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1.
Anaerobe ; 74: 102540, 2022 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-35219837

RESUMO

Clostridioides difficile is the predominant pathogen responsible for antimicrobial associated diarrhea (AAD) and health care facility-associated infectious diarrhea. The role of C. difficile in China and its impact on public health have gained attention in recent years. Most clinical C. difficile isolates in China belong to multilocus sequence type clade 1 with sequence types (STs) 3, 35 and 54 predominating. Of note, the proportion of C. difficile isolates from clade 4, especially ST37 (PCR ribotype 17), is much higher in China than in other areas. In China, the antimicrobial-resistance profile of C. difficile is similar to that of other countries, demonstrating a higher resistance rate to erythromycin, clindamycin, and fluoroquinolones (ciprofloxacin, levofloxacin, and moxifloxacin). In general, susceptibility to vancomycin and metronidazole of clinical C. difficile in China is high, however, some resistance to metronidazole have recently been reported. Preclinical research on C. difficile in animals in China is limited, and different studies have reported varied isolation rates and antimicrobial resistance profiles. The diverse molecular types of C. difficile in China merit further epidemiological, genomic and evolutionary investigation. While the use of probiotics in preventing C. difficile infection (CDI) have received both support and opposition, the discovery of new probiotics and new formulations are showing promising results in combating the threat posed by CDI.


Assuntos
Anti-Infecciosos , Clostridioides difficile , Infecções por Clostridium , Infecção Hospitalar , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , China/epidemiologia , Clostridioides difficile/genética , Infecções por Clostridium/tratamento farmacológico , Infecções por Clostridium/epidemiologia , Infecção Hospitalar/tratamento farmacológico , Diarreia/tratamento farmacológico , Farmacorresistência Bacteriana/genética , Humanos , Metronidazol/farmacologia , Metronidazol/uso terapêutico , Testes de Sensibilidade Microbiana , Ribotipagem
2.
BMC Microbiol ; 20(1): 70, 2020 03 30.
Artigo em Inglês | MEDLINE | ID: mdl-32228454

RESUMO

BACKGROUND: It has been performed worldwidely to explore the potential of animals that might be a reservoir for community associated human infections of Clostridioides difficile. Several genetically undistinguished PCR ribotypes of C. difficile from animals and human have been reported, illustrating potential transmission of C. difficile between them. Pig and calf were considered as the main origins of C. difficile with predominant RT078 and RT033, respectively. As more investigations involved, great diversity of molecular types from pig and calf were reported in Europe, North American and Australia. However, there were quite limited research on C. difficile isolates from meat animals in China, leading to non-comprehensive understanding of molecular epidemiology of C. difficile in China. RESULTS: A total of 55 C. difficile were isolated from 953 animal stool samples, within which 51 strains were from newborn dairy calf less than 7 days in Shandong Province. These isolates were divided into 3 STs and 6 RTs, of which ST11/RT126 was predominant type, and responsible for majority antibiotic resistance isolates. All the isolates were resistant to at least one tested antibiotics, however, only two multidrug resistant (MDR) isolates were identified. Furthermore, erythromycin (ERY) and clindamycin (CLI) were the two main resistant antibiotics. None of the isolates were resistant to vancomycin (VAN), metronidazole (MTZ), tetracycline (TET), and rifampin (RIF). CONCLUSIONS: In this study, we analyzed the prevalence, molecular characters and antibiotic resistance of C. difficile from calf, sheep, chicken, and pig in China. Some unique features were found here: first, RT126 not RT078 were the dominant type from baby calf, and none isolates were got from pig; second, on the whole, isolates from animals display relative lower resistant rate to these 11 tested antibiotics, compared with isolates from human in China in our previous report. Our study helps to deep understanding the situation of C. difficile from economic animals in China, and to further study the potential transmission of C. difficile between meat animals and human.


Assuntos
Antibacterianos/farmacologia , Clostridioides difficile/classificação , Infecções por Clostridium/epidemiologia , Farmacorresistência Bacteriana , Animais , Animais Recém-Nascidos , Bovinos , Galinhas , China/epidemiologia , Clindamicina/farmacologia , Clostridioides difficile/efeitos dos fármacos , Clostridioides difficile/genética , Clostridioides difficile/isolamento & purificação , Infecções por Clostridium/microbiologia , Infecções por Clostridium/veterinária , Eritromicina/farmacologia , Fezes/microbiologia , Testes de Sensibilidade Microbiana , Epidemiologia Molecular , Tipagem Molecular , Prevalência , Ovinos , Suínos
3.
BMC Genomics ; 20(1): 796, 2019 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-31666016

RESUMO

BACKGROUND: Clade 5 Clostridioides difficile diverges significantly from the other clades and is therefore, attracting increasing attention due its great heterogeneity. In this study, we used third-generation sequencing techniques to sequence the complete whole genomes of three ST11 C. difficile isolates, RT078 and another two new ribotypes (RTs), obtained from three independent hospitalized elderly patients undergoing antibiotics treatment. Mobile genetic elements (MGEs), antibiotic-resistance, drug resistance genes, and virulent-related genes were analyzed and compared within these three isolates. RESULTS: Isolates 10,010 and 12,038 carried a distinct deletion in tcdA compared with isolate 21,062. Furthermore, all three isolates had identical deletions and point-mutations in tcdC, which was once thought to be a unique characteristic of RT078. Isolate 21,062 (RT078) had a unique plasmid, different numbers of transposons and genetic organization, and harboring special CRISPR spacers. All three isolates retained high-level sensitivity to 11 drugs and isolate 21,062 (RT078) carried distinct drug-resistance genes and loss of numerous flagellum-related genes. CONCLUSIONS: We concluded that capillary electrophoresis based PCR-ribotyping is important for confirming RT078. Furthermore, RT078 isolates displayed specific MGEs, indicating an independent evolutionary process. In the further study, we could testify these findings with more RT078 isolates of divergent origins.


Assuntos
Clostridioides difficile/genética , Elementos de DNA Transponíveis , Evolução Molecular , Genoma Bacteriano , Clostridioides difficile/isolamento & purificação , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Farmacorresistência Bacteriana/genética , Sequenciamento Completo do Genoma
4.
Anaerobe ; 60: 102094, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31499177

RESUMO

It is known that antibiotic usage is associated with the development of Clostridioides difficile infection (CDI), especially clindamycin, third-generation cephalosporins, and fuoroquinolones. Antibiotic resistance rates to many antibiotics varies a lot by study. We performed a study focused on antibiotic resistance in clinical isolates of C. difficile from more widespread geographic regions across China. Of 319 C. difficile isolates tested against 11 antibiotics, 313 (98.1%) were resistant to at least one antibiotic. The highest rate of resistance was to ciprofloxacin, clindamycin, and erythromycin across all age groups, similar to previous studies. However, all isolates were susceptible to metronidazole and vancomycin. Overall the resistance rate to tested antibiotics was lower than other reports in China except for chloramphenicol and meropenem. Genotype ST37/RT017 in clade 4 was resistant to more antibiotics than other types. Unexpectedly, RT078 isolates in this study were susceptible to almost all tested antibiotics. In addition, the proportion of multi-drug resistant (MDR) isolates observed (17%) in this study was much lower than several European studies (up to 55%) and a previous study in China (78%). Although isolates from patients aged between 65 and 85 were more resistant to antibiotics in comparison to other age groups, MDR isolates were still detected in children below 2-years of age. The highest percentage of MDR isolates was determined in South China, an area that is most developed economically. The clade 4, RT017 (ST37) has been associated with outbreaks in Europe and North America and is responsible for most C. difficile infections (CDIs) in Asia. In addition, RT017 is often clindamycin and fluoroquinolone resistant. This study provided a relatively comprehensive description of antibiotic resistance of C. difficile in China, and further elucidates the epidemiology and antibiotic resistance of clinical isolates of C. difficile in China at a national level.


Assuntos
Antibacterianos/farmacologia , Clostridioides difficile/efeitos dos fármacos , Infecções por Clostridium/epidemiologia , Infecções por Clostridium/microbiologia , Farmacorresistência Bacteriana , Adolescente , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Toxinas Bacterianas/genética , Criança , Pré-Escolar , China/epidemiologia , Clostridioides difficile/classificação , Clostridioides difficile/genética , Clostridioides difficile/isolamento & purificação , Farmacorresistência Bacteriana Múltipla , Genótipo , Geografia Médica , Humanos , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Epidemiologia Molecular , Vigilância em Saúde Pública , Ribotipagem , Adulto Jovem
5.
Se Pu ; 42(3): 304-308, 2024 Mar 08.
Artigo em Chinês | MEDLINE | ID: mdl-38503708

RESUMO

To solve the problems of the lack of property research in organic synthesis experiments and the relative independence of instrumental analytical methods in experiments, we designed a comprehensive undergraduate experiment based on mechanofluorochromic materials. In this project, 4-[bis(4-methylphenyl)amino] benzaldehyde was synthesized via the Vilsmeier-Haack reaction using 4,4'-dimethyltriphenylamine as the raw material. The product was then characterized by mass spectrometry, infrared absorption spectroscopy, and nuclear magnetic resonance spectroscopy. The solvatofluorochromism and mechanofluorochromism of the target material were studied using ultraviolet-visible absorption spectroscopy, fluorescence spectroscopy, etc. Furthermore, the mechanism of mechanofluorochromism was determined using powder X-ray diffraction. Organic synthesis and a series of instrumental analytical methods were combined to form an integrated experiment. The experiment is interesting, scientific, and comprehensive for undergraduates as a creative exercise; moreover, it can inspire their interest in chemical research, cultivate a variety of experimental operation abilities, improve creative-thinking skills, and encourage the development of effective solutions to existing problems in chemical experiments.

6.
3 Biotech ; 13(2): 54, 2023 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-36685319

RESUMO

This study developed a new single-tube multiplex real-time PCR method for detecting toxigenic C. difficile directly from fecal samples using tcdA, tcdB, cdtB, and internal gene tpi as targets, which could be performed on kinds of polymerase chain reaction device including point-of-care testing (POCT), with improved detection efficiency. The specificity, sensitivity, and repeatability of each gene was evaluated using 69 C. difficile isolates and 74 fecal samples. Results were compared with established PCR, qPCR, and ELISA methods. Interspecies specificity was 100% based on six common intestinal pathogens (Escherichia coli, Enterococcus Faecium, Enterococcus faecalis, Clostridium perfringens, Bacteroides fragilis, Clostridium botulinum). The lower detection limit (LDL) for tcdA, tcdB, and cdtB with pure C. difficile DNA was 101,100, and 100 copies/µL, respectively, the coefficients of variation among different experimental batches and within each experimental batch were both less than 3%, which shows that this method has strong repeatability. And the LDL of fecal DNA was 5 × 100, 5 × 103, and 5 × 102 colony-forming units (CFU)/g, respectively. In addition, the efficiency for detection of tcdA was compared with established PCR and real-time PCR methods, demonstrating high consistency (98.4%) and similar sensitivity. ELISA was used to confirm inconsistent results, which were identical with our method. The sensitivity and specificity for detecting toxigenic C. difficile in fecal samples were 96.49% and 94.12% compared with the toxigenic culture (TC). This method effectively identified the toxigenic and non-toxigenic strains with high specificity, sensitivity, and repeatability, and could reduce the false positive rate of tcdA, and accurately identify the typical Asian strain RT017, making it potentially contribute to the surveillance of CDI in China. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-022-03434-6.

7.
Front Cell Infect Microbiol ; 13: 1109153, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36992688

RESUMO

Introduction: Clostridioides difficile (C. difficile) is a nosocomial bacterial pathogen that causes antibiotic-associated diarrhea mediated by cellular exotoxins secreted into the intestine during bacterial growth. Multilocus sequence typing (MLST) and PCR ribotyping are the main molecular typing for C. difficile. Whole genome sequencing (WGS) core genome multilocus sequence typing (cgMLST) was developed for genetic evolution and outbreak investigation of C. difficile with higher precision and accuracy. Methods: A total of 699 whole (complete and draft) genome sequences of distinct C. difficile strains were used in this study to identify core gene set (2469 core genes) and the cgMLST scheme for the phylogeny analysis of C. difficile. This cgMLST pipeline was then carried the Chinese Pathogen Identification Net (China PIN) for surveillance of C. difficile in China. Within the China PIN, 195 WGS of C. difficile and an outbreak of CDI with 12 WGS of C. difficile were used to evaluate the cgMLST pipeline. Results: The result displayed that mostly tested C. difficile isolates could be successfully divided into 5 classic clades and the outbreak event was also successfully identified. Discussion: The results are meaningful and offer a practicable pipeline for a national-wide surveillance of C. difficile in China.


Assuntos
Clostridioides difficile , Tipagem de Sequências Multilocus/métodos , Clostridioides difficile/genética , Genoma Bacteriano , Clostridioides , Filogenia , China/epidemiologia
8.
J Zhejiang Univ Sci B ; 21(1): 53-63, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31898442

RESUMO

In manned deep-space exploration, extremely isolated environments may adversely affect the mood and cognition of astronauts. Horticultural plants and activities have been proven to be effective in improving their physical, psychological, and cognitive states. To assess the effects of applying horticultural plants and activities in isolated environments, this study investigated the influence of viewing strawberry plants on the mood of people in a laboratory experiment as indicated by heart rate, salivary cortisol, and psychological scales. The results showed that heart rate and salivary cortisol were significantly decreased after viewing strawberry plants for 15 min. "Tension" and "confusion" scored using the Profile of Mood States negative mood subscales, and anxiety levels measured using the State-Trait Anxiety Inventory scale were also significantly reduced. This study further explored the impact of viewing strawberry plants on cognition. A notable reduction of the subjects' reaction time after 15-min plant viewing was observed. Based on these findings, a long-duration isolated experiment in a bioregenerative life support system-"Lunar Palace I"-was conducted. A similar trend was obtained that crew members' mood states were improved by viewing the strawberry plants, but no significant change was observed. This study provided some experimental evidence for the benefits of interacting with strawberry plants in isolated environments.


Assuntos
Afeto , Cognição , Fragaria , Adulto , Emoções , Meio Ambiente , Feminino , Frequência Cardíaca , Humanos , Hidrocortisona/análise , Masculino , Saliva/química
9.
Chin Med J (Engl) ; 132(4): 446-453, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30707165

RESUMO

BACKGROUND: Adhesion, biofilm formation, yeast-hyphal transition, secretion of enzymes, and hemolytic activity are all considered important factors in Candida tropicalis infection. However, DNA sequence data for this pathogen are limited. In this study, the polymorphism and heterogeneity of genes agglutinin-like sequences (ALS)2, Lipase (LIP)1, LIP4, and secretory aspartyl proteinase tropicalis (SAPT)1-4 as well as the relationship between phenotype and genotype were analyzed. METHODS: This study started in August 2013, and ended in July 2017. The complete length of ALS2, LIP1, LIP4, and SAPT1-4 of 68 clinical C. tropicalis isolates was sequenced. Single nucleotide polymorphisms (SNPs) as well as insertions and deletions (indels) were identified within these genes. In addition, phenotypic characteristics of the virulent factors, including adhesion and the secretion of aspartyl proteinases and phospholipases, were determined. RESULTS: There were 73, 24, 17, 16, 13, and 180 SNPs in the genes LIP1, LIP4, SAPT1, SAPT2, SAPT3, and SAPT4, respectively. Furthermore, 209 SNPs were identified in total for the gene ALS2. Interestingly, large fragment deletions and insertions were also found in ALS2. Isolate FXCT 01 obtained from blood had deletions on all 4 sites and showed the lowest adhesion ability on the polymethylpentene surface. In addition, isolates with deletions in the regions 1697 to 1925 and 2073 to 2272 bp displayed relatively low abilities for adhesion and biofilm formation, and this phenotype correlated with the deletions found in ALS2. LIP1, SAPT4, and ALS2 displayed great heterogeneity among the isolates. Large deletions found in gene ALS2 appeared to be associated with the low ability of adhesion and biofilm formation of C. tropicalis. CONCLUSION: This study might be useful for deeper explorations of gene function and studying the virulent mechanisms of C. tropicalis.


Assuntos
Candida tropicalis/genética , Polimorfismo de Nucleotídeo Único , Aderência Bacteriana , Biofilmes , Candida tropicalis/patogenicidade , Lipase/genética , Virulência/genética
10.
mSystems ; 4(2)2019.
Artigo em Inglês | MEDLINE | ID: mdl-30944881

RESUMO

Horizontal gene transfer of mobile genetic elements (MGEs) accounts for the mosaic genome of Clostridium difficile, leading to acquisition of new phenotypes, including drug resistance and reconstruction of the genomes. MGEs were analyzed according to the whole-genome sequences of 37 C. difficile isolates with a variety of sequence types (STs) within clade 4 from China. Great diversity was found in each transposon even within isolates with the same ST. Two novel transposons were identified in isolates ZR9 and ZR18, of which approximately one third to half of the genes showed heterogenous origins compared with the usual intestinal bacterial genes. Most importantly, catD, known to be harbored by Tn4453a/b, was replaced by aac(6') aph(2'') in isolates 2, 7, and 28. This phenomenon illustrated the frequent occurrence of gene exchanges between C. difficile and other enterobacteria with individual heterogeneity. Numerous prophages and CRISPR arrays were identified in C. difficile isolates of clade 4. Approximately 20% of spacers were located in prophage-carried CRISPR arrays, providing a new method for typing and tracing the origins of closely related isolates, as well as in-depth studies of the mechanism underlying genome remodeling. The rates of drug resistance were obviously higher than those reported previously around the world, although all isolates retained high sensitivity to vancomycin and metronidazole. The increasing number of C. difficile isolates resistant to all antibiotics tested here suggests the ease with which resistance is acquired in vivo. This study gives insights into the genetic mechanism of microevolution within clade 4. IMPORTANCE Mobile genetic elements play a key role in the continuing evolution of Clostridium difficile, resulting in the emergence of new phenotypes for individual isolates. On the basis of whole-genome sequencing analysis, we comprehensively explored transposons, CRISPR, prophage, and genetic sites for drug resistance within clade 4 C. difficile isolates with different sequence types. Great diversity in MGEs and a high rate of multidrug resistance were found within this clade, including new transposons, Tn4453a/b with aac(6') aph(2'') instead of catD, and a relatively high rate of prophage-carried CRISPR arrays. These findings provide important new insights into the mechanism of genome remodeling within clade 4 and offer a new method for typing and tracing the origins of closely related isolates.

11.
Front Microbiol ; 9: 845, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29760687

RESUMO

Clostridium difficile infection (CDI) has become a worldwide public health problem causing high mortality and a large disease burden. Molecular typing and analysis is important for surveillance and infection control of CDI. However, molecular characterization of C. difficile across China is extremely rare. Here, we report on the toxin profiles, molecular subtyping with multilocus sequence typing (MLST) and PCR ribotyping, and epidemiological characteristics of 199 C. difficile isolates collected between 2010 through 2015 from 13 participating centers across China. We identified 35 STs and 27 ribotypes (RTs) among the 199 C. difficile isolates: ST35 (15.58%), ST3 (15.08%), ST37 (12.06%), and RT017 (14.07%), RT001 (12.06%), RT012 (11.56%) are the most prevalent. One isolate with ST1 and 8 isolates with ST 11 were identified. We identified a new ST in this study, denoted ST332. The toxin profile tcdA+tcdB+tcdC+tcdR+tcdE+CDT- (65.83%) was the predominant profile. Furthermore, 11 isolates with positive binary toxin genes were discovered. According to the PCR ribotyping, one isolate with RT 027, and 6 isolates with RT 078 were confirmed. The epidemiological characteristics of C. difficile in China shows geographical differences, and both the toxin profile and molecular types exhibit great diversity across the different areas.

12.
Zhonghua Bing Li Xue Za Zhi ; 36(3): 155-9, 2007 Mar.
Artigo em Chinês | MEDLINE | ID: mdl-17535680

RESUMO

OBJECTIVE: To study the expression of E-cadherin and beta-catenin in neuroblastomas of various degrees of differentiation, and to investigate their molecular mechanisms in correlation with clinicopathologic parameters. METHODS: Immunohistochemistry EnVision method was used to detect E-cadherin and beta-catenin expression in 90 paraffin-embedded tissue samples of neuroblastomas. The methylation status of CpG islands of E-cadherin promoter was investigated by MSP in 7 fresh tissue and 24 paraffin-embedded tissue samples. The mutation status of exon 3 of beta-catenin gene was studied by PCR in 7 fresh tissue samples. Statistical analysis of the data was performed by SPSS software. RESULTS: E-cadherin and beta-catenin were abnormally expressed in neuroblastomas in general. The expression of beta-catenin in well-differentiated neuroblastoms was markedly higher (47/70, 67.1%) than that of the poorly differentiated tumors (8/20, 40.0%). There was a markedly decreased expression of both genes in tumors with lymph node metastasis than those without. Demethylation was seen in some regions of the promoter of E-cadherin in 31 cases of nuroblatomas. PCR of the exon 3 of beta-catenin followed by DNA sequencing demonstrated rearrangements and mutations in 7 cases, including 2 cases harboring identical point mutation at gene position 27184, leading to a T-->A alteration. CONCLUSIONS: The abnormal over-expression of E-cadherin in neuroblastomas is independent of the methylation status of their promoter sequences. The abnormal expression of beta-catenin may be related to mutational changes at exon 3 of the gene.


Assuntos
Caderinas/metabolismo , Neoplasias do Mediastino/metabolismo , Neuroblastoma/metabolismo , Neoplasias Retroperitoneais/metabolismo , beta Catenina/metabolismo , Caderinas/genética , Criança , Pré-Escolar , Ilhas de CpG/genética , Metilação de DNA , DNA de Neoplasias/genética , Éxons , Feminino , Ganglioneuroblastoma/genética , Ganglioneuroblastoma/metabolismo , Ganglioneuroblastoma/patologia , Rearranjo Gênico , Humanos , Lactente , Metástase Linfática , Masculino , Neoplasias do Mediastino/genética , Neoplasias do Mediastino/patologia , Neuroblastoma/genética , Neuroblastoma/patologia , Mutação Puntual , Regiões Promotoras Genéticas/genética , Neoplasias Retroperitoneais/genética , Neoplasias Retroperitoneais/patologia , Análise de Sequência de DNA , beta Catenina/genética
13.
Medicine (Baltimore) ; 96(32): e7634, 2017 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28796044

RESUMO

The aim of the study was to evaluate the potential role of CD40/CD40 ligand (CD40L) and CD134/CD134 ligand (CD134L) in the development of coronary heart disease (CHD) via the performance of a case-control study.The research objects were 234 cases of CHD patients and 120 cases of well-matched normal controls. Following the separation of peripheral blood mononuclear cells (PBMCs), real-time quantitative PCR (qRT-PCR), Western blot, immunohistochemistry, and flow cytometry were applied for the detection of mRNA levels and expression levels of CD40/CD40L and CD134/CD134L; meanwhile, intercellular adhesion molecule-1 (ICAM-1) and Fas protein mRNA levels were detected using qRT-PCR.There was no statistical difference in the comparison of baseline characteristics between groups, indicating comparability between groups. qRT-PCR and Western blot analysis indicated that CD40/CD40L and CD134/CD134L mRNA and protein expression levels were all increased in the CHD group than those in the control group. Flow cytometry further confirmed the similar tendency. Meanwhile, ICAM-1 and Fas protein mRNA levels were elevated in the CHD group and positively correlated with the above parameters. Furthermore, CD40/CD40L expression rates were negatively correlated with gender and different types of CHD. Meanwhile, CD134/CD134L expressions were also higher in male patients, in patients with family history, previous history of hypertension, diabetes, and cerebrovascular diseases.CD40/CD40L and CD134/CD134L are increased and may have potential correlation with clinical pathological features of patients with CHD. Further in-depth exploration of costimulatory molecules for CHD guidance as well as intrinsic mechanisms are needed combined with in vivo and in vitro experiments.


Assuntos
Antígenos CD40/biossíntese , Ligante de CD40/biossíntese , Doença das Coronárias/fisiopatologia , Ligante OX40/biossíntese , Receptores OX40/biossíntese , Adulto , Idoso , Estudos de Casos e Controles , Feminino , Citometria de Fluxo , Humanos , Molécula 1 de Adesão Intercelular/biossíntese , Masculino , Pessoa de Meia-Idade , RNA Mensageiro , Reação em Cadeia da Polimerase em Tempo Real , Receptor fas/biossíntese
14.
Hepatobiliary Pancreat Dis Int ; 4(2): 281-4, 2005 May.
Artigo em Inglês | MEDLINE | ID: mdl-15908330

RESUMO

BACKGROUND: Playing a central role in hepatic fibrosis, hepatic stellate cell (HSC) has made itself the major target of research. The limited supply of HSC, however, can not meet the ever growing need of experiment. Establishment and identification of novel immortalized HSC line thus may be urgently required. METHODS: Primary HSCs were isolated from a normal adult male Sprague-Dawley rat by in situ perfusion with collagenase IV and pronase E, and then were purified by single-step density gradient centrifugation with nycodenz. Once they reached total activation in culture, a new immortalized myofibroblast-like HSC line was established through cellular cloning. Its characteristics were identified by means of immunocytochemical staining, light microscopy, transmission electron microscopy, and growth curve analysis. RESULTS: The novel HSC line, termed HSC-PQ, had a doubling time of about 75 hours in the Dulbecco's modified Eagle medium (DMEM) containing 20% fetal bovine serum. Most of the main morphological characteristics of the differentiated primary HSC could be detected in HSC-PQ cell, while functional features of activated HSC such as alpha-smooth muscle actin, desmin, collagen type I, collagen type III, fibronectin, laminin and other extracellular matrix proteins could also be found in it except for collagen type IV. In contrast, fat droplets and autofluorescence of vitamin A disappeared in the HSC-PQ line. This cell line had been maintained in culture for over 30 passages and more than 1 year with little alternation in biological characteristics. CONCLUSION: A new rat HSC line (HSC-PQ) has been successfully established. It consistently retains the characteristics of activated primary HSC, and has proved to be immortalized.


Assuntos
Linhagem Celular , Hepatócitos/citologia , Hepatócitos/fisiologia , Análise de Variância , Animais , Apoptose/fisiologia , Linhagem Celular Transformada , Proliferação de Células , Citometria de Fluxo , Imuno-Histoquímica , Cirrose Hepática/patologia , Cirrose Hepática/fisiopatologia , Masculino , Microscopia Eletrônica de Varredura , Probabilidade , Ratos , Ratos Sprague-Dawley , Sensibilidade e Especificidade , Coleta de Tecidos e Órgãos
15.
World J Gastroenterol ; 8(3): 476-9, 2002 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-12046073

RESUMO

AIM: To investigate the influence of hepatic arterial blockage on blood perfusion of transplanted cancer in rat liver and the expression of vascular endothelial growth factor (VEGF) and matrix metalloproteinase-1 (MMP-1), and to explore the mechanisms involved in transarterial embolization (TAE)-induced metastasis of liver cancer preliminarily. METHODS: Walker 256 carcinosarcoma was transplanted into rat liver to establish the liver cancer model. Hepatic arterial ligation (HAL) was used to block the hepatic arterial blood supply and simulate TAE. Blood perfusion of tumor in control, laparotomy control, and HAL group was analyzed by Hoechst 33342 labeling assay, the serum VEGF level was assayed by ELISA, the expression of VEGF and MMP-1 mRNA was detected by in situ hybridization. RESULTS: Two days after HAL, the number of Hoechst 33342 labeled cells which represent the blood perfusion of tumor directly and hypoxia of tumor indirectly in HAL group decreased significantly compared with that in control group (329+/-29 vs 384+/-19, P<0.01). The serum VEGF level in the HAL group increased significantly as against that of the control group (93 ng.L(-1)+/-44 ng.L(-1) vs 55 ng.L(-1)+/-19 ng.L(-1), P<0.05). The expression of VEGF and MMP-1 mRNA in the tumor tissue of the HAL group increased significantly compared with that of the control and the laparotomy control groups (P<0.05). The blood perfusion data of the tumor, represented by the number of Hoechst 33342 labeled cells, showed a good linear inverse correlation with the serum VEGF level (r=-0.606, P<0.05) and the expression of VEGF mRNA in the tumor tissue ( r =-0.338, P<0.01). CONCLUSION: Blockage of hepatic arterial blood supply results in decreased blood perfusion and increased expression of metastasis-associated genes VEGF and MMP-1 of transplanted liver cancer in rats. Decreased blood perfusion and hypoxia may be the major cause of up-regulated expression of VEGF.


Assuntos
Fatores de Crescimento Endotelial/genética , Neoplasias Hepáticas Experimentais/irrigação sanguínea , Neoplasias Hepáticas Experimentais/genética , Linfocinas/genética , Metaloproteinase 1 da Matriz/genética , Animais , Carcinoma 256 de Walker/irrigação sanguínea , Carcinoma 256 de Walker/genética , Carcinoma 256 de Walker/secundário , Embolização Terapêutica/efeitos adversos , Fatores de Crescimento Endotelial/sangue , Expressão Gênica , Artéria Hepática , Ligadura , Linfocinas/sangue , Masculino , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Neoplásico/genética , RNA Neoplásico/metabolismo , Ratos , Ratos Wistar , Fator A de Crescimento do Endotélio Vascular , Fatores de Crescimento do Endotélio Vascular
16.
World J Gastroenterol ; 9(10): 2154-9, 2003 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-14562368

RESUMO

AIM: Ets1 proto-oncogene is a transcription factor involved in the activation of several genes of tumor invasion and metastasis. We aimed to determine the relationship between the extent and intensity of Ets1 expression and patients' clinicopathological factors in gastric carcinoma. METHODS: Immunohistochemical analysis was performed for gastric tumor paraffin-embedded sections, followed by image analysis. RESULTS: Ets1 was not expressed in the normal gastric epithelium and its surrounding cells. The percentage of Ets1 expressing cells detected increased significantly in both epithelial tumor and stromal cells from high T classification, lymph node metastasis positive, clinical advanced-stage groups (P<0.001). The level of Ets1 staining in epithelial tumor cells also reflected the degree of cell differentiation. The percentage of epithelial and stromal cells expressing Ets1 was significantly correlated with the presence of lymph node metastasis (P=0.014 and P<0.001 respectively). Ets1 expression was not observed in tissue samples from patients with benign gastric ulcers. CONCLUSION: Ets1 protein expression in epithelial tumor cells reflects the degree of differentiation, and the percentage of Ets1 positive tumor and stromal cells correlates with lymph node metastasis. Thus Ets1 is a valuable marker of malignant potential in terms of invasiveness and metastasis of gastric carcinoma. It is also possible that inhibition of Ets1 is a potential avenue for therapy in gastric cancer.


Assuntos
Biomarcadores Tumorais/metabolismo , Carcinoma de Células em Anel de Sinete/metabolismo , Carcinoma de Células em Anel de Sinete/secundário , Proteínas Proto-Oncogênicas/metabolismo , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia , Fatores de Transcrição/metabolismo , Adenocarcinoma/metabolismo , Adenocarcinoma/secundário , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma/metabolismo , Carcinoma/secundário , Feminino , Humanos , Metástase Linfática , Masculino , Pessoa de Meia-Idade , Proto-Oncogene Mas , Proteína Proto-Oncogênica c-ets-1 , Proteínas Proto-Oncogênicas c-ets
17.
Hepatogastroenterology ; 50(50): 349-53, 2003.
Artigo em Inglês | MEDLINE | ID: mdl-12749218

RESUMO

BACKGROUND/AIMS: To observe if octreotide can augment the effects of hepatic arterial occlusion for transplanted cancer in rat's liver. METHODOLOGY: Walker 256 carcinosarcoma was transplanted into rat's liver to create the liver cancer model. Hepatic arterial ligation was used to block the hepatic arterial blood supply. Rats bearing tumor were divided into three groups: control group, HAL (hepatic arterial ligation) group, and HAL plus octreotide group. Change of tumor volume and tumor growth inhibiting rate after therapy were evaluated. Hoechst 33342 labeling assay was used to analyze the blood perfusion of tumor (the labeled cells' number presenting blood perfusion). Expression of vascular endothelial growth factor and matrix metalloproteinase-1 mRNA was detected by in situ hybridization, and the level of serum vascular endothelial growth factor was assayed by ELISA. RESULTS: Six days after hepatic arterial ligation, the mean tumor volume in control group, HAL group, and HAL plus octreotide group was 0.103 +/- 0.043 cm3, 0.030 +/- 0.018 cm3, and 0.016 +/- 0.005 cm3, respectively. The tumor volume in the two behind groups was smaller than that in the control group (P < 0.01), and the tumor growth-inhibiting rate was 70.8%, and 84.5%, respectively. Compared with the HAL group, the tumor volume in HAL plus octreotide group decreased significantly (P < 0.05). Hoechst 33342 labeled cells' number in control group, HAL group, and HAL plus octreotide group was 369.7 +/- 30.2, 344.1 +/- 26.0, and 323.2 +/- 40.4, respectively. The number in HAL combined with octreotide group decreased significantly compared with that in control group (P < 0.05), which suggested that the blood perfusion of tumor in HAL plus octreotide group decreased significantly. The expression of vascular endothelial growth factor and matrix metalloprotenase-1 mRNA decreased slightly, but not significantly in HAL plus octreotide group compared with that in HAL group (P > 0.05). CONCLUSIONS: The results suggest that octreotide can promote the effects of hepatic arterial occlusion therapy for transplanted cancer in rat's liver. Decreasing the blood perfusion of tumor after hepatic arterial blockage maybe one of its major mechanisms.


Assuntos
Antineoplásicos Hormonais/uso terapêutico , Carcinoma 256 de Walker/terapia , Artéria Hepática/cirurgia , Neoplasias Hepáticas Experimentais/terapia , Octreotida/uso terapêutico , Animais , Modelos Animais de Doenças , Fatores de Crescimento Endotelial/sangue , Peptídeos e Proteínas de Sinalização Intercelular/sangue , Ligadura , Linfocinas/sangue , Masculino , Metaloproteinase 1 da Matriz/sangue , Distribuição Aleatória , Ratos , Ratos Wistar , Fator A de Crescimento do Endotélio Vascular , Fatores de Crescimento do Endotélio Vascular
18.
Hepatogastroenterology ; 51(56): 381-6, 2004.
Artigo em Inglês | MEDLINE | ID: mdl-15086165

RESUMO

BACKGROUND/AIMS: This study investigates the influence of hepatic arterial occlusion (HAO) on blood perfusion of transplanted cancer in rat's liver, and the expression of vascular endothelial growth factor (VEGF) and matrix metalloproteinase-1 (MMP-1) and explores the mechanisms involved in transcatheter arterial embolization (TAE)-induced metastasis of liver cancer preliminarily. METHODOLOGY: Walker 256 carcinosarcoma was transplanted into rat's liver to create the liver cancer model. Hepatic arterial ligation (HAL) was used to block the hepatic arterial blood supply and simulate TAE. Rats bearing tumor were divided into three groups: control, laparotomy control, and HAL groups. Blood perfusion of tumor was analyzed by a Hoechst 33342 labeling assay. The level of serum VEGF was assayed by ELISA; and the expression of VEGF and MMP-1 mRNA was detected by in situ hybridization. RESULTS: Two days after HAL, the number of Hoechst 33342 labeled cells which represent the blood perfusion of the tumor directly and hypoxia of tumor indirectly in the HAL group decreased significantly compared with that in the control group (329.1+/-29.3 vs. 383.6+/-19.2, P<0.01). The level of serum VEGF in the HAL group increased significantly compared with that of the control group (92.5+/-43.9 pg/mL vs. 54.9+/-19.3 pg/mL, P<0.05). The expression of VEGF and MMP-1 mRNA in the tumor tissue of the HAL group increased significantly compared with that of the control and the laparotomy control groups (P<0.05). The blood perfusion data of the tumor, represented by number of Hoechst 33342 labeled cells, showed an inverse correlation with the expression of VEGF mRNA in tumor tissue (P<0.05). While 6 days after HAL, the blood perfusion of tumor in HAL group decreased and the expression of VEGF and MMP-1 increased only slightly, not significantly, compared with that in the control group. CONCLUSIONS: In conclusion, blockage of hepatic arterial blood supply results in transient decreased blood perfusion and increased expression of metastasis-associated genes VEGF and MMP-1 of transplanted liver cancer in rats. Decreased blood perfusion and hypoxia may be the major reason for up-regulated expression of VEGF. Better understanding of the mechanisms involved with TAE-induced metastasis may lead to the enhancement of the long-term effects of TAE for liver cancer.


Assuntos
Carcinoma 256 de Walker/metabolismo , Carcinoma 256 de Walker/terapia , Embolização Terapêutica/efeitos adversos , Neoplasias Hepáticas Experimentais/metabolismo , Neoplasias Hepáticas Experimentais/terapia , Metaloproteinase 1 da Matriz/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Carcinoma 256 de Walker/irrigação sanguínea , Carcinoma 256 de Walker/secundário , Artéria Hepática/cirurgia , Hibridização In Situ , Ligadura , Neoplasias Hepáticas Experimentais/irrigação sanguínea , Neoplasias Hepáticas Experimentais/patologia , Masculino , Distribuição Aleatória , Ratos , Ratos Endogâmicos , Regulação para Cima
19.
Di Yi Jun Yi Da Xue Xue Bao ; 21(12): 940-941, 2001.
Artigo em Inglês | MEDLINE | ID: mdl-12426173

RESUMO

OBJECTIVE: To evaluate the efficacy of myocardial contrast echocardiography (MCE) in the assessment of myocardial viability, with positron emission tomography (PET) as the golden standard. METHODS: Eleven patients with anterior wall Q wave myocardial infarction were enrolled in this study, who received successful percutaneous transluminal coronary angioplasty (PTCA) 3 to 19 months prior to the examinations by PET and MCE that were completed within 2 d. RESULTS: MCE score for the segments of necrotic myocardium, viable myocardium and normal myocardium was mostly 0, 0.5 and 1 respectively, and there was a significant difference between the grades of MCE score in identifying myocardial viability. In terms of diagnosing myocardium survival, MCE result was closely correlated with that of PET (with the correlation index rp of 0.78). CONCLUSION: Myocardial perfusion evaluation can be effectively accomplished by MCE, which might serve as a new approach to assess myocardial viability in clinical practice.

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