The Open Biochemistry Journal最新文献

{"title":"New Perspectives of \"omics\" Applications in Melanoma Research.","authors":"Carmen Rodríguez-Cerdeira,&nbsp;Alberto Molares-Vila","doi":"10.2174/1874091X01105010060","DOIUrl":"https://doi.org/10.2174/1874091X01105010060","url":null,"abstract":"<p><strong>Background: </strong>Oncoproteomics is the study of proteins and their interactions in a cancer cell by proteomic technologies and has the potential to revolutionize clinical practice, including cancer diagnosis. Recent technological advances in the analysis of the human genome have opened the door to improving our primitive understanding of the gene expression patterns in cancer. The examination of the phenotypic and (epi) genetic changes in cutaneous melanoma has identified several genes deemed central to the development and progression of melanoma.</p><p><strong>Methods: </strong>A review of the literature was performed to determine the role of epigenetic modifications in human melanoma. The role of array-based high-throughput gene expression analysis in understanding the specific genes involved as well as the pathways and the comparative gene expression patterns of primary and metastatic melanoma. The development and clinical application of selective pharmacologic agents are also discussed.</p><p><strong>Results: </strong>We identified several articles that have extensively studied the role of epigenetics in melanoma, further elucidating the complex processes involved in gene regulation and expression. Other studies utilizing gene microarray analysis and other whole genome approaches reveal a wide array of genes and expression patterns in human melanoma. Several genes have been identified as potential prognostic markers of tumor progression and overall clinical outcome.</p><p><strong>Conclusions: </strong>High-throughput gene expression analysis has had a major impact in melanoma research. Several gene expression platforms have provided insight into the gene expression patterns in melanoma. Such data will provide foundations for the future development of prognostic markers and improved targeted therapies for patients with melanoma.</p>","PeriodicalId":515405,"journal":{"name":"The Open Biochemistry Journal","volume":"5 ","pages":"60-6"},"PeriodicalIF":0.0,"publicationDate":"2011-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.2174/1874091X01105010060","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"30393110","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Regions which are Responsible for Swapping are also Responsible for Folding and Misfolding.","authors":"Oxana V Galzitskaya","doi":"10.2174/1874091X01105010027","DOIUrl":"https://doi.org/10.2174/1874091X01105010027","url":null,"abstract":"<p><p>Domain swapping is a term used to describe a process when two or more protein chains exchange identical structural elements. Some cases of amyloid formation can be explained through a domain swapping mechanism therefore this deserves theoretical consideration and studying. It has been demonstrated that diverse proteins in sequence and structure are able to oligomerize via domain swapping. This allows us to suggest that the exchangeable regions are important in folding and misfolding processes of proteins, i.e. the residues from the swapping regions are typically incorporated into the native structure early during its formation. The modeling of folding of the proteins with swapped domains demonstrates that the regions exchanged in the oligomeric form in most cases are also responsible for folding and misfolding. For 11 out of 17 proteins, swapping regions intersect with the predicted amyloidogenic regions. Moreover, for 10 out of 17 proteins, high Φ-values (>0.5) belong to residues from the swapping regions. Our data confirm that the exchangeable regions are important in folding, misfolding, and domain swapping processes of the proteins, therefore the suggestion that domain swapping can serve as a mechanism for functional interconversion between monomers and oligomers is likely to be correct.</p>","PeriodicalId":515405,"journal":{"name":"The Open Biochemistry Journal","volume":"5 ","pages":"27-36"},"PeriodicalIF":0.0,"publicationDate":"2011-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/e8/52/TOBIOCJ-5-27.PMC3134983.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"30014921","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Thalidomide Analogues Suppress Lipopolysaccharide-Induced Synthesis of TNF-α and Nitrite, an Intermediate of Nitric Oxide, in a Cellular Model of Inflammation.","authors":"David Tweedie,&nbsp;Kathryn A Frankola,&nbsp;Weiming Luo,&nbsp;Yazhou Li,&nbsp;Nigel H Greig","doi":"10.2174/1874091X01105010037","DOIUrl":"https://doi.org/10.2174/1874091X01105010037","url":null,"abstract":"<p><p>An unregulated neuroinflammation accompanies numerous chronic and acute neurodegenerative disorders and it is postulated that such a neuroinflammatory component likely exacerbates disease progression. A key player in brain inflammation is the microglial cell; a vital soluble factor synthesized by activated microglial cells is the key cytokine, tumor necrosis factor-alpha (TNF-α). Additionally, microglial cells release IL-1α/β, reactive oxygen species (ROS), such as superoxide (O(2) (-)) and reactive nitrogen species (RNS) like nitric oxide (NO). Nitric oxide reactive oxygen species can undergo various forms of interactions in cells whereby the synthesis of RNS / ROS intermediates are generated that can damage cell membranes. The presence of oxidative damaged cells is implicated with the abnormal cellular activity in brain or in the spinal cord, and is a classical feature of neurodegenerative disorders. To aid characterize this process, a quantitative analysis of nitrite generation was undertaken on agents developed to lower TNF-α levels in cell culture. Nitrite is a stable end product of nitric oxide metabolism and, thereby, acts as a surrogate measure of the highly unstable nitric oxide. Utilizing a RAW 264.7 cellular model of lipopolysaccharide-induced inflammation that induces high levels of TNF-α protein accompanied by a robust generation of nitrite, the properties of a series of thalidomide-based TNF-α synthesis inhibitors were evaluated to reduce the levels of both. Specific analogues of thalidomide effectively suppressed the generation of both TNF-α and nitrite at well-tolerated doses.</p>","PeriodicalId":515405,"journal":{"name":"The Open Biochemistry Journal","volume":"5 ","pages":"37-44"},"PeriodicalIF":0.0,"publicationDate":"2011-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.2174/1874091X01105010037","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29892601","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Distinct metal ion requirements for the phosphomonoesterase and phosphodiesterase activities of calf intestinal alkaline phosphatase.","authors":"Adedoyin Igunnu,&nbsp;Dunsin S Osalaye,&nbsp;Olufunso O Olorunsogo,&nbsp;Sylvia O Malomo,&nbsp;Femi J Olorunniji","doi":"10.2174/1874091X01105010067","DOIUrl":"https://doi.org/10.2174/1874091X01105010067","url":null,"abstract":"<p><p>The roles of Mg(2+) and Zn(2+) ions in promoting phosphoryl transfer catalysed by alkaline phosphatase are yet to be fully characterised. We investigated the divalent metal ion requirements for the monoesterase and diesterase activities of calf intestinal alkaline phosphatase. The synergistic effect of Mg(2+) and Zn(2+) in promoting the hydrolysis of para-nitrophenyl phosphate (monoesterase reaction) by alkaline phosphatase is not observed in the hydrolysis of the diesterase substrate, bis-para-nitrophenyl phosphate. Indeed, the diesterase reaction is inhibited by concentrations of Mg(2+) that were optimal for the monoesterase reaction. This study reveals that the substrate specificities of alkaline phosphatases and related bimetalloenzymes are subject to regulation by changes in the nature and availability of cofactors, and the different cofactor requirements of the monoesterase and diesterase reactions of mammalian alkaline phosphatases could have significance for the biological functions of the enzymes.</p>","PeriodicalId":515405,"journal":{"name":"The Open Biochemistry Journal","volume":"5 ","pages":"67-72"},"PeriodicalIF":0.0,"publicationDate":"2011-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/4d/cb/TOBIOCJ-5-67.PMC3258565.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"30402498","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Systematic comparison of crystal and NMR protein structures deposited in the protein data bank.","authors":"Kresimir Sikic,&nbsp;Sanja Tomic,&nbsp;Oliviero Carugo","doi":"10.2174/1874091X01004010083","DOIUrl":"https://doi.org/10.2174/1874091X01004010083","url":null,"abstract":"Nearly all the macromolecular three-dimensional structures deposited in Protein Data Bank were determined by either crystallographic (X-ray) or Nuclear Magnetic Resonance (NMR) spectroscopic methods. This paper reports a systematic comparison of the crystallographic and NMR results deposited in the files of the Protein Data Bank, in order to find out to which extent these information can be aggregated in bioinformatics. A non-redundant data set containing 109 NMR – X-ray structure pairs of nearly identical proteins was derived from the Protein Data Bank. A series of comparisons were performed by focusing the attention towards both global features and local details. It was observed that: (1) the RMDS values between NMR and crystal structures range from about 1.5 Å to about 2.5 Å; (2) the correlation between conformational deviations and residue type reveals that hydrophobic amino acids are more similar in crystal and NMR structures than hydrophilic amino acids; (3) the correlation between solvent accessibility of the residues and their conformational variability in solid state and in solution is relatively modest (correlation coefficient = 0.462); (4) beta strands on average match better between NMR and crystal structures than helices and loops; (5) conformational differences between loops are independent of crystal packing interactions in the solid state; (6) very seldom, side chains buried in the protein interior are observed to adopt different orientations in the solid state and in solution.","PeriodicalId":515405,"journal":{"name":"The Open Biochemistry Journal","volume":"4 ","pages":"83-95"},"PeriodicalIF":0.0,"publicationDate":"2010-09-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/75/f1/TOBIOCJ-4-83.PMC3032220.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29648075","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Leukocyte Count versus C-Reactive Protein Levels in Obese Portuguese Patients Aged 6-12 Years Old.","authors":"Henrique Nascimento,&nbsp;Susana Rocha,&nbsp;Carla Rego,&nbsp;Helena Ferreira Mansilha,&nbsp;Alexandre Quintanilha,&nbsp;Alice Santos-Silva,&nbsp;Luís Belo","doi":"10.2174/1874091X01004010072","DOIUrl":"https://doi.org/10.2174/1874091X01004010072","url":null,"abstract":"<p><strong>Objectives: </strong>to evaluate whether total and differential WBC counts are altered in young obese patients (aged 6-12 years) and if a relationship exists between WBC counts and the severity of obesity as well as with CRP level.</p><p><strong>Materials and methods: </strong>a group of 77 obese patients [32 males and 45 females] and 19 controls [7 males and 12 females] were studied. Total WBC count was performed by using an automatic blood cell counter. Blood cell morphology and WBC differential count were evaluated in Wright stained blood films. The plasma levels of CRP were evaluated by immunoturbidimetry.</p><p><strong>Results: </strong>obese participants presented with a statistically significant higher neutrophil percentage and CRP levels when compared to controls; the median CRP value was about 5 times higher than that observed in controls. Absolute neutrophil count and neutrophil/lymphocyte ratio were also higher in patients, though without statistical significance. The parameters that were statistically significant related with adiposity markers were neutrophil count and CRP levels. The neutrophil count was positively and statistically correlated with body mass index (BMI), BMI z-score, waist circumference and waist/height ratio, and also with CRP levels. In multiple regression analysis, the only variable that remained statistically associated with neutrophil count was CRP (neutrophil count = 2.612 + 0.439lnCRP; standardised coefficient/beta: 0.384, P=0.001). When performing multiple regression without CRP, the only variable that remained statistically associated with neutrophil count was BMI.</p><p><strong>Conclusions: </strong>our results demonstrated in obese patients aged 6-12 years, a significant change in the differential leukocyte count towards neutrophilia, together with a significant higher CRP concentration, and that absolute neutrophil count correlates with obesity markers and with CRP levels. Our data also indicate that neutrophil count, a current clinically used low-cost parameter, may be used as an obesity-related inflammatory marker in young obese patients.</p>","PeriodicalId":515405,"journal":{"name":"The Open Biochemistry Journal","volume":"4 ","pages":"72-6"},"PeriodicalIF":0.0,"publicationDate":"2010-06-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/68/51/TOBIOCJ-4-72.PMC2911598.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29161626","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Predominantly Cytoplasmic Localization in Yeast of ASR1, a Non-Receptor Transcription Factor from Plants.","authors":"Nicolás Urtasun,&nbsp;Susana Correa García,&nbsp;Norberto D Iusem,&nbsp;Mariana Bermúdez Moretti","doi":"10.2174/1874091X01004010068","DOIUrl":"https://doi.org/10.2174/1874091X01004010068","url":null,"abstract":"<p><p>The Asr gene family (named after abscisic acid, stress and ripening), currently classified as a novel group of the LEA superfamily, is exclusively present in the genomes of seed plants, except for the Brassicaceae family. It is associated with water-deficit stress and is involved in adaptation to dry climates. Motivated by separate reports depicting ASR proteins as either transcription factors or chaperones, we decided to determine the intracellular localization of ASR proteins. For that purpose, we employed an in vivo eukaryotic expression system, the heterologous model Saccharomyces cerevisiae, including wild type strains as well as mutants in which the variant ASR1 previously proved to be functionally protective against osmotic stress. Our methodology involved immunofluorescence-based confocal microscopy, without artificially altering the native structure of the protein under study. Results show that, in both normal and osmotic stress conditions, recombinant ASR1 turned out to localize mainly to the cytoplasm, irrespective of the genotype used, revealing a scattered distribution in the form of dots or granules. The results are discussed in terms of a plausible dual (cytoplasmic and nuclear) role of ASR proteins.</p>","PeriodicalId":515405,"journal":{"name":"The Open Biochemistry Journal","volume":"4 ","pages":"68-71"},"PeriodicalIF":0.0,"publicationDate":"2010-05-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/ee/d0/TOBIOCJ-4-68.PMC2908927.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29143236","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Conformational States and kinetics of the calcium binding domain of NADPH oxidase 5.","authors":"Chin-Chuan Wei,&nbsp;Nicole Motl,&nbsp;Kelli Levek,&nbsp;Liu Qi Chen,&nbsp;Ya-Ping Yang,&nbsp;Tremylla Johnson,&nbsp;Lindsey Hamilton,&nbsp;Dennis J Stuehr","doi":"10.2174/1874091X01004010059","DOIUrl":"https://doi.org/10.2174/1874091X01004010059","url":null,"abstract":"<p><p>Superoxide generated by human NADPH oxidase 5 (NOX5) is of growing importance for various physiological and pathological processes. The activity of NOX5 appears to be regulated by a self-contained Ca(2+) binding domain (CaBD). Recently Bánfi et al. suggest that the conformational change of CaBD upon Ca(2+) binding is essential for domain-domain interaction and superoxide production. The authors studied its structural change using intrinsic Trp fluorescence and hydrophobic dye binding; however, their conformational study was not thorough and the kinetics of metal binding was not demonstrated. Here we generated the recombinant CaBD and an E99Q/E143Q mutant to characterize them using fluorescence spectroscopy. Ca(2+) binding to CaBD induces a conformational change that exposes hydrophobic patches and increases the quenching accessibilities of its Trp residues and AEDANS at Cys107. The circular dichroism spectra indicated no significant changes in the secondary structures of CaBD upon metal binding. Stopped-flow spectrometry revealed a fast Ca(2+) dissociation from the N-terminal half, followed by a slow Ca(2+) dissociation from the C-terminal half. Combined with a chemical stability study, we concluded that the C-terminal half of CaBD has a higher Ca(2+) binding affinity, a higher chemical stability, and a slow Ca(2+) dissociation. The Mg(2+)-bound CaBD was also investigated and the results indicate that its structure is similar to the apo form. The rate of Mg(2+) dissociation was close to that of Ca(2+) dissociation. Our data suggest that the N- and C-terminal halves of CaBD are not completely structurally independent.</p>","PeriodicalId":515405,"journal":{"name":"The Open Biochemistry Journal","volume":"4 ","pages":"59-67"},"PeriodicalIF":0.0,"publicationDate":"2010-05-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/2f/0e/TOBIOCJ-4-59.PMC2905766.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29137287","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The impact of the level of the intestinal short chain Fatty acids in inflammatory bowel disease patients versus healthy subjects.","authors":"N Huda-Faujan,&nbsp;A S Abdulamir,&nbsp;A B Fatimah,&nbsp;O Muhammad Anas,&nbsp;M Shuhaimi,&nbsp;A M Yazid,&nbsp;Y Y Loong","doi":"10.2174/1874091X01004010053","DOIUrl":"https://doi.org/10.2174/1874091X01004010053","url":null,"abstract":"<p><p>The aim of this study was to determine the changes of short chain fatty acids (SCFAs) in faeces of inflammatory bowel disease (IBD) patients compared to healthy subjects. SCFAs such as pyruvic, lactic, formic, acetic, propionic, isobutyric and butyric acids were analyzed by using high performance liquid chromatography (HPLC). This study showed that the level of acetic, 162.0 micromol/g wet faeces, butyric, 86.9 micromol/g wet faeces, and propionic acids, 65.6 micromol/g wet faeces, decreased remarkably in IBD faecal samples when compared with that of healthy individuals, 209.7, 176.0, and 93.3 micromol/g wet faeces respectively. On the contrary, lactic and pyruvic acids showed higher levels in faecal samples of IBD than in healthy subjects. In the context of butyric acid level, this study also found that the molar ratio of butyric acid was higher than propionic acid in both faecal samples. This might be due to the high intake of starch from rice among Malaysian population. It was concluded that the level of SCFAs differ remarkably between faecal samples in healthy subjects and that in IBD patients providing evidence that SCFAs more likely play an important role in the pathogenesis of IBD.</p>","PeriodicalId":515405,"journal":{"name":"The Open Biochemistry Journal","volume":"4 ","pages":"53-8"},"PeriodicalIF":0.0,"publicationDate":"2010-05-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.2174/1874091X01004010053","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29067707","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Cardiovascular dementia - a different perspective.","authors":"Udhaya Kumari,&nbsp;Klaus Heese","doi":"10.2174/1874091X01004010029","DOIUrl":"https://doi.org/10.2174/1874091X01004010029","url":null,"abstract":"<p><p>The number of dementia patients has been growing in recent years and dementia represents a significant threat to aging people all over the world. Recent research has shown that the number of people affected by Alzheimer's disease (AD) and dementia is growing at an epidemic pace. The rapidly increasing financial and personal costs will affect the world's economies, health care systems, and many families. Researchers are now exploring a possible connection among AD, vascular dementia (VD), diabetes mellitus (type 2, T2DM) and cardiovascular diseases (CD). This correlation may be due to a strong association of cardiovascular risk factors with AD and VD, suggesting that these diseases share some biologic pathways. Since heart failure is associated with an increased risk of AD and VD, keeping the heart healthy may prove to keep the brain healthy as well. The risk for dementia is especially high when diabetes mellitus is comorbid with severe systolic hypertension or heart disease. In addition, the degree of coronary artery disease (CAD) is independently associated with cardinal neuropathological lesions of AD. Thus, the contribution of T2DM and CD to AD and VD implies that cardiovascular therapies may prove useful in preventing AD and dementia.</p>","PeriodicalId":515405,"journal":{"name":"The Open Biochemistry Journal","volume":"4 ","pages":"29-52"},"PeriodicalIF":0.0,"publicationDate":"2010-03-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/0f/e1/TOBIOCJ-4-29.PMC2864432.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"28968948","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}