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1.
Molecules ; 29(13)2024 Jun 23.
Artigo em Inglês | MEDLINE | ID: mdl-38998934

RESUMO

Oral malodor still constitutes a major challenge worldwide. A strong effort is invested in eliminating volatile sulfur compound-producing oral bacteria through organic natural products such as essential oils. Fusobacterium nucleatum is a known volatile sulfur compound-producing bacteria that inspires oral malodor. The aim of the present study was to test the effect of lavender essential oil on the bacterium's ability to produce volatile sulfide compounds, the principal components of oral malodor. Lavender (Lavandula angustifolia) essential oil was extracted by hydrodistillation and analyzed using GC-MS. The minimal inhibitory concentration (MIC) of lavender essential oil on Fusobacterium nucleatum was determined in a previous trial. Fusobacterium nucleatum was incubated anaerobically in the presence of sub-MIC, MIC, and above MIC concentrations of lavender essential oil, as well as saline and chlorhexidine as negative and positive controls, respectively. Following incubation, volatile sulfur compound levels were measured using GC (Oralchroma), and bacterial cell membrane damage was studied using fluorescence microscopy. Chemical analysis of lavender essential oil yielded five main components, with camphor being the most abundant, accounting for nearly one-third of the total lavender essential oil volume. The MIC (4 µL/mL) of lavender essential oil reduced volatile sulfur compound secretion at a statistically significant level compared to the control (saline). Furthermore, the level of volatile sulfur compound production attributed to 1 MIC of lavender essential oil was in the range of the positive control chlorhexidine with no significant difference. When examining bacterial membrane damage, 2 MIC of lavender essential oil (i.e., 8 µL/mL) demonstrated the same, showing antibacterial membrane damage values comparative to chlorhexidine. Since lavender essential oil was found to be highly effective in hindering volatile sulfur compound production by Fusobacterium nucleatum through the induction of bacterial cell membrane damage, the results suggest that lavender essential oil may be a suitable alternative to conventional chemical-based anti-malodor agents.


Assuntos
Fusobacterium nucleatum , Halitose , Lavandula , Testes de Sensibilidade Microbiana , Óleos Voláteis , Óleos Voláteis/farmacologia , Óleos Voláteis/química , Fusobacterium nucleatum/efeitos dos fármacos , Fusobacterium nucleatum/metabolismo , Halitose/microbiologia , Halitose/tratamento farmacológico , Halitose/metabolismo , Lavandula/química , Sulfetos/farmacologia , Sulfetos/química , Humanos , Óleos de Plantas/farmacologia , Óleos de Plantas/química , Cromatografia Gasosa-Espectrometria de Massas , Compostos Orgânicos Voláteis/farmacologia , Compostos Orgânicos Voláteis/química , Antibacterianos/farmacologia , Antibacterianos/química
2.
Carbohydr Res ; 541: 109172, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38823062

RESUMO

Sialic acid metabolism in oral bacteria is a complex process involving nutrient acquisition, immune evasion, cell surface modification, and the production of metabolites that contribute to bacterial persistence and virulence in the oral cavity. In addition to causing various periodontal diseases, certain oral pathogenic bacteria, such as Porphyromonas gingivalis, Tannerella forsythia, and Fusobacterium nucleatum, can induce inflammatory reactions and influence the immunity of host cells. These associations with host cells are linked to various diseases, particularly colorectal cancer and Alzheimer's disease. Sialic acid can be found in the host oral mucosa, saliva, or food residues in the oral cavity, and it may promote the colonization of oral bacteria and contribute to disease development. This review aims to summarize the role of sialic acid metabolism in oral bacteria and discuss its effect on the pathogenesis of colorectal cancer and Alzheimer's disease.


Assuntos
Doença de Alzheimer , Neoplasias Colorretais , Ácido N-Acetilneuramínico , Humanos , Doença de Alzheimer/metabolismo , Doença de Alzheimer/microbiologia , Ácido N-Acetilneuramínico/metabolismo , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/microbiologia , Boca/microbiologia , Bactérias/metabolismo , Bactérias/patogenicidade , Fusobacterium nucleatum/metabolismo , Fusobacterium nucleatum/patogenicidade , Animais
3.
Front Cell Infect Microbiol ; 14: 1413787, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38836053

RESUMO

Background: Trimethylamine-N-oxide (TMAO) is produced by hepatic flavin-containing monooxygenase 3 (FMO3) from trimethylamine (TMA). High TMAO level is a biomarker of cardiovascular diseases and metabolic disorders, and it also affects periodontitis through interactions with the gastrointestinal microbiome. While recent findings indicate that periodontitis may alter systemic TMAO levels, the specific mechanisms linking these changes and particular oral pathogens require further clarification. Methods: In this study, we established a C57BL/6J male mouse model by orally administering Porphyromonas gingivalis (P. gingivalis, Pg), Fusobacterium nucleatum (F. nucleatum, Fn), Streptococcus mutans (S. mutans, Sm) and PBS was used as a control. We conducted LC-MS/MS analysis to quantify the concentrations of TMAO and its precursors in the plasma and cecal contents of mice. The diversity and composition of the gut microbiome were analyzed using 16S rRNA sequencing. TMAO-related lipid metabolism and enzymes in the intestines and liver were assessed by qPCR and ELISA methods. We further explored the effect of Pg on FMO3 expression and lipid molecules in HepG2 cells by stimulating the cells with Pg-LPS in vitro. Results: The three oral pathogenic bacteria were orally administered to the mice for 5 weeks. The Pg group showed a marked increase in plasma TMAO, betaine, and creatinine levels, whereas no significant differences were observed in the gut TMAO level among the four groups. Further analysis showed similar diversity and composition in the gut microbiomes of both the Pg and Fn groups, which were different from the Sm and control groups. The profiles of TMA-TMAO pathway-related genera and gut enzymes were not significantly different among all groups. The Pg group showed significantly higher liver FMO3 levels and elevated lipid factors (IL-6, TG, TC, and NEFA) in contrast to the other groups. In vitro experiments confirmed that stimulation of HepG2 cells with Pg-LPS upregulated the expression of FMO3 and increased the lipid factors TC, TG, and IL-6. Conclusion: This study conclusively demonstrates that Pg, compared to Fn and Sm, plays a critical role in elevating plasma TMAO levels and significantly influences the TMA-TMAO pathway, primarily by modulating the expression of hepatic FMO3 and directly impacting hepatic lipid metabolism.


Assuntos
Microbioma Gastrointestinal , Metilaminas , Camundongos Endogâmicos C57BL , Oxigenases , Porphyromonas gingivalis , Animais , Masculino , Metilaminas/metabolismo , Metilaminas/sangue , Humanos , Camundongos , Oxigenases/metabolismo , Porphyromonas gingivalis/metabolismo , Fusobacterium nucleatum/metabolismo , Redes e Vias Metabólicas , Células Hep G2 , Metabolismo dos Lipídeos , Modelos Animais de Doenças , Periodontite/microbiologia , Periodontite/metabolismo , Fígado/metabolismo , RNA Ribossômico 16S/genética , Espectrometria de Massas em Tandem , Boca/microbiologia
4.
Int J Mol Sci ; 25(8)2024 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-38673822

RESUMO

Complex microbial communities have been reported to be involved in endodontic infections. The microorganisms invade the dental pulp leading to pulpitis and initiating pulp inflammation. Fusobacterium nucleatum is a dominant bacterium implicated in both primary and secondary endodontic infections. Drugs targeting the molecular machinery of F. nucleatum will minimize pulp infection. LpxA and LpxD are early acyltransferases involved in the formation of lipid A, a major component of bacterial membranes. The identification of leads which exhibit preference towards successive enzymes in a single pathway can also prevent the development of bacterial resistance. A stringent screening strategy utilizing physicochemical and pharmacokinetic parameters along with a virtual screening approach identified two compounds, Lomefloxacin and Enoxacin, with good binding affinity towards the early acyltransferases LpxA and LpxD. Lomefloxacin and Enoxacin, members of the fluoroquinolone antibiotic class, exhibit wide-ranging activity against diverse bacterial strains. Nevertheless, their effectiveness in the context of endodontic treatment requires further investigation. This study explored the potential of Lomefloxacin and Enoxacin to manage endodontic infections via computational analysis. Moreover, the compounds identified herein serve as a foundation for devising novel combinatorial libraries with enhanced efficacy for endodontic therapeutic strategies.


Assuntos
Antibacterianos , Fusobacterium nucleatum , Lipopolissacarídeos , Fusobacterium nucleatum/efeitos dos fármacos , Fusobacterium nucleatum/metabolismo , Humanos , Antibacterianos/farmacologia , Antibacterianos/química , Lipopolissacarídeos/metabolismo , Simulação de Acoplamento Molecular , Simulação por Computador , Infecções por Fusobacterium/tratamento farmacológico , Infecções por Fusobacterium/microbiologia , Enoxacino/farmacologia , Proteínas de Bactérias/metabolismo , Pulpite/tratamento farmacológico , Pulpite/metabolismo , Pulpite/microbiologia
5.
Zhongguo Zhong Yao Za Zhi ; 49(5): 1266-1274, 2024 Mar.
Artigo em Chinês | MEDLINE | ID: mdl-38621974

RESUMO

This paper investigates the intervention effect and mechanism of Banxia Xiexin Decoction(BXD) on colitis-associated colorectal cancer(CAC) infected with Fusobacterium nucleatum(Fn). C57BL/6 mice were randomly divided into a control group, Fn group, CAC group [azoxymethane(AOM)/dextran sulfate sodium salt(DSS)](AOM/DSS), model group, and BXD group. Except for the control and AOM/DSS groups, the mice in the other groups were orally administered with Fn suspension twice a week. The AOM/DSS group, model group, and BXD group were also injected with a single dose of 10 mg·kg~(-1) AOM combined with three cycles of 2.5% DSS taken intragastrically. The BXD group received oral administration of BXD starting from the second cycle until the end of the experiment. The general condition and weight changes of the mice were monitored during the experiment, and the disease activity index(DAI) was calculated. At the end of the experiment, the colon length and weight of the mice in each group were compared. Hematoxylin-eosin(HE) staining was used to observe the pathological changes in the colon tissue. Enzyme-linked immunosorbent assay(ELISA) was used to detect the levels of interleukin(IL)-2, IL-4, and IL-6 inflammatory factors in the serum. Immunohistochemistry(IHC) was used to detect the expression of Ki67, E-cadherin, and ß-catenin in the colon tissue. Western blot was used to detect the protein content of Wnt3a, ß-catenin, E-cadherin, annexin A1, cyclin D1, and glycogen synthase kinase-3ß(GSK-3ß) in the colon tissue. The results showed that compared with the control group, the Fn group had no significant lesions. The mice in the AOM/DSS group and model group had decreased body weight, increased DAI scores, significantly increased colon weight, and significantly shortened colon length, with more significant lesions in the model group. At the same time, the colon histology of the model group showed more severe adenomas, inflammatory infiltration, and cellular dysplasia. The levels of IL-4 and IL-6 in the serum were significantly increased, while the IL-2 content was significantly decreased. The IHC results showed low expression of E-cadherin and high expression of Ki67 and ß-catenin in the model group, with a decreased protein content of E-cadherin and GSK-3ß and an increased protein content of Wnt3a, ß-catenin, annexin A1, and cyclin D1. After intervention with BXD, the body weight of the mice increased; the DAI score decreased; the colon length increased, and the tumor decreased. The histopathology showed reduced tumor proliferation and reduced inflammatory infiltration. The levels of IL-6 and IL-4 in the serum were significantly decreased, while the IL-2 content was increased. Meanwhile, the expression of E-cadherin was upregulated, and that of Ki67 and ß-catenin was downregulated. The protein content of E-cadherin and GSK-3ß increased, while that of Wnt3a, ß-catenin, annexin A1, and cyclin D1 decreased. In conclusion, BXD can inhibit CAC infected with Fn, and its potential mechanism may be related to the inhibition of Fn binding to E-cadherin, the decrease in annexin A1 protein level, and the regulation of the Wnt/ß-catenin pathway.


Assuntos
Anexina A1 , Neoplasias Associadas a Colite , Colite , Medicamentos de Ervas Chinesas , Camundongos , Animais , Colite/complicações , Colite/tratamento farmacológico , Colite/genética , beta Catenina/genética , beta Catenina/metabolismo , Ciclina D1/metabolismo , Fusobacterium nucleatum/metabolismo , Glicogênio Sintase Quinase 3 beta/metabolismo , Interleucina-6/genética , Interleucina-6/metabolismo , Antígeno Ki-67/metabolismo , Interleucina-2/metabolismo , Interleucina-4/metabolismo , Camundongos Endogâmicos C57BL , Caderinas/metabolismo , Peso Corporal , Sulfato de Dextrana/efeitos adversos , Modelos Animais de Doenças , Azoximetano
6.
mBio ; 15(4): e0353623, 2024 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-38436569

RESUMO

The oral commensal Fusobacterium nucleatum can spread to extra-oral sites, where it is associated with diverse pathologies, including pre-term birth and cancer. Due to the evolutionary distance of F. nucleatum to other model bacteria, we lack a deeper understanding of the RNA regulatory networks that allow this bacterium to adapt to its various niches. As a first step in that direction, we recently showed that F. nucleatum harbors a global stress response governed by the extracytoplasmic function sigma factor, σE, which displays a striking functional conservation with Proteobacteria and includes a noncoding arm in the form of a regulatory small RNA (sRNA), FoxI. To search for putative additional σE-dependent sRNAs, we comprehensively mapped the 5' and 3' ends of transcripts in the model strain ATCC 23726. This enabled the discovery of FoxJ, a ~156-nucleotide sRNA previously misannotated as the 5' untranslated region (UTR) of ylmH. FoxJ is tightly controlled by σE and activated by the same stress conditions as is FoxI. Both sRNAs act as mRNA repressors of the abundant porin FomA, but FoxJ also regulates genes that are distinct from the target suite of FoxI. Moreover, FoxJ differs from other σE-dependent sRNAs in that it also positively regulates genes at the post-transcriptional level. We provide preliminary evidence for a new mode of sRNA-mediated mRNA activation, which involves the targeting of intra-operonic terminators. Overall, our study provides an important resource through the comprehensive annotation of 5' and 3' UTRs in F. nucleatum and expands our understanding of the σE response in this evolutionarily distant bacterium.IMPORTANCEThe oral microbe Fusobacterium nucleatum can colonize secondary sites, including cancer tissue, and likely deploys complex regulatory systems to adapt to these new environments. These systems are largely unknown, partly due to the phylogenetic distance of F. nucleatum to other model organisms. Previously, we identified a global stress response mediated by σE that displays functional conservation with the envelope stress response in Proteobacteria, comprising a coding and noncoding regulatory arm. Through global identification of transcriptional start and stop sites, we uncovered the small RNA (sRNA) FoxJ as a novel component of the noncoding arm of the σE response in F. nucleatum. Together with its companion sRNA FoxI, FoxJ post-transcriptionally modulates the synthesis of envelope proteins, revealing a conserved function for σE-dependent sRNAs between Fusobacteriota and Proteobacteria. Moreover, FoxJ activates the gene expression for several targets, which is a mode of regulation previously unseen in the noncoding arm of the σE response.


Assuntos
Neoplasias , Pequeno RNA não Traduzido , Humanos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fusobacterium nucleatum/genética , Fusobacterium nucleatum/metabolismo , Transcriptoma , Filogenia , Pequeno RNA não Traduzido/genética , Pequeno RNA não Traduzido/metabolismo , Bactérias/genética , RNA Bacteriano/genética , RNA Bacteriano/metabolismo , Regulação Bacteriana da Expressão Gênica
7.
J Ethnopharmacol ; 328: 117932, 2024 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-38382652

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Colitis is an important risk factor for the occurrence of colorectal cancer (CRC), and the colonization of Fusobacterium nucleatum (Fn) in the intestines accelerates this transformation process. Banxia Xiexin Decoction (BXD), originating from Shanghanlun, is a classic prescription for treating gastrointestinal diseases. Current researches indicate that BXD can effectively delay the colitis-to-cancer transition, but it is still unclear whether it can inhibit Fn colonization to achieve this delaying effect. AIM OF STUDY: This study explored the effect and mechanism of BXD in inhibiting Fn intestinal colonization to delay colitis-to-cancer transition. MATERIALS AND METHODS: We constructed a mouse model of colitis-to-cancer transition by regularly gavaging Fn combined with azoxymethane (AOM)/dextran sodium sulfate (DSS), and administered BXD by gavage. We monitored the body weight of mice, measured the length and weight of their colons, and calculated the disease activity index (DAI) score. The growth status of colon tumors was observed by hematoxylin and eosin (H&E) staining, and the changes in gut microbiota in each group of mice were detected by 16S rDNA analysis. Immunohistochemistry was used to detect the expression of E-cadherin and ß-catenin in colon tissues, and immunofluorescence was used to observe the infiltration of M2 macrophages in colon tissues. In cell experiments, we established a co-culture model of Fn and colon cancer cells and intervened with BXD-containing serum. Malignant behaviors such as cell proliferation, invasion, and migration were detected, as well as changes in their cell cycle. We examined the protein levels of E-cadherin, ß-catenin, Axin2, and Cyclin D1 in each group were detected by Western blot. We used US1 strain (fadA-) as a control and observed the effects of BXD-containing serum on Fn attachment and invasion of colon cancer cells through attachment and invasion experiments. RESULTS: BXD can inhibit the colitis-to-cancer transition in mice infected with Fn, reduce crypt structure damage, improve gut microbiota dysbiosis, upregulate E-cadherin and decrease ß-catenin expression, and reduce infiltration of M2 macrophages, thus inhibiting the process of colitis-to-cancer transition. Cell experiments revealed that BXD-containing serum can inhibit the proliferation, migration, and invasion of colon cancer cells infected with Fn and regulate their cell cycle. More importantly, we found that BXD-containing serum can inhibit the binding of Fn's FadA adhesin to E-cadherin, reduce Fn's attachment and invasion of colon cancer cells, thereby downregulating the E-cadherin/ß-catenin signaling pathway. CONCLUSIONS: These findings show that BXD can inhibit Fn colonization by interfering with the binding of FadA to E-cadherin, reducing the activation of the E-cadherin/ß-catenin signaling pathway, and ultimately delaying colitis-to-cancer transition.


Assuntos
Colite , Neoplasias do Colo , Medicamentos de Ervas Chinesas , Animais , Camundongos , beta Catenina/metabolismo , Fusobacterium nucleatum/metabolismo , Transdução de Sinais , Colite/induzido quimicamente , Colite/tratamento farmacológico , Colite/metabolismo , Caderinas/genética , Caderinas/metabolismo , Neoplasias do Colo/induzido quimicamente , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/metabolismo , Sulfato de Dextrana/toxicidade , Modelos Animais de Doenças , Camundongos Endogâmicos C57BL , Colo
8.
J Nanobiotechnology ; 22(1): 62, 2024 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-38360615

RESUMO

BACKGROUND: A large number of Fusobacterium nucleatum (Fn) are present in colorectal cancer (CRC) tissues of patients who relapse after chemotherapy, and Fn has been reported to promote oxaliplatin and 5-FU chemoresistance in CRC. Pathogens such as bacteria and parasites stimulate exosome production in tumor cells, and the regulatory mechanism of exosomal circRNA in the transmission of oxaliplatin and 5-FU chemotherapy resistance in Fn-infected CRC remains unclear. METHODS: Hsa_circ_0004085 was screened by second-generation sequencing of CRC tissues. The correlation between hsa_circ_0004085 and patient clinical response to oxaliplatin/5-FU was analyzed. Exosome tracing experiments and live imaging systems were used to test the effect of Fn infection in CRC on the distribution of hsa_circ_0004085. Colony formation, ER tracking analysis and immunofluorescence were carried out to verify the regulatory effect of exosomes produced by Fn-infected CRC cells on chemotherapeutic resistance and ER stress. RNA pulldown, LC-MS/MS analysis and RIP were used to explore the regulatory mechanism of downstream target genes by hsa_circ_0004085. RESULTS: First, we screened out hsa_circ_0004085 with abnormally high expression in CRC clinical samples infected with Fn and found that patients with high expression of hsa_circ_0004085 in plasma had a poor clinical response to oxaliplatin/5-FU. Subsequently, the circular structure of hsa_circ_0004085 was identified. Fn infection promoted hsa_circ_0004085 formation by hnRNP L and packaged hsa_circ_0004085 into exosomes by hnRNP A1. Exosomes produced by Fn-infected CRC cells transferred hsa_circ_0004085 between cells and delivered oxaliplatin/5-FU resistance to recipient cells by relieving ER stress. Hsa_circ_0004085 enhanced the stability of GRP78 mRNA by binding to RRBP1 and promoted the nuclear translocation of ATF6p50 to relieve ER stress. CONCLUSIONS: Plasma levels of hsa_circ_0004085 are increased in colon cancer patients with intracellular Fn and are associated with a poor response to oxaliplatin/5-FU. Fn infection promoted hsa_circ_0004085 formation by hnRNP L and packaged hsa_circ_0004085 into exosomes by hnRNP A1. Exosomes secreted by Fn-infected CRC cells deliver hsa_circ_0004085 between cells. Hsa_circ_0004085 relieves ER stress in recipient cells by regulating GRP78 and ATF6p50, thereby delivering resistance to oxaliplatin and 5-FU.


Assuntos
Neoplasias do Colo , Neoplasias Colorretais , Exossomos , Ribonucleoproteínas Nucleares Heterogêneas Grupo L , MicroRNAs , Humanos , Oxaliplatina/farmacologia , Oxaliplatina/uso terapêutico , Oxaliplatina/metabolismo , Fusobacterium nucleatum/genética , Fusobacterium nucleatum/metabolismo , Ribonucleoproteína Nuclear Heterogênea A1/metabolismo , Neoplasias Colorretais/metabolismo , Exossomos/metabolismo , Cromatografia Líquida , Chaperona BiP do Retículo Endoplasmático , Ribonucleoproteínas Nucleares Heterogêneas Grupo L/metabolismo , Espectrometria de Massas em Tandem , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/metabolismo , Fluoruracila/farmacologia , Fluoruracila/uso terapêutico , MicroRNAs/metabolismo , Proliferação de Células
9.
Mol Oral Microbiol ; 39(1): 1-11, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38171827

RESUMO

Breast cancer is among the most prevalent malignancies in women worldwide. Epidemiological findings suggested that periodontal diseases may be associated with breast cancer, among which Fusobacterium nucleatum is considered an important cross-participant. In this work, we comprehensively summarize the known mechanisms of how F. nucleatum translocates to, colonizes in mammary tumors, and promotes the carcinogenesis. Specifically, F. nucleatum translocates to mammary tissue through the mammary-intestinal axis, direct nipple contact, and hematogenous transmission. Subsequently, F. nucleatum takes advantage of fusobacterium autotransporter protein 2 to colonize breast cancer and uses virulence factors fusobacterium adhesin A and lipopolysaccharide to promote proliferation. Moreover, the upregulated matrix metalloproteinase-9 induced by F. nucleatum does not only trigger the inflammatory response but also facilitates the tumor-promoting microenvironment. Aside from the pro-inflammatory effect, F. nucleatum may also be engaged in tumor immune evasion, which is achieved through the action of virulence factors on immune checkpoint receptors highly expressed on T cells, natural killer cells, and tumor-infiltrating lymphocytes. Taking breast cancer as an example, more relevant research studies may expand our current knowledge of how oral microbes affect systemic health. Hopefully, exploring these mechanisms in depth could provide new strategies for safer and more effective biologic and targeted therapies targeted at breast cancer.


Assuntos
Neoplasias da Mama , Neoplasias Colorretais , Infecções por Fusobacterium , Humanos , Feminino , Fusobacterium nucleatum/metabolismo , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Adesinas Bacterianas/metabolismo , Fatores de Virulência/metabolismo , Infecções por Fusobacterium/microbiologia , Infecções por Fusobacterium/patologia , Microambiente Tumoral
10.
Nucleic Acids Res ; 52(7): 3950-3970, 2024 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-38281181

RESUMO

The common oral microbe Fusobacterium nucleatum has recently drawn attention after it was found to colonize tumors throughout the human body. Fusobacteria are also interesting study systems for bacterial RNA biology as these early-branching species encode many small noncoding RNAs (sRNAs) but lack homologs of the common RNA-binding proteins (RBPs) CsrA, Hfq and ProQ. To search for alternate sRNA-associated RBPs in F. nucleatum, we performed a systematic mass spectrometry analysis of proteins that co-purified with 19 different sRNAs. This approach revealed strong enrichment of the KH domain proteins KhpA and KhpB with nearly all tested sRNAs, including the σE-dependent sRNA FoxI, a regulator of several envelope proteins. KhpA/B act as a dimer to bind sRNAs with low micromolar affinity and influence the stability of several of their target transcripts. Transcriptome studies combined with biochemical and genetic analyses suggest that KhpA/B have several physiological functions, including being required for ethanolamine utilization. Our RBP search and the discovery of KhpA/B as major RBPs in F. nucleatum are important first steps in identifying key players of post-transcriptional control at the root of the bacterial phylogenetic tree.


Assuntos
Proteínas de Bactérias , Fusobacterium nucleatum , RNA Bacteriano , Pequeno RNA não Traduzido , Proteínas de Ligação a RNA , Fusobacterium nucleatum/genética , Fusobacterium nucleatum/metabolismo , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/genética , Pequeno RNA não Traduzido/metabolismo , Pequeno RNA não Traduzido/genética , Pequeno RNA não Traduzido/química , RNA Bacteriano/metabolismo , RNA Bacteriano/genética , Regulação Bacteriana da Expressão Gênica , Ligação Proteica , Espectrometria de Massas
11.
Arch Oral Biol ; 160: 105897, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38290225

RESUMO

OBJECTIVE: Resveratrol is a natural phytoalexin that has anti-inflammatory properties, reverses doxorubicin resistance, and inhibits epithelial-mesenchymal transition (EMT) in many types of cancer cells. Fusobacterium nucleatum is reportedly enriched in oral squamous cell carcinoma (OSCC) tissues compared to adjacent normal tissues, sparking interest in the relationship between F. nucleatum and OSCC. Recently, F. nucleatum was shown to be associated with EMT in OSCC. In the present study, we aimed to investigate the effects of the natural plant compound resveratrol on F. nucleatum-induced EMT in OSCC. DESIGN: F. nucleatum was co-cultured with OSCC cells, with a multiplicity of infection (MOI) of 300:1. Resveratrol was used at a concentration of 10 µM. Cell Counting Kit-8 and wound healing assays were performed to examine the viability and migratory ability of OSCC cells. Subsequently, real-time RT-PCR was performed to investigate the gene expression of EMT-related markers. Western blotting and immunofluorescence analyses were used to further analyze the expression of the epithelial marker E-cadherin and the EMT transcription factor SNAI1. RESULTS: Co-cultivation with F. nucleatum did not significantly enhance cell viability. The co-cultured cells displayed similarities to the positive control of EMT, exhibiting enhanced migration and expression changes in EMT-related markers. SNAI1 was significantly upregulated, whereas E-cadherin, was significantly downregulated. Notably, resveratrol inhibited F. nucleatum-induced cell migration, decreasing the expression of SNAI1. CONCLUSIONS: Resveratrol inhibited F. nucleatum-induced EMT by downregulating SNAI1, which may provide a target for OSCC treatment.


Assuntos
Carcinoma de Células Escamosas , Neoplasias de Cabeça e Pescoço , Neoplasias Bucais , Humanos , Carcinoma de Células Escamosas/genética , Resveratrol/farmacologia , Fusobacterium nucleatum/metabolismo , Polifenóis/farmacologia , Neoplasias Bucais/genética , Linhagem Celular Tumoral , Caderinas/metabolismo , Transição Epitelial-Mesenquimal , Carcinoma de Células Escamosas de Cabeça e Pescoço , Movimento Celular , Neoplasias de Cabeça e Pescoço/tratamento farmacológico
12.
Mol Oral Microbiol ; 39(2): 29-39, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-37718958

RESUMO

The prevalence of periodontitis increases with physiological aging. However, whether bacteria associated with periodontal diseases foster aging and the mechanisms by which they may do so are unknown. Herein, we hypothesize that Fusobacterium nucleatum, a microorganism associated with periodontitis and several other age-related disorders, triggers senescence, a chief hallmark of aging responsible to reduce tissue repair capacity. Our study analyzed the senescence response of gingival epithelial cells and their reparative capacity upon long-term exposure to F. nucleatum. Specifically, we assessed (a) cell cycle arrest by analyzing the cyclin-dependent kinase inhibitors p16INK4a and p14ARF and their downstream cascade (pRb, p53, and p21) at both gene and protein levels, (b) lysosomal mediated dysfunction by using assays targeting the expression and activity of the senescence-associated ß-galactosidase (SA-ß-Gal) enzyme, and (c) nuclear envelope breakdown by assessing the expression of Lamin-B1. The consequences of the senescence phenotype mediated by F. nucleatum were further assessed using wound healing assays. Our results revealed that prolonged exposure to F. nucleatum promotes an aging-like phenotype as evidenced by the increased expression of pro-senescence markers (p16INK4a , p21, and pRb) and SA-ß-Gal activity and reduced expression of the counter-balancing cascade (p14ARF and p53) and Lamin-B1. Furthermore, we also noted impaired wound healing capacity of gingival epithelial cells upon prolong bacterial exposure, which was consistent with the senescence-induced phenotype. Together, our findings provide a proof-of-concept evidence that F. nucleatum triggers a pro-senescence response in gingival epithelial cells. This might affect periodontal tissue homeostasis by reducing its repair capacity and, consequently, increasing susceptibility to periodontitis during aging.


Assuntos
Fusobacterium nucleatum , Periodontite , Humanos , Fusobacterium nucleatum/metabolismo , Proteína Supressora de Tumor p14ARF/metabolismo , Senescência Celular/fisiologia , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Células Epiteliais/metabolismo , Fenótipo , Laminas/metabolismo
13.
Gut Microbes ; 16(1): 2295384, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38126163

RESUMO

The anaerobic bacterium Fusobacterium nucleatum is significantly associated with human colorectal cancer (CRC) and is considered a significant contributor to the disease. The mechanisms underlying the promotion of intestinal tumor formation by F. nucleatum have only been partially uncovered. Here, we showed that F. nucleatum releases a metabolite into the microenvironment that strongly activates NF-κB in intestinal epithelial cells via the ALPK1/TIFA/TRAF6 pathway. Furthermore, we showed that the released molecule had the biological characteristics of ADP-heptose. We observed that F. nucleatum induction of this pathway increased the expression of the inflammatory cytokine IL-8 and two anti-apoptotic genes known to be implicated in CRC, BIRC3 and TNFAIP3. Finally, it promoted the survival of CRC cells and reduced 5-fluorouracil chemosensitivity in vitro. Taken together, our results emphasize the importance of the ALPK1/TIFA pathway in Fusobacterium induced-CRC pathogenesis, and identify the role of ADP-H in this process.


Assuntos
Neoplasias Colorretais , Microbioma Gastrointestinal , Humanos , Fusobacterium nucleatum/metabolismo , Composição de Bases , Filogenia , RNA Ribossômico 16S , Análise de Sequência de DNA , Neoplasias Colorretais/patologia , Heptoses/metabolismo , Microambiente Tumoral
14.
FASEB J ; 38(1): e23357, 2024 01.
Artigo em Inglês | MEDLINE | ID: mdl-38085169

RESUMO

Bacterial infection is the main cause of pulpitis. However, whether a dominant bacteria can promote the progression of pulpitis and its underlying mechanism remains unclear. We provided a comprehensive assessment of the microbiota alteration in pulpitis using 16S rRNA sequencing. Fusobacterium nucleatum was the most enriched in pulpitis and played a pathogenic role accelerating pulpitis progression in rat pulpitis model. After odontoblast-like cells cocultured with F. nucleatum, the stimulator of interferon genes (STING) pathway and autophagy were activation. There was a float of STING expression during F. nucleatum stimulation. STING was degraded by autophagy at the early stage. At the late stage, F. nucleatum stimulated mitochondrial Reactive Oxygen Species (ROS) production, mitochondrial dysfunction and then mtDNA escape into cytosol. mtDNA, which escaped into cytosol, caused more cytosolic mtDNA binds to cyclic GMP-AMP synthase (cGAS). The release of IFN-ß was dramatically reduced when mtDNA-cGAS-STING pathway inhibited. STING-/- mice showed milder periapical bone loss and lower serum IFN-ß levels compared with wildtype mice after 28 days F. nucleatum-infected pulpitis model establishment. Our data demonstrated that F. nucleatum exacerbated the progression of pulpitis, which was mediated by the STING-dependent pathway.


Assuntos
Fusobacterium nucleatum , Pulpite , Camundongos , Ratos , Animais , Fusobacterium nucleatum/genética , Fusobacterium nucleatum/metabolismo , Transdução de Sinais , RNA Ribossômico 16S , Nucleotidiltransferases/metabolismo , DNA Mitocondrial/genética
15.
J Am Chem Soc ; 145(48): 26296-26307, 2023 12 06.
Artigo em Inglês | MEDLINE | ID: mdl-37987621

RESUMO

Immunotherapy of triple-negative breast cancer (TNBC) has an unsatisfactory therapeutic outcome due to an immunologically "cold" microenvironment. Fusobacterium nucleatum (F. nucleatum) was found to be colonized in triple-negative breast tumors and was responsible for the immunosuppressive tumor microenvironment and tumor metastasis. Herein, we constructed a bacteria-derived outer membrane vesicle (OMV)-coated nanoplatform that precisely targeted tumor tissues for dual killing of F. nucleatum and cancer cells, thus transforming intratumor bacteria into immunopotentiators in immunotherapy of TNBC. The as-prepared nanoparticles efficiently induced immunogenic cell death through a Fenton-like reaction, resulting in enhanced immunogenicity. Meanwhile, intratumoral F. nucleatum was killed by metronidazole, resulting in the release of pathogen-associated molecular patterns (PAMPs). PAMPs cooperated with OMVs further facilitated the maturation of dendritic cells and subsequent T-cell infiltration. As a result, the "kill two birds with one stone" strategy warmed up the cold tumor environment, maximized the antitumor immune response, and achieved efficient therapy of TNBC as well as metastasis prevention. Overall, this strategy based on a microecology distinction in tumor and normal tissue as well as microbiome-induced reversal of cold tumors provides new insight into the precise and efficient immune therapy of TNBC.


Assuntos
Neoplasias de Mama Triplo Negativas , Humanos , Neoplasias de Mama Triplo Negativas/metabolismo , Adjuvantes Imunológicos , Moléculas com Motivos Associados a Patógenos/metabolismo , Moléculas com Motivos Associados a Patógenos/uso terapêutico , Imunoterapia/métodos , Fusobacterium nucleatum/metabolismo , Linhagem Celular Tumoral , Microambiente Tumoral
16.
Biochem Biophys Res Commun ; 688: 149151, 2023 12 25.
Artigo em Inglês | MEDLINE | ID: mdl-37951156

RESUMO

Fusobacterium nucleatum (F. nucleatum) is an anaerobic gram-negative bacterium that was previously thought to be related to the progression of colorectal cancer. In F. nucleatum, thiolase participates in fatty acid metabolism, and it can catalyse the transfer of an acetyl group from acetyl-CoA to another molecule, typically a fatty acid or another molecule in the synthesis of lipids. To gain deeper insight into the molecular mechanism governing the function of thiolase in F. nucleatum (Fn0495), we herein report the structure of Fn0495. The monomer of Fn0495 consists of three subdomains, namely, the N-terminal domain (residues 1-117 and 252-270), the C-terminal domain (residues 273-393), and the loop domain (residues 118-251). Fn0495 shows a unique difference in the charge and structure of the substrate binding pocket compared with homologous proteins. This research found three conserved residues (Cys88, His357, and Cys387) in Fn0495 arranged near a potential substrate binding pocket. In this study, the conformational changes between the covering loop, catalytic cysteine loop, regulatory determinant region, and homologous protein were compared. These results will enhance our understanding of the molecular characteristics and roles of the thiolase family.


Assuntos
Acetil-CoA C-Acetiltransferase , Fusobacterium nucleatum , Fusobacterium nucleatum/metabolismo , Acetil-CoA C-Acetiltransferase/química , Acetilcoenzima A , Cisteína/metabolismo , Ácidos Graxos
17.
Int J Mol Sci ; 24(20)2023 Oct 12.
Artigo em Inglês | MEDLINE | ID: mdl-37894788

RESUMO

Microbial resistance to antibiotics poses a tremendous challenge. Bacteriophages may provide a useful alternative or adjunct to traditional antibiotics. To be used in therapy, bacteriophages need to be purified from endotoxins and tested for their effects on human immune cells. Interleukin-1 Receptor Associated Kinase-3 (IRAK3) is a negative regulator of inflammation and may play a role in the modulation of immune signalling upon bacteriophage exposure to immune cells. This study aimed to investigate the immune effects of crude and purified bacteriophage FNU1, a bacteriophage that targets the oral pathobiont Fusobacterium nucleatum, on wildtype and IRAK3 knockout THP-1 monocytic cell lines. The IRAK3 knockout cell line was also used to develop a novel endotoxin detection assay. Exposure to crude FNU1 increased the production of pro-inflammatory cytokines (Tumour necrosis factor - alpha (TNF-α) and Interleukin 6 (IL-6)) compared to purified FNU1 in wildtype and IRAK3 knockout THP-1 monocytes. In the IRAK3 knockout THP-1 cells, exposure to crude FNU1 induced a higher immune response than the wildtype monocytes, supporting the suggestion that the inhibitory protein IRAK3 regulates reactions to endotoxins and impurities in bacteriophage preparations. Finally, the novel endotoxin detection assay generated here provides a robust and accurate method for determining endotoxin concentrations.


Assuntos
Bacteriófagos , Citocinas , Humanos , Citocinas/metabolismo , Monócitos/metabolismo , Fusobacterium nucleatum/metabolismo , Endotoxinas/metabolismo , Bacteriófagos/genética , Bacteriófagos/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Antibacterianos/metabolismo , Quinases Associadas a Receptores de Interleucina-1/genética , Quinases Associadas a Receptores de Interleucina-1/metabolismo
18.
J Control Release ; 363: 43-56, 2023 11.
Artigo em Inglês | MEDLINE | ID: mdl-37734673

RESUMO

The gut microbiota is closely associated with the progression of colorectal cancer (CRC) in which Fusobacterium nucleatum (F. nucleatum) was found to induce cancer resistance to chemotherapeutics. To relieve F. nucleatum-induced drug resistance, herein, we found that short-chain fatty acid butyrate can inhibit the growth, enrichment and adhesion of F. nucleatum in colorectal cancer tissues by downregulating the expression of adhesion-associated outer membrane proteins, including RadD, FomA, and FadA, to reduce the colonization and invasion of F. nucleatum and relieve the chemoresistance induced by F. nucleatum. Leveraging the killing effect of butyrate on F. nucleatum, sodium butyrate (NaBu) was encapsulated in liposomes or prepared as NaBu tablets with Eudragit S100 coating and administered by intravenous injection or oral administration, respectively. Interestingly, both intravenous administration of NaBu liposomes and oral delivery of NaBu tablets could effectively inhibit the proliferation of F. nucleatum and significantly improve the therapeutic efficacy of oxaliplatin in mice with subcutaneous colorectal tumors, orthotopic colorectal tumors and even spontaneously formed colorectal tumors. Thus, our work provides a simple but effective formulation of NaBu to relieve F. nucleatum-induced chemoresistance, exhibiting ideal clinical application prospects.


Assuntos
Neoplasias Colorretais , Infecções por Fusobacterium , Animais , Camundongos , Fusobacterium nucleatum/metabolismo , Butiratos , Resistencia a Medicamentos Antineoplásicos , Lipossomos/metabolismo , Infecções por Fusobacterium/complicações , Infecções por Fusobacterium/metabolismo , Infecções por Fusobacterium/microbiologia , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/metabolismo
19.
Gut Microbes ; 15(1): 2240035, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37550944

RESUMO

Fusobacterium nucleatum (Fn) infection is known to exacerbate ulcerative colitis (UC). However, the link between Fn-infected intestinal epithelial cell (IEC)-derived exosomes (Fn-Exo) and UC progression has not been investigated. Differentially expressed miRNAs in Fn-Exo and non-infected IECs-derived exosomes (Con-Exo) were identified by miRNA sequencing. Then, the biological role and mechanism of Fn-Exo in UC development were determined in vitro and in vivo. We found that exosomes delivered miR-129-2-3p from Fn-infected IECs into non-infected IECs, exacerbating epithelial barrier dysfunction and experimental colitis. Mechanically, Fn-Exo induces DNA damage via the miR-129-2-3p/TIMELESS axis and subsequently activates the ATM/ATR/p53 pathway, ultimately promoting cellular senescence and colonic inflammation. In conclusion, Exo-miR-129-2-3p/TIMELESS/ATM/ATR/p53 pathway aggravates cellular senescence, barrier damage, and experimental colitis. The current study revealed a previously unknown regulatory pathway in the progression of Fn-infectious UC. Furthermore, Exosomal-miR-129-2-3p in serum and TIMELESS may function as novel potential diagnostic biomarkers for UC and Fn-high-UC.


Assuntos
Colite Ulcerativa , Colite , Microbioma Gastrointestinal , MicroRNAs , Humanos , Fusobacterium nucleatum/genética , Fusobacterium nucleatum/metabolismo , Transdução de Sinais/fisiologia , Proteína Supressora de Tumor p53/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Colite/genética , Colite Ulcerativa/genética , Colite Ulcerativa/microbiologia , Células Epiteliais/metabolismo , Senescência Celular
20.
Eur J Immunol ; 53(11): e2350455, 2023 11.
Artigo em Inglês | MEDLINE | ID: mdl-37471504

RESUMO

Caspase activation results in pyroptosis, an inflammatory cell death that contributes to several inflammatory diseases by releasing inflammatory cytokines and cellular contents. Fusobacterium nucleatum is a periodontal pathogen frequently detected in human cancer and inflammatory bowel diseases. Studies have reported that F. nucleatum infection leads to NLRP3 activation and pyroptosis, but the precise activation process and disease association remain poorly understood. This study demonstrated that F. nucleatum infection exacerbates acute colitis in mice and activates pyroptosis through caspase-11-mediated gasdermin D cleavage in macrophages. Furthermore, F. nucleatum infection in colitis mice induces the enhancement of IL-1⍺ secretion from the colon, affecting weight loss and severe disease activities. Neutralization of IL-1⍺ protects F. nucleatum infected mice from severe colitis. Therefore, F. nucleatum infection facilitates inflammation in acute colitis with IL-1⍺ from colon tissue by activating noncanonical inflammasome through gasdermin D cleavage.


Assuntos
Colite , Inflamassomos , Humanos , Animais , Camundongos , Inflamassomos/metabolismo , Fusobacterium nucleatum/metabolismo , Gasderminas , Colite/induzido quimicamente , Caspases/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo
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