RESUMEN
Inverted organic solar cells (OSCs) have garnered significant interest due to their remarkable stability. In this study, the efficiency and stability of inverted OSCs are enhanced via the in situ self-organization (SO) of an interfacial modification material Phen-NaDPO onto tin oxide (SnO2). During the device fabrication, Phen-NaDPO is spin-coated with the active materials all together on SnO2. Driven by the interactions with SnO2 and the thermodynamic forces due to its high surface energy and the convection flow, Phen-NaDPO spontaneously migrates to the SnO2 interface, resulting in the formation of an in situ modification layer on SnO2. This self-organization of Phen-NaDPO not only effectively reduces the work function of SnO2, but also enhances the ordered molecular stacking and manipulates the vertical morphology of the active layer, which suppress the surface trap-assisted recombination and minimize the charge extraction. As a result, the SO devices based on PM6:Y6 exhibit significantly improved photovoltaic performance with an enhanced power conversion efficiency of 17.62%. Moreover, the stability of the SO device is also improved. Furthermore, the SO ternary devices based on PM6:D18:L8-BO achieved an impressive PCE of 18.87%, standing as one of the highest values for single-junction inverted organic solar cells to date.
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Exposing C-section infants to the maternal vaginal microbiome, coined "vaginal seeding", partially restores microbial colonization. However, whether vaginal seeding decreases metabolic disease risk is unknown. Therefore, we assessed the effect of vaginal seeding of human infants on adiposity in a murine model. Germ-free mice were colonized with transitional stool from human infants who received vaginal seeding or control (placebo) seeding in a double-blind randomized trial. There was a reduction in intraabdominal adipose tissue (IAAT) volume in male mice that received stool from vaginally seeded infants compared to control infants. Higher levels of isoleucine and lower levels of nucleic acid metabolites were observed in controls and correlated with increased IAAT. This suggests that early changes in the gut microbiome and metabolome caused by vaginal seeding have a positive impact on metabolic health.
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Adiposidad , Trasplante de Microbiota Fecal , Heces , Microbioma Gastrointestinal , Vagina , Animales , Humanos , Femenino , Ratones , Masculino , Vagina/microbiología , Heces/microbiología , Heces/química , Método Doble Ciego , Grasa Intraabdominal/metabolismo , Lactante , Recién NacidoRESUMEN
Introduction: Obesity, an independent risk factor for breast cancer growth and metastatic progression, is also closely intertwined with gut dysbiosis; and both obese state and dysbiosis promote each other. Enteric abundance of Bacteroides fragilis is strongly linked with obesity, and we recently discovered the presence of B. fragilis in malignant breast cancer. Given that enterotoxigenic B. fragilis or ETBF, which secretes B. fragilis toxin (BFT), has been identified as a procarcinogenic microbe in breast cancer, it is necessary to examine its impact on distant metastasis and underlying systemic and localized alterations promoting metastatic progression of breast cancer. Methods: We used syngeneic mammary intraductal (MIND) model harboring gut colonization with ETBF to query distant metastasis of breast cancer cells. Alterations in the immune network and cytokines/chemokines in the tumor microenvironment and distant metastatic sites were examined using flow cytometry, immunohistochemistry, and multiplex arrays. Results: ETBF infection initiates a systemic inflammation aiding in the establishment of the premetastatic niche formation in vital organs via increased proinflammatory and protumorigenic cytokines like IL17A, IL17E, IL27p28, IL17A/F, IL6, and IL10 in addition to creating a prometastatic immunosuppressive environment in the liver and lungs rich in myeloid cells, macrophages, and T regulatory cells. It induces remodeling of the tumor microenvironment via immune cell and stroma infiltration, increased vasculogenesis, and an EMT-like response, thereby encouraging early metastatic dissemination ready to colonize the conducive environment in liver and lungs of the breast tumor-bearing mice. Discussion: In this study, we show that enteric ETBF infection concomitantly induces systemic inflammation, reshapes the tumor immune microenvironment, and creates conducive metastatic niches to potentiate early dissemination and seeding of metastases to liver and lung tissues in agreement with the "seed and soil hypothesis." Our results also support the ETBF-induced "parallel model" of metastasis that advocates for an early dissemination of tumor cells that form metastatic lesions independent of the primary tumor load.
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Toxinas Bacterianas , Neoplasias Hepáticas , Neoplasias Pulmonares , Ratones , Animales , Disbiosis , Inflamación , Citocinas , Pulmón , Obesidad , Microambiente TumoralRESUMEN
Defining the complex role of the microbiome in colorectal cancer and the discovery of novel, protumorigenic microbes are areas of active investigation. In the present study, culturing and reassociation experiments revealed that toxigenic strains of Clostridioides difficile drove the tumorigenic phenotype of a subset of colorectal cancer patient-derived mucosal slurries in germ-free ApcMin/+ mice. Tumorigenesis was dependent on the C. difficile toxin TcdB and was associated with induction of Wnt signaling, reactive oxygen species, and protumorigenic mucosal immune responses marked by the infiltration of activated myeloid cells and IL17-producing lymphoid and innate lymphoid cell subsets. These findings suggest that chronic colonization with toxigenic C. difficile is a potential driver of colorectal cancer in patients. SIGNIFICANCE: Colorectal cancer is a leading cause of cancer and cancer-related deaths worldwide, with a multifactorial etiology that likely includes procarcinogenic bacteria. Using human colon cancer specimens, culturing, and murine models, we demonstrate that chronic infection with the enteric pathogen C. difficile is a previously unrecognized contributor to colonic tumorigenesis. See related commentary by Jain and Dudeja, p. 1838. This article is highlighted in the In This Issue feature, p. 1825.
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Toxinas Bacterianas , Clostridioides difficile , Neoplasias del Colon , Neoplasias Colorrectales , Animales , Toxinas Bacterianas/genética , Toxinas Bacterianas/metabolismo , Carcinogénesis , Clostridioides , Humanos , Inmunidad Innata , Linfocitos/metabolismo , RatonesRESUMEN
Delineating the molecular and morphological changes that cancer cells undergo in response to extracellular stimuli is crucial for identifying factors that promote tumor progression. Label-free optical imaging offers a potentially promising route for retrieving such single-cell information by generating detailed visualization of the morphology and determining alterations in biomolecular composition. The potential of such nonperturbative morphomolecular microscopy for analyzing microbiota-cancer cell interactions has been surprisingly underappreciated, despite the growing evidence of the critical role of dysbiosis in malignant transformations. Here, using a model system of breast cancer cells, we show that label-free Raman microspectroscopy and quantitative phase microscopy can detect biomolecular and morphological changes in single cells exposed to Bacteroides fragilis toxin (BFT), a toxin secreted by enterotoxigenicB. fragilis. Remarkably, using machine learning to elucidate subtle, but consistent, cellular differences, we found that the morphomolecular differences between BFT-exposed and control breast cancer cells became more accentuated after in vivo passage, corroborating our findings that a short-term BFT exposure imparts a long-term effect on cancer cells and promotes a more invasive phenotype. Complementing more classical labeling techniques, our label-free platform offers a global detection approach with measurements representative of the overall cellular phenotype, paving the way for further investigations into the multifaceted interactions between the cancer cell and the microbiota.
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Bacteroides fragilis , Neoplasias de la Mama , Bacteroides fragilis/genética , Recuento de Células , Femenino , Humanos , Microscopía , VibraciónRESUMEN
Enterotoxigenic Bacteroides fragilis (ETBF) is consistently found at higher frequency in individuals with sporadic and hereditary colorectal cancer (CRC) and induces tumorigenesis in several mouse models of CRC. However, whether specific mutations induced by ETBF lead to colon tumor formation has not been investigated. To determine if ETBF-induced mutations impact the Apc gene, and other tumor suppressors or proto-oncogenes, we performed whole-exome sequencing and whole-genome sequencing on tumors isolated after ETBF and sham colonization of Apcmin/+ and Apcmin/+Msh2fl/flVC mice, as well as whole-genome sequencing of organoids cocultured with ETBF. Our results indicate that ETBF-induced tumor formation results from loss of heterozygosity (LOH) of Apc, unless the mismatch repair system is disrupted, in which case, tumor formation results from new acquisition of protein-truncating mutations in Apc. In contrast to polyketide synthase-positive Escherichia coli (pks+ E. coli), ETBF does not produce a unique mutational signature; instead, ETBF-induced tumors arise from errors in DNA mismatch repair and homologous recombination DNA damage repair, established pathways of tumor formation in the colon, and the same genetic mechanism accounting for sham tumors in these mouse models. Our analysis informs how this procarcinogenic bacterium may promote tumor formation in individuals with inherited predispositions to CRC, such as Lynch syndrome or familial adenomatous polyposis (FAP). IMPORTANCE Many studies have shown that microbiome composition in both the mucosa and the stool differs in individuals with sporadic and hereditary colorectal cancer (CRC). Both human and mouse models have established a strong association between particular microbes and colon tumor induction. However, the genetic mechanisms underlying putative microbe-induced colon tumor formation are not well established. In this paper, we applied whole-exome sequencing and whole-genome sequencing to investigate the impact of ETBF-induced genetic changes on tumor formation. Additionally, we performed whole-genome sequencing of human colon organoids exposed to ETBF to validate the mutational patterns seen in our mouse models and begin to understand their relevance in human colon epithelial cells. The results of this study highlight the importance of ETBF colonization in the development of sporadic CRC and in individuals with hereditary tumor conditions, such as Lynch syndrome and familial adenomatous polyposis (FAP).
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Poliposis Adenomatosa del Colon , Infecciones Bacterianas , Neoplasias del Colon , Neoplasias Colorrectales Hereditarias sin Poliposis , Neoplasias Colorrectales , Poliposis Adenomatosa del Colon/genética , Poliposis Adenomatosa del Colon/patología , Animales , Infecciones Bacterianas/patología , Bacteroides fragilis/genética , Bacteroides fragilis/metabolismo , Colon/microbiología , Neoplasias del Colon/genética , Neoplasias del Colon/microbiología , Neoplasias del Colon/patología , Neoplasias Colorrectales/microbiología , Neoplasias Colorrectales Hereditarias sin Poliposis/genética , Neoplasias Colorrectales Hereditarias sin Poliposis/patología , Modelos Animales de Enfermedad , Escherichia coli/genética , Genes APC , Ratones , MutaciónRESUMEN
Objective: The opportunistic pathogen Streptococcus gallolyticus is one of the few intestinal bacteria that has been consistently linked to colorectal cancer (CRC). This study aimed to identify novel S. gallolyticus-induced pathways in colon epithelial cells that could further explain how S. gallolyticus contributes to CRC development. Design and Results: Transcription profiling of in vitro cultured CRC cells that were exposed to S. gallolyticus revealed the specific induction of oxidoreductase pathways. Most prominently, CYP1A and ALDH1 genes that encode phase I biotransformation enzymes were responsible for the detoxification or bio-activation of toxic compounds. A common feature is that these enzymes are induced through the Aryl hydrocarbon receptor (AhR). Using the specific inhibitor CH223191, we showed that the induction of CYP1A was dependent on the AhR both in vitro using multiple CRC cell lines as in vivo using wild-type C57bl6 mice colonized with S. gallolyticus. Furthermore, we showed that CYP1 could also be induced by other intestinal bacteria and that a yet unidentified diffusible factor from the S. galloltyicus secretome (SGS) induces CYP1A enzyme activity in an AhR-dependent manner. Importantly, priming CRC cells with SGS increased the DNA damaging effect of the polycyclic aromatic hydrocarbon 3-methylcholanthrene. Conclusion: This study shows that gut bacteria have the potential to modulate the expression of biotransformation pathways in colonic epithelial cells in an AhR-dependent manner. This offers a novel theory on the contribution of intestinal bacteria to the etiology of CRC by modifying the capacity of intestinal epithelial or (pre-)cancerous cells to (de)toxify dietary components, which could alter intestinal susceptibility to DNA damaging events.
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Neoplasias Colorrectales , Streptococcus gallolyticus , Animales , Biotransformación , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP1A1/metabolismo , Células Epiteliales/metabolismo , Ratones , Ratones Endogámicos C57BL , Receptores de Hidrocarburo de Aril/genética , Receptores de Hidrocarburo de Aril/metabolismo , Streptococcus gallolyticus/metabolismoRESUMEN
G protein-coupled receptor (GPR)35 is highly expressed in the gastro-intestinal tract, predominantly in colon epithelial cells (CEC), and has been associated with inflammatory bowel diseases (IBD), suggesting a role in gastrointestinal inflammation. The enterotoxigenic Bacteroides fragilis (ETBF) toxin (BFT) is an important virulence factor causing gut inflammation in humans and animal models. We identified that BFT signals through GPR35. Blocking GPR35 function in CECs using the GPR35 antagonist ML145, in conjunction with shRNA knock-down and CRISPRcas-mediated knock-out, resulted in reduced CEC-response to BFT as measured by E-cadherin cleavage, beta-arrestin recruitment and IL-8 secretion. Importantly, GPR35 is required for the rapid onset of ETBF-induced colitis in mouse models. GPR35-deficient mice showed reduced death and disease severity compared to wild-type C57Bl6 mice. Our data support a role for GPR35 in the CEC and mucosal response to BFT and underscore the importance of this molecule for sensing ETBF in the colon.
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Toxinas Bacterianas/administración & dosificación , Bacteroides fragilis/patogenicidad , Colitis/patología , Colon/patología , Células Epiteliales/patología , Tracto Gastrointestinal/patología , Metaloendopeptidasas/administración & dosificación , Receptores Acoplados a Proteínas G/fisiología , Animales , Bacteroides fragilis/genética , Bacteroides fragilis/metabolismo , Colitis/etiología , Colitis/metabolismo , Colon/efectos de los fármacos , Colon/metabolismo , Colon/microbiología , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Células Epiteliales/microbiología , Tracto Gastrointestinal/efectos de los fármacos , Tracto Gastrointestinal/metabolismo , Tracto Gastrointestinal/microbiología , Ratones , Ratones Endogámicos C57BLRESUMEN
The existence of distinct breast microbiota has been recently established, but their biological impact in breast cancer remains elusive. Focusing on the shift in microbial community composition in diseased breast compared with normal breast, we identified the presence of Bacteroides fragilis in cancerous breast. Mammary gland as well as gut colonization with enterotoxigenic Bacteroides fragilis (ETBF), which secretes B. fragilis toxin (BFT), rapidly induces epithelial hyperplasia in the mammary gland. Breast cancer cells exposed to BFT exhibit "BFT memory" from the initial exposure. Intriguingly, gut or breast duct colonization with ETBF strongly induces growth and metastatic progression of tumor cells implanted in mammary ducts, in contrast to nontoxigenic Bacteroides fragilis. This work sheds light on the oncogenic impact of a procarcinogenic colon bacterium ETBF on breast cancer progression, implicates the ß-catenin and Notch1 axis as its functional mediators, and proposes the concept of "BFT memory" that can have far-reaching biological implications after initial exposure to ETBF. SIGNIFICANCE: B. fragilis is an inhabitant of breast tissue, and gut or mammary duct colonization with ETBF triggers epithelial hyperplasia and augments breast cancer growth and metastasis. Short-term exposure to BFT elicits a "BFT memory" with long-term implications, functionally mediated by the ß-catenin and Notch1 pathways.This article is highlighted in the In This Issue feature, p. 995.
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Bacteroides fragilis , Neoplasias de la Mama/patología , Colon/microbiología , Animales , Carcinogénesis/patología , Transformación Celular Neoplásica/patología , Modelos Animales de Enfermedad , Femenino , Humanos , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos NOD , Ratones SCID , Metástasis de la Neoplasia , beta Catenina/metabolismoRESUMEN
Fusobacteria are commonly associated with human colorectal cancer (CRC), but investigations are hampered by the absence of a stably colonized murine model. Further, Fusobacterium nucleatum subspecies isolated from human CRC have not been investigated. While F. nucleatum subspecies are commonly associated with CRC, their ability to induce tumorigenesis and contributions to human CRC pathogenesis are uncertain. We sought to establish a stably colonized murine model and to understand the inflammatory potential and virulence genes of human CRC F. nucleatum, representing the 4 subspecies, animalis, nucleatum, polymorphum, and vincentii. Five human CRC-derived and two non-CRC derived F. nucleatum strains were tested for colonization, tumorigenesis, and cytokine induction in specific-pathogen-free (SPF) and/or germfree (GF) wild-type and ApcMin/+ mice, as well as in vitro assays and whole-genome sequencing (WGS). SPF wild-type and ApcMin/+ mice did not achieve stable colonization with F. nucleatum, whereas certain subspecies stably colonized some GF mice but without inducing colon tumorigenesis. F. nucleatum subspecies did not form in vivo biofilms or associate with the mucosa in mice. In vivo inflammation was inconsistent across subspecies, whereas F. nucleatum induced greater cytokine responses in a human colorectal cell line, HCT116. While F. nucleatum subspecies displayed genomic variability, no distinct virulence genes associated with human CRC strains were identified that could reliably distinguish these strains from non-CRC clinical isolates. We hypothesize that the lack of F. nucleatum-induced tumorigenesis in our model reflects differences in human and murine biology and/or a synergistic role for F. nucleatum in concert with other bacteria to promote carcinogenesis. IMPORTANCE Colon cancer is a leading cause of cancer morbidity and mortality, and it is hypothesized that dysbiosis in the gut microbiota contributes to colon tumorigenesis. Fusobacterium nucleatum, a member of the oropharyngeal microbiome, is enriched in a subset of human colon tumors. However, it is unclear whether this genetically varied species directly promotes tumor formation, modulates mucosal immune responses, or merely colonizes the tumor microenvironment. Mechanistic studies to address these questions have been stymied by the lack of an animal model that does not rely on daily orogastric gavage. Using multiple murine models, in vitro assays with a human colon cancer cell line, and whole-genome sequencing analysis, we investigated the proinflammatory and tumorigenic potential of several F. nucleatum clinical isolates. The significance of this research is development of a stable colonization model of F. nucleatum that does not require daily oral gavages in which we demonstrate that a diverse library of clinical isolates do not promote tumorigenesis.
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Neoplasias del Colon , Neoplasias Colorrectales , Animales , Humanos , Ratones , Carcinogénesis , Citocinas , Modelos Animales de Enfermedad , Fusobacterium nucleatum/genética , Inflamación/complicaciones , Microambiente TumoralRESUMEN
Enterotoxigenic Bacteroides fragilis (ETBF) is a commensal bacterium of great importance to human health due to its ability to induce colitis and cause colon tumor formation in mice through the production of B. fragilis toxin (BFT). The formation of tumors is dependent on a pro-inflammatory signaling cascade, which begins with the disruption of epithelial barrier integrity through cleavage of E-cadherin. Here, we show that BFT increases levels of glucosylceramide, a vital intestinal sphingolipid, both in mice and in colon organoids (colonoids) generated from the distal colons of mice. When colonoids are treated with BFT in the presence of an inhibitor of glucosylceramide synthase (GCS), the enzyme responsible for generating glucosylceramide, colonoids become highly permeable, lose structural integrity, and eventually burst, releasing their contents into the extracellular matrix. By increasing glucosylceramide levels in colonoids via an inhibitor of glucocerebrosidase (GBA, the enzyme that degrades glucosylceramide), colonoid permeability was reduced, and bursting was significantly decreased. In the presence of BFT, pharmacological inhibition of GCS caused levels of tight junction protein 1 (TJP1) to decrease. However, when GBA was inhibited, TJP1 levels remained stable, suggesting that BFT-induced production of glucosylceramide helps to stabilize tight junctions. Taken together, our data demonstrate a glucosylceramide-dependent mechanism by which the colon epithelium responds to BFT.
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Toxinas Bacterianas/toxicidad , Bacteroides fragilis/metabolismo , Colon/efectos de los fármacos , Células Epiteliales/efectos de los fármacos , Glucosilceramidas/metabolismo , Metaloendopeptidasas/toxicidad , Transducción de Señal/efectos de los fármacos , Animales , Colitis/inducido químicamente , Colitis/metabolismo , Colon/metabolismo , Células Epiteliales/metabolismo , Glucosilceramidasa/metabolismo , Glucosiltransferasas/metabolismo , Mucosa Intestinal/efectos de los fármacos , Mucosa Intestinal/metabolismo , Ratones , Ratones Endogámicos C57BL , Permeabilidad/efectos de los fármacos , Proteína de la Zonula Occludens-1/metabolismoRESUMEN
ZIP8 is a metal transporter with a role in manganese (Mn) homeostasis. A common genetic variant in ZIP8 (rs13107325; A391T) ranks in the top 10 of pleiotropic SNPs identified in GWAS; A391T has associations with an increased risk of schizophrenia, obesity, Crohn's disease, and reduced blood Mn. Here, we used CRISPR/Cas9-mediated knockin (KI) to generate a mouse model of ZIP8 A391T (Zip8 393T-KI mice). Recapitulating the SNP association with blood Mn, blood Mn was reduced in Zip8 393T-KI mice. There was restricted abnormal tissue Mn homeostasis, with decreases in liver and kidney Mn and a reciprocal increase in biliary Mn, providing in vivo evidence of hypomorphic Zip8 function. Upon challenge in a chemically induced colitis model, male Zip8 393T-KI mice exhibited enhanced disease susceptibility. ZIP8 391-Thr associated with reduced triantennary plasma N-glycan species in a population-based cohort to define a genotype-specific glycophenotype hypothesized to be linked to Mn-dependent glycosyltransferase activity. This glycophenotype was maintained in a cohort of patients with Crohn's disease. These data and the pleiotropic disease associations with ZIP8 391-Thr suggest underappreciated roles of Mn homeostasis in complex human disease.
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Proteínas de Transporte de Catión/genética , Enfermedad de Crohn/genética , Riñón/metabolismo , Manganeso/metabolismo , Animales , Sistemas CRISPR-Cas/genética , Colitis/inducido químicamente , Colitis/genética , Colitis/patología , Enfermedad de Crohn/metabolismo , Enfermedad de Crohn/patología , Sulfato de Dextran/toxicidad , Predisposición Genética a la Enfermedad , Estudio de Asociación del Genoma Completo , Homeostasis/genética , Humanos , Riñón/patología , Hígado/metabolismo , Hígado/patología , Masculino , Manganeso/sangre , Ratones , Polimorfismo de Nucleótido Simple/genéticaRESUMEN
Objective: The human intestinal microbiome plays an important role in inflammatory bowel disease (IBD) and colorectal cancer (CRC) development. One of the first discovered bacterial mediators involves Bacteroides fragilis toxin (BFT, also named as fragilysin), a metalloprotease encoded by enterotoxigenic Bacteroides fragilis (ETBF) that causes barrier disruption and inflammation of the colon, leads to tumorigenesis in susceptible mice, and is enriched in the mucosa of IBD and CRC patients. Thus, targeted inhibition of BFT may benefit ETBF carrying patients. Design: By applying two complementary in silico drug design techniques, drug repositioning and molecular docking, we predicted potential BFT inhibitory compounds. Top candidates were tested in vitro on the CRC epithelial cell line HT29/c1 for their potential to inhibit key aspects of BFT activity, being epithelial morphology changes, E-cadherin cleavage (a marker for barrier function) and increased IL-8 secretion. Results: The primary bile acid and existing drug chenodeoxycholic acid (CDCA), currently used for treating gallstones, cerebrotendinous xanthomatosis, and constipation, was found to significantly inhibit all evaluated cell responses to BFT exposure. The inhibition of BFT resulted from a direct interaction between CDCA and BFT, as confirmed by an increase in the melting temperature of the BFT protein in the presence of CDCA. Conclusion: Together, our results show the potential of in silico drug discovery to combat harmful human and microbiome-derived proteins and more specifically suggests a potential for retargeting CDCA to inhibit the pro-oncogenic toxin BFT.
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Carcinógenos/metabolismo , Transformación Celular Neoplásica , Descubrimiento de Drogas , Reposicionamiento de Medicamentos , Microbioma Gastrointestinal , Toxinas Biológicas , Transformación Celular Neoplásica/inducido químicamente , Transformación Celular Neoplásica/efectos de los fármacos , Endotoxinas/química , Endotoxinas/farmacología , Microbioma Gastrointestinal/efectos de los fármacos , Humanos , Metaloendopeptidasas/química , Metaloendopeptidasas/farmacología , Conformación Molecular , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Unión Proteica , Relación Estructura-Actividad , Toxinas Biológicas/efectos adversos , Toxinas Biológicas/biosíntesisRESUMEN
Mucus-invasive bacterial biofilms are identified on the colon mucosa of approximately 50% of colorectal cancer (CRC) patients and approximately 13% of healthy subjects. Here, we test the hypothesis that human colon biofilms comprise microbial communities that are carcinogenic in CRC mouse models. Homogenates of human biofilm-positive colon mucosa were prepared from tumor patients (tumor and paired normal tissues from surgical resections) or biofilm-positive biopsies from healthy individuals undergoing screening colonoscopy; homogenates of biofilm-negative colon biopsies from healthy individuals undergoing screening colonoscopy served as controls. After 12 weeks, biofilm-positive, but not biofilm-negative, human colon mucosal homogenates induced colon tumor formation in 3 mouse colon tumor models (germ-free ApcMinΔ850/+;Il10-/- or ApcMinΔ850/+ and specific pathogen-free ApcMinΔ716/+ mice). Remarkably, biofilm-positive communities from healthy colonoscopy biopsies induced colon inflammation and tumors similarly to biofilm-positive tumor tissues. By 1 week, biofilm-positive human tumor homogenates, but not healthy biopsies, displayed consistent bacterial mucus invasion and biofilm formation in mouse colons. 16S rRNA gene sequencing and RNA-Seq analyses identified compositional and functional microbiota differences between mice colonized with biofilm-positive and biofilm-negative communities. These results suggest human colon mucosal biofilms, whether from tumor hosts or healthy individuals undergoing screening colonoscopy, are carcinogenic in murine models of CRC.
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Biopelículas , Carcinogénesis , Colon/microbiología , Neoplasias del Colon/microbiología , Microbioma Gastrointestinal , Neoplasias Experimentales/microbiología , Animales , Colon/metabolismo , Neoplasias del Colon/genética , Neoplasias del Colon/metabolismo , Neoplasias del Colon/patología , Humanos , Ratones , Ratones Noqueados , Neoplasias Experimentales/genética , Neoplasias Experimentales/metabolismo , Neoplasias Experimentales/patologíaRESUMEN
Polysaccharide A (PSA), an immunogenic capsular component of non-toxigenic Bacteroides fragilis (NTBF) strain NCTC 9343, is reported to promote mucosal immune development and suppress colitis. Contrastingly, enterotoxigenic Bacteroides fragilis (ETBF) is highly associated with inflammatory bowel disease (IBD) and colorectal cancer (CRC), rapidly inducing IL-17-dependent murine colitis and tumorigenesis. In specific-pathogen-free (SPF) C57BL/6 wild-type (WT) and multiple intestinal neoplasia (MinApc716+/-) mice, we show that sequential treatment of the NTBF strain, 9343, followed by the ETBF strain, 86-5443-2-2 (86), diminished colitis and tumorigenesis. Mice treated simultaneously with 9343 and 86 exhibited both severe colitis and tumorigenesis. Abrogated disease severity in sequentially treated mice was attributed to 9343 strain dominance and decreased IL-17A, but 86 colonization prior to or simultaneous with 9343 mitigated the anti-inflammatory effect of 9343. Remarkably, 9343-mediated protection was independent of PSA, as sequentially treated mice receiving ΔPSA 9343 exhibited similar protection. Further, SPF WT and Min mice colonized with PSA-competent or PSA-deficient 9343 exhibited similar IL-10, IL-17, and IFN-γ responses. Treatment of 86-colonized mice with 9343 failed to disrupt 86 pathogenesis. Our findings demonstrate that 9343 colonization, independent of PSA, offers prophylaxis against colitis-inducing 86 but may not be a valid therapy once colitis is established.
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Bacteroides fragilis/inmunología , Colitis/inmunología , Neoplasias Colorrectales/inmunología , Enfermedades Inflamatorias del Intestino/inmunología , Mucosa Intestinal/inmunología , Células Th17/inmunología , Proteína de la Poliposis Adenomatosa del Colon/genética , Animales , Bacteroides fragilis/patogenicidad , Carcinogénesis , Células Cultivadas , Colitis/inducido químicamente , Modelos Animales de Enfermedad , Humanos , Interleucina-17/metabolismo , Lipopolisacáridos/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ácido TrinitrobencenosulfónicoRESUMEN
We present the first, most compact, ultrahigh-resolution, high-speed, distal scanning optical coherence tomography (OCT) endoscope operating at 800 nm. Achieving high speed imaging while maintaining an ultrahigh axial resolution is one of the most significant challenges with endoscopic OCT at 800 nm. Maintaining an ultrahigh axial resolution requires preservation of the broad spectral bandwidth of the light source throughout the OCT system. To overcome this critical limitation we implemented a distal scanning endoscope with diffractive optics to minimize loss in spectral throughput. In this paper, we employed a customized miniature 900 µm diameter DC micromotor fitted with a micro reflector to scan the imaging beam. We integrated a customized diffractive microlens into the imaging optics to reduce chromatic focal shift over the broad spectral bandwidth of the Ti:Sapphire laser of an approximately 150 nm 3dB bandwidth, affording a measured axial resolution of 2.4 µm (in air). The imaging capability of this high-speed, ultrahigh-resolution distal scanning endoscope was validated by performing 3D volumetric imaging of mouse colon in vivo at 50 frames-per-second (limited only by the A-scan rate of linear CCD array in the spectral-domain OCT system and sampling requirements). The results demonstrated that fine microstructures of colon could be clearly visualized, including the boundary between the absorptive cell layer and colonic mucosa as well the crypt patterns. Furthermore, this endoscope was employed to visualize morphological changes in an enterotoxigenic Bacteriodes fragilis (ETBF) induced colon tumor model. We present the results of our feasibility studies and suggest the potential of this system for visualizing time dependent morphological changes associated with tumorigenesis on murine models in vivo.
RESUMEN
Pro-carcinogenic bacteria have the potential to initiate and/or promote colon cancer, in part via immune mechanisms that are incompletely understood. Using ApcMin mice colonized with the human pathobiont enterotoxigenic Bacteroides fragilis (ETBF) as a model of microbe-induced colon tumorigenesis, we show that the Bacteroides fragilis toxin (BFT) triggers a pro-carcinogenic, multi-step inflammatory cascade requiring IL-17R, NF-κB, and Stat3 signaling in colonic epithelial cells (CECs). Although necessary, Stat3 activation in CECs is not sufficient to trigger ETBF colon tumorigenesis. Notably, IL-17-dependent NF-κB activation in CECs induces a proximal to distal mucosal gradient of C-X-C chemokines, including CXCL1, that mediates the recruitment of CXCR2-expressing polymorphonuclear immature myeloid cells with parallel onset of ETBF-mediated distal colon tumorigenesis. Thus, BFT induces a pro-carcinogenic signaling relay from the CEC to a mucosal Th17 response that results in selective NF-κB activation in distal colon CECs, which collectively triggers myeloid-cell-dependent distal colon tumorigenesis.
Asunto(s)
Toxinas Bacterianas/inmunología , Bacteroides fragilis/inmunología , Carcinogénesis/patología , Colon/inmunología , Neoplasias Colorrectales/etiología , Células Epiteliales/inmunología , Interleucina-17/inmunología , Metaloendopeptidasas/inmunología , Factor de Transcripción ReIA/metabolismo , Proteína de la Poliposis Adenomatosa del Colon/genética , Animales , Toxinas Bacterianas/metabolismo , Bacteroides fragilis/patogenicidad , Línea Celular Tumoral , Colon/citología , Colon/microbiología , Neoplasias Colorrectales/microbiología , Neoplasias Colorrectales/patología , Activación Enzimática/inmunología , Femenino , Eliminación de Gen , Células HT29 , Humanos , Inflamación/inmunología , Inflamación/microbiología , Interleucina-17/genética , Masculino , Metaloendopeptidasas/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Células Mieloides/inmunología , Receptores de Interleucina-17/genética , Receptores de Interleucina-17/inmunología , Receptores de Interleucina-8B/genética , Factor de Transcripción STAT3/metabolismoRESUMEN
Individuals with sporadic colorectal cancer (CRC) frequently harbor abnormalities in the composition of the gut microbiome; however, the microbiota associated with precancerous lesions in hereditary CRC remains largely unknown. We studied colonic mucosa of patients with familial adenomatous polyposis (FAP), who develop benign precursor lesions (polyps) early in life. We identified patchy bacterial biofilms composed predominately of Escherichia coli and Bacteroides fragilis Genes for colibactin (clbB) and Bacteroides fragilis toxin (bft), encoding secreted oncotoxins, were highly enriched in FAP patients' colonic mucosa compared to healthy individuals. Tumor-prone mice cocolonized with E. coli (expressing colibactin), and enterotoxigenic B. fragilis showed increased interleukin-17 in the colon and DNA damage in colonic epithelium with faster tumor onset and greater mortality, compared to mice with either bacterial strain alone. These data suggest an unexpected link between early neoplasia of the colon and tumorigenic bacteria.
Asunto(s)
Poliposis Adenomatosa del Colon/microbiología , Poliposis Adenomatosa del Colon/patología , Bacteroides fragilis/patogenicidad , Biopelículas , Carcinogénesis , Colon/microbiología , Neoplasias del Colon/microbiología , Escherichia coli/patogenicidad , Interleucina-17/análisis , Animales , Toxinas Bacterianas/genética , Bacteroides fragilis/genética , Bacteroides fragilis/aislamiento & purificación , Colon/patología , Neoplasias del Colon/patología , Daño del ADN , Escherichia coli/genética , Escherichia coli/aislamiento & purificación , Microbioma Gastrointestinal , Humanos , Mucosa Intestinal/química , Mucosa Intestinal/microbiología , Mucosa Intestinal/patología , Metaloendopeptidasas/genética , Metaloendopeptidasas/metabolismo , Ratones , Péptidos/genética , Péptidos/metabolismo , Policétidos , Lesiones Precancerosas/microbiologíaRESUMEN
Bacteroides fragilis is an extensively studied anaerobic bacterium comprising the normal flora of the human gut. B. fragilis is known to be one of the most commonly isolated species from clinical samples and has been shown to cause a wide range of pathologies in humans [1, 2]. As an opportunistic pathogen B. fragilis can cause abscess formation and bacteremia [2]. Additionally in its enterotoxigenic form, B. fragilis is a known cause of diarrheal illness, is associated with inflammatory bowel disease, and has been recently characterized in patients with colon cancer [3 - 5]. As research in the field of the gut microbiome continues to expand at an ever increasing rate due to advances in the availability of next generation sequencing and analysis tools it is important to outline various molecular methods that can be employed in quickly detecting and isolating relevant strains of B. fragilis. This review outlines methods that are routinely employed in the isolation and detection of B. fragilis, with an emphasis on characterizing enterotoxigenic B. fragilis (ETBF) strains.