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1.
J Biosci Bioeng ; 127(1): 121-127, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30072117

RESUMO

Many volatile compounds, such as isoprene, a precursor used in the synthesis of natural rubber, have been produced through fermentation using genetically engineered microorganisms. Despite this biotechnological success, measuring the concentrations of volatile compounds during fermentation is difficult because of their high volatility. In current systems, off-line analytical methods usually lead to product loss, whereas on-line methods raise the production cost due to the requirement of complex devices. Here, we developed a novel on-line gas chromatography (GC)-based system for analyzing the concentration of isoprene with the aim to minimize the cost and requirement for devices as compared to current strategies. In this system, a programmable logic controller is used to combine conventional GC with a syringe pump module (SPM) directly connected to the exhaust pipe of the fermentor, and isoprene-containing samples are continuously pumped from the SPM into the GC using an air cylinder recycle stream. We showed that this novel system enables isoprene analysis during fermentation with convenient equipment and without the requirement of an expensive desorption tube. Furthermore, this system may be extended to the detection of other volatile organic compounds in fermentation or chemical processes.


Assuntos
Eletrocromatografia Capilar , Fermentação/fisiologia , Compostos Orgânicos Voláteis/química , Compostos Orgânicos Voláteis/metabolismo , Aerobiose , Reatores Biológicos , Butadienos/química , Butadienos/metabolismo , Eletrocromatografia Capilar/instrumentação , Eletrocromatografia Capilar/métodos , Cromatografia Gasosa/instrumentação , Cromatografia Gasosa/métodos , Cromatografia Gasosa-Espectrometria de Massas/instrumentação , Cromatografia Gasosa-Espectrometria de Massas/métodos , Hemiterpenos/química , Hemiterpenos/metabolismo , Borracha/química , Volatilização
2.
PLoS One ; 12(7): e0181933, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28732091

RESUMO

OBJECTIVE: To determine if perfusion in surgical cavity wall enhancement (SCWE) obtained in early post-treatment MR imaging can stratify time-to-progression (TTP) in glioblastoma. MATERIALS AND METHODS: This study enrolled 60 glioblastoma patients with more than 5-mm-thick SCWEs as detected on contrast-enhanced MR imaging after concurrent chemoradiation therapy. Two independent readers categorized the shape and perfusion state of SCWEs as nodular or non-nodular and as having positive or negative perfusion compared with the contralateral grey matter on arterial spin labeling (ASL). The perfusion fraction on ASL within the contrast-enhancing lesion was calculated. The independent predictability of TTP was analyzed using the Kaplan-Meier method and Cox proportional hazards modelling. RESULTS: The perfusion fraction was higher in the non-progression group, significantly for reader 2 (P = 0.03) and borderline significantly for reader 1 (P = 0.08). A positive perfusion state and (P = 0.02) a higher perfusion fraction of the SCWE were found to become an independent predictor of longer TTP (P = 0.001 for reader 1 and P < 0.001 for reader 2). The contrast enhancement pattern did not become a TTP predictor. CONCLUSION: Assessment of perfusion in early post-treatment MR imaging can stratify TTP in patients with glioblastoma for adjuvant temozolomide therapy. Positive perfusion in SCWEs can become a predictor of a longer TTP.


Assuntos
Neoplasias Encefálicas/patologia , Glioblastoma/patologia , Idoso , Quimiorradioterapia/métodos , Terapia Combinada/métodos , Meios de Contraste/administração & dosagem , Dacarbazina/análogos & derivados , Dacarbazina/uso terapêutico , Progressão da Doença , Feminino , Glioblastoma/tratamento farmacológico , Glioblastoma/radioterapia , Humanos , Interpretação de Imagem Assistida por Computador/métodos , Estimativa de Kaplan-Meier , Imageamento por Ressonância Magnética/métodos , Masculino , Perfusão/métodos , Marcadores de Spin , Temozolomida
3.
J Biol Chem ; 287(16): 13170-81, 2012 Apr 13.
Artigo em Inglês | MEDLINE | ID: mdl-22303020

RESUMO

Bacterial lipoproteins/lipopeptides inducing host innate immune responses are sensed by mammalian Toll-like receptor 2 (TLR2). These bacterial lipoproteins are structurally divided into two groups, diacylated or triacylated lipoproteins, by the absence or presence of an amide-linked fatty acid. The presence of diacylated lipoproteins has been predicted in low-GC content gram-positive bacteria and mycoplasmas based on the absence of one modification enzyme in their genomes; however, we recently determined triacylated structures in low-GC gram-positive Staphylococcus aureus, raising questions about the actual lipoprotein structure in other low-GC content gram-positive bacteria. Here, through intensive MS analyses, we identified a novel and unique bacterial lipoprotein structure containing an N-acyl-S-monoacyl-glyceryl-cysteine (named the lyso structure) from low-GC gram-positive Enterococcus faecalis, Bacillus cereus, Streptococcus sanguinis, and Lactobacillus bulgaricus. Two of the purified native lyso-form lipoproteins induced proinflammatory cytokine production from mice macrophages in a TLR2-dependent and TLR1-independent manner but with a different dependence on TLR6. Additionally, two other new lipoprotein structures were identified. One is the "N-acetyl" lipoprotein structure containing N-acetyl-S-diacyl-glyceryl-cysteine, which was found in five gram-positive bacteria, including Bacillus subtilis. The N-acetyl lipoproteins induced the proinflammatory cytokines through the TLR2/6 heterodimer. The other was identified in a mycoplasma strain and is an unusual diacyl lipoprotein structure containing two amino acids before the lipid-modified cysteine residue. Taken together, our results suggest the existence of novel TLR2-stimulating lyso and N-acetyl forms of lipoproteins that are conserved in low-GC content gram-positive bacteria and provide clear evidence for the presence of yet to be identified key enzymes involved in the bacterial lipoprotein biosynthesis.


Assuntos
Bactérias Gram-Positivas/imunologia , Macrófagos Peritoneais/imunologia , Macrófagos Peritoneais/microbiologia , Receptor 2 Toll-Like/imunologia , Receptor 2 Toll-Like/metabolismo , Animais , Bacillus cereus/imunologia , Bacillus cereus/metabolismo , Bacillus subtilis/imunologia , Bacillus subtilis/metabolismo , Enterococcus faecalis/imunologia , Enterococcus faecalis/metabolismo , Geobacillus/imunologia , Geobacillus/metabolismo , Bactérias Gram-Positivas/metabolismo , Lactobacillus/imunologia , Lactobacillus/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Pneumonia por Mycoplasma/imunologia , Pneumonia por Mycoplasma/metabolismo , Streptococcus sanguis/imunologia , Streptococcus sanguis/metabolismo , Receptor 1 Toll-Like/genética , Receptor 1 Toll-Like/imunologia , Receptor 1 Toll-Like/metabolismo , Receptor 2 Toll-Like/genética , Receptor 6 Toll-Like/genética , Receptor 6 Toll-Like/imunologia , Receptor 6 Toll-Like/metabolismo
4.
BMB Rep ; 44(5): 335-40, 2011 May.
Artigo em Inglês | MEDLINE | ID: mdl-21615989

RESUMO

In this study, we attempted to isolate novel mast cell-stimulating molecules from Staphylococcus aureus. Water-soluble extract of S. aureus cell lysate strongly induced human interleukin- 8 in human mast cell line-1 and mouse interleukin-6 in mouse bone marrow-derived mast cells. The active molecule was purified to homogeneity through a C(18) reverse phase HPLC column. By determination of its structure by MALDITOF and (1)H- and (13)C-NMR, adenosine was revealed to be responsible for the observed cytokine induction activities. Further studies using 8-sulfophenyl theophylline, a selective adenosine receptor blocker, verified that purified adenosine can induce interleukin-8 production via adenosine receptors on mast cells. Moreover, adenosine was purified from S. aureusengulfed RAW264.7 cells, a murine macrophage cell line, used to induce phagocytosis of S. aureus. These results show a novel view of the source of exogenous adenosine in vivo and provide a mechanistic link between inflammatory disease and bacterial infection.


Assuntos
Adenosina/imunologia , Citocinas/biossíntese , Citocinas/imunologia , Inflamação/metabolismo , Macrófagos/imunologia , Mastócitos/metabolismo , Staphylococcus aureus/imunologia , Animais , Linhagem Celular , Humanos , Interleucina-6/imunologia , Interleucina-8/imunologia , Macrófagos/citologia , Mastócitos/citologia , Camundongos , Camundongos Endogâmicos C57BL , Fagocitose/fisiologia , Antagonistas de Receptores Purinérgicos P1/metabolismo , Receptores Purinérgicos P1/metabolismo
5.
FEBS J ; 278(5): 716-28, 2011 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-21205202

RESUMO

Bacterial lipoproteins are known to be diacylated or triacylated and activate mammalian immune cells via Toll-like receptor 2/6 or 2/1 heterodimer. Because the genomes of low G+C content gram-positive bacteria, such as Staphylococcus aureus, do not contain Escherichia coli-type apolipoprotein N-acyltransferase, an enzyme converting diacylated lipoproteins into triacylated forms, it has been widely believed that native lipoproteins of S. aureus are diacylated. However, we recently demonstrated that one lipoprotein SitC purified from S. aureus RN4220 strain was triacylated. Almost simultaneously, another group reported that another lipoprotein SA2202 purified from S. aureus SA113 strain was diacylated. The determination of exact lipidated structures of S. aureus lipoproteins is thus crucial for elucidating the molecular basis of host-microorganism interactions. Toward this purpose, we intensively used MS-based analyses. Here, we demonstrate that SitC lipoprotein of S. aureus RN4220 strain has two lipoprotein lipase-labile O-esterified fatty acids and one lipoprotein lipase-resistant fatty acid. Further MS/MS analysis of the lipoprotein lipase digest revealed that the lipoprotein lipase-resistant fatty acid was acylated to α-amino group of the N-terminal cysteine residue of SitC. Triacylated forms of SitC with various length fatty acids were also confirmed in cell lysate of the RN4220 and Triton X-114 phase in three other S. aureus strains, including SA113 strain and one Staphylococcus epidermidis strain. Moreover, four other major lipoproteins including SA2202 in S. aureus strains were identified as N-acylated. These results strongly suggest that lipoproteins of S. aureus are mainly in the N-acylated triacyl form.


Assuntos
Proteínas de Bactérias/química , Lipoproteínas HDL/química , Staphylococcus aureus/metabolismo , Espectrometria de Massas em Tandem
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