RESUMO
Methyl glycolate (MG) is an important biodegradable PGA plastic monomer. Herein, a green approach to synthesize MG by methanolysis of glucose is proposed, in which the subcritical methanol and phenol/quinone redox system were combined to promote the reversible C-C cleavage and oxidation during the cascade reaction of glucose to MG.
Assuntos
Glucose , Metanol , Glicolatos , Fenol , FenóisRESUMO
Background and Aims: Irritable bowel syndrome (IBS) is a common functional gastrointestinal disorder. However, the underlying mechanism of IBS is not fully understood. The aim of this study was to investigate potential mechanism and novel biomarkers of IBS through evaluation of the metabolomic and microbiologic profile. Methods: Fecal samples were collected from 15 irritable bowel syndrome patients and 15 healthy controls. By using gas chromatography coupled to time-of-flight mass spectrometry (GC-TOFMS) and 16S rDNA amplicon sequencing, fecal metabolites and microbiota of healthy controls and the IBS patients were measured. Results: IBS patients had a significantly differential metabolite profile as compared to healthy controls, and 4 clusters with 31 metabolites, including a group of amino acids and fatty acids, were significantly up-regulated as compared to the healthy controls. In addition, 19 microbes were significantly up-regulated, and 12 microbes were down-regulated in the IBS group, when compared with the healthy controls. Some clusters of fecal metabolites or microorganisms were significantly correlated with the severity of IBS symptoms, such as the frequency of abdominal pain/discomfort and the number of bowel movements. Correlation of the metabolite levels with abundances of microbial genera showed some statistically significant metabolite-microbe associations. Four differentially abundant amino acids clustered together were positively correlated with some microbes, including Lachnospira, Clostridium, and so on. Conclusion: The finding of this study puts a global perspective on metabolomics and microbiota profiling in IBS patients and provides a theoretical basis for future research on pathophysiology of IBS.
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Microbioma Gastrointestinal , Síndrome do Intestino Irritável/etiologia , Síndrome do Intestino Irritável/metabolismo , Metaboloma , Metabolômica , Estudos de Casos e Controles , Biologia Computacional/métodos , Suscetibilidade a Doenças , Metabolismo Energético , Fezes/microbiologia , Feminino , Humanos , Síndrome do Intestino Irritável/diagnóstico , Masculino , Metabolômica/métodos , Metagenômica/métodos , RNA Ribossômico 16S/genética , Índice de Gravidade de Doença , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por MatrizRESUMO
BACKGROUND: Only a paucity of large-scale perspective and cross-sectional studies on H. pylori infection in China have been published. The purpose of this study was to investigate the prevalence and risk factors for H. pylori infection among residents of Jidong community located in Hebei Province of China. METHODS: A perspective, cross-sectional study was conducted in Jidong community. Questionnaires and 13C-urea breath test were performed, and 10-ml blood samples were obtained for laboratory tests. RESULTS: Four thousand seven hundred ninety-six subjects were enrolled in this study, and 2506 (52.25%) were H. pylori positive. There was no difference in prevalence between both sexes (P = 0.5974). Age (P = 0.004) and education level (P = 0.0128) were significantly associated with H. pylori infection, and there were statistical trends in the prevalence across five age subgroups (χ2 test for trend = 23.5; P < 0.001) and education levels (χ2 test for trend = 19.50; P < 0.001). H. pylori infection was also associated with marital status (P = 0.0243), source of drinking water (P = 0.0433), frequency of eating raw garlic (P = 0.0310), alcohol drinking (P = 0.0207), knowledge about H. pylori transmission route (P = 0.0125) and related diseases (P = 0.0257). Age, alcohol drinking and knowledge about transmission route were found to be independent predictors of H. pylori infection. CONCLUSIONS: More than half of the population was infected with H. pylori in Jidong community. The socio-demographic profiles, socio-economic factors and lifestyle are worthy taking into consideration to prevent diseases associated with H. pylori infection. Understanding the prevalence and risk patterns for H. pylori infection in China will help in prioritizing public health efforts to better manage the H. pylori infection.
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Infecções por Helicobacter , Helicobacter pylori/isolamento & purificação , Adulto , Testes Respiratórios/métodos , China/epidemiologia , Estudos Transversais , Feminino , Infecções por Helicobacter/diagnóstico , Infecções por Helicobacter/epidemiologia , Humanos , Estilo de Vida , Masculino , Pessoa de Meia-Idade , Prevalência , Fatores de Risco , Fatores SocioeconômicosRESUMO
Irritable bowel syndrome (IBS) is a common gastrointestinal disorder that is associated with psychological stress. However, the full landscape of IBSrelated epigenetic factors remains unveiled and needs to be elucidated. The wateravoidance stress (WAS) method was used to induce a rat IBS model. Each rat was monitored, and its defecation and behavior were recorded. Total colon RNA was isolated and subjected to Affymetrix GeneChip analysis. Reduced Representation Bisulfate Sequencing (RRBS) was applied to determine the genomewide methylation pattern in both IBS and control rats. Rats with IBS egested a significantly increased amount of dry and loose stools compared with the control animals, without significant changes in body weight. Compared with the control group, 309 genes were upregulated and 224 genes were downregulated in the colon of the IBS rats. Notch signaling and focal adhesion were increased in the differentially expressed genes (DEGs). A total of 541 genes had significant lower methylation level and 626 genes had significantly higher methylation level in their promoter regions. Adherens junction and leukocyte transendothelial migration were enriched in the differentially methylated genes (DMGs). Few genes were identified in common in both DEGs and DMGs, suggesting that gene expression was not altered by promoter methylation. Reverse transcriptionquantitative polymerase chain reaction validation revealed that the mRNA levels of SSX2IP, PARD3 and VCL were significantly downregulated in the IBS group, in accordance with hypermethylation of their promoters. In summary, the present study used a WASinduced IBS rat model to provide transcriptome and methylome profiling. Most DEGs were associated with Notch signaling and focal adhesion, and only a few were altered by promoter methylation. The present results demonstrated that psychological stress could influence the integrity of the intestinal mucosa barrier and regulate inflammatory response.
Assuntos
Metilação de DNA/genética , Síndrome do Intestino Irritável/genética , Transcriptoma/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Peso Corporal/genética , Peso Corporal/fisiologia , Masculino , Proteínas dos Microfilamentos/genética , Regiões Promotoras Genéticas/genética , RNA Mensageiro/genética , Distribuição Aleatória , Ratos , Ratos Endogâmicos WKYRESUMO
BACKGROUND: Biomarkers for esophageal squamous cell carcinoma (ESCC) identification with high sensitivity are not well established. Since abnormal expression of cadherins has been widely reported in cancer, we explored its feasibility as an ESCC biomarker. METHODS: Expression of E-cadherin and N-cadherin were detected in 150 esophageal tissues by immunohistochemistry. Staining strength and percentage in different subcellular structures of each specimen were evaluated by 2 independent pathologists. A logistic regression-based classifier derived from E-cadherin and N-cadherin staining was generated. RESULTS: E-cadherin exhibited decreased membrane expression in ESCC, while N-cadherin exhibited decreased expression in the nucleus but elevated expression in the cytoplasm. Both E-cadherin and N-cadherin could distinguish ESCC tissues from non-cancerous tissues (area under the curve (AUC) = 0.748, 0.801, respectively). E-cadherin and N-cadherin staining scores could be merged into a cadherin (CDH) logistic index, which showed better discrimination (AUC = 0.909) than E-cadherin or N-cadherin alone. Further investigation indicated that the CDH logistic index was significantly correlated with tumor size and differentiation in ESCC. CONCLUSION: Both E-cadherin and N-cadherin had a strong expression shift in ESCC compared with non-cancerous tissues. The CDH logistic index, a parameter integrating the expression data of both cadherins, could be used as a marker with high sensitivity and specificity in the identification of ESCC.
Assuntos
Biomarcadores Tumorais/biossíntese , Caderinas/biossíntese , Carcinoma de Células Escamosas/metabolismo , Neoplasias Esofágicas/metabolismo , Esôfago/metabolismo , Idoso , Carcinoma de Células Escamosas/diagnóstico , Neoplasias Esofágicas/diagnóstico , Feminino , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Sensibilidade e EspecificidadeRESUMO
OBJECTIVE: This study aimed to investigate the association of ideal cardiovascular behaviors and factors with the incidence of nonalcoholic fatty liver disease (NAFLD) prospectively. PATIENTS AND METHODS: We analyzed 25 278 (21 433 men and 11 895 women) participants in the study. Participants were divided into four categories according to the number of ideal cardiovascular behaviors and factors: 0-2, 3, 4, and 5-7 groups. Multivariate logistic regression was used to calculate the odds ratios with 95% confidence intervals (CIs). RESULTS: After adjustment for confounding factors, the multivariate logistic regression model showed that the risk of NAFLD among the groups with 3, 4, 5-7 ideal factors was lower than the 0-2 group; after adjustment for age, sex, income, education level, and other confounders, the odds ratios were 0.74 (95% CI: 0.68-0.80), 0.49 (95% CI: 0.45-0.53), and 0.37 (95% CI: 0.33-0.41), respectively. CONCLUSION: The incidence of NAFLD decreased gradually with increasing ideal cardiovascular health behaviors and factors.
Assuntos
Doenças Cardiovasculares/prevenção & controle , Comportamentos Relacionados com a Saúde , Hepatopatia Gordurosa não Alcoólica/epidemiologia , Adulto , China/epidemiologia , Feminino , Humanos , Incidência , Masculino , Pessoa de Meia-Idade , Hepatopatia Gordurosa não Alcoólica/etiologia , Hepatopatia Gordurosa não Alcoólica/prevenção & controle , Estudos Prospectivos , Fatores de Risco , Fatores Sexuais , Fatores SocioeconômicosRESUMO
Neurocysticercosis (NCC) caused by Taeniasolium is one of the most common parasitic diseases of the central nervous system. Inflammation and apoptosis are two main responses involved in NCC pathogenesis. We aimed to examine apoptosis by the TUNEL assay and apoptosis-associated sFas and sFasL levels in the cerebrospinal fluid (CSF) and serum of patients with NCC. Brain biopsy (n = 1), CSF (n = 14), and serum (n = 36) of patients with NCC and uninfected controls (n = 14 and 24 for CSF and serum, respectively) were collected together with clinical data. Residual brain tissue was analyzed by the TUNEL assay. sFas and sFasL in CSF samples and sFas, sFasL, and p53 in serum samples were measured by ELISA. Immunohistochemistry of the biopsy indicated the presence of vimentin-positive arachnoid tissue in the TUNEL-positive region. Compared to controls, sFas was significantly reduced in CSF samples of patients with NCC (P = 0.018), especially among those without inflammation, but significantly increased in the serum samples of the vesicular(P = 0.011), moderate(P = 0.025), and non-epilepsy(P = 0.049) subgroups of patients with NCC. sFasL was elevated in the CSF (P < 0.0001), as well as in the calcified subgroup (P = 0.031), but sFasL levels in CSF were similar among patients with NCC with and without inflammation. These findings support a role of sFas and sFasL in the induction of apoptosis in patients with NCC, with sFas probably being involved in the inflammation phase of NCC and depending on host factors such as parasite stage, disease severity, and symptoms, and sFasL being involved in the inflammation, non-inflammation, and calcification phase of the disease.
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Apoptose , Proteína Ligante Fas/sangue , Proteína Ligante Fas/líquido cefalorraquidiano , Neurocisticercose/sangue , Neurocisticercose/líquido cefalorraquidiano , Receptor fas/sangue , Adulto , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Marcação In Situ das Extremidades Cortadas , Inflamação/sangue , Inflamação/líquido cefalorraquidiano , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Adulto JovemRESUMO
Cystic echinococcosis (CE) is a global parasitic zoonosis caused by Echinococcus granulosus. The disease is highly endemic in western China, especially in Tibetan areas, because of poor economic development and hygiene conditions, limited community knowledge of CE, a large scale of dogs, and home slaughtering of livestock. Although many researchers have analyzed risk factors of CE transmission in Tibetan Plateau, there are rare reports of knowledge, attitude, and practice (KAP) of residents about CE in Tibetan communities. In our current study, community based cross-sectional study was conducted in three townships in Xiahe County, Gannan Tibetan Autonomous Prefectures of Gansu Province from May to September 2013. A total of 972 participants originating from Tibetan communities of 31 villages in the 3 townships were registered and data were collected using structured questionnaires. From the total of 972 study participants (457 males and 515 females), 65.9% heard of the disease CE. Most of them (96.1%) would like to accept CE inspection. About half of the peoples feed their dogs often and major of them do not play with the dogs. Risk factors included resident, knowing dog could be infected, knowing eating could be route of infection, oldest dog's age, usually feed your dog by self, feed dogs with internal organs. In general our findings showed that most of residents had positive attitude toward treatments of the disease, but their practice about disease prevention and control was low. Therefore, our study called for continued and strengthened education of changing the life style, especially the behaviors related to dogs.
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Equinococose/epidemiologia , Conhecimentos, Atitudes e Prática em Saúde , Higiene , Gado , Animais de Estimação , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Criança , Pré-Escolar , China/epidemiologia , Estudos Transversais , Cães , Echinococcus granulosus , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Prevalência , Fatores de Risco , Inquéritos e Questionários , Adulto Jovem , ZoonosesRESUMO
Pneumocystis jirovecii is a fungus that causes Pneumocystis pneumonia in immuno-compromised patients. To analyze the genetic diversity of P. jirovecii in HIV-negative patients in China, respiratory specimens were obtained from 105 patients who tested PCR-positive for the presence of the P. jirovecii mitochondrial large subunit ribosomal RNA (mtLSU rRNA) between 2011 and 2013. P. jirovecii isolates were genotyped based on the upstream conserved sequence (UCS) of the major surface glycoprotein (MSG) gene and the internal transcribed spacer (ITS) of nuclear rRNA operon. Eighty-one of the 105 isolates showed a positive PCR for the UCS region. We identified six different patterns comprising two, three, four, or five UCS repeats, including 1, 2, 3 (69.14%), 1, 2, 3, 3 (22.22%), 1, 2 (3.7%), 1, 1, (2.47%), 2, 2, 3, 3 (1.23%), and 1, 1, 2, 3, 3 (1.23%). In regard to the ITS region, 58 of the 105 isolates were cloned and sequenced successfully. Six known ITS1 alleles (A, B, DEL1, E, N, and SYD1), two new alleles (designated as BTM3 and BTM4), six known ITS2 alleles (a, b, i, g, h and O) and one new allele (designated as btm6) were observed. A total of 19 P. jirovecii ITS haplotypes were identified. The most frequent type was Bi (25.9%), followed by Ai (13.8%), Eb (10.3%), and SYD1g (6.9%). Among the 58 specimens examined, 49 (84.5%) were found to contain a single type of P. jirovecii, while 9 (15.5%) contained multiple genotypes. A total of 34 allelic profiles were observed in 58 isolates when the two loci were combined with each other. A Fisher's exact test revealed that there was no statistically significant (P=0.330) association between the most frequent UCS and ITS genotypes. An analysis of the phylogenetic relationship between different patient groups identified two major groups based on the sequence variations of concatenated UCS and ITS sequences in 49 isolates. Our results demonstrated the high genetic variability of P. jirovecii in HIV-negative patients in China.
Assuntos
Genótipo , Soronegatividade para HIV , Pneumocystis carinii/genética , Pneumonia por Pneumocystis/microbiologia , Adolescente , Adulto , Idoso , Sequência de Bases , China , Sequência Conservada , DNA Fúngico , DNA Intergênico , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Filogenia , Pneumocystis carinii/classificação , Pneumocystis carinii/isolamento & purificação , Pneumonia por Pneumocystis/epidemiologia , Alinhamento de Sequência , Análise de Sequência de DNA , Adulto JovemRESUMO
Inflammation is the predominant characteristic of pneumonia. The present study aimed to to identify a faster and more reliable novel inflammatory marker for the diagnosis of pneumonia. The expression of the S100A12 gene was analyzed by reverse transcription quantitative polymerase chain reaction in samples obtained from 46 patients with bacterial pneumonia and other infections, compared with samples from 20 healthy individuals, using the 2ΔΔCt method. The expression levels of S100A12 were increased in 12 patients with bacterial pneumonia. Compared with clinical inflammatory data, a positive correlation was observed between the expression of the S100A12 gene and levels of white blood cells, Creactive protein (CRP), thrombocytocrit, neutrophils, erythrocyte sedimentation and soterocytes, and an inverse correlation was observed with the width of red blood cell volume distribution and platelet distribution, monocytes and hemoglobin, using Pearson's productmoment correlation method. The Pvalue of CRP and erythrocyte sedimentation were revealed to be statistically significant (P<0.05). A sporadic distribution of S100A12 was observed in a heatmap among the patients with different infections and bacterial pneumonia. Furthermore, the expression of S100A12 occurred in parallel to the number of clumps of inflamed tissue observed in chest computed tomography and Xray. The value of gene expression of S100A12 (>1.0) determined using the 2ΔΔCt method was associated with more severe respiratory diseases in the patients compromised by bacterial pneumonia, sepsis and pancreatitis. These findings suggested that S100A12 is an effective marker for inflammatory diseases.
Assuntos
Expressão Gênica , Mediadores da Inflamação/metabolismo , Pneumonia Bacteriana/genética , Pneumonia Bacteriana/metabolismo , Proteína S100A12/genética , Adulto , Idoso , Estudos de Casos e Controles , Análise por Conglomerados , Feminino , Perfilação da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Pneumonia Bacteriana/diagnóstico por imagem , Tomografia Computadorizada por Raios X , Adulto JovemRESUMO
OBJECTIVE: To identify the protein from host macrophages which interacted with GRA7 dense granule protein of Toxoplasma gondii, and reveal the relationship between protein interaction and infection process. METHODS: The recombinant GRA7 protein with N-terminal GST tag were used as a bait in in vitro GST Pull-down experiment, the proteins of THP-1 monocytic macrophage cell line were captured and identified by LC-MS/MS proteomics method. The in vivo protein interaction was verified by Co-IP experiment The overexpression of the target host protein by pcDNA3.1 (+) vector in THP-1 macrophage was further used to analyze the relationship between protein interaction and infection process. RESULTS: The captured THP-1 cell protein was about Mr 29000, which was identified as human carbonic anhydrase 1 (hCA1). The significant in vivo protein-protein interaction between GRA7 and hCA1 was verified by Co-IP assay. The overexpression of hCA1 gene in THP-1 macrophage induced a higher propagation speed of Tgondii and the formation of the parasitophorous vacuole, but did nmt influence the number of the parasite. CONCLUSION: There is a significant protein interaction between Toxoplasma GRA7 dense granule protein and hCA1 enzyme from host macrophages, which is positively related with the propagation speed of T. gondii.
Assuntos
Antígenos de Protozoários/metabolismo , Macrófagos/metabolismo , Proteínas de Protozoários/metabolismo , Toxoplasma/metabolismo , Humanos , Proteínas Recombinantes , Espectrometria de Massas em TandemRESUMO
Leptospirosis is a re-emerging tropical infectious disease caused by pathogenic Leptospira spp. The different host innate immune responses are partially related to the different severities of leptospirosis. In this study, we employed transcriptomics and cytokine arrays to comparatively calculate the responses of murine peritoneal macrophages (MPMs) and human peripheral blood monocytes (HBMs) to leptospiral infection. We uncovered a series of different expression profiles of these two immune cells. The percentages of regulated genes in several biological processes of MPMs, such as antigen processing and presentation, membrane potential regulation, and the innate immune response, etc., were much greater than those of HBMs (>2-fold). In MPMs and HBMs, the caspase-8 and Fas-associated protein with death domain (FADD)-like apoptosis regulator genes were significantly up-regulated, which supported previous results that the caspase-8 and caspase-3 pathways play an important role in macrophage apoptosis during leptospiral infection. In addition, the key component of the complement pathway, C3, was only up-regulated in MPMs. Furthermore, several cytokines, e.g. interleukin 10 (IL-10) and tumor necrosis factor alpha (TNF-alpha), were differentially expressed at both mRNA and protein levels in MPMs and HBMs. Some of the differential expressions were proved to be pathogenic Leptospira-specific regulations at mRNA level or protein level. Though it is still unclear why some animals are resistant and others are susceptible to leptospiral infection, this comparative study based on transcriptomics and cytokine arrays partially uncovered the differences of murine resistance and human susceptibility to leptospirosis. Taken together, these findings will facilitate further molecular studies on the innate immune response to leptospiral infection.
Assuntos
Interações Hospedeiro-Patógeno , Leptospira/imunologia , Macrófagos/imunologia , Macrófagos/microbiologia , Análise em Microsséries , Animais , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB CRESUMO
OBJECTIVE: To separate and identify the surface proteins and secreted proteins of Toxoplasma gondii tachyzoites of RH strain. METHODS: T. gondii tachyzoites were cultured in Vero cells, and purified by filtration and Percoll cell separation solution. The biotin-labeled tachyzoites were lysed, and the surface and secreted proteins were separated by NeutrAvidin agarose beads. After condensation and SDS-PAGE, the protein were collected, digested and identified by LC/MS-MS. RESULTS: A total of 785 T. gondii proteins were identified, 81 (10.3%) of which were originally annotated as the surface or secreted proteins. Among the highly-expressed (PSM>10) 65 proteins, 43 (66%) were originally annotated as surface or secreted proteins, while the others were predicted unknown proteins. CONCLUSION: The surface and secreted proteins of T. gondii are separated by biotin labeling and avidin chromatography, among which some potential new surface or secreted proteins of T. gondii are identified.
Assuntos
Proteínas de Membrana , Proteômica/métodos , Proteínas de Protozoários , Toxoplasma/metabolismo , Animais , Biotina , Chlorocebus aethiops , Proteínas de Membrana/isolamento & purificação , Proteínas de Membrana/metabolismo , Proteínas de Protozoários/isolamento & purificação , Proteínas de Protozoários/metabolismo , Células VeroRESUMO
BACKGROUND: There is a significant association between obesity and breast cancer, which is possibly due to the expression of leptin. Therefore, it is important to clarify the role of leptin/ObR (leptin receptor) signaling during the progression of human breast cancer. METHODS: Nude mice with xenografts of MCF-7 human breast cancer cells were administered recombinant human leptin subcutaneous via injection around the tumor site. Mice in the experimental group were intratumorally injected with ObR-RNAi-lentivirus, while negative control group mice were injected with the same dose of negative-lentivirus. Tumor size was blindly measured every other day, and mRNA and protein expression levels of ObR, estrogen receptor a (ERa), and vascular endothelial growth factor (VEGF) for each group were determined. RESULTS: Knockdown of ObR-treated xenografted nude mice with a high leptin microenvironment was successfully established. Local injection of ObR-RNAi-lentivirus significantly suppressed the established tumor growth in nude mice. ObR level was significantly lower in the experimental group than in the negative control group, while the amounts of ERa and VEGF expression were significantly lower in the leptin group than in the control group (P < 0.01 for all). CONCLUSIONS: Inhibition of leptin/ObR signaling is essential to breast cancer proliferation and possible crosstalk between ObR and ERa, and VEGF, and may lead to novel therapeutic treatments aiming at targeting ObR in breast cancers.
Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/terapia , Lentivirus/genética , Receptores para Leptina/genética , Animais , Neoplasias da Mama/metabolismo , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Feminino , Humanos , Células MCF-7 , Camundongos , Camundongos Nus , Interferência de RNA/fisiologia , Receptores para Leptina/metabolismo , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: It is important to identify the multiple sites of leptin activity in obese women with breast cancer. In this study, we examined the effect of exogenous human leptin on heat shock protein 70 (HSP70) expression in MCF-7 human breast cancer cells and in a breast carcinoma xenograft model of nude mice. METHODS: We cultured MCF-7 human breast cancer cells and established nude mice bearing xenografts of these cells, and randomly divided them into experimental and control groups. The experimental group was treated with human leptin, while the control group was treated with the same volume of normal saline. A real-time reverse transcriptase-polymerase chain reaction (RT-PCR) assay was developed to quantify the mRNA expression of HSP70 in the MCF-7 human breast cancer cells and in tumor tissues. Western blotting analysis was applied to quantify the protein expression of HSP70 in the MCF-7 cells. Immunohistochemical staining was done to assess the positive rate of HSP70 expression in the tumor tissues. RESULTS: Leptin activated HSP70 in a dose-dependent manner in vitro: leptin upregulated significantly the expression of HSP70 at mRNA and protein levels in MCF-7 human breast cancer cells (P < 0.001). There was no significant difference in expression of HSP70 mRNA in the implanted tumors between the leptin-treated group and the control group (P > 0.05). Immunohistochemical staining revealed no significant difference in tumor HSP70 expression between the leptin-treated group and the control group (P > 0.05). CONCLUSIONS: A nude mouse xenograft model can be safely and efficiently treated with human leptin by subcutaneous injections around the tumor. HSP70 may be target of leptin in breast cancer. Leptin can significantly upregulate the expression of HSP70 in a dose-dependent manner in vitro.
Assuntos
Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Proteínas de Choque Térmico HSP70/metabolismo , Leptina/uso terapêutico , Animais , Western Blotting , Linhagem Celular Tumoral , Feminino , Proteínas de Choque Térmico HSP70/genética , Humanos , Imuno-Histoquímica , Leptina/farmacologia , Camundongos , Camundongos Nus , Reação em Cadeia da Polimerase em Tempo Real , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: Telomerase plays an important role in cell proliferation and carcinogenesis and is believed to be a good target for anti-cancer drugs. Elimination of template function of telomerase RNA may repress the telomerase activity. METHODS: A pseudo-knotted HDV ribozyme (g.RZ57) directed against the RNA component of human telomerase (hTR) was designed and synthesized. An in vitro transcription plasmid and a eukaryotic expression plasmid of ribozyme were constructed. The eukaryotic expression plasmid was induced into heptocellular carcinoma 7402 cells, colon cancer HCT116 cells and L02 hepatocytes respectively. Then we determine the cleavage activity of ribozyme against human telomerase RNA component (hTR) both in vitro and in vivo, and detect telomerase activity continuously. RESULTS: HDV ribozyme showed a specific cleavage activity against the telomerase RNA in vitro. The maximum cleavage ratio reached about 70.4%. Transfection of HDV ribozyme into 7402 cells and colon cancer cells HCT116 led to growth arrest and the spontaneous apoptosis of cells, and the telomerase activity dropped to 10% of that before. CONCLUSION: HDV ribozyme (g.RZ57) is an effective strategy for gene therapy.
Assuntos
Carcinoma Hepatocelular/patologia , Neoplasias do Colo/patologia , Vírus Delta da Hepatite/enzimologia , Neoplasias Hepáticas/patologia , RNA Catalítico/genética , Telomerase/metabolismo , Apoptose/efeitos dos fármacos , Carcinoma Hepatocelular/enzimologia , Ciclo Celular/efeitos dos fármacos , Neoplasias do Colo/enzimologia , Hepatócitos/enzimologia , Humanos , Neoplasias Hepáticas/enzimologia , Conformação de Ácido Nucleico , RNA/metabolismo , RNA Catalítico/síntese química , RNA Catalítico/metabolismo , Telomerase/antagonistas & inibidores , Telomerase/genética , Transfecção , Células Tumorais CultivadasRESUMO
The purpose of this study was to investigate the mechanism of expression of matrix metalloproteinase-13 (MMP-13) induced by nitric oxide (NO). Human chondrocytes (HCs) were stimulated with a NO donor (MAHMA-NONOate), then mitogen-activated protein kinases' (MAPKs) and nuclear factor κB' (NF-κB) activations and MMP-13' expression were assayed by Western blot analysis. Additionally, the intracellular signalling of NO was investigated using the inhibitors of MAPKs and NF-κB. NO-induced MMP-13 expression was not suppressed by extracellular signal-regulated kinase (ERK) inhibitor (PD98059) or inhibitors of p38 kinase (SB203580), but was inhibited by a c-jun terminal kinase (JNK) inhibitor (SP600125) and inhibitors of NF-κB (SN-50). Additionally, SP600125 treatment reduced NF-κB activation, but SN-50 treatment did not significantly affect JNK activation. These results suggest that NO induces MMP-13 expression by JNK and NF-κB activation in HCs.
Assuntos
Condrócitos/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Metaloproteinase 13 da Matriz/metabolismo , NF-kappa B/metabolismo , Óxido Nítrico/metabolismo , Western Blotting , Cartilagem Articular/citologia , Cartilagem Articular/efeitos dos fármacos , Células Cultivadas , Condrócitos/efeitos dos fármacos , Indução Enzimática/fisiologia , Inibidores Enzimáticos/farmacologia , Humanos , Hidrazinas/farmacologia , Inibidores de Metaloproteinases de MatrizRESUMO
BACKGROUND: In our previous studies, we found the expression of 14-kD phosphohistidine phosphatase (PHPT1) was associated with lung cancer cells migration and invasion, and PHPT1 mRNA expression level in lung cancer tissues clinically correlated with lymph node metastasis. In the present study, we aimed to further investigate the expression of PHPT1 protein in lung cancer. METHODS: Expression of PHPT1 protein in tissue samples from 146 lung cancers and 30 normal tissues adjacent to lung cancers was assessed using immunohistochemical method. Fisher's exact test was used to analyze expression patterns of PHPT1 protein in these tissue types. Meanwhile, we studied the correlation between expression of PHPT1 protein and clinicopathological features in lung cancer. RESULTS: Significantly higher expression levels of PHPT1 protein were found in lung cancer samples (53.42%) than in normal tissues adjacent to lung cancer (23.33%) (P = 0.003). Fisher's exact test showed that lung cancer stage positively correlated with expression of PHPT1 protein (P = 0.02), and lung cancer samples with lymph node metastasis showed higher PHPT1 protein expression (P = 0.016) than the samples without lymph node metastasis. CONCLUSIONS: The results of this study agree with findings from our previous study of PHPT1 mRNA expression in lung cancer tissues, and strongly suggest that PHPT1 protein is closely associated with the carcinogenesis and metastasis of lung cancer. Thus, therapy targeting PHPT1 (inhibition or silencing) could be potentially benefited for lung cancer patients.
Assuntos
Neoplasias Pulmonares/metabolismo , Monoéster Fosfórico Hidrolases/metabolismo , Western Blotting , Regulação Neoplásica da Expressão Gênica , Humanos , Imuno-Histoquímica , Técnicas In VitroRESUMO
BACKGROUND: It is essential to clarify the interactions of hormones during the progression of human breast cancer. This study examined the effects of exogenous human leptin on estrogen receptor (ER) alpha and beta in human breast tumor tissue in a nude mouse xenograft model. METHODS: We created nude mice xenografts of MCF-7 human breast cancer cells, and randomly divided them into an experimental group and a control group. The mice in experimental group were injected subcutaneously around tumors with human leptin, while the control group were injected with the same dose of normal saline. A real-time RT-PCR assay was developed to quantify the mRNA of ERalpha, beta in the tumor tissues. Western blotting analyses were used to assess the relative quantities of the ERalpha, beta proteins. RESULTS: Leptin-treated xenografted nude mice were successfully established. The amount of ERalpha mRNA was significantly higher in the leptin group than in the control group (P < 0.01), while the amount of ERbeta mRNA was significantly lower in the leptin group than in the control group (P < 0.01). Western blotting analyses revealed that the ERalpha protein level was significantly higher in the leptin group than in the control group (P < 0.01), while the ERbeta protein level was significantly lower in the leptin group than in the control group (P < 0.01). CONCLUSIONS: Nude mouse xenograft model can be safely and serviceably treated with human leptin by subcutaneous injections around tumor. ERalpha, beta were both targets of leptin in breast cancer. Leptin can up-regulate the expression of ERalpha and down-regulate the expression of the ERbeta in human breast tumor.
Assuntos
Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/metabolismo , Leptina/uso terapêutico , Animais , Western Blotting , Neoplasias da Mama/genética , Linhagem Celular Tumoral , Receptor alfa de Estrogênio/genética , Receptor beta de Estrogênio/genética , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Camundongos , Camundongos Nus , Reação em Cadeia da Polimerase , RNA Mensageiro/genética , Distribuição Aleatória , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
This article investigates the mechanism of activation of matrix metalloproteinase-13 induced by nitric oxide. SW1353 cells were stimulated with S-nitroso-N-acetyl-D,L-penicillamine, expressions and activities of metalloproteinase-13, and membrane type-1 metalloproteinase were assayed, and a proteolytic activation of recombinant human metalloproteinase-13 was measured in the presence of recombinant human membrane type-1 metalloproteinase. Nitric oxide increased expressions of both matrix metalloproteinases and stimulated the proteolytic processing of metalloproteinase-13 from the pro-enzyme to the final active form. Recombinant human membrane type-1 metalloproteinase was able to process recombinant human metalloproteinase-13 to fully active enzyme. S-nitroso-N-acetyl-D,L-penicillamine had no effect on recombinant human metalloproteinase-13 activity. Nitric oxide scavenger (OxyHb) strongly attenuated nitric oxide-induced proteolytic activation of metalloproteinase-13. Furthermore, tissue inhibitor of metalloproteinase-2 markedly reduced the activation of metalloproteinase-13 in response to nitric oxide. These studies identify membrane type-1 metalloproteinase as a target for nitric oxide, which may be critical for the nitric oxide-induced activation of metalloproteinase-13.