RESUMO
BACKGROUND: Integrin-linked kinase (ILK) is crucial in solid tumors by regulating the Hippo-Yes-associated protein 1 (YAP) pathway. This study aimed to uncover how Helicobacter pylori influences ILK levels and its role in regulating YAP during H. pylori-induced gastric cancer. MATERIALS AND METHODS: GES-1 cells with stable Ilk knockdown and overexpression and a mouse carcinogenesis model for H. pylori infection were constructed. And ILK, the phosphorylated mammalian STE20-like protein kinase 1 (MST1), large tumor suppressor 1 (LATS1; S909, T1079), and YAP (S109, S127) were detected in cells, and mice by western blotting, as well as fluorescence intensity of YAP were assayed by immunofluorescence. YAP downstream genes Igfbp4 and Ctgf, the pathological changes and tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6), interleukin-1beta (IL-1ß), and nitric oxide (NO) levels in mice gastric tissues were detected by real-time PCR, H&E, and ELISA assays. RESULTS: In this study, stable Ilk knockdown cells exhibited significantly higher phosphorylated levels of MST1, LATS1, and YAP, as well as increased YAP in the nuclei of GES-1 cells. Conversely, cells with Ilk overexpression showed opposite results. H. pylori infection led to decreased ILK levels in gastric epithelial cells but increased ILK levels in gastric cancer cell lines (MGC803, SGC7901) and gastric cancer tissues in mice. Treatment with the ILK inhibitor OST-T315 elevated the phosphorylated MST, LATS1, and YAP levels, and inhibited the mRNA levels of Igfbp4 and Ctgf at 44, 48 week-aged mice. OST-T315 also reduced the release of TNF-α, IL-6, IL-1ß, and NO, as well as the progression of gastric cancer caused by H. pylori and N-Nitroso-N-methylurea (NMU) treatment. CONCLUSION: Upon initiation of gastric tumorigenesis signals, H. pylori increases ILK levels and suppresses Hippo signaling, thereby promoting YAP activation and gastric cancer progression. ILK can serve as a potential prevention target to impede H. pylori-induced gastric cancer.
Assuntos
Infecções por Helicobacter , Helicobacter pylori , Proteínas Serina-Treonina Quinases , Neoplasias Gástricas , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Serina-Treonina Quinases/genética , Animais , Neoplasias Gástricas/microbiologia , Neoplasias Gástricas/patologia , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/genética , Infecções por Helicobacter/microbiologia , Infecções por Helicobacter/patologia , Camundongos , Humanos , Modelos Animais de Doenças , Linhagem Celular , MasculinoRESUMO
CagA is a significant oncogenic factor injected into host cells by Helicobacter pylori, which is divided into two subtypes: East Asian type (CagAE), characterized by the EPIYA-D motif, and western type (CagAW), harboring the EPIYA-C motif. CagAE has been reported to have higher carcinogenicity than CagAW, although the underlying reason is not fully understood. SHIP2 is an intracellular phosphatase that can be recruited by CagA to perturb the homeostasis of intracellular signaling pathways. In this study, we found that SHIP2 contributes to the higher oncogenicity of CagAE. Co-Immunoprecipitation and Pull-down assays showed that CagAE bind more SHIP2 than CagAW. Immunofluorescence staining showed that a higher amount of SHIP2 recruited by CagAE to the plasma membrane catalyzes the conversion of PI(3,4,5)P3 into PI(3,4)P2. This alteration causes higher activation of Akt signaling, which results in enhanced IL-8 secretion, migration, and invasion of the infected cells. SPR analysis showed that this stronger interaction between CagAE and SHIP2 stems from the higher affinity between the EPIYA-D motif of CagAE and the SH2 domain of SHIP2. Structural analysis revealed the crucial role of the Phe residue at the Y + 5 position in EPIYA-D. After mutating Phe of CagAE into Asp (the corresponding residue in the EPIYA-C motif) or Ala, the activation of downstream Akt signaling was reduced and the malignant transformation of infected cells was alleviated. These findings revealed that CagAE hijacks SHIP2 through its EPIYA-D motif to enhance its carcinogenicity, which provides a better understanding of the higher oncogenic risk of H. pylori CagAE.
Assuntos
Motivos de Aminoácidos , Antígenos de Bactérias , Proteínas de Bactérias , Infecções por Helicobacter , Helicobacter pylori , Fosfatidilinositol-3,4,5-Trifosfato 5-Fosfatases , Humanos , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/genética , Helicobacter pylori/genética , Helicobacter pylori/patogenicidade , Fosfatidilinositol-3,4,5-Trifosfato 5-Fosfatases/genética , Fosfatidilinositol-3,4,5-Trifosfato 5-Fosfatases/metabolismo , Antígenos de Bactérias/metabolismo , Antígenos de Bactérias/genética , Infecções por Helicobacter/microbiologia , Transdução de Sinais , Carcinogênese , Ligação Proteica , População do Leste AsiáticoRESUMO
BACKGROUND: SHP1 has been documented as a tumor suppressor and it was thought to play an antagonistic role in the pathogenesis of Helicobacter pylori infection. In this study, the exact mechanism of this antagonistic action was studied. MATERIALS AND METHODS: AGS, MGC803, and GES-1 cells were infected with H. pylori, intracellular distribution changes of SHP1 were first detected by immunofluorescence. SHP1 overexpression and knockdown were then constructed in these cells to investigate its antagonistic roles in H. pylori infection. Migration and invasion of infected cells were detected by transwell assay, secretion of IL-8 was examined via ELISA, the cells with hummingbird-like alteration were determined by microexamination, and activation of JAK2/STAT3, PI3K/Akt, and ERK pathways were detected by immunoblotting. Mice infection model was established and gastric pathological changes were evaluated. Finally, the SHP1 activator sorafenib was used to analyze the attenuating effect of SHP1 activation on H. pylori pathogenesis in vitro and in vivo. RESULTS: The sub-localization of SHP1 changed after H. pylori infection, specifically that the majority of the cytoplasmic SHP1 was transferred to the cell membrane. SHP1 inhibited H. pylori-induced activation of JAK2/STAT3 pathway, PI3K/Akt pathway, nuclear translocation of NF-κB, and then reduced EMT, migration, invasion, and IL-8 secretion. In addition, SHP1 inhibited the formation of CagA-SHP2 complex by dephosphorylating phosphorylated CagA, reduced ERK phosphorylation and the formation of CagA-dependent hummingbird-like cells. In the mice infection model, gastric pathological changes were observed and increased IL-8 secretion, indicators of cell proliferation and EMT progression were also detected. By activating SHP1 with sorafenib, a significant curative effect against H. pylori infection was obtained in vitro and in vivo. CONCLUSIONS: SHP1 plays an antagonistic role in H. pylori pathogenesis by inhibiting JAK2/STAT3 and PI3K/Akt pathways, NF-κB nuclear translocation, and CagA phosphorylation, thereby reducing cell EMT, migration, invasion, IL-8 secretion, and hummingbird-like changes.
Assuntos
Infecções por Helicobacter , Helicobacter pylori , Animais , Camundongos , Proteínas de Bactérias/metabolismo , Antígenos de Bactérias/metabolismo , Helicobacter pylori/fisiologia , NF-kappa B/metabolismo , Interleucina-8/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Infecções por Helicobacter/patologia , Sorafenibe/metabolismo , Células Epiteliais/metabolismoRESUMO
Micro(nano)plastics are prevalent in the environment, and prolonged exposure to them represents a threat to human health. The goal of this study is to assess the health risk of long-term exposure to nanoplastics (NPs) at environmental concentrations on the intestinal mechanical and immune barrier in mice. In this study, mice were provided drinking water containing polystyrene NPs (PS-NPs; 0.1, 1, and 10 mg·L-1) for 32 consecutive weeks. The levels of endocytosis proteins caveolin and clathrin and of tight junctional proteins claudin-1, occludin, and ZO-1, and morphological changes, proportion of lymphocytes B in MLNs and lymphocytes T in IELs and LPLs were determined by immunohistochemistry, hematoxylin-eosin, and flow cytometry assays in the intestinal tissues of mice at 28 weeks. The activities or concentrations of ROS, SOD, MDA, and GSH-Px and inflammatory factors (IL-1ß, IL-6, and TNF-α) in the intestinal tissues of mice were measured by ELISA at 12, 16, 20, 24, and 32 weeks. Compared with the control group, oral ingested PS-NPs entered the intestinal tissues of mice and upregulated expression levels of the clathrin and caveolin. The intestinal tissue structure of mice in the PS-NPs (1 and 10 mg·L-1) exposure groups showed significant abnormalities, such as villus erosion, decreased of crypts numbers and large infiltration of inflammatory cells. Exposure to 0.1 mg·L-1 PS-NPs decreased occludin protein levels, but not claudin-1 and ZO-1 levels. The levels of these three tight junction proteins decreased significantly in the 1 and 10 mg·L-1 PS-NPs exposed groups. Exposure to PS-NPs led to a significant time- and dose-dependent increase in ROS and MDA levels, and concurrently decreased GSH-Px and SOD contents. Exposure to PS-NPs increased the proportion of B cells in MLNs, and decreased the proportion of CD8+ T cells in IELs and LPLs. The levels of pro-inflammatory cytokines IL-6, TNF-α and IL-1ß were markedly elevated after PS-NPs exposure. Long-term PS-NPs exposure impaired intestinal mechanical and immune barrier, and indicate a potentially significant threat to human health.
Assuntos
Nanopartículas , Poliestirenos , Humanos , Poliestirenos/toxicidade , Microplásticos , Linfócitos T CD8-Positivos , Interleucina-6 , Ocludina , Espécies Reativas de Oxigênio , Fator de Necrose Tumoral alfa , Caveolinas , Clatrina , Superóxido DismutaseRESUMO
YAP participates in autophagy associated with many diseases. In this study, we demonstrate that YAP promotes autophagy by interacting with beclin 1, upregulating beclin 1 and LC3B-II protein expression, and promoting autophagosome formation after H. pylori infection in a vacuolating cytotoxin A-dependent manner. The protein levels of ß-catenin in the cytoplasm and nuclei of GES-1 cells and the mRNA levels of Axin2, Myc, Lgr5, and Ccnd1 were increased in H. pylori-infected cells or YAP-overexpressed cells, but were decreased in YAP-silenced cells. The ß-catenin inhibitor XAV939 significantly downregulated autophagy, whereas the activator LiCl showed opposite effects. An H. pylori-infected mouse model of gastric carcinoma was successfully established. The mouse model showed that H. pylori infection, when combined with NMU, promoted the tumorigenesis of gastric tissues; increased IL-1ß, IL-6, and TNF-α levels; promoted NO release; and increased the expression of beclin 1, LC3B-II more than NMU alone. Chloroquine inhibited these phenomena, but did not completely attenuate the effects of H. pylori. These results demonstrate that chloroquine can be used as a drug for the treatment of H. pylori-related gastric cancer, but the treatment should simultaneously remove H. pylori.
Assuntos
Infecções por Helicobacter , Helicobacter pylori , Neoplasias Gástricas , Camundongos , Animais , beta Catenina/metabolismo , Cloroquina/farmacologia , Cloroquina/metabolismo , Proteína Beclina-1/metabolismo , Proteína Beclina-1/farmacologia , Carcinogênese/metabolismo , Carcinogênese/patologia , Neoplasias Gástricas/genética , Autofagia , Modelos Animais de Doenças , Infecções por Helicobacter/metabolismo , Mucosa Gástrica/patologiaRESUMO
Klebsiella pneumoniae is one of the most common opportunistic pathogens causing hospital- and community-acquired infections. Antibiotic resistance in K. pneumoniae has emerged as a major clinical and public health threat. Persisters are specific antibiotic-tolerant bacterial cells. Studies on the mechanism underlying their formation mechanism and growth status are scarce. Therefore, it is urgent to explore the key genes and signalling pathways involved in the formation and recovery process of K. pneumoniae persisters to enhance the understanding and develop relevant treatment strategies. In this study, we treated K. pneumoniae with a lethal concentration of levofloxacin. It resulted in a distinct plateau of surviving levofloxacin-tolerant persisters. Subsequently, we obtained bacterial samples at five different time points during the formation and recovery of K. pneumoniae persisters to perform transcriptome analysis. ptsH gene was observed to be upregulated during the formation of persisters, and down-regulated during the recovery of the persisters. Further, we used CRISPR-Cas9 to construct ΔptsH, the ptsH-knockout K. pneumoniae strain, and to investigate the effect of ptsH on the persister formation. We observed that ptsH can promote the formation of persisters, reduce accumulation of reactive oxygen species, and enhance antioxidant capacity by reducing cyclic adenosine monophosphate (cAMP) levels. To the best of our knowledge, this is the first study to report that ptsH plays a vital role in forming K. pneumoniae persisters. This study provided important insights to further explore the mechanism underlying the formation of K. pneumoniae persisters and provided a potential target for treating infection with K. pneumoniae persisters.
Assuntos
Infecções por Klebsiella , Klebsiella pneumoniae , Humanos , Levofloxacino/farmacologia , Antibacterianos/farmacologia , Monofosfato de Adenosina , Fosfotransferases/farmacologia , Infecções por Klebsiella/microbiologiaRESUMO
A novel Gram-negative, motile, aerobic, spiral-shaped bacterium designated D5T, was isolated from a coastal sediment collected in the Yellow Sea. Optimal growth occurred at 30 °C, pH 7.0-8.0 and in the presence of 1-3% (w/v) NaCl. Strain D5T contained ubiquinone 8 (Q-8) as the predominant respiratory quinone. The major fatty acids (> 10%) were C16:0, C16:1 ω7c/C16:1 ω6c and C18:1w7c/C18:1w6c. The main polar lipids were phosphatidylglycerol and phosphatidylethanolamine. The draft genome is 5.6 Mb in length, and DNA G + C content is 47.2 mol%. 16S rRNA gene sequences showed that strain D5T is most closely related to Oceanospirillum beijerinckii NBRC 15445T (97.8%, sequence similarity). However, the digital DNA-DNA hybridization (dDDH) value and average nucleotide identity (ANI) between strain D5T and O. beijerinckii is only 27.8% and 77.1%. Phylogenetic trees based on 16S rRNA gene sequences and whole genomes all indicated that strain D5T formed a separate branch in the genus Oceanospirillum. Combined results of the polyphasic analyses suggested that strain D5T represents a novel species in the genus Oceanospirillum, for which the name Oceanospirillum sediminis sp. nov. is proposed. The type strain is D5T (= MCCC 1K06061T = KCTC 62987T).
Assuntos
Sedimentos Geológicos , Oceanospirillaceae , Filogenia , Água do Mar , Técnicas de Tipagem Bacteriana , DNA Bacteriano/genética , Ácidos Graxos/química , Sedimentos Geológicos/microbiologia , Oceanospirillaceae/classificação , Oceanospirillaceae/isolamento & purificação , Fosfolipídeos/química , RNA Ribossômico 16S/genética , Água do Mar/microbiologia , Análise de Sequência de DNA , Ubiquinona/químicaRESUMO
Helicobacter pylori infection is an essential factor in the development of human gastric diseases, but its pathogenic mechanism is still unclear. In this work we have showed that, the LC3II levels were increased and ß1-integrin levels were decreased in H. pylori-positive human gastric tissue samples and H. pylori co-cultured GES-1 cells. There was significant upregulation of LC3II levels and downregulation of P62 levels in GES-1 cells after ß1-integrin knockdown co-cultured with H. pylori. This indicated that ß1-integrin downregulation promoted autophagy in GES-1 cells after H. pylori infection. The cell apoptosis rate and poly ADP-ribose polymerase (PARP) and caspase-3 activities were increased in GES-1 cells pretreated with 3-methyladenine (3-MA ) after H. pylori infection. Furthermore, there was a significant decrease in apoptosis of ß1-integrin knockdown GES-1 cells co-cultured with H. pylori; apoptosis was also downregulated in ß1-integrin knockdown- and 3-MA-treated GES-1 cells co-cultured with H. pylori. Correspondingly, PARP and caspase-3 activities were decreased in ß1-integrin knockdown cells co-cultured with H. pylori and ß1-integrin knockdown-3-MA-treated-1 cells with H. pylori infection. Thus, ß1-integrin is a novel autophagy and apoptosis regulator during H. pylori infection. However, inhibition of autophagy did not reverse the decrease in apoptosis caused by downregulation of ß1-integrin.
Assuntos
Apoptose , Autofagia , Células Epiteliais/metabolismo , Infecções por Helicobacter/metabolismo , Helicobacter pylori , Integrina beta1/metabolismo , Proteínas Associadas aos Microtúbulos/metabolismo , Caspase 3/metabolismo , Linhagem Celular , Mucosa Gástrica/metabolismo , Regulação da Expressão Gênica , Infecções por Helicobacter/microbiologia , Interações Hospedeiro-Patógeno , Humanos , Poli(ADP-Ribose) Polimerases/metabolismoRESUMO
The ability of Helicobacter pylori to manipulate host autophagy is an important pathogenic mechanism. We found an inverse correlation between the expression of ILK and the autophagy marker protein LC3B in H. pylori-positive human samples, H. pylori-infected mice models and H. pylori-infected GES-1 cell lines. When the ILK-knockdown GES-1 cells were infected by H. pylori, CagA were significantly degraded, autophagosomes accumulation and autolysosomes formation were significantly increased, and LC3B protein levels and ratio of LC3BII to LC3BI were also remarkably upregulated. And chloroquine treatment increased LC3B levels in ILK-knockdown GES-1 cells. The expression levels of both Rac1 and RhoA were downregulated in GES-1 cells after H. pylori infection and were decreased in ILK-knockdown GES-1 cells. The mRNA and protein levels of PAK1, MLC, and LIMK were significantly decreased and cofilin mRNA and protein levels were significantly increased in GES-1 cells treated with the Rac1 inhibitor NSC 23766. The mRNA and protein levels of ROCK1, ROCK2, MLC, and LIMK1 were significantly reduced and cofilin mRNA and protein levels were significantly increased in GES-1 cells treated with the RhoA inhibitor CCG-1423. F-actin was significantly reduced in Rac1- or RhoA-inhibited GES-1 cells. F-actin depolymerization induced autophagosomes accumulation, autolysosomes formation, and the increase of LC3B levels in GES-1 cells. Therefore, these findings revealed that ILK could serve as a novel regulator to affect Rac1/PAK1 and RhoA/ROCKs signaling pathways, thereby influencing H. pylori-induced autophagy.
Assuntos
Autofagia , Células Epiteliais/microbiologia , Infecções por Helicobacter , Helicobacter pylori , Animais , Células Cultivadas , Células Epiteliais/enzimologia , Mucosa Gástrica , Humanos , Camundongos , Proteínas Serina-Treonina Quinases , Transdução de Sinais , Proteínas rac1 de Ligação ao GTP , Quinases Associadas a rho , Proteína rhoA de Ligação ao GTPRESUMO
A novel Gram-stain-negative, facultative anaerobic, motile bacterium, designated as 404T, was isolated from a marine sediment sample in the Bohai Gulf, China. Growth was observed at 10-35 °C (optimum, 20-25 °C) and in the presence of 1.0-6.0% (w/v) NaCl (optimum, 1.0-3.0%). Phylogenetic analysis based on 16S rRNA gene sequences indicated that the strain 404T belonged to the genus Vibrio, showing the highest sequence similarity to Vibrio renipiscarius KCTC 42287T (97.6%). The draft genome is 4.5 Mb in length, containing 4278 protein-coding genes, 60 tRNA genes and 9 rRNA genes, and DNA G+C content is 44.1 mol%. Strain 404T contains phosphatidylglycerol, phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylcholine, and phospholipid as the main polar lipids, and the predominant quinone is ubiquinone Q-8. The major cellular fatty acids (>8.0%) are C16â:â1ω6c and/or C16â:â1ω7c, C16â:â0, C18â:â1ω6c and/or C18â:â1ω7c. Strain 404T shows some typical characteristics among the members of genus Vibrio, while it can be clearly distinguished from the closely related type strains through genome analysis (average nucleotide identity and digital DNA-DNA hybridization values), fatty acid composition and a series of physiological and biochemical characteristics. On the basis of the polyphasic analysis, strain 404T is considered to represent a novel species of the genus Vibrio, for which the name Vibrio marinisediminis sp. nov., is proposed. The type strain is 404T (= MCCC 1H00367T = KCTC 62958T).
Assuntos
Sedimentos Geológicos , Vibrio , Técnicas de Tipagem Bacteriana , China , DNA Bacteriano/genética , Ácidos Graxos , Fosfolipídeos , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Vibrio/genéticaRESUMO
A novel Gram-stain-negative, moderately halophilic bacterium, designated strain 204T, was isolated from a marine sediment sample in the Bohai Gulf, Yellow Sea, China. Cells of strain 204T are aerobic, motile, cocci or short rods with two lateral flagella. Growth was observed at 15-40 °C (optimum, 30 °C), pH 6.0-9.0 (optimum, 7.0-7.5) and in the presence of 1.0-18.0% (w/v) NaCl (optimum, 3.0-8.0%). Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain 204T belonged to the genus Halomonas, showing highest sequence similarity to Halomonas alimentaria YKJ-16T (98.08%), followed by Halomonas sediminicola (97.47%), Halomonas fontilapidosi (97.14%), Halomonas halodenitrificans (96.98%), Halomonas ventosae (96.92%), and Halomonas shengliensis (96.85%). The draft genome is 3.8 Mb in length, containing 3673 protein-coding genes, 62 tRNA genes and 10 rRNA genes, and DNA G+C content is 62.7 mol%. Strain 204T contains phosphatidylglycerol, phosphatidylethanolamine, and diphosphatidylglycerol as the main polar lipids, and the predominant respiratory quinone was ubiquinone Q-9. The major cellular fatty acids (> 5%) are C18:1ω7c, C16:1ω7c and/or C16:1ω6c, C16:0 and C12:03-OH. Strain 204T was clearly distinguished from the closely related type strains through phylogenetic analysis, dDNA-DNA hybridization, fatty acid composition data and a range of physiological and biochemical characteristics comparisons. On the basis of the polyphasic analysis, strain 204T is considered to represent a novel species of the genus Halomonas, for which the name Halomonas marinisediminis sp. nov. is proposed. The type strain is 204T (= MCCC 1H00366T = KCTC 62957T).
Assuntos
Halomonas , Técnicas de Tipagem Bacteriana , China , DNA Bacteriano/genética , Ácidos Graxos , Halomonas/genética , Fosfolipídeos , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNARESUMO
BACKGROUND: Helicobacter pylori (H. pylori) is a common human pathogen, which is closely correlated with gastric cancer (GC). However, the mechanism of H. pylori-related GC has not been elucidated. This study aimed to explore the role of H. pylori infection in GC and find biomarkers for early diagnosis of H. pylori-related GC. METHODS: We identified differentially expressed microRNAs (DEMs) and genes (DEGs) from the Gene Expression Omnibus (GEO) dataset, constructed microRNA-(miRNA-)mRNA expression networks, analyzed the function and signal pathway of cross-genes, analyzed the relations between cross-genes and GC prognosis with the Cancer Genome Atlas (TCGA) data, and verified the expression of cross-genes in patients with H. pylori infection. RESULTS: 22 DEMs and 68 DEGs were identified in GSE197694 and GSE27411 dataset. 16 miRNAs and 509 genes were involved in the expression network, while the cross-genes of the network were mainly enriched in MAP kinase (MAPK) signaling pathway and TGF-beta signaling pathway. Patients with higher expression of hsa-miR-196b-3p, CALML4, or SMAD6 or lower expression of PITX2 or TGFB2 had better outcomes than those with lower expression of hsa-miR-196b-3p, CALML4, or SMAD6 or higher expression of PITX2 or TGFB2 (P < 0.05). Patients with H. pylori infection had a higher expression of hsa-miR-196b-3p and CALML4 than those without H. pylori infection (P < 0.05). CONCLUSION: The study of miRNA-mRNA expression network would provide molecular support for early diagnosis and treatment of H. pylori-related GC.
RESUMO
Helicobacter pylori infection can cause a wide range of digestive diseases. Gene hp0788 encodes an outer membrane protein HofF, which can reduce the bacterial adherence to the GES-1â¯cells and affect pathogenesis of H. pylori. In this study, the role of hp0788 in H. pylori infection was further analyzed. RNA-seq data showed that two genes (hp0523 and hp0539), located on the cagPAI, were down-regulated in Δ0788 mutant. The changes were confirmed through qRT-PCR, and the expression of these two genes will be almost recovered to the normal level in complemented strain. These two genes, hp0523 and hp0539, are known to be necessary for integrated T4SS, which related to CagA translocation and IL-8 induction. In H. pylori infected assay, lower amount of phosphorylated CagA and lower induction of IL-8 were both detected in GES-1â¯cells infected by Δ0788 mutant, compared with the wild type strain. Meanwhile, these reductions almost recovered to the wild-type level in complemented strain. These results reveal that there is a correlation between hp0788 disruption and CagA/IL-8 decline. Deletion of CagA-encoding gene (hp0547) in Δ0788 mutant was further constructed. The double deleted mutant shows lower IL-8-inducing capability than Δ0788 mutant, indicated the correlation between deficiency of CagA and reduced IL-8 production. These results together imply that disruption of hp0788 might affect the efficiency of T4SS and CagA injection, then weaken the induction of IL-8 in infected GES-1â¯cells.
Assuntos
Proteínas de Bactérias/genética , Células Epiteliais/microbiologia , Infecções por Helicobacter/microbiologia , Helicobacter pylori/genética , Gastropatias/microbiologia , Antígenos de Bactérias/genética , Aderência Bacteriana/genética , Linhagem Celular Tumoral , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/fisiologia , Humanos , Interleucina-8/metabolismoRESUMO
AIMS AND METHODS: Retrospectively, this paper compared the differences of the Epstein-Barr virus (EBV)-encoded small RNAs (EBERs), protein expression and gene mutations of tumor suppressor gene p53 (TP53) in keratinized nasopharyngeal squamous cell carcinoma (KNSCC) and nonKNSCC, and the relationships between pathological features and the prognosis of patients were analyzed. RESULTS: The positive rate of EBERs hybridization and TP53 expressions was 76.3% and 52.2%, respectively, while the mutation rate of TP53 gene was 39.6%. Logistic regression analysis showed direct relationships between the subtypes of nasopharyngeal squamous cell carcinoma (NPSCC) and EBERs-positive, or frequent consumption of pickled food. Overall survival rates of patients with positive TP53 expression, the TP53 gene mutations, vascular invasions, organ metastases, lymph node metastasis, and clinical recurrence were significantly lower than those of patients without those symptoms. The poorer prognosis was related to regularly drinking and the advanced age. According to the Cox regression analysis, we found that the main prognostic factors of NPSCC patients were the aging, recurrence, TP53 gene mutations, especially exon 7 or 8 mutations. CONCLUSIONS: We concluded that there were the correlations between NPSCC subtypes with EBV infection and frequent intaking of pickled food, while aging, clinical recurrence, and TP53 gene mutations were independent predictors for the poor prognosis of nasopharyngeal carcinoma.
Assuntos
Carcinoma de Células Escamosas/etiologia , Carcinoma de Células Escamosas/mortalidade , Infecções por Vírus Epstein-Barr/complicações , Herpesvirus Humano 4 , Neoplasias Nasofaríngeas/etiologia , Neoplasias Nasofaríngeas/mortalidade , Proteína Supressora de Tumor p53/genética , Adulto , Idoso , Biomarcadores Tumorais , Carcinoma de Células Escamosas/diagnóstico , Linhagem Celular Tumoral , Suscetibilidade a Doenças , Infecções por Vírus Epstein-Barr/virologia , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Imuno-Histoquímica , Hibridização In Situ , Masculino , Pessoa de Meia-Idade , Mutação , Neoplasias Nasofaríngeas/diagnóstico , Gradação de Tumores , Prognóstico , Recidiva , Fatores de Risco , Carga TumoralRESUMO
A novel Gram-stain-negative bacterium, designated strain BH-SD16T, was isolated from a marine sediment sample collected in the Bohai Sea. Cells of strain BH-SD16T are aerobic, non-flagellated oval-shaped rods, showing oxidase- and catalase-positive activities. Growth occurs between 15-45 °C (optimum, 30 °C), at pH 6.0-9.0 (pH 7.0-7.5) and with 1-10â% (w/v) NaCl (3.0â%). Strain BH-SD16T contains C18â:â1ω7c (49.2â%), C16â:â0 (17.7â%) and C18â:â1ω7c 11-methyl (16.6â%) as the predominant fatty acids and ubiquinone-10 as the major respiratory quinone. The major polar lipids comprise phosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine, diphosphatidylglycerol and two glycolipids. The size of the draft genome is 3â442â538 bp, including 3213 protein-coding genes, 40 tRNA genes and three rRNA genes, and the DNA G+C content is 63.4 mol%. Strain BH-SD16T shows the highest 16S rRNA gene sequence similarity to Pseudooctadecabacter jejudonensis (95.7â%), strains of the genus Octadecabacter(95.4-95.6â%) and strains of the genus Loktanella(93.8-95.4â%). Phylogenetic trees based on 16S rRNA gene sequences show that strain BH-SD16T forms a distinct lineage within the family Hyphomonadaceae, which is also confirmed in the multigenic phylogenetic tree calculated by RAxML. Based on the results of phenotypic, chemotaxonomic and phylogenetic analysis, strain BH-SD16T is considered to represent a novel genus and species in the family Hyphomonadaceae, for which the name Thalassorhabdomicrobium marinisediminis gen. nov., sp. nov. is proposed. The type strain is BH-SD16T (=CCTCC AB 2017073T=KCTC 62201T).
Assuntos
Alphaproteobacteria/classificação , Sedimentos Geológicos/microbiologia , Filogenia , Água do Mar/microbiologia , Alphaproteobacteria/isolamento & purificação , Técnicas de Tipagem Bacteriana , Composição de Bases , China , DNA Bacteriano/genética , Ácidos Graxos/química , Glicolipídeos/química , Oceanos e Mares , Fosfolipídeos/química , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Ubiquinona/químicaRESUMO
A Gram-stain-negative bacterium, strain BH-SD19T, that was isolated from a marine sediment sample collected from the Bohai Sea, was subjected to a polyphasic taxonomic study. Cells of BH-SD19T are non-flagellated, non-gliding, oval-shaped rods, 0.5-1.0 µm wide and 1.0-2.0 µm long. BH-SD19T is strictly aerobic, and oxidase- and catalase-positive. Growth occurs at 15-40 °C (optimum 35 °C), at pH 6.0-8.5 (optimum 7.0-7.5) and with 1-10â% (w/v) NaCl (optimum 2â%). The predominant fatty acids are C19â:â0cyclo ω8c (46.5â%), C16â:â0 (20.3â%) and C18â:â1ω7c and/or C18â:â1ω6c (10.6â%). The major respiratory quinone is Q-10. The major polar lipids are phosphatidylglycerol, phosphatidylethanolamine and an unidentified phospholipid. The DNA G+C content is 64.0 mol%. BH-SD19T shows the highest 16S rRNA sequence similarity to Pontibaca methylaminivorans (95.2â%) and strains of species of the genus Roseovarius(93.4-95.2â%). Sequence similarity values between BH-SD19T and other phylogenetically related species are all below 95.0â%. Phylogenetic trees based on 16S rRNA gene sequences indicate that BH-SD19T forms a distinct lineage and does not join any known genera in the trees. Phenotypic, chemotaxonomic and phylogenetic data indicate that BH-SD19T represents a novel genus and species in the family Rhodobacteraceae, for which the name Pelagivirga sediminicola gen. nov., sp. nov. is proposed. The type strain is BH-SD19T (=CCTCC AB 2017074T=KCTC 62202T).
Assuntos
Sedimentos Geológicos/microbiologia , Filogenia , Rhodobacteraceae/classificação , Técnicas de Tipagem Bacteriana , Composição de Bases , DNA Bacteriano/genética , Ácidos Graxos/química , Oceanos e Mares , Fosfolipídeos/química , RNA Ribossômico 16S/genética , Rhodobacteraceae/genética , Rhodobacteraceae/isolamento & purificação , Análise de Sequência de DNA , Ubiquinona/análogos & derivados , Ubiquinona/químicaRESUMO
Epidermal growth factor receptor (EGFR) is an important gene in the development of lung adenocarcinoma. However, there is controversy regarding the association between EGFR mutations and survival time of patients with lung adenocarcinoma. In the present study, tissue specimens and clinical data were collected from 219 patients with lung adenocarcinoma who had not undergone prior radiotherapy or chemotherapy. EGFR mutations were detected using a fluorescence polymerase chain reaction method, and the association between EGFR mutations and clinicopathological characteristics was analyzed. Overall survival (OS) curves were constructed using the Kaplan-Meier method and the influence of clinicopathological characteristics on OS was analyzed using the Cox regression model. The EGFR mutation rate was 50.7%, and the most common mutations were the L858R substitution mutation in exon 21 (L858R; 54.9%) and the deletion mutation in exon 19 (19-Del; 36%). The presence of EGFR mutations varied significantly with sex, smoking history, T stage, vascular invasion and adenocarcinoma subtypes (P<0.05). The survival time was significantly longer for female, young (<60 years-old), non-smokers or patients exhibiting EGFR mutations (G719X, 19-Del, L858R and L861Q). The survival time was also significantly longer for patients with a 19-Del mutation, early stage tumors, tyrosine kinase inhibitors targeted therapy-treated patients, for those not exhibiting nerve or vascular invasion, and for those without disease recurrence (P<0.05). Multivariate analysis revealed that tumor pathological Tumor-Node-Metastasis (pTNM) stage, nerve invasion, vascular invasion, EGFR mutation and the 19-Del mutation were independent predictors (P<0.05). Therefore, tumor pTNM stage, nerve invasion, vascular invasion and EGFR mutation status, particularly that of 19-Del, were independent prognostic factors for patients with lung adenocarcinoma.
RESUMO
Human leukocyte-associated immunoglobulin-like receptor-1 (LAIR-1), an immunoinhibitory receptor, is expressed on most types of hematopoietic cells and some tumor cells. LAIR-1 plays an inhibitory role in immune cell maturation, differentiation, and activation. LAIR-1 is also involved in some autoimmune diseases and tumors. However, the mechanism controlling the regulation of the LAIR-1 gene is still unknown. In order to elucidate the molecular mechanisms involved in LAIR-1 regulation, in the present study, we cloned and characterized the promoter region of LAIR-1 gene using a series of truncated promoter plasmids in luciferase reporter assays. Our results show that the basic core promoter of LAIR-1 is located within the region -256/-8 relative to the translational start site. Our further studies indicate that five ETS transcription factors: ELF-1, ETV-4, ETV-3L, ETS-1 and ETS-2, can up-regulate the LAIR-1 basic promoter activity. Of these, ETS-2 is the most effective transcription factor. Moreover, ETS-2 was confirmed to interact directly with the basic promoter of LAIR-1. This study presents the first description of regions/factors capable of up-regulation the promoter activity of LAIR-1. This new knowledge contributes to understanding of the molecular mechanisms involved in LAIR-1 associated immune regulation and diseases.
Assuntos
Proteínas Proto-Oncogênicas c-ets/metabolismo , Receptores Imunológicos/metabolismo , Proteínas E1A de Adenovirus/genética , Proteínas E1A de Adenovirus/metabolismo , Efrina-A2/genética , Efrina-A2/metabolismo , Regulação da Expressão Gênica/fisiologia , Células HEK293 , Humanos , Regiões Promotoras Genéticas , Proteína Proto-Oncogênica c-ets-1/genética , Proteína Proto-Oncogênica c-ets-1/metabolismo , Proteína Proto-Oncogênica c-ets-2/genética , Proteína Proto-Oncogênica c-ets-2/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-ets/genética , Receptores Imunológicos/genética , Regulação para CimaRESUMO
Many virulence genes have been reported to play important roles in Helicobacter pylori pathogenesis. However the detailed mechanisms of many of them have not been completely clear. In this study, we found gene hp0169, encoding a putative collagenase (HpPrtC), was involved in pathogenesis of H. pylori. Recombinant HpPrtC shows activities to both native and heat-denatured collagens. This result indicated that HpPrtC may act as a virulence factor to help the bacterium colonize in their host stomach by degrading surrounding collagens. hp0169 was deleted by homologous recombination to study its function in bacterium-host cell interaction. For the pathogenic functions on the host cells, the hp0169 mutant exhibits no significant changes on inducing apoptosis of GES-1 cells. However, the viability and proliferation rate of GES-1 cells infected with mutant strain were higher than the cells infected with wild-type strain. These results indicated that except for its collagenolytic activity, HpPrtC might participate in H. pylori pathogenesis through an additional pathway. Functional studies on hp0169 involved in pathogenesis would shed light on deep understanding of the pathogenic mechanism of H. pylori.
Assuntos
Colagenases/metabolismo , Helicobacter pylori/patogenicidade , Interações Hospedeiro-Patógeno , Fatores de Virulência/metabolismo , Apoptose , Linhagem Celular , Proliferação de Células , Colágeno/metabolismo , Colagenases/genética , Células Epiteliais/microbiologia , Células Epiteliais/fisiologia , Deleção de Genes , Técnicas de Inativação de Genes , Helicobacter pylori/enzimologia , Helicobacter pylori/genética , Humanos , Hidrólise , Fatores de Virulência/genéticaRESUMO
Helicobacter pylori infection represents a key factor in the etiology of various gastro-duodenal diseases, ranging from chronic gastritis to the development of peptic ulcer disease and end-stage gastric cancer. In the present study, the 26695 and SS1 strains of H. pylori were used to study the differential functional profiles of gastric epithelial cells infected with H. pylori. The apoptosis rates in GES-1 cells were significantly increased 3, 12 and 24 h after H. pylori 26695 and SS1 infection. Moreover, apoptosis by cells infected with the H. pylori 26695 strain was significantly higher than cells infected with the SS1 strain of H. pylori. No significant changes in the proliferation rates of GES-1 cells were observed after H. pylori 26695 or SS1 infection at any time during the experimental period. Exposure to H. pylori 26695 and SS1 induced a significant decline in the adhesion rates of GES-1 cells in a time-dependent manner. Furthermore, H. pylori 26695 infection increased migration of GES-1 cells every hour during the whole experimental period compared with control cells. However, GES-1 cells infected with the H. pylori SS1 strain exhibited migration rates almost stable and comparable to those of control cells. These results indicate that the gastric epithelial cells respond differently depending on the H. pylori strains. This study indicates that the development of different gastric-related diseases may be a H. pylori strain-specific response.