secretory proteins (ESPs), which play important roles in the parasite-host interactions, to establish successful infection and ensure its own survival. Myoglobin, a hemoprotein, is present in high concentrations in trematodes and ESPs. To further understand the biological function of CsMb and its putative roles in the interactions of C. sinensis with its host, we explored the molecular characterization of CsMb in this paper.
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Identification, immunolocalization, and immunological characterization of nitric oxide synthase-interacting protein from Clonorchis sinensis.
Parasitol. Res.
PUBLISHED: 02-14-2014
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Recently, accumulating evidences indicate that nitric oxide (NO) is a potent mediator with diverse roles in regulating cellular functions, signaling pathways, and variety of pathological processes. In the present study, using data from the published genomic for Clonorchis sinensis (C. sinensis), we investigated a gene encoding nitric oxide synthase-interacting protein (NOSIP) of C. sinensis. Recombinant CsNOSIP (rCsNOSIP) was expressed and purified from Escherichia coli BL21. The open reading frame of CsNOSIP comprises 867 bp which encodes 289 amino acids and shares 72.9, 45.2, 47, 46.4, and 45.8% identity with NOSIP from Schistosoma mansoni, Xenopus laevis, Rattus norvegicus, Mus musculus, and Homo sapiens, respectively. Bioinformatics analysis suggested that the full-length sequence contains an eNOS-interacting domain and numerous B-cell epitopes. Quantitative RT-PCR indicated that CsNOSIP differentially transcribed throughout the adult worms, metacercariae, and egg stages of C. sinensis, and were highly expressed in the adult worms. Moreover, western blot analysis showed that the rCsNOSIP could be detected by the serum from BALB/c mice infected with C. sinensis and the serum from BALB/c mice immunized with excretory/secretory products (ESPs). Furthermore, immunolocalization assay showed that CsNOSIP was specifically localized in the intestine, vitellarium, and eggs of adult worm. Both immunoblot and immunolocalization results demonstrated that CsNOSIP was one component of ESPs of C. sinensis, which could be supported by SignalP analysis. Moreover, analysis of the antibody subclass and cytokine profile demonstrated that subcutaneously immunized BALB/c mice with rCsNOSIP could significantly enhance serum IgG1 level and up-regulate expression of IL-4 and IL-6 in the splenocytes. Our results suggested that CsNOSIP was an important antigen exposed to host immune system and probably involved in immune regulation of host by inducing Th2-polarized immune response.
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Molecular characterization and serological reactivity of a vacuolar ATP synthase subunit ?-like protein from Clonorchis sinensis.
Parasitol. Res.
PUBLISHED: 01-28-2014
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The vacuolar ATPase enzyme complex (V-ATPase) pumps protons across membranes, energized by hydrolysis of ATP. Extensive investigations on structural and biochemical features of these molecules have implied their importance in the physiological process. In this study, a full-length sequence encoding a vacuolar ATP synthase subunit ?-like protein of Clonorchis sinensis (CsATP-?) was isolated from our cDNA library. The hypothetical 226 amino acid sequence shared 76% identity with ATP-? proteins of Schistosoma japonicum and above 55% identity with ATP-? proteins from human and other eukaryotes. Characteristic Asp??? amino acid residues and seven B-cell epitopes were predicted in this sequence. The complete coding sequence of the gene was expressed in Escherichia coli. Recombinant CsATP-? (rCsATP-?) protein could be probed by anti-rCsATP-? rat serum and C.sinensis-infected human serum in Western blotting experiment, indicating that it is an antigen of strong antigenicity. The high level of antibody titers (1:204,800) showed that CsATP-? has a powerful immunogenicity. Both the increased level and the change trend of IgG1/IgG2a subtypes in serum showed that the rCsATP-? can induce strong combined Th1/Th2 immune responses in rats and stimulate the immune response changes to the dominant Th2 from Th1 along with long time infection. The results of immunoblot and immunolocalization demonstrated that CsATP-? was consecutively expressed at various developmental stages of the parasite, which was supported by real-time PCR analysis. In immunohistochemistry, CsATP-? was localized on the intestine, vitellarium, and testicle of an adult worm and excretory bladder of metacercaria, implying that CsATP-? may relate to energy intake and metabolism. This fundamental study would contribute to further researches that are related to growth and development and immunomodulation of C. sinensis.
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Surface display of Clonorchis sinensis enolase on Bacillus subtilis spores potentializes an oral vaccine candidate.
Vaccine
PUBLISHED: 01-06-2014
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Clonorchis sinensis (C. sinensis) infections remain the common public health problem in freshwater fish consumption areas. New effective prevention strategies are still the urgent challenges to control this kind of foodborne infectious disease. The biochemical importance and biological relevance render C. sinensis enolase (Csenolase) as a potential vaccine candidate. In the present study, we constructed Escherichia coli/Bacillus subtilis shuttle genetic engineering system and investigated the potential of Csenolase as an oral vaccine candidate for C. sinensis prevention in different immunization routes. Our results showed that, compared with control groups, both recombinant Csenolase protein and nucleic acid could induce a mixed IgG1/IgG2a immune response when administrated subcutaneously (P<0.001), intraperitoneally (P<0.01) and intramuscularly (P<0.001) with worm reduction rate of 56.29%, 15.38% and 37.42%, respectively. More importantly, Csenolase could be successfully expressed as a fusion protein (55kDa) on B. subtilis spore indicated by immunoblot and immunofluorescence assays. Killed spores triggered reactive Th1/Th2 immune response and exhibited protective efficacy against C. sinensis infection. Csenolase derived oral vaccine conferred worm reduction rate and egg reduction rate at 60.07% (P<0.001) and 80.67% (P<0.001), respectively. The shuttle genetic engineering system facilitated the development of oral vaccine with B. subtilis stably overexpressing target protein. Comparably vaccinal trails with Csenolase in different immunization routes potentialize Csenolase an oral vaccine candidate in C. sinensis prevention.
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RNAi-mediated silencing of enolase confirms its biological importance in Clonorchis sinensis.
Parasitol. Res.
PUBLISHED: 01-03-2014
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Clonorchis sinensis (C. sinensis) infection is still a common public health problem in freshwater fish consumption areas in Asian countries. More molecular evidence are required to speed up the prevention strategies to control this kind of infectious disease. In the present study, to confirm the biological importance of Csenolase followed by our previous observations of the key metabolic enzyme, we explored the RNA silence effect of the Csenolase-derived RNA interference (RNAi) in C. sinensis. The extramembranous region aa105-226 was selected as the target sequence of RNA silence. Csenolase-derived double strand RNA (dsRNA-Csenolase, 366 bp) was synthetized and delivered into C. sinensis by soaking approach. The penetration of dsRNA into adult worms and metacercariae was tracked using fluorescently labeled RNA. Western blotting and qRT-PCR experiments were performed to determine dsRNA-Csenolase-silencing effect. Our results showed that, after incubating for 120 h, dsRNA-Csenolase could effectively target and downregulate the expression of Csenolase in both adult worms (P < 0.001) and metacercariae (P < 0.01), resulting in a remarkable killing effect on C. sinensis adult worms (P < 0.01). Fluorescent Cy3-labeled dsRNA was mostly deposited in the uterus and vitellarium of adult worm and in the cyst wall of metacercaria. The present study is the first report of RNAi trials in C. sinensis, allowing further applications in identifying functional genes in C. sinensis.
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25-hydroxyvitamin d3-deficiency enhances oxidative stress and corticosteroid resistance in severe asthma exacerbation.
PLoS ONE
PUBLISHED: 01-01-2014
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Oxidative stress plays a significant role in exacerbation of asthma. The role of vitamin D in oxidative stress and asthma exacerbation remains unclear. We aimed to determine the relationship between vitamin D status and oxidative stress in asthma exacerbation. Severe asthma exacerbation patients with 25-hydroxyvitamin D3-deficiency (V-D deficiency) or 25-hydroxyvitamin D-sufficiency (V-D sufficiency) were enrolled. Severe asthma exacerbation with V-D-deficiency showed lower forced expiratory volume in one second (FEV1) compared to that with V-D-sufficiency. V-D-deficiency intensified ROS release and DNA damage and increased TNF-?, OGG1 and NF?B expression and NF?B phosphorylation in severe asthma exacerbation. Supplemental vitamin D3 significantly increased the rates of FEV1 change and decreased ROS and DNA damage in V-D-deficiency. Vitamin D3 inhibited LPS-induced ROS and DNA damage and were associated with a decline in TNF-? and NF?B in epithelial cells. H2O2 reduces nuclear translocation of glucocorticoid receptors in airway epithelial cell lines. V-D pretreatment enhanced the dexamethasone-induced nuclear translocation of glucocorticoid receptors in airway epithelial cell lines and monocytes from 25-hydroxyvitamin D3-deficiency asthma patients. These findings indicate that V-D deficiency aggravates oxidative stress and DNA damage, suggesting a possible mechanism for corticosteroid resistance in severe asthma exacerbation.
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Sequence analysis and molecular characterization of Clonorchis sinensis hexokinase, an unusual trimeric 50-kDa glucose-6-phosphate-sensitive allosteric enzyme.
PLoS ONE
PUBLISHED: 01-01-2014
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Clonorchiasis, which is induced by the infection of Clonorchis sinensis (C. sinensis), is highly associated with cholangiocarcinoma. Because the available examination, treatment and interrupting transmission provide limited opportunities to prevent infection, it is urgent to develop integrated strategies to prevent and control clonorchiasis. Glycolytic enzymes are crucial molecules for trematode survival and have been targeted for drug development. Hexokinase of C. sinensis (CsHK), the first key regulatory enzyme of the glycolytic pathway, was characterized in this study. The calculated molecular mass (Mr) of CsHK was 50.0 kDa. The obtained recombinant CsHK (rCsHK) was a homotrimer with an Mr of approximately 164 kDa, as determined using native PAGE and gel filtration. The highest activity was obtained with 50 mM glycine-NaOH at pH 10 and 100 mM Tris-HCl at pH 8.5 and 10. The kinetics of rCsHK has a moderate thermal stability. Compared to that of the corresponding negative control, the enzymatic activity was significantly inhibited by praziquantel (PZQ) and anti-rCsHK serum. rCsHK was homotropically and allosterically activated by its substrates, including glucose, mannose, fructose, and ATP. ADP exhibited mixed allosteric effect on rCsHK with respect to ATP, while inorganic pyrophosphate (PPi) displayed net allosteric activation with various allosteric systems. Fructose behaved as a dose-dependent V activator with the substrate glucose. Glucose-6-phosphate (G6P) displayed net allosteric inhibition on rCsHK with respect to ATP or glucose with various allosteric systems in a dose-independent manner. There were differences in both mRNA and protein levels of CsHK among the life stages of adult worm, metacercaria, excysted metacercaria and egg of C. sinensis, suggesting different energy requirements during different development stages. Our study furthers the understanding of the biological functions of CsHK and supports the need to screen for small molecule inhibitors of CsHK to interfere with glycolysis in C. sinensis.
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Molecular Characterization of Severin fromClonorchis sinensis Excretory/Secretory Products and Its Potential Anti-apoptotic Role in Hepatocarcinoma PLC Cells.
PLoS Negl Trop Dis
PUBLISHED: 12-01-2013
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Clonorchiasis, caused by the infection of Clonorchis sinensis (C. sinensis), is a kind of neglected tropical disease, but it is highly related to cholangiocarcinoma and hepatocellular carcinoma (HCC). It has been well known that the excretory/secretory products of C. sinensis (CsESPs) play key roles in clonorchiasis associated carcinoma. From genome and transcriptome of C. sinensis, we identified one component of CsESPs, severin (Csseverin), which had three putative gelsolin domains. Its homologues are supposed to play a vital role in apoptosis resistance of tumour cell.
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Molecular characterization and immune modulation properties of Clonorchis sinensis-derived RNASET2.
Parasit Vectors
PUBLISHED: 09-04-2013
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Clonorchis sinensis (C. sinensis, Cs) is a trematode parasite that often causes chronic cumulative infections in the hepatobiliary ducts of the host and can lead to pathological changes by continuously released excretory/secretory proteins (ESPs). A T2 ribonuclease in trematode ESPs, has been identified as a potent regulator of dendritic cell (DCs) modulation. We wondered whether there was a counterpart present in CsESPs with similar activity. To gain a better understanding of CsESPs associated immune responses, we identified and characterized RNASET2 of C. sinensis (CsRNASET2) in this paper.
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Role of TGF-?1 in production of fibronectin in vascular smooth muscle cells cultured under high-phosphate conditions.
J. Nephrol.
PUBLISHED: 08-24-2013
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Hyperphosphatemia is associated with up-regulation of the extracellular matrix formation in vascular smooth muscle cells (VSMCs) and increased risk of cardiovascular disease. In the present study, the role of transforming growth factor-?1 (TGF-?1) in the production of fibronectin (FN) in vascular smooth muscle cells in high-phosphate environments was evaluated.
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High-glucose-based peritoneal dialysis solution induces the upregulation of VEGF expression in human peritoneal mesothelial cells: The role of pleiotrophin.
Int. J. Mol. Med.
PUBLISHED: 06-10-2013
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The aim of the present study was to investigate the effect of a high-glucose-based peritoneal dialysis solution (HGPDS) on the expression of pleiotrophin (PTN) and vascular endothelial growth factor (VEGF) in human peritoneal mesothelial cells (HPMCs) and the mechanisms through which fluvastatin (Flu) protects the peritoneal membrane in continuous ambulatory peritoneal dialysis (CAPD). HPMCs were cultured with HGPDS, Flu (10-8?10-6 mol/l) and PTN (10?30 nmol/l). The expression of PTN and VEGF was examined at the mRNA and protein level. To define the role of PTN in the regulation of VEGF expression, HPMCs were cultured with HGPDS in the presence or absence of the blocking peptide of PTN. The signaling pathways involved in PTN synthesis induced by HGPDS were also characterized. The phenotypic characteristics of HPMCs were observed under a light microscope. Cell viability was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) colorimetry and the mRNA and protein expression of PTN, VEGF and ERK1/2 was assessed by RT?PCR and the western blot analysis, respectively. Following incubation with HGPDS for 48 h, the morphology of the HPMCs changed from a typical cobblestone?like appearance to a fibroblast?like phenotype. The same alteration in the morphology of the HPMCs also occurred following incubation with 20 nmol/l PTN. Flu (10-6 mol/l), GSK650394 [a competitive inhibitor of serum/glucocorticoid-regulated kinase 1 (SGK1), 10-5 mol/l] and PD98059 (a competitive inhibitor of ERK1/2, 10-5 mol/l) improved the negative changes in cell morphology induced by HGPDS. The results of MTT assay revealed that the reduction in HPMC viability occurred in the groups treated with HGPDS and this reduction was partially restored by Flu, GSK650394 and PD98059. A significant improvement in cell viability, which had been decreased by HGPDS, was observed following treatment with Flu (10-6 mol/l), PD98059 (10-5 mol/l) or GSK650394 (10-5 mol/l) (P<0.05). Compared with the control, the mRNA and protein expression of PTN and VEGF significantly increased in the HPMCs treated with HGPDS (P<0.05). GSK650394 and PD98059 significantly decreased the high mRNA and protein expression levels of PTN and VEGF induced by HGPDS (P<0.05) and Flu had the same inhibitory effect as GSK650394 and PD98059 in a dose?dependent manner (P<0.05). The mRNA and protein expression of VEGF increased following the incubation of HPMCs with 20 nmol/l PTN. By contrast, the mRNA and protein expression levels of VEGF in the HPMCs decreased in the presence of the blocking peptide of PTN. The results from the present study indicated that HGPDS increased the expression of PTN and VEGF in the HPMCs, and this increase was attenuated by Flu, GSK650394 and PD98059. The protein expression of phosphorylated ERK1/2 (p-ERK1/2) was decreased by GSK650394 in the HPMCs treated with HGPDS. Taken together, the protective effects of Flu in HPMCs may be partially achieved through the SGK1?ERK1/2 signaling pathway.
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QTL analysis for early-maturing traits in cotton using two upland cotton (Gossypium hirsutum L.) crosses.
Breed. Sci.
PUBLISHED: 06-01-2013
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Making use of the markers linked closely to QTL for early-maturing traits for MAS (Marker-assisted selection) is an effective method for the simultaneous improvement of early maturity and other properties in cotton. In this study, two F2 populations and their F2:3 families were generated from the two upland cotton (Gossypium hirsutum L.) crosses, Baimian2 × TM-1 and Baimian2 × CIR12. QTL for early-maturing traits were analyzed using F2:3 families. A total of 54 QTL (31 suggestive and 23 significant) were detected. Fourteen significant QTL had the LOD scores not only > 3 but also exceeding permutation threshold. At least four common QTL, qBP-17 for bud period (BP), qGP-17a/qGP-17b (qGP-17) for growth period (GP), qYPBF-17a/qYPBF-17b (qYPBF-17) for yield percentage before frost (YPBF) and qHFFBN-17 for height of first fruiting branch node (HFFBN), were found in both populations. These common QTL should be reliable and could be used for MAS to facilitate early maturity. The common QTL, qBP-17, had a LOD score not only > 3 but also exceeding permutation threshold, explaining 12.6% of the phenotypic variation. This QTL should be considered preferentially in MAS. Early-maturing traits of cotton are primarily controlled by dominant and over-dominant effects.
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Discovering RNA-protein interactome by using chemical context profiling of the RNA-protein interface.
Cell Rep
PUBLISHED: 03-04-2013
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RNA-protein (RNP) interactions generally are required for RNA function. At least 5% of human genes code for RNA-binding proteins. Whereas many approaches can identify the RNA partners for a specific protein, finding the protein partners for a specific RNA is difficult. We present a machine-learning method that scores a proteins binding potential for an RNA structure by utilizing the chemical context profiles of the interface from known RNP structures. Our approach is applicable even when only a single RNP structure is available. We examined 801 mammalian proteins and find that 37 (4.6%) potentially bind transfer RNA (tRNA). Most are enzymes involved in cellular processes unrelated to translation and were not known to interact with RNA. We experimentally tested six positive and three negative predictions for tRNA binding in vivo, and all nine predictions were correct. Our computational approach provides a powerful complement to experiments in discovering new RNPs.
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Polymorphisms of the IL-23R gene are associated with primary immune thrombocytopenia but not with the clinical outcome of pulsed high-dose dexamethasone therapy.
Ann. Hematol.
PUBLISHED: 02-16-2013
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Primary immune thrombocytopenia (ITP) is an autoimmune heterogeneous disorder that is characterized by decreased platelet count. The interleukin-23 receptor (IL-23R) has been identified as a susceptibility gene for the development of multiple autoimmune diseases. To investigate the possible association of IL-23R gene single-nucleotide polymorphisms (SNPs) with ITP and the association with the clinical outcome of pulsed high-dose dexamethasone (HD-DXM) therapy, four SNPs in the IL-23R gene, rs10889677, rs1884444, rs7517847, and rs11209032, were tested in a cohort of 75 ITP subjects and 81 controls by direct sequencing. IL-23R rs1884444 GT/TT variant genotypes were observed to be associated with significantly increased risk of ITP as compared with controls (GT/TT vs. GG: odds ratio (OR) 2.776, 95 % confidence intervals (CI) 1.086-7.090, p?=?0.028). However, other three SNPs revealed no statistically significant differences between patients and controls (rs10889677 CA/AA vs. CC: OR 2.200, 95 % CI 0.727-6.661, p?=?0.155; rs11209032 GA/AA vs. GG: OR 0.747, 95 % CI 0.379-1.472, p?=?0.399; rs7517847 TG/GG vs. TT: OR 1.031, 95 % CI 0.544-1.956, p?=?0.925). Furthermore, IL-23R SNPs revealed no association with clinical outcome of HD-DXM therapy. This study suggests that polymorphism in the IL-23R gene, rs1884444, indicates a significant association with susceptibility to ITP in a recessive genetic model but does not have association with the clinical outcome of HD-DXM therapy.
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Budesonide suppresses pulmonary antibacterial host defense by down-regulating cathelicidin-related antimicrobial peptide in allergic inflammation mice and in lung epithelial cells.
BMC Immunol.
PUBLISHED: 02-04-2013
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Glucocorticoids are widely regarded as the most effective treatment for asthma. However, the direct impact of glucocorticoids on the innate immune system and antibacterial host defense during asthma remain unclear. Understanding the mechanisms underlying this process is critical to the clinical application of glucocorticoids for asthma therapy. After sensitization and challenge with ovalbumin (OVA), BALB/c mice were treated with inhaled budesonide and infected with Pseudomonas aeruginosa (P. aeruginosa). The number of viable bacteria in enflamed lungs was evaluated, and levels of interleukin-4 (IL-4) and interferon-? (IFN-?) in serum were measured. A lung epithelial cell line was pretreated with budesonide. Levels of cathelicidin-related antimicrobial peptide (CRAMP) were measured by immunohistochemistry and western blot analysis. Intracellular bacteria were observed in lung epithelial cells.
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The carcinogenic liver fluke, Clonorchis sinensis: new assembly, reannotation and analysis of the genome and characterization of tissue transcriptomes.
PLoS ONE
PUBLISHED: 01-30-2013
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Clonorchis sinensis (C. sinensis), an important food-borne parasite that inhabits the intrahepatic bile duct and causes clonorchiasis, is of interest to both the public health field and the scientific research community. To learn more about the migration, parasitism and pathogenesis of C. sinensis at the molecular level, the present study developed an upgraded genomic assembly and annotation by sequencing paired-end and mate-paired libraries. We also performed transcriptome sequence analyses on multiple C. sinensis tissues (sucker, muscle, ovary and testis). Genes encoding molecules involved in responses to stimuli and muscle-related development were abundantly expressed in the oral sucker. Compared with other species, genes encoding molecules that facilitate the recognition and transport of cholesterol were observed in high copy numbers in the genome and were highly expressed in the oral sucker. Genes encoding transporters for fatty acids, glucose, amino acids and oxygen were also highly expressed, along with other molecules involved in metabolizing these substrates. All genes involved in energy metabolism pathways, including the ?-oxidation of fatty acids, the citrate cycle, oxidative phosphorylation, and fumarate reduction, were expressed in the adults. Finally, we also provide valuable insights into the mechanism underlying the process of pathogenesis by characterizing the secretome of C. sinensis. The characterization and elaborate analysis of the upgraded genome and the tissue transcriptomes not only form a detailed and fundamental C. sinensis resource but also provide novel insights into the physiology and pathogenesis of C. sinensis. We anticipate that this work will aid the development of innovative strategies for the prevention and control of clonorchiasis.
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Identification and immunological characterization of thioredoxin transmembrane-related protein from Clonorchis sinensis.
Parasitol. Res.
PUBLISHED: 01-29-2013
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Thioredoxin transmembrane related protein (TMX), a member of thioredoxin superfamily, is localized to the endoplasmic reticulum and possesses a thioredoxin-like domain that plays an important role as an oxidoreductase. The functions of TMX in Clonorchis sinensis remain to be elucidated. In this study, we cloned and characterized a novel TMX of C. sinensis (CsTMX). The CsTMX cDNA sequence contained a 414-nucleotide open-reading frame encoding a protein of 137 amino acids. A thioredoxin domain was found in the position of aa21-117 and contained the putative active-site motif Cys-Pro-Ala-Cys. BLASTx analysis showed that CsTMX shared 39-57% amino acid identities with TMX of other organisms. Quantitative RT-PCR analysis demonstrated that CsTMX was differentially transcribed, with the highest level of expression in the adult worm stage and the lowest expression in egg stage. In addition, immunofluorescence assay showed CsTMX was localized in the tegument, vitelline gland, intestine, and intrauterine eggs of adult worm. Besides, immunoblot assay revealed that the recombinant CsTMX (rCsTMX) could be recognized by the sera from rats infected with C. sinensis and the sera from rats immunized by excretory-secretory products. Furthermore, analysis of the antibody isotype profile revealed that rats subcutaneously immunized with rCsTMX developed rCsTMX-specific antibody, which is dominance of IgG2a in sera. Meanwhile, production of IFN-? was elevated strongly in the supernatants of spleen cell. The results collectively indicated that CsTMX might play an important role in the host-parasite interaction, as well as CsTMX probably involved in immunoregulation of host by inducing Th1-type dominated immune response in rats.
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Biochemical characterization and functional analysis of fructose-1,6-bisphosphatase from Clonorchis sinensis.
Mol. Biol. Rep.
PUBLISHED: 01-12-2013
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Fructose-1,6-bisphosphatase (FBPase), a key regulatory enzyme of gluconeogenesis, plays an essential role in metabolism and development of most organisms. To the wealth of available knowledge about FBPase from Clonorchis sinensis (CsFBPase), in this study, the characteristics of CsFBPase and its potential role in pathogenesis of clonorchiasis were investigated. The Km value of CsFBPase was calculated to be 41.9 uM. The optimal temperature and pH of CsFBPase were 37 °C and pH 7.5-8.0, respectively. In addition, Mg(2+) or K(+) played a regulatory role in enzyme activity of CsFBPase. Both transcriptional and translational level of CsFBPase were higher in metacercariae (one of larva stages) than those in adult worm (P < 0.05). CsFBPase were observed to extensively express in the intestine, vitellaria and tegument of adult worms and ubiquitously in metacercariae. Moreover, CsFBPase was confirmed as a component of excretory/secretory products. Consequently, the translocation of CsFBPase could be detected on epithelial cells of bile duct in liver of C. sinensis infected rat. Recombinant CsFBPase can specifically bind to the membrane of human hepatic stellate cell line LX-2 by immunofluorescence analysis and stimulated proliferation and activation of LX-2 which demonstrated by Cell Counting Kit-8 and upregulation of key fibrosis-related factors, such as ?-smooth muscle actin, collagen I and collagen III using qRT-PCR. Thus, we predicated that CsFBPase might be a multifunctional enzyme which played as both regulatory enzyme and virulence factor in pathogenesis of C. sinensis infection.
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Whole transcriptome sequencing enables discovery and analysis of viruses in archived primary central nervous system lymphomas.
PLoS ONE
PUBLISHED: 01-01-2013
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Primary central nervous system lymphomas (PCNSL) have a dramatically increased prevalence among persons living with AIDS and are known to be associated with human Epstein Barr virus (EBV) infection. Previous work suggests that in some cases, co-infection with other viruses may be important for PCNSL pathogenesis. Viral transcription in tumor samples can be measured using next generation transcriptome sequencing. We demonstrate the ability of transcriptome sequencing to identify viruses, characterize viral expression, and identify viral variants by sequencing four archived AIDS-related PCNSL tissue samples and analyzing raw sequencing reads. EBV was detected in all four PCNSL samples and cytomegalovirus (CMV), JC polyomavirus (JCV), and HIV were also discovered, consistent with clinical diagnoses. CMV was found to express three long non-coding RNAs recently reported as expressed during active infection. Single nucleotide variants were observed in each of the viruses observed and three indels were found in CMV. No viruses were found in several control tumor types including 32 diffuse large B-cell lymphoma samples. This study demonstrates the ability of next generation transcriptome sequencing to accurately identify viruses, including DNA viruses, in solid human cancer tissue samples.
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Low Divergence of Clonorchis sinensis in China Based on Multilocus Analysis.
PLoS ONE
PUBLISHED: 01-01-2013
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Clonorchis sinensis, an ancient parasite that infects a number of piscivorous mammals, attracts significant public health interest due to zoonotic exposure risks in Asia. The available studies are insufficient to reflect the prevalence, geographic distribution, and intraspecific genetic diversity of C. sinensis in endemic areas. Here, a multilocus analysis based on eight genes (ITS1, act, tub, ef-1a, cox1, cox3, nad4 and nad5 [4.986 kb]) was employed to explore the intra-species genetic construction of C. sinensis in China. Two hundred and fifty-six C. sinensis isolates were obtained from environmental reservoirs from 17 provinces of China. A total of 254 recognized Multilocus Types (MSTs) showed high diversity among these isolates using multilocus analysis. The comparison analysis of nuclear and mitochondrial phylogeny supports separate clusters in a nuclear dendrogram. Genetic differentiation analysis of three clusters (A, B, and C) showed low divergence within populations. Most isolates from clusters B and C are geographically limited to central China, while cluster A is extraordinarily genetically diverse. Further genetic analyses between different geographic distributions, water bodies and hosts support the low population divergence. The latter haplotype analyses were consistent with the phylogenetic and genetic differentiation results. A recombination network based on concatenated sequences showed a concentrated linkage recombination population in cox1, cox3, nad4 and nad5, with spatial structuring in ITS1. Coupled with the history record and archaeological evidence of C. sinensis infection in mummified desiccated feces, these data point to an ancient origin of C. sinensis in China. In conclusion, we present a likely phylogenetic structure of the C. sinensis population in mainland China, highlighting its possible tendency for biogeographic expansion. Meanwhile, ITS1 was found to be an effective marker for tracking C. sinensis infection worldwide. Thus, the present study improves our understanding of the global epidemiology and evolution of C. sinensis.
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Cordycepin inhibits renal interstitial myofibroblast activation probably by inducing hepatocyte growth factor expression.
J. Pharmacol. Sci.
PUBLISHED: 12-02-2011
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Renal interstitial fibrosis is the common end point of progressive renal diseases leading to the deterioration and eventual loss of renal function. This study investigated the effect and potential mechanism of cordycepin on activation of renal interstitial fibroblast cells. The time and dose-responses of cordycepin in rat renal interstitial fibroblast (NRK-49F) cells were analyzed. The proliferation of NRK-49F and the expression of ?-smooth muscle actin (?-SMA) and fibronectin (FN) were examined. The expression and translocation of Smad proteins also were measured by western blot and indirect immunofluorescence staining. The mRNA level of hepatocyte growth factor (HGF) and the expression of HGF receptor c-Met and its phosphorylation (p-Met) were also detected. Cordycepin suppressed the proliferation of NRK-49F and the expression of ?-SMA and FN induced by transforming growth factor-?1 (TGF-?1). The pretreatment of cordycepin markedly attenuated the nuclear translocation and accumulation of activated Smad2/3 in NRK-49F cells. Furthermore, cordycepin not only increased HGF expression, but also induced HGF secretion, as well as HGF receptor phosphorylation in NRK-49F cells. Cordycepin possesses renoprotective activity through suppression myofibroblast activation. This action is mediated, at least in part, by blocking nuclear translocation and accumulation of activated Smad2/3 protein and up-regulating anti-fibrotic HGF expression and secretion and HGF receptor activation.
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[Clinical study on intervention of spleen-restoring decoction integrating with dormancy hygiene education on subhealthy insomnia of deficiency of both heart and spleen pattern].
Zhongguo Zhong Yao Za Zhi
PUBLISHED: 11-22-2011
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To appraise the clinical efficacy, safety and compliance of the intervention of spleen-restoring decoction combined with dormancy hygiene education and the intervention of spleen-restoring decoction alone on sub-healthy insomnia of deficiency of both the heart and spleen pattern.
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Molecular characterization and expression of a cysteine protease from Clonorchis sinensis and its application for serodiagnosis of clonorchiasis.
Parasitol. Res.
PUBLISHED: 11-02-2011
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Cysteine proteases play essential roles in parasite physiology as well as in host-parasite interactions through their modulation of various biological and pathobiological events. In the present study, a full-length sequence encoding cysteine protease of Clonorchis sinensis (CsCP) was isolated from our adult cDNA library. The open reading frame contains 984 bp encoding 327 amino acids. The present amino acid sequence shared 68% identity with two known CsCP genes and 29-49% identity with that of other species. Bioinformatics analysis showed that conserved domains and characteristic amino acid residues of cysteine proteases were observed in this sequence. Real-time PCR experiments revealed that CsCP was consecutively transcribed in various developmental stages of the parasite, including adult worm, excysted juvenile, metacercaria and egg. Recombinant CsCP (rCsCP) could be probed by rat anti-CsCP serum, rabbit anti-excretory-secretory products (ESP) serum and serum from human infected with Clonorchis sinensis in Western blot. The result of immunolocalization showed that CsCP was mainly located in the oral sucker, excretory bladder and tegument of cercariae and metacercariae, as well as the intestine of adult worm. The rCsCP-based IgG and its isotypes were all detected in sera from human infected with C. sinensis by enzyme-linked immunosorbent assay, and the level of IgG1 is the highest. The receiver-operating characteristic (ROC) analysis was used to determine the most appropriate cut-off value that yielded the high sensitivity (86.96%) and specificity (70.42%). These results revealed that CsCP may play an important role in the biology of C. sinensis and could be a diagnostic candidate for clonorchiasis.
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Preventive effects and mechanisms of rhein on renal interstitial fibrosis in obstructive nephropathy.
Biol. Pharm. Bull.
PUBLISHED: 08-02-2011
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Renal interstitial fibrosis is a common outcome of a variety of chronic renal diseases. Here we evaluated the therapeutic efficacy of rhein on renal interstitial fibrosis induced by unilateral ureteral obstruction (UUO) and investigated the potential mechanisms. Mice underwent UUO, followed by orally administrated rhein (150 mg/kg/d) or control vehicle. Renal interstitial injury and the degree of fibrosis were evaluated by pathological staining and Western blot. The possible mechanisms were studied by Western blot, indirect immune-fluorescence and enzyme-linked immunosorbent assay. Our results showed that rhein therapy markedly ameliorated renal interstitial fibrotic lesions, reduced ?-smooth muscle actin (?-SMA) expression, attenuated deposition of fibronectin (FN). Rhein also suppressed transforming growth factor-?1 (TGF-?1) and its type I receptor expression in obstructed kidneys. In vitro, rhein abolished the ?-SMA and fibronectin expression of rat kidney interstitial fibroblasts cells (NRK-49F) induced by TGF-?1. These observations strongly suggest that rhein is a potent inhibitor of renal interstitial fibrosis, and its therapeutic mechanism is, at least in part, blocking interstitial fibroblasts cells activation.
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Molecular expression and characterization of a novel protein phosphatase 2A gene from Clonorchis sinensis.
Parasitol. Res.
PUBLISHED: 08-01-2011
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Reversible phosphorylation of proteins is a critical mechanism involved in physiological function of organisms, including Clonorchis sinensis. In the present study, One cDNA clone encoding protein phosphatase 2A (CsPP2A) was isolated from a C. sinensis adult cDNA plasmid library. The open reading frame of the novel gene contains 924 bp and encoded a putative protein of 307 amino acids. A similarity analysis showed high homology with Schistosoma japonicum (76.3%) and Homo sapiens (84.4%), respectively. Recombinant CsPP2A (rCsPP2A) was expressed and purified from Escherichia coli BL21 using pET28a (+) as an expression vector. CsPP2A showed higher transcript level in adult worm but excysted metacercaria (P > 0.05), metacercaria (P < 0.05), and egg (P < 0.05) using real-time RT-PCR. Western blotting analysis showed that rCsPP2A could be identified by anti-rCsPP2A rat serum, C. sinensis-infected rat serum, and the serum from the rats immunized with excretory-secretory products of C. sinensis. Immunohistochemical assay showed that CsPP2A was deposited at the egg, the vitellarium of adult worm, and the excretory bladder of metacercaria. Collectively, the results of this study suggested that CsPP2A may be involved in the development of adult and metacercaria of C. sinensis.
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CHFR suppression by hypermethylation sensitizes endometrial cancer cells to paclitaxel.
Int. J. Gynecol. Cancer
PUBLISHED: 07-28-2011
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Impairment of a cell cycle checkpoint is often associated with sensitivity to chemotherapeutic drugs. Here, we studied the correlations between the checkpoint with forkhead-associated and ring finger (CHFR) gene expression and responses to paclitaxel in endometrial cancer cells.
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Molecular cloning and characterization of a novel ras-related protein (rap2) from Clonorchis sinensis.
Parasitol. Res.
PUBLISHED: 07-16-2011
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Ras are key components of diverse signal transduction pathways and play important roles in growth and development. To know about growth regulation in Clonorchis sinensis, we have identified a full-length sequence encoding a ras-related protein (rap2) from our adult cDNA library. The open reading frame contains 561 bp encoding 186 amino acids. The hypothetical amino acid sequence shared high identities with rap2 proteins from Schistosoma japonicum and Homo sapiens. Conserved domains of small guanosine triphosphate-binding proteins and characteristic amino acid residues of rap2 proteins were observed in this sequence. Reverse transcription polymerase chain reaction experiments revealed that rap2 transcribed in adult worm, metacercaria, and eggs of C. sinensis. Recombinant rap2 protein was expressed and purified from Escherichia coli. rap2 could be probed by C. sinensis-infected rat serum in western blotting experiment. By immunohistochemistry, rap2 was localized on the tegument of adult worm and metacercaria of C. sinensis. This fundamental study might contribute to further researches in signaling systems that are related to growth control and development of C. sinensis and other parasites.
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A new electrochemical biosensor for DNA detection based on molecular recognition and lead sulfide nanoparticles.
Anal. Biochem.
PUBLISHED: 05-12-2011
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In this paper, we constructed a new electrochemical biosensor for DNA detection based on a molecule recognition technique. In this sensing protocol, a novel dual-labeled DNA probe (DLP) in a stem-loop structure was employed, which was designed with dabcyl labeled at the 3 end as a guest molecule, and with a Pb nanoparticle labeled at the 5 end as electrochemical tag to indicate hybridization. One ?-cyclodextrin-modified electrode (?-CD/MCNT/GCE) was used for capturing the DNA hybridization. Initially, the DLP was in the "closed" state in the absence of the target, which shielded dabcyl from the bulky ?-CD/MCNT/GCE conjugate due to a steric effect. After hybridization, the loop sequence (16 bases) formed a rigid duplex with the target, breaking the relatively shorter stem duplex (6 bases). Consequently, dabcyl was forced away from the Pb nanoparticle and became accessible by the electrode. Therefore, the target hybridization event can be sensitively transduced via detecting the electrochemical reduction current signal of Pb. Using this method, as low as 7.1×10(-10)M DNA target had been detected with excellent differentiation ability for even a single mismatch.
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Identification and molecular characterization of a novel signaling molecule 14-3-3 epsilon in Clonorchis sinensis excretory/secretory products.
Parasitol. Res.
PUBLISHED: 04-21-2011
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Increasing evidence shows that 14-3-3 proteins are involved in many biology events in addition to signal transduction. Extensive investigations on structural and biochemical features of these signaling molecules have implied their importance in the biological process. In the present study, we have identified and characterized the 14-3-3 epsilon (Cs14-3-3) in Clonorchis sinensis that causes human clonorchiasis. Recombinant protein was expressed in Escherichia coli (E. coli) and identified by MALDI-TOF/TOF. Immunoblot results revealed that Cs14-3-3 was a component of excretory/secretory products. Ligand blot assay indicated that 14-3-3 epsilon could bind C. sinensis MAPKAPK 2 in a nonphosphorylation-dependent manner. This protein could be detected at four stages of the life cycle by RT-PCR experiments and immunolocalization showed that Cs14-3-3 was extensively distributed in C. sinensis, especially at the outer surface and the sucker of adult worm and cyst wall of metacercaria. Taken together, 14-3-3 epsilon might play some roles in the development of the parasites. In addition, Cs14-3-3 epsilon should be addressed for the diagnostic value in C. sinensis infection in consideration of high sensitivity and specificity. As an immune stimulus, C. sinensis 14-3-3 epsilon was found to provoke a Th1/Th2 balanced immune response by inducing high levels of both IgG1 and IgG2a. Recombinant Cs14-3-3 conferred effective protection both in worm reduction rate and egg reduction rate, suggesting that the signaling molecule Cs14-3-3 was a promising vaccine candidate against C. sinensis infection.
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Serine/threonine kinase gene Stpk-V, a key member of powdery mildew resistance gene Pm21, confers powdery mildew resistance in wheat.
Proc. Natl. Acad. Sci. U.S.A.
PUBLISHED: 04-20-2011
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Powdery mildew resistance gene Pm21, located on the chromosome 6V short arm of Haynaldia villosa and transferred to wheat as a 6VS·6AL translocation (T6VS·6AL), confers durable and broad-spectrum resistance to wheat powdery mildew. Pm21 has become a key gene resource for powdery mildew resistance breeding all over the world. In China, 12 wheat varieties containing Pm21 have been planted on more than 3.4 million hectares since 2002. Pm21 has been intractable to molecular genetic mapping because the 6VS does not pair and recombine with the 6AS. Moreover, all known accessions of H. villosa are immune to powdery mildew fungus. Pm21 is still defined by cytogenetics as a locus. In the present study, a putative serine and threonine protein kinase gene Stpk-V was cloned and characterized with an integrative strategy of molecular and cytogenetic techniques. Stpk-V is located on the Pm21 locus. The results of a single cell transient expression assay showed that Stpk-V could decrease the haustorium index dramatically. After the Stpk-V was transformed into a susceptible wheat variety Yangmai158, the characterized transgenic plants showed high and broad-spectrum powdery mildew resistance similar to T6VS·6AL. Silencing of the Stpk-V by virus-induced gene silencing in both T6VS·6AL and H. villosa resulted in their increased susceptibility. Stpk-V could be induced by Bgt and exogenous H(2)O(2), but it also mediated the increase of endogenous H(2)O(2), leading to cell death and plant resistance when the plant was attacked by Bgt.
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Effect of coupled plasma filtration adsorption on endothelial cell function in patients with multiple organ dysfunction syndrome.
Int J Artif Organs
PUBLISHED: 04-12-2011
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The purpose of our study was to investigate the effect of coupled plasma filtration adsorption (CPFA) on endothelial cell (EC) function in patients with multiple organ dysfunction syndrome (MODS).
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Molecular characterization of cathepsin B from Clonorchis sinensis excretory/secretory products and assessment of its potential for serodiagnosis of clonorchiasis.
Parasit Vectors
PUBLISHED: 03-23-2011
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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 Clonorchis sinensis (CsCB) and to investigate its diagnostic value for human helminthiases.
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Enhanced antitumor efficacy of a novel fiber chimeric oncolytic adenovirus expressing p53 on hepatocellular carcinoma.
Cancer Lett.
PUBLISHED: 03-15-2011
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Oncolytic adenoviruses may offer a new treatment option and improve the prognosis for patients with hepatocellular carcinoma (HCC). However, the antitumor efficacy of oncolytic adenoviruses on HCC cells is compromised due to low expression of the adenovirus serotype 5 (Ad5) receptor on the target cells. In this study we showed that all HCC cell lines and clinical samples expressed high level of CD46, the receptor for Adenovirus 35 (Ad35) and constructed new fiber chimeric oncolytic adenoviruses with or without a p53 gene expression cassette, SG635-p53 and SG635, respectively. These variants were derived from the previously described Ad5 vectors SG600-p53 and SG600 by replacing the Ad5 fiber with a chimeric Ad5/35 fiber. It was found that the 5/35 fiber chimeric adenovirus vector (Ad5/35-EGFP) demonstrated significantly improved transduction in all tested HCC cell lines compared with the Ad5 vector (Ad5-EGFP). The new fiber chimeric oncolytic adenoviruses produced more progeny viruses in HCC cells than did the Ad5-based viruses but replicated weakly in normal fibroblast BJ cells. In addition, SG635-p53 mediated a higher level of transgenic expression than SG600-p53 in Hep3B and Huh7 cells and showed a markedly enhanced antitumor effect on HCC cells in vitro compared with SG635 or SG600-p53 without causing significant cytotoxicity to normal cells. Antitumor activity of SG635-p53 was shown in Hep3B subcutaneous xenograft tumor models following intratumoral injection, resulting in significant inhibition of tumor growth and prolonged survival of animals. Our data suggest that SG635-p53, as a fiber chimeric oncolytic adenovirus in combination with p53 expression, may serve as a novel, promising and safe anticancer agent for the treatment of HCC.
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Clonorchis sinensis enolase: identification and biochemical characterization of a glycolytic enzyme from excretory/secretory products.
Mol. Biochem. Parasitol.
PUBLISHED: 02-21-2011
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Enolase plays a key role in energy metabolism and development of most organisms. We isolated a gene encoding enolase from Clonorchis sinensis (C. sinensis) adult cDNA library and expressed the recombinant protein in Escherichia coli. C. sinensis enolase (Csenolase) was identified as both an excretory/secretory product and a tegumental component of C. sinensis by western blot analysis. The transcriptional level of Csenolase was examined at adult worm, metacercaria, cercaria and egg of C. sinensis, and results showed that Csenolase is transcribed at the four life stages of C. sinensis while showing a significant higher expression level at the stage of adult worm. Immunohistochemical localization indicated that Csenolase was specifically deposited on the tegument of adult worm and cyst wall of metacercaria. Ligand blot assay revealed a specific characteristic of dose-dependent plasminogen-binding activity of Csenolase and kinetic parameters were explored using 2-phospho-D-glycerate (2-PGA) as the primary substrate by monitoring the conversion of nicotinamide-adenine dinucleotide (NADH) into nicotinamide adenine dinucleotide (NAD). In addition, Csenolase exhibited active enzyme activity in catalytic reactions while the anti-Csenolase serum inhibited the enzyme activity. In vitro incubation experiments revealed that Csenolase might play key roles in the growth of the parasites. In conclusion, Csenolase is an important glycolytic enzyme required for the development of C. sinensis, and may be a potential vaccine candidate and drug target against C. sinensis infection.
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Functional identification of hyoscyamine 6?-hydroxylase from Anisodus acutangulus and overproduction of scopolamine in genetically-engineered Escherichia coli.
Biotechnol. Lett.
PUBLISHED: 02-21-2011
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Hyoscyamine 6?-hydroxylase (H6H; EC 1.14.11.11) converts hyoscyamine to scopolamine in the last step of scopolamine biosynthetic pathway. The gene encoding H6H in Anisodus acutangulus was cloned and expressed in Escherichia coli and the recombinant proteins fused with His-tag or GST-tag at its N-terminal were purified and then confirmed by Western bolt analysis. The biofunctional assay revealed that the His-AaH6H and GST-AaH6H converted hyoscyamine (40 mg/l) to scopolamine at 32 and 31 mg/l, respectively. This is the first report on AaH6H expression, purification and functional characterization facilitates further genetic improvement of scopolamine yield in A. acutangulus.
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Cyclooxygenase 2 promotes parathyroid hyperplasia in ESRD.
J. Am. Soc. Nephrol.
PUBLISHED: 02-18-2011
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Hyperplasia of the PTG underlies the secondary hyperparathyroidism (SHPT) observed in CKD, but the mechanism underlying this hyperplasia is incompletely understood. Because aberrant cyclooxygenase 2 (COX2) expression promotes epithelial cell proliferation, we examined the effects of COX2 on the parathyroid gland in uremia. In patients with ESRD who underwent parathyroidectomy, clusters of cells within the parathyroid glands had increased COX2 expression. Some COX2-positive cells exhibited two nuclei, consistent with proliferation. Furthermore, nearly 78% of COX2-positive cells expressed proliferating cell nuclear antigen (PCNA). In the 5/6-nephrectomy rat model, rats fed a high-phosphate diet had significantly higher serum PTH levels and larger parathyroid glands than sham-operated rats. Compared with controls, the parathyroid glands of uremic rats exhibited more PCNA-positive cells and greater COX2 expression in the chief cells. Treatment with COX2 inhibitor celecoxib significantly reduced PCNA expression, attenuated serum PTH levels, and reduced the size of the glands. In conclusion, COX2 promotes the pathogenesis of hyperparathyroidism in ESRD, suggesting that inhibiting the COX2 pathway could be a potential therapeutic target.
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Membrane topology and mutational analysis of Mycobacterium tuberculosis VKOR, a protein involved in disulfide bond formation and a homologue of human vitamin K epoxide reductase.
Antioxid. Redox Signal.
PUBLISHED: 02-18-2011
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We have presented evidence that a homologue of vertebrate membrane protein vitamin K epoxide reductase (VKOR) is an important component of the protein disulfide bond-forming pathway in many bacteria. Bacterial VKOR appears to take the place of the nonhomologous DsbB found in Escherichia coli. We also determined the structure of a VKOR from a Cyanobacterium and showed that two or four conserved cysteines are required, according to different reductants for activity in an in vitro assay. Here we present evidence for the topologic arrangement in the cytoplasmic membrane of the VKOR from Mycobacterium tuberculosis (Mtb). The results show that Mtb VKOR is a membrane protein that spans the membrane 5 times with its N-terminus in the cytoplasm, C-terminus in the periplasm, and all four cysteines facing the periplasm. The essentiality of the four conserved cysteine residues has also been demonstrated in promoting disulfide bond formation in vivo and a mixed disulfide between a cysteine of DsbA of E. coli, and one of the cysteines (Cys(57)) of the VKOR homologue has been identified to be a likely intermediate in the disulfide bond-forming pathway. These studies may inform future resolution of issues surrounding the functioning of human VKOR.
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The draft genome of the carcinogenic human liver fluke Clonorchis sinensis.
Genome Biol.
PUBLISHED: 01-31-2011
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Clonorchis sinensis is a carcinogenic human liver fluke that is widespread in Asian countries. Increasing infection rates of this neglected tropical disease are leading to negative economic and public health consequences in affected regions. Experimental and epidemiological studies have shown a strong association between the incidence of cholangiocarcinoma and the infection rate of C. sinensis. To aid research into this organism, we have sequenced its genome.
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Down-regulation of IGF-1R expression inhibits growth and enhances chemosensitivity of endometrial carcinoma in vitro.
Mol. Cell. Biochem.
PUBLISHED: 01-24-2011
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The type-1 insulin-like growth factor receptor (IGF-1R) is over-expressed by endometrial carcinoma, level of IGF-1R has been correlated with tumor progression, and high IGF-1R expression has been found to be an important prognostic factor. In the study, we used lentivirus-mediated shRNA targeting IGF-1R to silence its expression, then assessed the effect of down-regulation of this receptor on cell growth and chemosensitivity to cisplatin. Lentivirus-mediate shRNA was designed and transfected to the endometrial carcinoma HEC-1B cell. The IGF-1R mRNA and related protein expression, cell proliferation ability, cell apoptosis, and cell cycle change were detected. Cell proliferation inhibition rates, cell apoptosis, and level of cleaved caspase-9 were measured in various concentrations of cisplatin. The mRNA and protein level of IGF-1R, and the phosphorylated protein p-Akt, p-Erk were all suppressed after transfection. Cell proliferation was inhibited in successive five days after transfection, the highest inhibition rate was 43.28 ± 3.55% on day 5. After transfection, 24.96 ± 1.05% cells were in G(2)/M phase, and cell apoptotic rate increased from 10.66 ± 0.08 to 19.92 ± 1.34%. In various concentrations of cisplatin, transfected cells proliferation was significantly inhibited which made the IC50 value drop from 21.85 uM to 10.58 uM. Incubation with different concentrations of cisplatin for 48 h, cells apoptotic rate increased to 41.92 ± 2.5, 31.13 ± 2.76, 22.21 ± 4.63%, respectively, which was accompanied with increased cleaved caspase-9 expression. Lentivirus-mediated shRNA targeting IGF-1R has the potential to develop as a clinical treatment method in advanced and chemoresistant endometrial carcinoma.
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Molecular cloning, expression, and immunolocalization of the NAD(+)-dependent glycerol 3-phosphate dehydrogenase (GPD) from Clonorchis sinensis.
Parasitol. Res.
PUBLISHED: 01-09-2011
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Glycerol 3-phosphate dehydrogenase (GPD) plays an important role in the energy metabolism and nutrition metabolism. In order to know about the biological functions of GPD of Clonorchis sinensis (C. sinensis), we identified a complete gene coding GPD from C. sinensis metacercaria cDNA library. This novel cDNA sequence contains 1,056 bp with a putative open reading frame of 351 amino acids and shares 74% identity with GPD from Schistosoma mansoni. Recombinant CsGPD was expressed and purified from Escherichia coli BL21 (DE3). Western blot analysis displayed that recombinant CsGPD can be recognized by anti-CsGPD serum and C. sinensis-infected serum. RT-PCR and immunolocalization analysis confirmed that GPD expressed both at the stage of adult worm and metacercaria of C. sinensis and immunolocated at the tegument of adult worm, tegument and tegumentary cells of metacercaria. Our current study has paved the way for the further researches about the biological functions involved in the growth of C. sinensis.
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Combination effects of paclitaxel with signaling inhibitors in endometrial cancer cells.
Asian Pac. J. Cancer Prev.
PUBLISHED: 01-01-2011
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This study was conducted to evaluate and compare molecular and cellular effects of paclitaxel in combination with epidermoid growth factor receptor (EGFR) or/and mammalian target of rapamycin( mTOR) inhibitors with two endometrial cancer lines HEC-1A and Ishikawa. Treatment was with the EGFR inhibitor RG14620, the mTOR inhibitor rapamycin, and the conventional cytotoxic drug paclitaxel, alone or in combination. The 50% inhibitory concentration (IC50) and cell viability were determined by the MTT assay. Multiple drug effect/ combination indexes (CI) analysis was applied to assess interactions between paclitaxel and the two inhibitors. Apoptosis and cell cycling were evaluated by flow cytometry analysis. Western blotting was performed to evaluate the related protein alteration in PI3K/AKT signaling pathway. RG14620, rapamycin and paclitaxel showed obvious dose-dependent growth inhibition with time. The IC50 of paclitaxel at 24 hours decreased significantly when pretreated with low doses of RG14620 and Rapamycin alone or in combination. Moreover, combination index (CI) of paclitaxel with each inhibitor was larger than 1, indicating a synergistic effect between pairs of drugs in these two cell lines. FACS analysis showed the cell apoptosis rate increased with a synergistic effect. On Western blotting, activation of PI3K/AKT pathway was detected in both two cell lines in the control case. When paclitaxel was used as a single-agent or in combinations, the protein expression of PI3K/AKT pathway totally abated, especially in HEC-1A cells, suggesting a role in chemoresistance. The combination of three drugs induced the greatest over-expression of caspase-3. Combining targeted inhibitors with cytotoxic chemotherapy appears to be a promising strategy for the effective treatment of endometrial cancer which merits further clinical investigation.
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A Case of Cutaneous Nocardiosis with Involvement of the Trachea, Anterior Mediastinum and Sternum.
Case Rep Dermatol
PUBLISHED: 10-21-2010
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Nocardiosis is a rare infectious disease due to Nocardia infections. In this report, we present a rare case of cutaneous nocardiosis with involvement of the trachea, anterior mediastinum and sternum. The strain of Nocardia has been isolated from bacterial culture of infected tissue. 16s rRNA sequencing confirmed that it contained the Nocardia genus. The patient was successfully treated with Co-SMZ.
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Population sequencing of two endocannabinoid metabolic genes identifies rare and common regulatory variants associated with extreme obesity and metabolite level.
Genome Biol.
PUBLISHED: 06-28-2010
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Targeted re-sequencing of candidate genes in individuals at the extremes of a quantitative phenotype distribution is a method of choice to gain information on the contribution of rare variants to disease susceptibility. The endocannabinoid system mediates signaling in the brain and peripheral tissues involved in the regulation of energy balance, is highly active in obese patients, and represents a strong candidate pathway to examine for genetic association with body mass index (BMI).
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Improvement of cytokine response and survival time by bioartificial kidney therapy in acute uremic pigs with multi-organ dysfunction.
Int J Artif Organs
PUBLISHED: 06-20-2010
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To explore whether bioartificial kidney (BAK) ameliorates cytokine response and biochemical indices, and prolongs the survival time in acute uremic pigs with multiple organ dysfunction syndrome (MODS).
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Derp2-mutant gene vaccine inhibits airway inflammation and up-regulates Toll-like receptor 9 in an allergic asthmatic mouse model.
Asian Pac. J. Allergy Immunol.
PUBLISHED: 05-15-2010
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A DNA vaccine encoding the whole segment of the Derp2 allergen could prevent allergic airway inflammation in a Derp2 allergen-induced allergic airway inflammation mouse model.
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Role of TGF-beta1 in bone matrix production in vascular smooth muscle cells induced by a high-phosphate environment.
Nephron Exp. Nephrol.
PUBLISHED: 04-24-2010
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Aims: We demonstrated a relationship between transforming growth factor-beta(1) (TGF-beta(1)) and production of bone matrix in vascular smooth muscle cells (VSMCs) induced by a high-phosphate environment. Methods: Rat VSMCs were incubated in a high-phosphate (2.5 or 3.5 mM) or TGF-beta(1) (2 or 5 ng/ml) environment. TGF-beta(1) monoclonal neutralization antibody (50 microg/ml) was added to inhibit the TGF-beta(1) signal in high-phosphate medium. Production of TGF-beta(1) was analyzed by Western blot and real-time PCR. Core binding factor a1 (Cbfa1), osteopontin (OP), collagen type I (Col I) and osteocalcin (OC) was investigated by Western blot and immunofluorescence staining. Mineral deposition was assessed by von Kossa staining and o-cresolphthalein complexone method. Results: VSMCs transformation induced by high phosphate occurs via an autocrine loop of TGF-beta(1). First, high phosphate stimulated the production of TGF-beta(1) in VSMCs. Second, TGF-beta(1) could induce increased expression of osteoblast-specific transcription factor Cbfa1 and osteogenic molecule in VSMCs. Third, the TGF-beta(1) neutralization antibody largely attenuated the upregulation of Cbfa1 and bone matrix in high-phosphate-stimulated cells. However, neutralization of TGF-beta(1) could not inhibit high-phosphate-induced VSMCs calcification, indicating that TGF-beta(1) was not necessary for the deposition of calcium. Conclusion: TGF-beta(1) plays a crucial role in bone matrix production but not calcium deposition in VSMCs induced by a high-phosphate environment, and the blockade of TGF-beta(1) signaling may thus be a therapeutic strategy for use with vascular disease in a high-phosphate environment.
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Expression and immune response analysis of Schistosoma japonicum VAL-1, a homologue of vespid venom allergens.
Parasitol. Res.
PUBLISHED: 01-31-2010
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Many parasites such as trematodes and nematodes have been found to express members of a gene family variously termed as venom allergen-like protein (VAL) or sperm-coating protein (SCP)-like protein. The molecular functions of these proteins remain unclear. We isolated the corresponding gene from Schistosoma japonicum, which we designated as Sj-VAL-1. The cDNA of Sj-VAL-1 contains an open reading frame encoding 221 amino acids, the first 25 residues being a putative secretion signal. RT-PCR analysis confirmed that Sj-VAL-1 was transcribed mainly in cercariae and eggs stages. Western blot analysis indicated that Sj-VAL-1 protein was an egg excretory-secretory products (ES products). Immunofluorescence showed it was secreted by eggs, head gland, and penetration glands of cercariae. In S. japonicum-infected mice, Sj-VAL-1-specific antibody significantly increased 6 weeks after infection and a higher level of IgG1 antibody contrast to IgG2a antibody indicated that a polarized Th2 immune response could be induced by Sj-VAL-1. These findings suggest that Sj-VAL-1 must play a role in the interaction between parasite and host. Its role as a potential modulator of immune function or as a pathogenic factor of granuloma formation in schistosomiasis needs further study.
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Stage and tissue specific differences in SjBMI1, a Polycomb protein in Schistosoma japonicum.
Parasitol. Res.
PUBLISHED: 01-04-2010
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Polycomb group protein BMI1, plays a central role in the stem cell pluripotency and development in metazoans. A gene encoding BMI1 homologue in the Schistosoma japonicum (SjBMI1) was cloned and identified. The deduced amino acid sequence shows high identity to the homologues from Schistosoma mansoni and Homo sapiens. Quantitative real time polymerase chain reaction (RT-PCR) and Western blot analysis revealed that the SjBMI1 is highly expressed in adult worms and eggs, not in cercariae. By immunofluorescent studies, SjBMI1 was localized to testes, ovaries of mixed sex infected adult worms, but not of single sex infected adult worms. The study reveals the SjBMI1 expression profile in developmental stages and localization characteristic and provides a clue that it may be associated with reproductive development of S. japonicum.
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Inhibition of NF-kappaB expression and allergen-induced airway inflammation in a mouse allergic asthma model by andrographolide.
Cell. Mol. Immunol.
PUBLISHED: 11-06-2009
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Andrographolide from traditional Chinese herbal medicines previously showed it possesses a strong anti-inflammatory activity. In present study, we investigated whether Andrographolide could inhibit allergen-induced airway inflammation and airways hyper-responsiveness and explored the mechanism of Andrographolide on allergen-induced airway inflammation and airways hyper-responsiveness. After sensitized and challenged by ovalbumin, the BALB/c mice were administered intraperitoneally with Andrographolide. Hyper-responsiveness was recorded. The lung tissues were assessed by histological examinations. NF-kappaB in lung was determined by immunofluorescence staining and Western blotting. Treatment of mice with Androqrapholide displayed lower Penh in response to asthma group mice. After treatment with Andrographolide, the extent of inflammation and cellular infiltration in the airway were reduced. Andrographolide interrupted NF-kappaB to express in cell nucleus. The level of NF-kappaB expression was inhibited by Andrographolide. The data indicate that Andrographolide from traditional Chinese herbal medicines could inhibit extensive infiltration of inflammatory cells in lung and decrease airway hyperreactivity. Andrographolide could inhibit NF-kappaB expression in lung and suppress NF-kappaB expressed in the nucleus of airway epithelial cells.
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Surgical treatment of hermaphroditism: experience with 25 cases.
Ann Plast Surg
PUBLISHED: 10-07-2009
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This article is a summary of our experience with surgical treatment of 25 patients with hermaphroditism. We treated 25 patients with hermaphroditism since 1985, including 12 with male pseudohermaphroditism, 9 with female pseudohermaphroditism and 4 with true hermaphroditism. Decision on sex reassignment for these patients was made according to their genetic sex, gonad sex, social sex, psychologic sex, and the request of the patients and their relatives. Of the 12 male pseudohermaphrodites, 1 was reassigned with the male sex and penile reconstruction was performed, and the other 11 had a female sex assignment and received undescended testis removal, clitoral reduction, labioplasty of the labia minora and labium majus, and vaginoplasty. Nine female pseudohermaphrodites with female sex assignment underwent clitoral reduction, labioplasty of the labia minora and labium majus, and vaginoplasty. The 4 true hermaphrodites also had female sex assignment, and received such procedures as resection of the ovotestes and undescended testes, clitoral reduction, and labioplasty of the labia minora; resection of the uterus and appendices was also performed in 1 case due to the identification of malignant cells. The reconstructed penis in the patient with male sex assignment had good appearance and allowed normal urination. In patients with female sex assignment, the reconstructed external genitalia had also good appearance and good sensation without obvious contraction. No serious complications occurred in these patients, who were satisfactory with the outcome and reported stable gender identity, and 3 married patients reported normal sexual life. Plastic surgery is still the primary option for correction of hermaphroditism after determination of sex assignment, and satisfactory effect can be achieved by surgical intervention with stable gender identity and minimal complications.
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Molecular cloning and characterization of a novel serpin gene of Clonorchis sinensis, highly expressed in the stage of metacercaria.
Parasitol. Res.
PUBLISHED: 09-22-2009
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The serpins are a superfamily of proteins (350-500 amino acids in size) that fold into a conserved structure. From about 3,475 unigenes of Clonorchis sinensis metacercaria, a novel gene-encoding serpin was identified and characterized. The opening reading frame is 1,149 bp encoding 382 amino acids. The deduced amino acid sequence shows high identity to previously reported serpins from C. sinensis and other helminthic parasites. A typical serpin signature was found by motif search. The recombinant C. sinensis serpin protein (rCsproSERPIN) was produced and purified. Semiquantitative analysis revealed that the transcripts of this serpin gene in metacercaria were much higher than that in adult worms and that the corresponding band of serpin protein in the crude soluble antigen of metacercaria probed by rat anti-CsproSERPIN serum was also much clearer compared with that of adult, suggesting that it plays an important role in the stage of C. sinensis metacercaria. Although we are not much clear about the detailed function of this serpin protein, the study that proteinase initiates metacercaria excystment gives a clue that it may participate in the encystment of cercaria.
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Experimental model in rats for study on transmission dynamics and evaluation of Clonorchis sinensis infection immunologically, morphologically, and pathologically.
Parasitol. Res.
PUBLISHED: 08-18-2009
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This study aims to gain a better insight into the transmission patterns and immunologic profile of Clonorchis sinensis infection and make a headway on the pathogenesis regarding cholangiocarcinoma and hepatic lesions. Experimental models orally infected by C . sinensis metacercariae were constructed in rats. Immunological assays were performed to measure serum level of IgA, IgE, IgG1, IgG2a, IFN-gamma, and IL-4. Infection parameters were assessed by worm recovery rate, eggs per gram faece and worm size. Pathological sections with livers were managed with immunofluorescence, hematoxylin, eosin, and Massons trichrome staining to evaluate the hepatic pathological changes. Interestingly, rats infected with only one C . sinensis metacercariae even gained a high worm recovery rate of 83.3% compared with rats infected with more metacercariae. Serological changes according to different infection doses indicated that immune response presented a tendency to Th2 type by expressing transient high level of IgG1, IL-4, and IgE. Hepatic tissues appeared inflammatory and fibrotic, revealed by different stainings. Intrahepatic bile ducts displayed cholangiectasis and proliferation with excreted/secreted antigen histologically located. C . sinensis, as a fish-borne zoonosis, presented novel transmission patterns which explained high infection rate in endemic areas; infection rate of C . sinensis was frequency-dependent and dose-related. Humoral immunity played a prevalent role in resisting to C . sinensis based on the rat models. C . sinensis infection played an undoubted role in biliary and hepatic diseases.
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C1 domain mediates CalDAGIII localization to the Golgi.
Mol. Biol. Rep.
PUBLISHED: 07-04-2009
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CalDAGs are a family of Ras guanyl exchange factors that contain calcium and DAG-binding domains. Among the four identified members of CalDAG family, CalDAGIII has been shown to play important role in B lymphocyte and endocrine cell functions. However, the mechanism underlining these functions remain to be determined. Here in the present study, we determined the subcellular localization of CalDAGIII and roles of calcium-binding and DAG-binding domains in its localization. We found that C1 domain but not EF hands is important for both CalDAGIII localization to the Golgi and p38 activation in B cells, indicating that CalDAGIII may be regulated by DAG but not calcium.
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Enrichment of sequencing targets from the human genome by solution hybridization.
Genome Biol.
PUBLISHED: 06-17-2009
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To exploit fully the potential of current sequencing technologies for population-based studies, one must enrich for loci from the human genome. Here we evaluate the hybridization-based approach by using oligonucleotide capture probes in solution to enrich for approximately 3.9 Mb of sequence target. We demonstrate that the tiling probe frequency is important for generating sequence data with high uniform coverage of targets. We obtained 93% sensitivity to detect SNPs, with a calling accuracy greater than 99%.
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Immunogenicity of self-adjuvanticity oral vaccine candidate based on use of Bacillus subtilis spore displaying Schistosoma japonicum 26 KDa GST protein.
Parasitol. Res.
PUBLISHED: 05-11-2009
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One of the promising approaches in mucosal immunization relies on live recombinant vaccine carriers. In this study, we used a six-extracellular protease-deficient Bacillus subtilis strain WB600 to express Schistosoma japonicum 26 kDa glutathione S-transferase (GST). Western blot, immunofluorescence, and flow cytometry analyses were used to identify SjGST expression on spore surface. SjGST recombinant spores were used for oral vaccination in mice and were shown to generate mucosal and systemic response. Both SjGST-specific secretory IgA in feces and IgG in serum augmented significantly on day 33 after oral administration. It seemed that surface display of recombinant S. japonicum SjGST on B. subtilis WB600 spores showed good immunogenicity, and B. subtilis spores could be used as potential mucosal delivery vehicles to provide more effective vaccination strategies for parasite prevention and control in the future.
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Evaluation of next generation sequencing platforms for population targeted sequencing studies.
Genome Biol.
PUBLISHED: 02-23-2009
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Next generation sequencing (NGS) platforms are currently being utilized for targeted sequencing of candidate genes or genomic intervals to perform sequence-based association studies. To evaluate these platforms for this application, we analyzed human sequence generated by the Roche 454, Illumina GA, and the ABI SOLiD technologies for the same 260 kb in four individuals.
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[Genetic modification of secondary metabolite biosynthesis in higher plants: a review].
Sheng Wu Gong Cheng Xue Bao
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Plants provide an immense reservoir of natural secondary metabolites. Secondary metabolites and those involved enzymes accumulate in various compartments in specific plant tissues. The biosynthesis of diverse groups of secondary metabolites is often complicated, tightly controlled via network interconnections, metabolite levels, metabolite channeling and multi-enzyme complexes, and so on. Secondary metabolite profiles could be genetically altered by two strategies, i.e. single gene modification and multiple gene modification; which thus has opened a feasible and prospective platform for secondary chemicals production in plant.
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