RESUMO
Basophils (BAs) are the least common granulocytes of all leukocytes, but they play an important role in orchestrating of chronic allergic inflammation. The Notch signaling pathway is a highly conserved pathway that influences cell lineage decisions and differentiation during various stages of development. However, the relationship between Notch signaling and BA remains to be elucidate. Here, we report that several Notch signaling molecules were found to be expressed in BAs. γ-secretase inhibitor (GSI) treatment increase BAs apoptosis, and suppress BAs proliferation. Furthermore, GSI reduced BAs in the S phase, with a concomitant accumulation in G1 and G2 phases. In addition, GSI also significantly down-regulated mRNA levels of cytokines IL-4, IL-6 and IL-13 induced by A23187, and this effect was dependent on MAPK pathway. Finally, IL-6 inhibition was specifically associated with ERK and IL-13 with JNK. Therefore, Notch signaling regulates BA biological function, at least partially via the modulation of MAPK.
Assuntos
Basófilos/imunologia , Hipersensibilidade/imunologia , Inflamação/imunologia , Receptores Notch/metabolismo , Transdução de Sinais , Animais , Calcimicina/farmacologia , Ciclo Celular , Células Cultivadas , Doença Crônica , Citocinas/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Mediadores da Inflamação/metabolismo , Camundongos , Oligopeptídeos/farmacologia , Receptores Notch/genéticaRESUMO
The biosynthesis of antibiotics in bacteria is usually believed to be an intracellular process, at the end of which the matured compounds are exported outside the cells. The biosynthesis of saframycinâ A (SFM-A), an antitumor antibiotic, requires a cryptic fatty acyl chain to guide the construction of a pentacyclic tetrahydroisoquinoline scaffold; however, the follow-up deacylation and deamination steps remain unknown. Herein we demonstrate that SfmE, a membrane-bound peptidase, hydrolyzes the fatty acyl chain to release the amino group; and SfmCy2, a secreted oxidoreductase covalently associated with FAD, subsequently performs an oxidative deamination extracellularly. These results not only fill in the missing steps of SFM-A biosynthesis, but also reveal that a FAD-binding oxidoreductase catalyzes an unexpected deamination reaction through an unconventional extracellular pathway in Streptmyces bacteria.
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Antibióticos Antineoplásicos/biossíntese , Oxirredutases/metabolismo , Pró-Fármacos/metabolismo , Antibióticos Antineoplásicos/química , Biocatálise , Desaminação , Flavina-Adenina Dinucleotídeo/química , Isoquinolinas/química , Isoquinolinas/metabolismo , Pró-Fármacos/química , Streptomyces/metabolismoRESUMO
Nonribosomal peptide synthetases (NRPSs) usually catalyze the biosynthesis of peptide natural products by sequential selection, activation, and condensation of amino acid precursors. It was reported that some fatty acids, α-ketoacids, and α-hydroxyacids originating from amino acid metabolism as well as polyketide-derived units can also be used by NRPS assembly lines as an alternative to amino acids. Ecteinascidin 743 (ET-743), naphthyridinomycin (NDM), and quinocarcin (QNC) are three important antitumor natural products belonging to the tetrahydroisoquinoline family. Although ET-743 has been approved as an anticancer drug, the origin of an identical two-carbon (C(2)) fragment among these three antibiotics has not been elucidated despite much effort in the biosynthetic research in the past 30 y. Here we report that two unexpected two-component transketolases (TKases), NapB/NapD in the NDM biosynthetic pathway and QncN/QncL in QNC biosynthesis, catalyze the transfer of a glycolaldehyde unit from ketose to the lipoyl group to yield the glycolicacyl lipoic acid intermediate and then transfer the C(2) unit to an acyl carrier protein (ACP) to form glycolicacyl-S-ACP as an extender unit for NRPS. Our results demonstrate a unique NRPS extender unit directly derived from ketose phosphates through (α,ß-dihydroxyethyl)-thiamin diphosphate and a lipoyl group-tethered ester intermediate catalyzed by the TKase-ACP platform in the context of NDM and QNC biosynthesis, all of which also highlights the biosynthesis of ET-743. This hybrid system and precursor are distinct from the previously described universal modes involving the NRPS machinery. They exemplify an alternate strategy in hybrid NRPS biochemistry and enrich the diversity of precursors for NRPS combinatorial biosynthesis.
Assuntos
Cetoses/metabolismo , Peptídeos/metabolismo , Streptomyces/metabolismo , Proteína de Transporte de Acila/genética , Proteína de Transporte de Acila/metabolismo , Sequência de Aminoácidos , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Vias Biossintéticas , Eletroforese em Gel de Poliacrilamida , Cetoses/química , Cinética , Espectroscopia de Ressonância Magnética , Modelos Químicos , Dados de Sequência Molecular , Estrutura Molecular , Família Multigênica , Mutação , Naftiridinas/química , Naftiridinas/metabolismo , Peptídeo Sintases/genética , Peptídeo Sintases/metabolismo , Peptídeos/química , Peptídeos/genética , Homologia de Sequência de Aminoácidos , Streptomyces/química , Streptomyces/genética , Especificidade por Substrato , Tetra-Hidroisoquinolinas/química , Tetra-Hidroisoquinolinas/metabolismo , Transcetolase/genética , Transcetolase/metabolismoRESUMO
BACKGROUND: The incidence of acute coronary syndrome caused by the rupture of atherosclerotic plaque and subsequent arterial thrombosis increases as the weather gets colder. However, the association between cold stress and atherosclerotic plaque rupture is currently unknown. METHODS: An atherosclerotic plaque model was established in rabbits by balloon injury and a high-fat diet with or without cold stress (4 °C, 1 hour per day, 20 weeks) at the onset of modeling. Additionally, oxidized low-density lipoprotein (ox-LDL) was applied to induce the formation of macrophage foam cells in vitro. RESULTS: Serum lipid profiles and inflammatory cytokines (ox-LDL, high-sensitivity C-reactive protein, and interleukin-8) were significantly higher in cold stress-exposed rabbits than in controls (P<0.05). Animals with atherosclerotic lesions that were exposed to cold stress had increased macrophages, foam cells, intima-media thickness, and neovascularization in the plaque, along with significantly thinned plaque fibrous caps. Moreover, we found that cold stress induced more apoptotic cells in the atherosclerotic plaques and up-regulated endoplasmic reticulum stress (ERS)-associated proteins CHOP, GRP78, and p-JNK (P<0.05). In addition, tunicamycin treatment promoted ox-LDL-induced apoptosis, expression of CHOP and GPR78, and the p-JNK level in macrophage foam cells, while JNK inhibitor sp600125 reduced cell apoptosis and the p-JNK level. The three main ERS sensors sensors phosphorylated extracellular signal-regulated kinase (PERK), activating transcription factor 6 (ATF6), and inositol-requiring enzyme1 (IRE1) declined significantly after ox-LDL treatment. CONCLUSIONS: Cold stress may enhance the instability of atherosclerotic plaques through activating ERS and enhancing cell apoptosis. Up-regulated CHOP levels mediated by PERK and ATF6 and the activated IRE1-XBP1-JNK pathway contributed to the apoptosis of foam cells.
Assuntos
Temperatura Baixa , Retículo Endoplasmático/fisiologia , Placa Aterosclerótica/fisiopatologia , Estresse Fisiológico , Animais , Apoptose/fisiologia , Sequência de Bases , Linhagem Celular , Primers do DNA , Chaperona BiP do Retículo Endoplasmático , Marcação In Situ das Extremidades Cortadas , Masculino , Camundongos , Coelhos , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Genetic polymorphisms of cytochrome P450 1A1 (CYP1A1) and glutathione S-transferase M1 (GSTM1) genes might contribute to the variability in individual susceptibility to lung cancer, but the reported results from individual studies are not always consistent. We therefore conducted a meta-analysis to systematically estimate the associations between polymorphisms of these two genes and risk of lung cancer. Twenty-one studies with 8,926 subjects were finally enrolled into this study. Meta-analysis was performed by RevMan 5.2. Odds ratio (OR) and its 95 % confidence interval (CI) were calculated to evaluate the susceptibility to lung cancer. Compared with the wild-type homozygous genotype, significantly elevated risk of lung cancer were associated with variant CYP1A1 MspI (m1/m2 + m2/m2 vs. m1/m1: OR = 1.27, 95 % CI = 1.12-1.43, P < 0.001) and deletion of GSTM1 (null vs. present: OR = 1.26, 95 % CI = 1.13-1.40, P < 0.001). Both the two genetic polymorphisms were independently associated with the risk of lung cancer. The pooled OR of lung cancer for population with both CYP1A1 MspI and GSTM1 mutations (MspI m1/m2 or m2/m2 and GSTM1 null) was 1.62 (95 % CI 1.27-2.07, P < 0.001) when compared with those without any of the above mutations, which is higher than single genetic polymorphism. In the stratified analysis, significantly higher risks of lung cancer associated with above genetic polymorphisms were found only in Asian population. This meta-analysis suggests that the CYP1A1 MspI and GSTM1 polymorphisms correlate with increased lung cancer susceptibility independently, and that there is an interaction between the two genes. However, the associations vary in different ethnic populations.
Assuntos
Citocromo P-450 CYP1A1/genética , Predisposição Genética para Doença/genética , Glutationa Transferase/genética , Neoplasias Pulmonares/genética , Polimorfismo Genético , Sítios de Ligação/genética , Estudos de Casos e Controles , Citocromo P-450 CYP1A1/metabolismo , Desoxirribonuclease HpaII/metabolismo , Frequência do Gene , Genótipo , Humanos , Neoplasias Pulmonares/metabolismo , Razão de Chances , Fatores de RiscoRESUMO
BACKGROUND AND OBJECTIVE: Previous studies have demonstrated that our recombinant bacille Calmette-Guerin (rBCG), which expresses Der p2 in house dust mite (Der p2 rBCG) suppresses asthmatic airway inflammation by regulating the phenotype and function of dendritic cells (DC) and reprogramming T helper (Th) 0 cell differentiation into different T cell (Th1/Th2/Treg) subtypes. However, the exact role of Der p2 rBCG in reprogramming Th17 differentiation and the relevant mechanisms are not known. The aim of this study was to examine whether Der p2 rBCG-mediated inhibition of allergic airway inflammation is mediated by regulating Th17 differentiation in a murine asthma model. METHODS: Primary mouse bone marrow-derived dendritic cells (BMDC) were infected with Der p2 rBCG and adoptively transferred to Der p2-intranasally sensitized mice. The role of Der p2 rBCG-BMDC on the regulation of airway inflammation and Th17 cell differentiation was assessed. RESULTS: Adoptive transfer of Der p2 rBCG-BMDC suppressed airway inflammation and mucin secretion. Der p2 rBCG-BMDC inhibited excessive Th17 immune responses but not BCG-BMDC. Furthermore, Der p2 rBCG decreased jagged-2 and increased delta-like-4 expressions on BMDC to a greater extent than BCG. CONCLUSIONS: These findings suggest that DC plays a key role in Der p2 rBCG-induced immunoregulation. Der p2 rBCG also displayed a potent inhibitory effect on Th17 differentiation, and these findings increase our understanding of the cellular basis of Der p2 BCG-mediated inhibition of asthma.
Assuntos
Antígenos de Dermatophagoides/genética , Proteínas de Artrópodes/genética , Asma/genética , Células da Medula Óssea/patologia , Células Dendríticas/imunologia , Regulação Bacteriana da Expressão Gênica , Mycobacterium bovis/metabolismo , Células Th17/imunologia , Animais , Antígenos de Dermatophagoides/biossíntese , Proteínas de Artrópodes/biossíntese , Asma/imunologia , Asma/metabolismo , Células da Medula Óssea/metabolismo , Células da Medula Óssea/microbiologia , Células Dendríticas/metabolismo , Modelos Animais de Doenças , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Mycobacterium bovis/imunologia , RNA/genética , Células Th17/metabolismoRESUMO
Background: Kirsten rat sarcoma viral oncogene homolog (KRAS) mutation seemingly suffered less effective therapeutic regimens in the absence of widely-accepted targeted drugs compared with other mutation types in non-small cell lung cancer (NSCLC). However, whether these non-selective therapy schedules for KRAS mutation matters is still under debate. Correspondingly, we aimed to compare the long term expectancy of indicated therapeutic regimes and further explore the optimal schemes of KRAS mutated NSCLC in the absence of targeted drugs in this retrospective study cohort. Methods: We conducted a single-center retrospective analysis among 66 patients diagnosed with KRAS-mutant advanced NSCLC from November 2018 to December 2020. These enrolled cases were divided into different subgroups in light of mutant isotypes, pathological characteristics, and therapeutic regimes to uncover indicated long-term survival benefits. Additionally, clinical outcomes of treatment schedules and interventional lines to KRAS-mutant NSCLC were described in detail. Results: This cohort enrolled 8 patients with stage IIIB (12.1%) and 58 patients with stage IV (87.9%) with the median age 62 years, ranging from 32 to 91 years old. Genetically, G12C conducted as the most common KRAS mutation type, accounting for 30.3%. Pemetrexed combined with platinum chemotherapy seemed to be a priority (72.7%), and chemotherapy combined with immunotherapy became an alternative (15.2%) in clinic. Performing further analysis of long-term survival of patients receiving different treatment methods indicated that the median overall survival (mOS) in first-line therapy with antiangiogenesis or untreated was 13 and 12 months, respectively (P=0.79). In the first-line regimen, median survival was 17 months for patients who received combined immune checkpoint inhibitors and 12 months for those who did not (P=0.34). The mOS was 20 months for those who had used immune checkpoint inhibitors and 12 months for those who had not (P=0.11). Survival analysis results of NSCLC patients with different KRAS mutation types showed the median survival time of patients with G12C mutation type and patients without with nonG12C mutation type was 19 and 12 months, respectively (P=0.37). Conclusions: In the absence of KRAS targeted drugs, available treatment plans failed to benefit KRAS mutant sufferers regardless of isotypes, making the KRAS-targeted drugs urgent.
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BACKGROUND: Neoadjuvant chemotherapy (NACT) combined with surgery is regarded as an effective treatment for advanced gastric cancer (AGC). Laparoscopic surgery represents the mainstream of minimally invasive surgery. Currently, surgeons focus more on surgical safety and oncological outcomes of laparoscopic gastrectomy after NACT. Thus, we sought to evaluate short- and long-term outcomes between laparoscopic total gastrectomy (LTG) and open total gastrectomy (OTG) after NACT. AIM: To compare the short and long-term outcomes between LTG and OTG for AGC after NACT. METHODS: We retrospectively collected the clinicopathological data of 136 patients who accepted gastrectomy after NACT from June 2012 to June 2019, including 61 patients who underwent LTG and 75 who underwent OTG. Clinicopathological characteristics between the LTG and OTG groups showed no significant difference. SPSS 26.0, R software, and GraphPad PRISM 8.0 were used to perform statistical analyses. RESULTS: Of the 136 patients included, eight acquired pathological complete response, and the objective response rate was 47.8% (65/136). The LTG group had longer operation time (P = 0.015), less blood loss (P = 0.003), shorter days to first flatus (P < 0.001), and shorter postoperative hospitalization days (P < 0.001). LTG spent more surgical cost than OTG (P < 0.001), while total hospitalized cost of LTG was less than OTG (P < 0.001). 21 (28.0%) patients in the OTG group and 14 (23.0%) in the LTG group had 30-d postoperative complications, but there was no significant difference between the two groups (P = 0.503). The 3-year overall survival (OS) rate was 60.6% and 64.6% in the LTG and OTG groups, respectively [hazard ratio (HR) = 0.859, 95% confidence interval (CI): 0.522-1.412, P = 0.546], while the 3-year disease-free survival (DFS) rate was 54.5% and 51.8% in the LTG and OTG group, respectively (HR = 0.947, 95%CI: 0.582-1.539, P = 0.823). Multivariate cox analysis showed that body mass index and pTNM stage were independent risk factors for OS while vascular invasion and pTNM stage were independent risk factors for DFS (P < 0.05). CONCLUSION: After NACT, LTG shows comparable 30-d postoperative morbidity as well as 3-year OS and DFS rate to OTG. We recommend that experienced surgeons select LTG other than OTG for proper AGC patients after NACT.
RESUMO
Background: It has been demonstrated that miRNAs play critical roles in the tumorigenesis and progression of various tumors.Objective: The purpose of this research was to determine the serum miR-632 levels in patients with laryngeal squamous cell carcinoma (LSCC) and to investigate its diagnostic and prognostic value.Materials and methods: We detected serum miR-632 levels in 162 LSCC patients and 42 healthy volunteers. The ROC curve was carried out to determine diagnostic accuracy.Results: We observed that serum miR-632 levels were upregulated in LSCC patients compared with healthy volunteers (p < .01). Subsequent results from ROC indicated that high sensitivity and specificity of serum miR-632 for diagnosing LSCC (area under the curve 0.8828). In addition, it was found that high expressions of serum miR-632 were significantly associated with advanced N stage (p = .020), histological grade (p = .001), and TNM stage (p = .014). Furthermore, patients with higher serum miR-632 expression had a shorter OS and DFS time than those with lower serum miR-632 levels.Conclusion: Our data revealed that serum miR-632 may be a potential noninvasive biomarker which may become a potential diagnostic and prognostic biomarker for LSCC patients.
Assuntos
Carcinoma de Células Escamosas/sangue , Neoplasias Laríngeas/sangue , MicroRNAs/sangue , Biomarcadores Tumorais/sangue , Carcinoma de Células Escamosas/diagnóstico , Carcinoma de Células Escamosas/mortalidade , Estudos de Casos e Controles , China/epidemiologia , Feminino , Humanos , Neoplasias Laríngeas/diagnóstico , Neoplasias Laríngeas/mortalidade , Masculino , Pessoa de Meia-IdadeRESUMO
Evidence has shown that Notch signaling modulates CD4(+)CD25(+) regulatory T-cells (Tregs). As transcription factor Foxp3 acts as a master molecule governing the development and function of Tregs, we investigated whether Notch signaling might directly regulate Foxp3 expression. Here, we provide evidence that Notch signaling can modulate the FOXP3 promoter through RBP-J- and Hes1-dependent mechanisms. A conserved RBP-J-binding site and N-box sites were identified within the FOXP3 promoter. We show that the Notch intracellular domain (NIC), the active form of Notch receptors, activates a reporter driven by the FOXP3 promoter. Dissection of the FOXP3 promoter revealed bipartite effects of the RBP-J-binding site and the N-boxes: the RBP-J-binding site positively, while the N-boxes negatively regulated the FOXP3 promoter activity. Moreover, in freshly isolated Tregs, NIC-RBP-J complex is bound to the FOXP3 promoter in Tregs. Our results suggest that Notch signaling might be involved in the development and function of Tregs through regulating Foxp3 expression.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Fatores de Transcrição Forkhead , Regulação da Expressão Gênica , Proteínas de Homeodomínio/metabolismo , Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/metabolismo , Regiões Promotoras Genéticas , Receptores Notch/metabolismo , Transdução de Sinais/fisiologia , Animais , Sequência de Bases , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/metabolismo , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismo , Genes Reporter , Células HeLa , Proteínas de Homeodomínio/genética , Humanos , Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/genética , Células Jurkat , Camundongos , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Receptores Notch/genética , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/metabolismo , Fatores de Transcrição HES-1RESUMO
The tetrahydroisoquinoline (THIQ) alkaloids are naturally occurring antibiotics isolated from a variety of microorganisms and marine invertebrates. This family of natural products exhibit broad spectrum antimicrobial and strong antitumor activities, and the potency of clinical application has been validated by the marketing of ecteinascidin 743 (ET-743) as anticancer drug. In the past 20 years, the biosynthetic gene cluster of six THIQ antibiotics has been characterized including saframycin Mx1 from Myxococcus xanthus, safracin-B from Pseudomonas fluorescens, saframycin A, naphthyridinomycin, and quinocarcin from Streptomyces, as well as ET-743 from Ecteinascidia turbinata. This review gives a brief summary of the current status in understanding the molecular logic for the biosynthesis of these natural products, which provides new insights on the biosynthetic machinery involved in the nonribosomal peptide synthetase system. The proposal of the THIQ biosynthetic pathway not only shows nature's route to generate such complex molecules, but also set the stage to develop a different process for production of ET-743 by synthetic biology.
Assuntos
Antibacterianos/biossíntese , Vias Biossintéticas , Tetra-Hidroisoquinolinas/metabolismo , Antibacterianos/química , Produtos Biológicos/química , Produtos Biológicos/metabolismo , Humanos , Conformação Molecular , Tetra-Hidroisoquinolinas/químicaRESUMO
Dipyridamole (DIP) inhibits thrombus formation when given chronically, and causes vasodilation over a short time. To date, DIP can increase the anticancer drugs (5-fluorouracil, methotrexate, piperidine, vincristine) concentration in cancer cells and hence enhance the efficacy of treatment cancer. The inhibition of DIP may result in increased 5-fluorouracil efficacy and diminish the drug side effects. But the actual molecular targets remain unknown. In this study, reverse protein-ligands docking, and quantum mechanics were used to search for the potential molecular targets of DIP. The quantum mechanics calculation was performed by using Gaussian 03 program package. Reverse pharmacophore mapping was used to search for potential molecular target candidates for a given small molecule. The docking study was used for exploring the potential anti-cancer targets of dipyridamole. The two predicted binders with the statistically significant prediction are dihydropyrimidine dehydrogenase (DPD) (PDB Id: 1GTE) and human spindle checkpoint kinase Bub1 (PDB Id: 3E7E). Structure analysis suggests that electrostatic interaction and hydrogen bonding play an important role in their binding process. The strong functional linkage of DIP and 5FU supports our prediction. In conclusion, these results generate a tractable set of anticancer proteins. The exploration of polypharmacology will provide us new opportunities in treating systematic diseases, such as the cancers. The results would generate a tractable set of anticancer target proteins for future experimental validations.
RESUMO
The noninvasive measurement of human blood glucose was achieved with NIR diffusion reflectance spectrum method. The thumb fingertip NIR diffusion reflectance spectra of six different age healthy volunteers were collected using Nexus-870 and its NIR fiber port smart accessory. The test was implemented with changing the blood glucose concentration for the limosis and satiation of every volunteer. The calibration model was set up using PLS method with the smoothing, baseline correction and first derivatives pretreatment spectrum in the 7500-8500 cm(-1) region for single volunteer, the same age combination and that of different age. When the spectrum was obtained, the actual blood glucose value of every spectrun sample was demarcated using ultraviolet spectrophotometer. The correlation between the calibration value and true value for single volunteer is better than that for the combination of volunteers, the correlative coefficients are all over 0.90471, RMSECs are all less than 0.171.
Assuntos
Glicemia/análise , Espectroscopia de Luz Próxima ao Infravermelho/métodos , Adulto , Calibragem , Difusão , Feminino , Humanos , Masculino , Modelos Teóricos , Adulto JovemAssuntos
Cânula , Insuficiência Respiratória , Adulto , Consenso , Humanos , Oxigênio , Oxigenoterapia , Insuficiência Respiratória/terapiaRESUMO
OBJECTIVE: To observe the inhibitive effect on airway mucus overproduction with DNA vaccine based on human calcium-activated chloride channel 1 (hCLCA1) in asthmatic mice, who own mCLCA3 being xenogeneic homology of hCLCA1 in airway goblet cell. METHODS: The DNA vaccine was made with hCLCA1 gene inserted into pSecTag2B, and then BALB/c mice were vaccinated by i.m. once every two weeks. When serum antibody showed binding activity to mCLCA3 with ELISA analysis, asthma will be induced with ovalbumin in the vaccinated mice. To detect mucus production, lung sections were PAS stained and their MUC5AC mRNA levels were investigated by reverse transcription polymerase chain reaction (RT-PCR). Mice in control groups were injected with pSecTag2B/mCLCA3, pSecTag2B and saline, respectively. RESULTS: Antiserum of vaccine group after three times vaccination showed good binding activity to three mCLCA3 extracellular domains (ED), and the activity to N-terminal C-terminal ED was stronger than middle-ED. After induced to asthma, the number of goblet cell and MUC5AC mRNA level in vaccine group were lower than these in control group. CONCLUSION: hCLCA1 DNA vaccine can induce mouse to produce serum antibody binding itself mCLCA3, and thus airway mucus overproduction of asthmatic mouse is effectively inhibited.
Assuntos
Asma/tratamento farmacológico , Canais de Cloreto/imunologia , Mucinas/biossíntese , Vacinas de DNA/uso terapêutico , Animais , Asma/metabolismo , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Expressão Gênica/efeitos dos fármacos , Pulmão/efeitos dos fármacos , Pulmão/imunologia , Pulmão/patologia , Camundongos , Mucina-5AC , Mucinas/genética , RNA Mensageiro/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
OBJECTIVE: To evaluate the inhibiting effect of niflumic acid (NFA), an inhibitor of calcium-activated chloride channel (ClCa) on airway epithelium, on the airway hyperresponsiveness in asthmatic mice. METHODS: BALB/c mice were randomly divided into an asthma group (A group), a NFA prevention asthmatic group (B group) and a sham-challenged group (C group). The airway pressure time index (APTI) and the content of ET-1 and NO in bronchoalveolar lavage fluid (BALF) in all groups were measured. With the isolated tracheal rings with integral epithelium or epithelium removed from the asthma group (A(1) group and A(2) group) and the sham-challenged group (C(1) group and C(2) group), the contractile responsiveness of various rings to methacholine (mACh) was examined, and its change was observed when the rings were exposed to NFA beforehand. RESULTS: Compared with A group (1.62 +/- 0.14), the APTI in B group (1.21 +/- 0.07) was reduced remarkably (P < 0.01), and the contents of ET-1 [(103 +/- 9) ng/L] and NO [(48.5 +/- 3.2) micromol/L] in BALF of A group were significantly higher than those in B group, [(53 +/- 5) ng/L, (23.7 +/- 2.5) micromol/L (P < 0.01), respectively]. The ratios of maximum contractility in A(1), A(2), C(1) and C(2) groups were (3.79 +/- 0.44), (2.15 +/- 0.21), (1.26 +/- 0.14) and (2.06 +/- 0.18), respectively. The contractility of A(1) group was highest among all groups (all P < 0.01), but could be effectively decreased by NFA. CONCLUSIONS: By inhibiting the special ClCa on the airway epithelium, NFA can inhibit the production of ET-1 and NO by epithelium and thus exert preventive effect on airway hyperresponsiveness in asthma.
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Asma/tratamento farmacológico , Hiper-Reatividade Brônquica/tratamento farmacológico , Ácido Niflúmico/uso terapêutico , Animais , Líquido da Lavagem Broncoalveolar/química , Endotelina-1/análise , Técnicas In Vitro , Masculino , Camundongos , Óxido Nítrico/análise , Traqueia/efeitos dos fármacosRESUMO
BACKGROUND: Platelet inhibition by clopidogrel is highly variable and the elevated platelet activity will increase the risk of major adverse cardiovascular events after percutaneous coronary intervention (PCI). CYP2C19 loss-of-function (LOF) alleles and risk factors of coronary heart disease (CAD) were reported to be associated with the low response of clopidogrel. PURPOSE: This study was carried out to analyze the contributions of CYP2C19 polymorphisms and risk factors to the various clopidogrel responses in Chinese patients with stable CAD after PCI. MATERIALS AND METHODS: The platelet reactivity index (PRI) was measured in 145 patients who underwent PCI using the vasodilator-stimulated phosphoprotein assay. Gene chip hybrid tests were used to analyze the genetic polymorphisms of CYP2C19. RESULTS: With a cutoff value of 50% in PRI, 20.67% (31/145) of the patients were defined to be clopidogrel resistant. With respect to the normal *1, *2, and *3 LOF CYP2C19 alleles, patients were classified into three metabolism phenotypes: 39.31% were extensive, 47.59% were intermediate, and 13.10% were poor metabolizers (PMs). Of the enrolled patients, 53.82 and 9.66%, respectively, were carriers of *2 and *3 alleles. There was a significant difference in PRI between PM and either extensive or intermediate metabolizers (P<0.05). In all, 36.84% of the patients with the PM phenotype were clopidogrel resistant. Carriers of two CYP2C19 LOF alleles, BMI, and the presence of type 2 diabetes were three independent risk factors for clopidogrel resistance. CONCLUSION: Genetic CYP2C19 polymorphisms and CAD risk factors - type 2 diabetes mellitus and BMI - synergistically affect the antiplatelet activity of clopidogrel and the occurrence of major adverse cardiovascular events after PCI.
Assuntos
Plaquetas/efeitos dos fármacos , Doença das Coronárias/terapia , Citocromo P-450 CYP2C19/genética , Intervenção Coronária Percutânea , Inibidores da Agregação Plaquetária/farmacocinética , Polimorfismo Genético , Ticlopidina/análogos & derivados , Idoso , Biomarcadores/sangue , Plaquetas/metabolismo , Índice de Massa Corporal , Moléculas de Adesão Celular/sangue , China , Clopidogrel , Doença das Coronárias/sangue , Doença das Coronárias/diagnóstico , Doença das Coronárias/mortalidade , Citocromo P-450 CYP2C19/metabolismo , Diabetes Mellitus Tipo 2/complicações , Resistência a Medicamentos/genética , Feminino , Frequência do Gene , Genótipo , Humanos , Masculino , Proteínas dos Microfilamentos/sangue , Pessoa de Meia-Idade , Análise de Sequência com Séries de Oligonucleotídeos , Intervenção Coronária Percutânea/efeitos adversos , Intervenção Coronária Percutânea/mortalidade , Farmacogenética , Fenótipo , Fosfoproteínas/sangue , Inibidores da Agregação Plaquetária/efeitos adversos , Testes de Função Plaquetária , Fatores de Risco , Ticlopidina/efeitos adversos , Ticlopidina/farmacocinética , Resultado do TratamentoRESUMO
Analysis of naphthyridinomycin gene cluster revealed that this antibiotic is generated by nonribosomal peptide synthetase (NRPS) machinery. However, four modules encoded by two genes do not correspond with the structural units in the final product. Genetic and biochemical characterization of the gene cluster suggested that the leader peptide mechanism for the NRPS assembly line was involved in biosynthesis of this tetrahydroisoquinoline alkaloid.
Assuntos
Alcaloides/química , Alcaloides/metabolismo , Peptídeo Sintases/química , Peptídeo Sintases/genética , Peptídeo Sintases/metabolismo , Peptídeos/química , Peptídeos/genética , Peptídeos/metabolismo , Tetra-Hidroisoquinolinas/química , Tetra-Hidroisoquinolinas/metabolismo , Sequência de Aminoácidos , Estrutura Molecular , Família Multigênica , Naftiridinas/química , Naftiridinas/metabolismoRESUMO
The proliferation, migration, and adhesion of vascular smooth muscle cells (VSMCs) and their interactions with extracellular matrix are key features of atherosclerosis and restenosis. Recently, there has been evidence that magnetic fields exert multiple effects on the biological performance of cells and may aid in the treatment of vascular disease. However, the effect of a static magnetic field (SMF) on human VSMCs still remains unknown. In this study, we aimed to determine the effects of low strength SMF on human VSMCs in an in vitro restenosis model. A SMF was established using neodymium-yttrium-iron permanent magnet. Human umbilical artery smooth muscle cells (hUASMCs) were isolated and seeded to a fibronectin-coated plate to form an in vitro restenosis model and then exposed to a vertically oriented field of 5 militesla (mT). MTT, transwell, and adhesion assays were used to demonstrate that the proliferation, migration, and adhesion potential of hUASMCs were significantly decreased after exposure to 5 mT SMF for 48 h compared with a non-treated group. Meanwhile, confocal microscopy analysis was used to demonstrate that integrin ß(1) clustering was inhibited by exposure to 5 mT SMF. Furthermore, the phosphorylation of focal adhesion kinase (FAK) was markedly inhibited, and the upregulated cytosolic free calcium had been reversed (p < 0.05). However, the biological effects of low strength SMF on hUASMCs could be blocked by the administration of GRGDSP-the blockade of integrins. In conclusion, a low strength SMF can influence the proliferation, migration, and adhesion of VSMCs by inhibiting the clustering of integrin ß1, decreasing cytosolic free calcium concentration, and inactivating FAK. With further validation, SMFs may aid in attenuating abnormal VSMCs biological performance and has potential to block atherogenesis and prevent restenosis.
Assuntos
Movimento Celular , Proliferação de Células , Oclusão de Enxerto Vascular/metabolismo , Integrinas/metabolismo , Campos Magnéticos , Modelos Biológicos , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Adesão Celular/efeitos dos fármacos , Quinase 1 de Adesão Focal/metabolismo , Oclusão de Enxerto Vascular/patologia , Humanos , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/patologia , Oligopeptídeos/farmacologia , Fosforilação/efeitos dos fármacos , Artérias Umbilicais/metabolismo , Artérias Umbilicais/patologiaRESUMO
Curcumin, a yellow pigment derived from Curcuma longa Linn, has been favored by the Eastern as dietary ingredients for centuries. During the past decade, extensive investigations have revealed curcumin sensitized various chemotherapeutic agents in human breast, colon, pancreas, gastric, liver, brain and hematological malignant disorders in vivo and in vitro. Several pathways and specific targets including NF-κB, STAT3, COX-2, Akt and multidrug resistant protein have been identified to facilitate curcumin as a chemosensitizer. Recent studies suggest HIF-1α participated in the development of drug resistance in cancer cells and targeting HIF-1α either by RNAi or siRNA successfully overcame chemotherapeutic resistance. To investigate the mechanism basis of curcumin as a chemosensitizer in lung cancer, we examined curcumin's effects on HIF-1α in cis-platin (DDP) sensitive A549 and resistant A549/DDP cell lines by RT-PCR and Western blot. HIF-1α in A549/DDP cells was found to be overexpressed at both mRNA and protein levels together with a poor response to DDP. Results from transient transfection and flow cytometry showed the HIF-1α abnormality contributed to DDP resistance in A549/DDP lung cancer cells. Combined curcumin and DDP treatment markedly inhibited A549/DDP cells proliferation, reversed DDP resistance and triggered apoptotic death by promoting HIF-1α degradation and activating caspase-3, respectively. Expression of HIF-1α-dependent P-gp also seemed to decrease as response to curcumin in a dose-dependent manner. Our findings shed light on drug resistant reversing effect of curcumin in lung cancer cells by inhibiting HIF-1α expression and activating caspase-3.