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Sample records for human intestinal bacterial

  1. Impact of Different Fecal Processing Methods on Assessments of Bacterial Diversity in the Human Intestine

    PubMed Central

    Hsieh, Yu-Hsin; Peterson, Courtney M.; Raggio, Anne; Keenan, Michael J.; Martin, Roy J.; Ravussin, Eric; Marco, Maria L.

    2016-01-01

    The intestinal microbiota are integral to understanding the relationships between nutrition and health. Therefore, fecal sampling and processing protocols for metagenomic surveys should be sufficiently robust, accurate, and reliable to identify the microorganisms present. We investigated the use of different fecal preparation methods on the bacterial community structures identified in human stools. Complete stools were collected from six healthy individuals and processed according to the following methods: (i) randomly sampled fresh stool, (ii) fresh stool homogenized in a blender for 2 min, (iii) randomly sampled frozen stool, and (iv) frozen stool homogenized in a blender for 2 min, or (v) homogenized in a pneumatic mixer for either 10, 20, or 30 min. High-throughput DNA sequencing of the 16S rRNA V4 regions of bacterial community DNA extracted from the stools showed that the fecal microbiota remained distinct between individuals, independent of processing method. Moreover, the different stool preparation approaches did not alter intra-individual bacterial diversity. Distinctions were found at the level of individual taxa, however. Stools that were frozen and then homogenized tended to have higher proportions of Faecalibacterium, Streptococcus, and Bifidobacterium and decreased quantities of Oscillospira, Bacteroides, and Parabacteroides compared to stools that were collected in small quantities and not mixed prior to DNA extraction. These findings indicate that certain taxa are at particular risk for under or over sampling due to protocol differences. Importantly, homogenization by any method significantly reduced the intra-individual variation in bacteria detected per stool. Our results confirm the robustness of fecal homogenization for microbial analyses and underscore the value of collecting and mixing large stool sample quantities in human nutrition intervention studies. PMID:27812352

  2. Pasteurization Procedures for Donor Human Milk Affect Body Growth, Intestinal Structure, and Resistance against Bacterial Infections in Preterm Pigs.

    PubMed

    Li, Yanqi; Nguyen, Duc Ninh; de Waard, Marita; Christensen, Lars; Zhou, Ping; Jiang, Pingping; Sun, Jing; Bojesen, Anders Miki; Lauridsen, Charlotte; Lykkesfeldt, Jens; Dalsgaard, Trine Kastrup; Bering, Stine Brandt; Sangild, Per Torp

    2017-03-15

    Background: Holder pasteurization (HP) destroys multiple bioactive factors in donor human milk (DM), and UV-C irradiation (UVC) is potentially a gentler method for pasteurizing DM for preterm infants.Objective: We investigated whether UVC-treated DM improves gut maturation and resistance toward bacterial infections relative to HP-treated DM.Methods: Bacteria, selected bioactive components, and markers of antioxidant capacity were measured in unpasteurized donor milk (UP), HP-treated milk, and UVC-treated milk (all from the same DM pool). Fifty-seven cesarean-delivered preterm pigs (91% gestation; ratio of males to females, 30:27) received decreasing volumes of parental nutrition (average 69 mL ⋅ kg(-1) ⋅ d(-1)) and increasing volumes of the 3 DM diets (n = 19 each, average 89 mL ⋅ kg(-1) ⋅ d(-1)) for 8-9 d. Body growth, gut structure and function, and systemic bacterial infection were evaluated.Results: A high bacterial load in the UP (6×10(5) colony forming units/mL) was eliminated similarly by HP and UVC treatments. Relative to HP-treated milk, both UVC-treated milk and UP showed greater activities of lipase and alkaline phosphatase and concentrations of lactoferrin, secretory immunoglobulin A, xanthine dehydrogenase, and some antioxidant markers (all P < 0.05). The pigs fed UVC-treated milk and pigs fed UP showed higher relative weight gain than pigs fed HP-treated milk (5.4% and 3.5%), and fewer pigs fed UVC-treated milk had positive bacterial cultures in the bone marrow (28%) than pigs fed HP-treated milk (68%) (P < 0.05). Intestinal health was also improved in pigs fed UVC-treated milk compared with those fed HP-treated milk as indicated by a higher plasma citrulline concentration (36%) and villus height (38%) (P < 0.05) and a tendency for higher aminopeptidase N (48%) and claudin-4 (26%) concentrations in the distal intestine (P < 0.08). The gut microbiota composition was similar among groups except for greater proportions of Enterococcus in pigs

  3. Prevalence of intestinal parasitic and bacterial pathogens in diarrhoeal and non-diarroeal human stools from Vhembe district, South Africa.

    PubMed

    Samie, A; Guerrant, R L; Barrett, L; Bessong, P O; Igumbor, E O; Obi, C L

    2009-12-01

    In the present study, a cross-sectional survey of intestinal parasitic and bacterial infections in relation to diarrhoea in Vhembe district and the antimicrobial susceptibility profiles of isolated bacterial pathogens was conducted. Stool samples were collected from 528 patients attending major public hospitals and 295 children attending two public primary schools and were analyzed by standard microbiological and parasitological techniques. Entamoeba histolytica/E. dispar (34.2%) and Cryptosporidium spp. (25.5%) were the most common parasitic causes of diarrhoea among the hospital attendees while Giardia lamblia (12.8%) was the most common cause of diarrhoea among the primary school children (p < 0.05). Schistosoma mansoni (14.4%) was more common in non-diarrhoeal samples at both hospitals (16.9%) and schools (17.6%). Campylobacter spp. (24.9%), Aeromonas spp. (20.8%), and Shigella spp. (8.5%) were the most common bacterial causes of diarrhoea among the hospital attendees while Campylobacter (12.8%) and Aeromonas spp. (12.8%) were most common in diarrhoeal samples from school children. Vibrio spp. was less common (3% in the hospitals) and were all associated with diarrhoea. Antimicrobial resistance was common among the bacterial isolates but ceftriaxone (91%) and ciprofloxacin (88.6%) showed stronger activities against all the organisms. The present study has demonstrated that E. histolytica/dispar, Cryptosporidium, Giardia, and Cyclospora are common parasitic causes of diarrhoea in Vhembe district while Campylobacter spp. and Aeromonas are the most common bacterial causes of diarrhoea in Vhembe district of South Africa.

  4. CAP-D3 Promotes Bacterial Clearance in Human Intestinal Epithelial Cells by Repressing Expression of Amino Acid Transporters

    PubMed Central

    Kemp, Jacqueline R.; Nickerson, Kourtney P.; Deutschman, Emily; Kim, Yeojung; West, Gail; Sadler, Tammy; Stylianou, Eleni; Krokowski, Dawid; Hatzoglou, Maria; de la Motte, Carol; Rubin, Brian P.; Fiocchi, Claudio

    2015-01-01

    BACKGROUND & AIMS Defects in colonic epithelial barrier defenses are associated with ulcerative colitis (UC). The proteins that regulate bacterial clearance in the colonic epithelium have not been completely identified. The chromosome-associated protein D3 (dCAP-D3), regulates responses to bacterial infection. We examined whether CAP-D3 promotes bacterial clearance in human colonic epithelium. METHODS Clearance of Salmonella or adherent-invasive Escherichia coli LF82 was assessed by gentamycin protection assays in HT-29 and Caco-2 cells expressing small hairpin RNAs against CAP-D3. We used immunoblot assays to measure levels of CAP-D3 in colonic epithelial cells from patients with UC and healthy individuals (controls). RNA sequencing identified genes activated by CAP-D3. We analyzed the roles of CAP-D3 target genes in bacterial clearance using gentamycin protection and immunofluorescence assays and studies with pharmacologic inhibitors. RESULTS CAP-D3 expression was reduced in colonic epithelial cells from patients with active UC. Reduced CAP-D3 expression decreased autophagy and impaired intracellular bacterial clearance by HT-29 and Caco-2 colonic epithelial cells. Lower levels of CAP-D3 increased transcription of genes encoding SLC7A5 and SLC3A2, whose products heterodimerize to form an amino acid transporter in HT-29 cells following bacterial infection; levels of SLC7A5–SLC3A2 were increased in tissues from patients with UC, compared with controls. Reduced CAP-D3 in HT-29 cells resulted in earlier recruitment of SLC7A5 to Salmonella-containing vacuoles, increased activity of mTORC1, and increased survival of bacteria. Inhibition of SLC7A5–SLC3A2 or mTORC1 activity rescued the bacterial clearance defects of CAP-D3– deficient cells. CONCLUSIONS CAP-D3 downregulates transcription of genes that encode amino acid transporters (SLC7A5 and SLC3A2) to promote bacterial autophagy by colon epithelial cells. Levels of CAP-D3 protein are reduced in patients with

  5. Upper intestinal bacterial flora during transpyloric feeding.

    PubMed Central

    Dellagrammaticas, H D; Duerden, B I; Milner, R D

    1983-01-01

    Samples from the pharynx, stomach, duodenum or jejunum, and faeces were collected on 7 days between 1st and 28th day from neonates weighing less than 1.5 kg at birth who were fed by transpyloric tube. These were cultured on selective and non-selective media, and the results were expressed in a semi-quantitative manner. The number of bacterial species and the density of their growth increased with the patient's age; this was particularly noticeable with Gram-negative bacteria and the ratio of Gram-negative to Gram-positive organisms increased steadily in specimens from all sites with increasing age. The upper small intestine was more heavily colonised than the stomach early in life and the microflora present was predominantly faecal in nature. The species isolated from all sites were mainly aerobes or facultative anaerobes; strict anaerobes did not form a significant proportion of the microflora in these infants. Necrotising enterocolitis developed only after heavy jejunal colonisation with Gram-negative bacilli. PMID:6402990

  6. Giardia duodenalis infection reduces granulocyte infiltration in an in vivo model of bacterial toxin-induced colitis and attenuates inflammation in human intestinal tissue.

    PubMed

    Cotton, James A; Motta, Jean-Paul; Schenck, L Patrick; Hirota, Simon A; Beck, Paul L; Buret, Andre G

    2014-01-01

    Giardia duodenalis (syn. G. intestinalis, G. lamblia) is a predominant cause of waterborne diarrheal disease that may lead to post-infectious functional gastrointestinal disorders. Although Giardia-infected individuals could carry as much as 106 trophozoites per centimetre of gut, their intestinal mucosa is devoid of overt signs of inflammation. Recent studies have shown that in endemic countries where bacterial infectious diseases are common, Giardia infections can protect against the development of diarrheal disease and fever. Conversely, separate observations have indicated Giardia infections may enhance the severity of diarrheal disease from a co-infecting pathogen. Polymorphonuclear leukocytes or neutrophils (PMNs) are granulocytic, innate immune cells characteristic of acute intestinal inflammatory responses against bacterial pathogens that contribute to the development of diarrheal disease following recruitment into intestinal tissues. Giardia cathepsin B cysteine proteases have been shown to attenuate PMN chemotaxis towards IL-8/CXCL8, suggesting Giardia targets PMN accumulation. However, the ability of Giardia infections to attenuate PMN accumulation in vivo and how in turn this effect may alter the host inflammatory response in the intestine has yet to be demonstrated. Herein, we report that Giardia infection attenuates granulocyte tissue infiltration induced by intra-rectal instillation of Clostridium difficile toxin A and B in an isolate-dependent manner. This attenuation of granulocyte infiltration into colonic tissues paralled decreased expression of several cytokines associated with the recruitment of PMNs. Giardia trophozoite isolates that attenuated granulocyte infiltration in vivo also decreased protein expression of cytokines released from inflamed mucosal biopsy tissues collected from patients with active Crohn's disease, including several cytokines associated with PMN recruitment. These results demonstrate for the first time that certain

  7. Alcohol, intestinal bacterial growth, intestinal permeability to endotoxin, and medical consequences: summary of a symposium.

    PubMed

    Purohit, Vishnudutt; Bode, J Christian; Bode, Christiane; Brenner, David A; Choudhry, Mashkoor A; Hamilton, Frank; Kang, Y James; Keshavarzian, Ali; Rao, Radhakrishna; Sartor, R Balfour; Swanson, Christine; Turner, Jerrold R

    2008-08-01

    This report is a summary of the symposium on Alcohol, Intestinal Bacterial Growth, Intestinal Permeability to Endotoxin, and Medical Consequences, organized by National Institute on Alcohol Abuse and Alcoholism, Office of Dietary Supplements, and National Institute of Diabetes and Digestive and Kidney Diseases of National Institutes of Health in Rockville, Maryland, October 11, 2006. Alcohol exposure can promote the growth of Gram-negative bacteria in the intestine, which may result in accumulation of endotoxin. In addition, alcohol metabolism by Gram-negative bacteria and intestinal epithelial cells can result in accumulation of acetaldehyde, which in turn can increase intestinal permeability to endotoxin by increasing tyrosine phosphorylation of tight junction and adherens junction proteins. Alcohol-induced generation of nitric oxide may also contribute to increased permeability to endotoxin by reacting with tubulin, which may cause damage to microtubule cytoskeleton and subsequent disruption of intestinal barrier function. Increased intestinal permeability can lead to increased transfer of endotoxin from the intestine to the liver and general circulation where endotoxin may trigger inflammatory changes in the liver and other organs. Alcohol may also increase intestinal permeability to peptidoglycan, which can initiate inflammatory response in liver and other organs. In addition, acute alcohol exposure may potentiate the effect of burn injury on intestinal bacterial growth and permeability. Decreasing the number of Gram-negative bacteria in the intestine can result in decreased production of endotoxin as well as acetaldehyde which is expected to decrease intestinal permeability to endotoxin. In addition, intestinal permeability may be preserved by administering epidermal growth factor, l-glutamine, oats supplementation, or zinc, thereby preventing the transfer of endotoxin to the general circulation. Thus reducing the number of intestinal Gram-negative bacteria

  8. Cohabitation in the Intestine: Interactions among Helminth Parasites, Bacterial Microbiota, and Host Immunity.

    PubMed

    Reynolds, Lisa A; Finlay, B Brett; Maizels, Rick M

    2015-11-01

    Both intestinal helminth parasites and certain bacterial microbiota species have been credited with strong immunomodulatory effects. Recent studies reported that the presence of helminth infection alters the composition of the bacterial intestinal microbiota and, conversely, that the presence and composition of the bacterial microbiota affect helminth colonization and persistence within mammalian hosts. This article reviews recent findings on these reciprocal relationships, in both human populations and mouse models, at the level of potential mechanistic pathways and the implications these bear for immunomodulatory effects on allergic and autoimmune disorders. Understanding the multidirectional complex interactions among intestinal microbes, helminth parasites, and the host immune system allows for a more holistic approach when using probiotics, prebiotics, synbiotics, antibiotics, and anthelmintics, as well as when designing treatments for autoimmune and allergic conditions.

  9. Fucose sensing regulates bacterial intestinal colonization.

    PubMed

    Pacheco, Alline R; Curtis, Meredith M; Ritchie, Jennifer M; Munera, Diana; Waldor, Matthew K; Moreira, Cristiano G; Sperandio, Vanessa

    2012-12-06

    The mammalian gastrointestinal tract provides a complex and competitive environment for the microbiota. Successful colonization by pathogens requires scavenging nutrients, sensing chemical signals, competing with the resident bacteria and precisely regulating the expression of virulence genes. The gastrointestinal pathogen enterohaemorrhagic Escherichia coli (EHEC) relies on inter-kingdom chemical sensing systems to regulate virulence gene expression. Here we show that these systems control the expression of a novel two-component signal transduction system, named FusKR, where FusK is the histidine sensor kinase and FusR the response regulator. FusK senses fucose and controls expression of virulence and metabolic genes. This fucose-sensing system is required for robust EHEC colonization of the mammalian intestine. Fucose is highly abundant in the intestine. Bacteroides thetaiotaomicron produces multiple fucosidases that cleave fucose from host glycans, resulting in high fucose availability in the gut lumen. During growth in mucin, B. thetaiotaomicron contributes to EHEC virulence by cleaving fucose from mucin, thereby activating the FusKR signalling cascade, modulating the virulence gene expression of EHEC. Our findings suggest that EHEC uses fucose, a host-derived signal made available by the microbiota, to modulate EHEC pathogenicity and metabolism.

  10. Flow and active mixing have a strong impact on bacterial growth dynamics in the proximal large intestine

    NASA Astrophysics Data System (ADS)

    Cremer, Jonas; Segota, Igor; Yang, Chih-Yu; Arnoldini, Markus; Groisman, Alex; Hwa, Terence

    2016-11-01

    More than half of fecal dry weight is bacterial mass with bacterial densities reaching up to 1012 cells per gram. Mostly, these bacteria grow in the proximal large intestine where lateral flow along the intestine is strong: flow can in principal lead to a washout of bacteria from the proximal large intestine. Active mixing by contractions of the intestinal wall together with bacterial growth might counteract such a washout and allow high bacterial densities to occur. As a step towards understanding bacterial growth in the presence of mixing and flow, we constructed an in-vitro setup where controlled wall-deformations of a channel emulate contractions. We investigate growth along the channel under a steady nutrient inflow. Depending on mixing and flow, we observe varying spatial gradients in bacterial density along the channel. Active mixing by deformations of the channel wall is shown to be crucial in maintaining a steady-state bacterial population in the presence of flow. The growth-dynamics is quantitatively captured by a simple mathematical model, with the effect of mixing described by an effective diffusion term. Based on this model, we discuss bacterial growth dynamics in the human large intestine using flow- and mixing-behavior having been observed for humans.

  11. A model for Vibrio cholerae colonization of the human intestine.

    PubMed

    Spagnuolo, Anna Maria; Dirita, Victor; Kirschner, Denise

    2011-11-21

    Vibrio cholerae is a strict human pathogen that causes the disease cholera. It is an old-world pathogen that has re-emerged as a new threat since the early 1990s. V. cholerae colonizes the upper, small intestine where it produces a toxin that leads to watery diarrhea, characterizing the disease (Kahn et al., 1988). The dynamics of colonization by the bacteria of the intestines are largely unknown. Although a large initial infectious dose is required for infection, data suggests that only a smaller sub-population colonizes a portion of the small bowel leading to disease. There are many barriers to colonization in the intestines including peristalsis, fluid wash-out, viscosity of the mucus layer, and pH. We are interested in identifying the mechanisms that allow this sub-population of bacteria to survive and colonize the intestines when faced with these barriers. To elaborate the dynamics of V. cholerae infection, we have developed a mathematical model based on a convection-diffusion-reaction-swimming equation capturing bacterial dynamics coupled with Stokes equations governing fluid velocity where we developed a novel non-local boundary condition. Our results indicate that both host and bacterial factors contribute to bacterial density in the gut. Host factors include intestinal diffusion and convection rates while bacterial factors include adherence, motility and growth rates. This model can ultimately be used to test therapeutic strategies against V. cholerae.

  12. Diagnosis and management of small intestinal bacterial overgrowth.

    PubMed

    Bohm, Matthew; Siwiec, Robert M; Wo, John M

    2013-06-01

    Small intestinal bacterial overgrowth (SIBO) can result from failure of the gastric acid barrier, failure of small intestinal motility, anatomic alterations, or impairment of systemic and local immunity. The current accepted criteria for the diagnosis of SIBO is the presence of coliform bacteria isolated from the proximal jejunum with >10(5) colony-forming units/mL. A major concern with luminal aspiration is that it is only one random sampling of the small intestine and may not always be representative of the underlying microbiota. A new approach to examine the underlying microbiota uses rapid molecular sequencing, but its clinical utilization is still under active investigation. Clinical manifestations of SIBO are variable and include bloating, flatulence, abdominal distention, abdominal pain, and diarrhea. Severe cases may present with nutrition deficiencies due to malabsorption of micro- and macronutrients. The current management strategies for SIBO center on identifying and correcting underlying causes, addressing nutrition deficiencies, and judicious utilization of antibiotics to treat symptomatic SIBO.

  13. Vitamin B12 uptake by intestinal microorganisms: mechanism and relevance to syndromes of intestinal bacterial overgrowth

    PubMed Central

    Giannella, R. A.; Broitman, S. A.; Zamcheck, N.

    1971-01-01

    The mechanism of bacterial uptake of vitamin B12, the spectrum of microorganisms capable of such uptake, and the factors involved were the subject of this study. Bacterial uptake of vitamin B12 was found to be at least a two stage process. A primary uptake phase which was rapid (1 min or less), pH dependent, nontemperature dependent, did not require viable organisms and was insensitive to either the metabolic inhibitor dinitrophenol or to the sulfhydryl inhibitor N-ethyl-maleimide. Protein denaturation (formalin treatment or autoclaving) abolished all B12 uptake. This primary uptake phase is thought to represent adsorption to binding or “receptor” sites on the cell wall. Second stage uptake was slower, pH and temperature dependent, required living bacteria, and was abolished by either dinitrophenol or N-ethyl-maleimide. This phase is dependent upon metabolic processes and may reflect transfer of B12 from surface “receptor” sites into the bacterial cell. Although differences among organisms were observed in total 1 hr uptake, number of surface “receptor” sites, and relative avidities for B12, all organisms except Streptococcus fecalis shared the two stage mechanism. Two Gram-positive organisms. Bacillus subtilis and Group A streptococcus, demonstrated the highest 1 hr vitamin B12 uptake values; Gram-negative bacteria required 2,000-10,000 the number of organisms for comparable uptake. Binding constants (Km) varied from 5.05 ±1.67 × 10-10M for B. subtilis to 6.18 ±3.08 × 10-9M for Klebsiella pneumoniae which approximate the Km for human intrinsic factor (0.38 × 10-10M). Competition between bacteria and intrinsic factor for vitamin B12 may be inferred from the similarity of these constants. These observations suggest that a variety of enteric and nonenteric organisms, not requiring exogenous B12, may play a role in the pathogenesis of the vitamin B12 malabsorption found in the intestinal bacterial overgrowth syndromes. PMID:4994753

  14. Intestinal mucin distribution in the germ-free rat and in the heteroxenic rat harbouring a human bacterial flora: effect of inulin in the diet.

    PubMed

    Fontaine, N; Meslin, J C; Lory, S; Andrieux, C

    1996-06-01

    A colorimetric method was used on water-soluble mucin extracted from mucosal scrapings and contents of the caecum and the colon of five germ-free (GF) rats and five heteroxenic (HE) rats harbouring a human flora (GF rats associated with a human flora). These rats were fed on a diet containing either 100 g sucrose/kg or 100 g inulin/kg. Histological stains, periodic acid-Schiff, alcian blue pH 2.5 and alcian blue pH 0.5 were used to discriminate between neutral, acidic and acidic sulphated mucins respectively. Spectrocolorimetric assays led to a calculated absorbance value for 1 mg of the initial mucin extract. Each mucin type was compared between treatments. The caecal contents of GF rats contained more acidic mucin than sulphomucin, which was present in the same proportion as neutral mucin. Their colonic contents contained more acidic mucins than sulphomucin, which in turn was more abundant than neutral mucin. Their caecal mucosa mucin distribution differed from that of the contents: very little acidic mucin was present and neutral and sulphomucin proportions were of the same order of magnitude. Inulin increased the amount of neutral mucin in the caecal contents and of sulphated mucins in the colonic contents and increased the amounts of neutral and acidic mucins in the caecal mucosa. Mucin distribution in the HE rats was very different from that in the GF rats: the caecal contents contained a high proportion of acidic mucins and very little sulphomucin. The same distribution of mucins was observed in the colonic contents. The caecal mucosa contained less acidic mucin and more sulphomucin than the caecal contents. Inulin decreased acidic mucins and increased sulphated mucins in the caecal contents and increased neutral and sulphated mucins in the colonic contents. Inulin increased sulphomucin in the caecal mucosa and decreased acidic mucin in the caecal and colonic mucosas. The very low amount of mucin that was recovered in the colonic mucosa suggests that, in the

  15. Bacterial Intestinal Superinfections in Inflammatory Bowel Diseases Beyond Clostridum difficile.

    PubMed

    Lobatón, Triana; Domènech, Eugeni

    2016-07-01

    Besides genetics and environmental factors, intestinal microbiota seem to play a major role in the pathogenesis of inflammatory bowel diseases. For many decades, it has been said that some enteropathogens may even trigger both inflammatory bowel disease development and disease flares. For this reason, stool testing had been performed in inflammatory bowel disease flares but current guidelines only recommend to rule out Clostridium difficile infection and there is no clear advice for other enteropathogens given that the scarce available evidence points at a low prevalence of this sort of intestinal superinfections with no clear impact on disease course. The present article reviews the current knowledge about the role of bacterial enteropathogens on disease pathogenesis and flares beyond C. difficile.

  16. Emerging insights on intestinal dysbiosis during bacterial infections.

    PubMed

    Pham, Tu Anh N; Lawley, Trevor D

    2014-02-01

    Infection of the gastrointestinal tract is commonly linked to pathological imbalances of the resident microbiota, termed dysbiosis. In recent years, advanced high-throughput genomic approaches have allowed us to examine the microbiota in an unprecedented manner, revealing novel biological insights about infection-associated dysbiosis at the community and individual species levels. A dysbiotic microbiota is typically reduced in taxonomic diversity and metabolic function, and can harbour pathobionts that exacerbate intestinal inflammation or manifest systemic disease. Dysbiosis can also promote pathogen genome evolution, while allowing the pathogens to persist at high density and transmit to new hosts. A deeper understanding of bacterial pathogenicity in the context of the intestinal microbiota should unveil new approaches for developing diagnostics and therapies for enteropathogens.

  17. Small Intestine Bacterial Overgrowth and Environmental Enteropathy in Bangladeshi Children

    PubMed Central

    Haque, Rashidul; Kirkpatrick, Beth D.; Alam, Masud; Lu, Miao; Kabir, Mamun; Kakon, Shahria Hafiz; Islam, Bushra Zarin; Afreen, Sajia; Musa, Abu; Khan, Shaila Sharmeen; Colgate, E. Ross; Carmolli, Marya P.; Ma, Jennie Z.

    2016-01-01

    ABSTRACT Recent studies suggest small intestine bacterial overgrowth (SIBO) is common among developing world children. SIBO’s pathogenesis and effect in the developing world are unclear. Our objective was to determine the prevalence of SIBO in Bangladeshi children and its association with malnutrition. Secondary objectives included determination of SIBO’s association with sanitation, diarrheal disease, and environmental enteropathy. We performed a cross-sectional analysis of 90 Bangladeshi 2-year-olds monitored since birth from an impoverished neighborhood. SIBO was diagnosed via glucose hydrogen breath testing, with a cutoff of a 12-ppm increase over baseline used for SIBO positivity. Multivariable logistic regression was performed to investigate SIBO predictors. Differences in concomitant inflammation and permeability between SIBO-positive and -negative children were compared with multiple comparison adjustment. A total of 16.7% (15/90) of the children had SIBO. The strongest predictors of SIBO were decreased length-for-age Z score since birth (odds ratio [OR], 0.13; 95% confidence interval [CI], 0.03 to 0.60) and an open sewer outside the home (OR, 4.78; 95% CI, 1.06 to 21.62). Recent or frequent diarrheal disease did not predict SIBO. The markers of intestinal inflammation fecal Reg 1β (116.8 versus 65.6 µg/ml; P = 0.02) and fecal calprotectin (1,834.6 versus 766.7 µg/g; P = 0.004) were elevated in SIBO-positive children. Measures of intestinal permeability and systemic inflammation did not differ between the groups. These findings suggest linear growth faltering and poor sanitation are associated with SIBO independently of recent or frequent diarrheal disease. SIBO is associated with intestinal inflammation but not increased permeability or systemic inflammation. PMID:26758185

  18. Human intestinal capillariasis in Thailand

    PubMed Central

    Saichua, Prasert; Nithikathkul, Choosak; Kaewpitoon, Natthawut

    2008-01-01

    Intestinal capillariasis caused by Capillaria philippinensis appeared first in the Philippines and subsequently in Thailand, Japan, Iran, Egypt and Taiwan; major outbreaks have occurred in the Philippines and Thailand. This article reviews the epidemiology, history and sources of C. philippinensis infection in Thailand. The annual epidemiological surveillance reports indicated that 82 accumulated cases of intestinal capillariasis were found in Thailand from 1994-2006. That made Thailand a Capillaria-prevalent area. Sisaket, in northeast Thailand, was the first province which has reported intestinal capillariasis. Moreover, Buri Ram presented a high prevalence of intestinal capillariasis, totaling 24 cases from 1994-2006. About half of all cases have consumed raw or undercooked fish. However, even if the numbers of the intestinal capillariasis cases in Thailand is reduced, C. philippinensis infection cases are still reported. The improvement of personal hygiene, specifically avoiding consumption of undercooked fish and promoting a health education campaign are required. These strategies may minimize or eliminate C. philippinensis infection in Thailand. PMID:18203280

  19. The Contributions of Human Mini-Intestines to the Study of Intestinal Physiology and Pathophysiology.

    PubMed

    Yu, Huimin; Hasan, Nesrin M; In, Julie G; Estes, Mary K; Kovbasnjuk, Olga; Zachos, Nicholas C; Donowitz, Mark

    2017-02-10

    The lack of accessibility to normal and diseased human intestine and the inability to separate the different functional compartments of the intestine even when tissue could be obtained have held back the understanding of human intestinal physiology. Clevers and his associates identified intestinal stem cells and established conditions to grow "mini-intestines" ex vivo in differentiated and undifferentiated conditions. This pioneering work has made a new model of the human intestine available and has begun making contributions to the understanding of human intestinal transport in normal physiologic conditions and the pathophysiology of intestinal diseases. However, this model is reductionist and lacks many of the complexities of normal intestine. Consequently, it is not yet possible to predict how great the advances using this model will be for understanding human physiology and pathophysiology, nor how the model will be modified to include multiple other intestinal cell types and physical forces necessary to more closely approximate normal intestine. This review describes recent studies using mini-intestines, which have readdressed previously established models of normal intestinal transport physiology and newly examined intestinal pathophysiology. The emphasis is on studies with human enteroids grown either as three-dimensional spheroids or two-dimensional monolayers. In addition, comments are provided on mouse studies in cases when human studies have not yet been described.

  20. Comparison of intestinal bacterial communities in grass carp, Ctenopharyngodon idellus, from two different habitats

    NASA Astrophysics Data System (ADS)

    Ni, Jiajia; Yu, Yuhe; Zhang, Tanglin; Gao, Lei

    2012-09-01

    The intestinal bacteria of vertebrates form a close relationship with their host. External and internal conditions of the host, including its habitat, affect the intestinal bacterial community. Similarly, the intestinal bacterial community can, in turn, influence the host, particularly with respect to disease resistance. We compared the intestinal bacterial communities of grass carp that were collected from farm-ponds or a lake. We conducted denaturing gradient gel electrophoresis of amplified 16S rRNA genes, from which 66 different operational taxonomic units were identified. Using both the unweighted pair-group method with arithmetic means clustering and principal component analysis ordination, we found that the intestinal bacterial communities from the two groups of pond fish were clustered together and inset into the clusters of wild fish, except for DF-7, and there was no significant correlation between genetic diversity of grass carp and their intestinal bacterial communities (Mantel one-tailed test, R=0.157, P=0.175). Cetobacterium appeared more frequently in the intestine of grass carp collected from pond. A more thorough understanding of the role played by intestinal microbiota on fish health would be of considerable benefit to the aquaculture industry.

  1. A geometric description of human intestine.

    PubMed

    Coşkun, Ihsaniye; Yildiz, Hüseyin; Arslan, Kadri; Yildiz, Bahri

    2007-01-01

    Mathematical models of natural phenomena play a central role in the physical sciences. Moreover, modeling of the organs draws from some beautiful areas of mathematics, such as nonlinear dynamics, multiscale transforms and stability analysis. In this study, a geometric recognition of the separate intestine sections (duodenum, jejunum, ileum, cecum and colon) of the human is presented. The human intestine was considered a tubular shape along a special curve and two male Turkish men were used for the modeling study. The length (cm) and diameter (mm) of the intestines were measured with a digital compass and formulated. These models were compared with their original photographs. It has been concluded that the geometric modeling and experimental work were consistent. These kinds of organ modeling techniques will also profit to medical lecturers to show 3-D figures to their students.

  2. Transformation of trollioside and isoquercetin by human intestinal flora in vitro.

    PubMed

    Yuan, Ming; Shi, Duo-Zhi; Wang, Teng-Yu; Zheng, Shi-Qi; Liu, Li-Jia; Sun, Zhen-Xiao; Wang, Ru-Feng; Ding, Yi

    2016-03-01

    The present study was designed to determine the intestinal bacterial metabolites of trollioside and isoquercetin and their antibacterial activities. A systematic in vitro biotransformation investigation on trollioside and isoquercetin, including metabolite identification, metabolic pathway deduction, and time course, was accomplished using a human intestinal bacterial model. The metabolites were analyzed and identified by HPLC and HPLC-MS. The antibacterial activities of trollioside, isoquercetin, and their metabolites were evaluated using the broth microdilution method with berberine as a positive control, and their potency was measured as minimal inhibitory concentration (MIC). Our results indicated that trollioside and isoquercetin were metabolized by human intestinal flora through O-deglycosylation, yielding aglycones proglobeflowery acid and quercetin, respectively The antibacterial activities of both metabolites were more potent than that of their parent compounds. In conclusion, trollioside and isoquercetin are totally and rapidly transformed by human intestinal bacteria in vitro and the transformation favors the improvement of the antibacterial activities of the parent compounds.

  3. Human Enteroids/Colonoids and Intestinal Organoids Functionally Recapitulate Normal Intestinal Physiology and Pathophysiology.

    PubMed

    Zachos, Nicholas C; Kovbasnjuk, Olga; Foulke-Abel, Jennifer; In, Julie; Blutt, Sarah E; de Jonge, Hugo R; Estes, Mary K; Donowitz, Mark

    2016-02-19

    Identification of Lgr5 as the intestinal stem cell marker as well as the growth factors necessary to replicate adult intestinal stem cell division has led to the establishment of the methods to generate "indefinite" ex vivo primary intestinal epithelial cultures, termed "mini-intestines." Primary cultures developed from isolated intestinal crypts or stem cells (termed enteroids/colonoids) and from inducible pluripotent stem cells (termed intestinal organoids) are being applied to study human intestinal physiology and pathophysiology with great expectations for translational applications, including regenerative medicine. Here we discuss the physiologic properties of these cultures, their current use in understanding diarrhea-causing host-pathogen interactions, and potential future applications.

  4. Tipping elements in the human intestinal ecosystem

    PubMed Central

    Lahti, Leo; Salojärvi, Jarkko; Salonen, Anne; Scheffer, Marten; de Vos, Willem M.

    2014-01-01

    The microbial communities living in the human intestine can have profound impact on our well-being and health. However, we have limited understanding of the mechanisms that control this complex ecosystem. Here, based on a deep phylogenetic analysis of the intestinal microbiota in a thousand western adults, we identify groups of bacteria that exhibit robust bistable abundance distributions. These bacteria are either abundant or nearly absent in most individuals, and exhibit decreased temporal stability at the intermediate abundance range. The abundances of these bimodally distributed bacteria vary independently, and their abundance distributions are not affected by short-term dietary interventions. However, their contrasting alternative states are associated with host factors such as ageing and overweight. We propose that the bistable groups reflect tipping elements of the intestinal microbiota, whose critical transitions may have profound health implications and diagnostic potential. PMID:25003530

  5. Tipping elements in the human intestinal ecosystem.

    PubMed

    Lahti, Leo; Salojärvi, Jarkko; Salonen, Anne; Scheffer, Marten; de Vos, Willem M

    2014-07-08

    The microbial communities living in the human intestine can have profound impact on our well-being and health. However, we have limited understanding of the mechanisms that control this complex ecosystem. Here, based on a deep phylogenetic analysis of the intestinal microbiota in a thousand western adults, we identify groups of bacteria that exhibit robust bistable abundance distributions. These bacteria are either abundant or nearly absent in most individuals, and exhibit decreased temporal stability at the intermediate abundance range. The abundances of these bimodally distributed bacteria vary independently, and their abundance distributions are not affected by short-term dietary interventions. However, their contrasting alternative states are associated with host factors such as ageing and overweight. We propose that the bistable groups reflect tipping elements of the intestinal microbiota, whose critical transitions may have profound health implications and diagnostic potential.

  6. Contrasting Ecological Processes and Functional Compositions Between Intestinal Bacterial Community in Healthy and Diseased Shrimp.

    PubMed

    Zhu, Jinyong; Dai, Wenfang; Qiu, Qiongfen; Dong, Chunming; Zhang, Jinjie; Xiong, Jinbo

    2016-11-01

    Intestinal bacterial communities play a pivotal role in promoting host health; therefore, the disruption of intestinal bacterial homeostasis could result in disease. However, the effect of the occurrences of disease on intestinal bacterial community assembly remains unclear. To address this gap, we compared the multifaceted ecological differences in maintaining intestinal bacterial community assembly between healthy and diseased shrimps. The neutral model analysis shows that the relative importance of neutral processes decreases when disease occurs. This pattern is further corroborated by the ecosphere null model, revealing that the bacterial community assembly of diseased samples is dominated by stochastic processes. In addition, the occurrence of shrimp disease reduces the complexity and cooperative activities of species-to-species interactions. The keystone taxa affiliated with Alphaproteobacteria and Actinobacteria in healthy shrimp gut shift to Gammaproteobacteria species in diseased shrimp. Changes in intestinal bacterial communities significantly alter biological functions in shrimp. Within a given metabolic pathway, the pattern of enrichment or decrease between healthy and deceased shrimp is correlated with its functional effects. We propose that stressed shrimp are more prone to invasion by alien strains (evidenced by more stochastic assembly and higher migration rate in diseased shrimp), which, in turn, disrupts the cooperative activity among resident species. These findings greatly aid our understanding of the underlying mechanisms that govern shrimp intestinal community assembly between health statuses.

  7. The impact of hypoxia on intestinal epithelial cell functions: consequences for invasion by bacterial pathogens.

    PubMed

    Zeitouni, Nathalie E; Chotikatum, Sucheera; von Köckritz-Blickwede, Maren; Naim, Hassan Y

    2016-12-01

    The maintenance of oxygen homeostasis in human tissues is mediated by several cellular adaptations in response to low-oxygen stress, called hypoxia. A decrease in tissue oxygen levels is initially counteracted by increasing local blood flow to overcome diminished oxygenation and avoid hypoxic stress. However, studies have shown that the physiological oxygen concentrations in several tissues are much lower than atmospheric (normoxic) conditions, and the oxygen supply is finely regulated in individual cell types. The gastrointestinal tract has been described to subsist in a state of physiologically low oxygen level and is thus depicted as a tissue in the state of constant low-grade inflammation. The intestinal epithelial cell layer plays a vital role in the immune response to inflammation and infections that occur within the intestinal tissue and is involved in many of the adaptation responses to hypoxic stress. This is especially relevant in the context of inflammatory disorders, such as inflammatory bowel disease (IBD). Therefore, this review aims to describe the intestinal epithelial cellular response to hypoxia and the consequences for host interactions with invading gastrointestinal bacterial pathogens.

  8. Small bowel bacterial overgrowth

    MedlinePlus

    Overgrowth - intestinal bacteria; Bacterial overgrowth - intestine; Small intestinal bacterial overgrowth; SIBO ... intestine does not have a high number of bacteria. Excess bacteria in the small intestine may use ...

  9. Macropinocytosis in Shiga toxin 1 uptake by human intestinal epithelial cells and transcellular transcytosis.

    PubMed

    Malyukova, Irina; Murray, Karen F; Zhu, Chengru; Boedeker, Edgar; Kane, Anne; Patterson, Kathleen; Peterson, Jeffrey R; Donowitz, Mark; Kovbasnjuk, Olga

    2009-01-01

    Shiga toxin 1 and 2 production is a cardinal virulence trait of enterohemorrhagic Escherichia coli infection that causes a spectrum of intestinal and systemic pathology. However, intestinal sites of enterohemorrhagic E. coli colonization during the human infection and how the Shiga toxins are taken up and cross the globotriaosylceramide (Gb3) receptor-negative intestinal epithelial cells remain largely uncharacterized. We used samples of human intestinal tissue from patients with E. coli O157:H7 infection to detect the intestinal sites of bacterial colonization and characterize the distribution of Shiga toxins. We further used a model of largely Gb3-negative T84 intestinal epithelial monolayers treated with B-subunit of Shiga toxin 1 to determine the mechanisms of non-receptor-mediated toxin uptake. We now report that E. coli O157:H7 were found at the apical surface of epithelial cells only in the ileocecal valve area and that both toxins were present in large amounts inside surface and crypt epithelial cells in all tested intestinal samples. Our in vitro data suggest that macropinocytosis mediated through Src activation significantly increases toxin endocytosis by intestinal epithelial cells and also stimulates toxin transcellular transcytosis. We conclude that Shiga toxin is taken up by human intestinal epithelial cells during E. coli O157:H7 infection regardless of the presence of bacterial colonies. Macropinocytosis might be responsible for toxin uptake by Gb3-free intestinal epithelial cells and transcytosis. These observations provide new insights into the understanding of Shiga toxin contribution to enterohemorrhagic E. coli-related intestinal and systemic diseases.

  10. Characterisation of the bacterial community structures in the intestine of Lampetra morii.

    PubMed

    Li, Yingying; Xie, Wenfang; Li, Qingwei

    2016-07-01

    The metagenomic analysis and 16S rDNA sequencing method were used to investigate the bacterial community in the intestines of Lampetra morii. The bacterial community structure in L. morii intestine was relatively simple. Eight different operational taxonomic units were observed. Chitinophagaceae_unclassified (26.5 %) and Aeromonas spp. (69.6 %) were detected as dominant members at the genus level. The non-dominant genera were as follows: Acinetobacter spp. (1.4 %), Candidatus Bacilloplasma (2.5 %), Enterobacteria spp. (1.5 %), Shewanella spp. (0.04 %), Vibrio spp. (0.09 %), and Yersinia spp. (1.8 %). The Shannon-Wiener (H) and Simpson (1-D) indexes were 0.782339 and 0.5546, respectively. The rarefaction curve representing the bacterial community richness and Shannon-Wiener curve representing the bacterial community diversity reached asymptote, which indicated that the sequence depth were sufficient to represent the majority of species richness and bacterial community diversity. The number of Aeromonas in lamprey intestine was two times higher after stimulation by lipopolysaccharide than PBS. This study provides data for understanding the bacterial community harboured in lamprey intestines and exploring potential key intestinal symbiotic bacteria essential for the L. morii immune response.

  11. Inflammatory Bowel Diseases: When Natural Friends Turn into Enemies—The Importance of CpG Motifs of Bacterial DNA in Intestinal Homeostasis and Chronic Intestinal Inflammation

    PubMed Central

    Obermeier, Florian; Hofmann, Claudia; Falk, Werner

    2010-01-01

    From numerous studies during the last years it became evident that bacteria and bacterial constituents play a decisive role both in the maintenance of intestinal immune homeostasis as well as in the development and perpetuation of chronic intestinal inflammation. In this review we focus on the role of bacterial DNA which is a potent immunomodulatory component of the bacterial flora. Bacterial DNA has been shown to be protective against experimental colitis. In contrast bacterial DNA essentially contributes to the perpetuation of an already established chronic intestinal inflammation in a Toll-like receptor (TLR)9-dependent manner. This dichotomic action may be explained by a different activation status of essential regulators of TLR signaling like Glycogen synthase kinase 3-β (GSK3-β) depending on the pre-activation status of the intestinal immune system. In this review we suggest that regulators of TLR signaling may be interesting therapeutic targets in IBD aiming at the restoration of intestinal immune homeostasis. PMID:21188217

  12. Composition, Diversity, and Origin of the Bacterial Community in Grass Carp Intestine

    PubMed Central

    Wu, Shangong; Wang, Guitang; Angert, Esther R.; Wang, Weiwei; Li, Wenxiang; Zou, Hong

    2012-01-01

    Gut microbiota has become an integral component of the host, and received increasing attention. However, for many domestic animals, information on the microbiota is insufficient and more effort should be exerted to manage the gastrointestinal bacterial community. Understanding the factors that influence the composition of microbial community in the host alimentary canal is essential to manage or improve the microbial community composition. In the present study, 16S rRNA gene sequence-based comparisons of the bacterial communities in the grass carp (Ctenopharyngodon idellus) intestinal contents and fish culture-associated environments are performed. The results show that the fish intestinal microbiota harbors many cellulose-decomposing bacteria, including sequences related to Anoxybacillus, Leuconostoc, Clostridium, Actinomyces, and Citrobacter. The most abundant bacterial operational taxonomic units (OTUs) in the grass carp intestinal content are those related to feed digestion. In addition, the potential pathogens and probiotics are important members of the intestinal microbiota. Further analyses show that grass carp intestine holds a core microbiota composed of Proteobacteria, Firmicutes, and Actinobacteria. The comparison analyses reveal that the bacterial community in the intestinal contents is most similar to those from the culture water and sediment. However, feed also plays significant influence on the composition of gut microbiota. PMID:22363439

  13. Bacterial Population in Intestines of the Black Tiger Shrimp (Penaeus monodon) under Different Growth Stages

    PubMed Central

    Rungrassamee, Wanilada; Klanchui, Amornpan; Chaiyapechara, Sage; Maibunkaew, Sawarot; Tangphatsornruang, Sithichoke; Jiravanichpaisal, Pikul; Karoonuthaisiri, Nitsara

    2013-01-01

    Intestinal bacterial communities in aquaculture have been drawn to attention due to potential benefit to their hosts. To identify core intestinal bacteria in the black tiger shrimp (Penaeus monodon), bacterial populations of disease-free shrimp were characterized from intestines of four developmental stages (15-day-old post larvae (PL15), 1- (J1), 2- (J2), and 3-month-old (J3) juveniles) using pyrosequencing, real-time PCR and denaturing gradient gel electrophoresis (DGGE) approaches. A total of 25,121 pyrosequencing reads (reading length = 442±24 bases) were obtained, which were categorized by barcode for PL15 (7,045 sequences), J1 (3,055 sequences), J2 (13,130 sequences) and J3 (1,890 sequences). Bacteria in the phyla Bacteroides, Firmicutes and Proteobacteria were found in intestines at all four growth stages. There were 88, 14, 27, and 20 bacterial genera associated with the intestinal tract of PL15, J1, J2 and J3, respectively. Pyrosequencing analysis revealed that Proteobacteria (class Gammaproteobacteria) was a dominant bacteria group with a relative abundance of 89% for PL15 and 99% for J1, J2 and J3. Real-time PCR assay also confirmed that Gammaproteobacteria had the highest relative abundance in intestines from all growth stages. Intestinal bacterial communities from the three juvenile stages were more similar to each other than that of the PL shrimp based on PCA analyses of pyrosequencing results and their DGGE profiles. This study provides descriptive bacterial communities associated to the black tiger shrimp intestines during these growth development stages in rearing facilities. PMID:23577162

  14. Robust bioengineered 3D functional human intestinal epithelium

    PubMed Central

    Chen, Ying; Lin, Yinan; Davis, Kimberly M.; Wang, Qianrui; Rnjak-Kovacina, Jelena; Li, Chunmei; Isberg, Ralph R.; Kumamoto, Carol A.; Mecsas, Joan; Kaplan, David L.

    2015-01-01

    Intestinal functions are central to human physiology, health and disease. Options to study these functions with direct relevance to the human condition remain severely limited when using conventional cell cultures, microfluidic systems, organoids, animal surrogates or human studies. To replicate in vitro the tissue architecture and microenvironments of native intestine, we developed a 3D porous protein scaffolding system, containing a geometrically-engineered hollow lumen, with adaptability to both large and small intestines. These intestinal tissues demonstrated representative human responses by permitting continuous accumulation of mucous secretions on the epithelial surface, establishing low oxygen tension in the lumen, and interacting with gut-colonizing bacteria. The newly developed 3D intestine model enabled months-long sustained access to these intestinal functions in vitro, readily integrable with a multitude of different organ mimics and will therefore ensure a reliable ex vivo tissue system for studies in a broad context of human intestinal diseases and treatments. PMID:26374193

  15. Robust bioengineered 3D functional human intestinal epithelium.

    PubMed

    Chen, Ying; Lin, Yinan; Davis, Kimberly M; Wang, Qianrui; Rnjak-Kovacina, Jelena; Li, Chunmei; Isberg, Ralph R; Kumamoto, Carol A; Mecsas, Joan; Kaplan, David L

    2015-09-16

    Intestinal functions are central to human physiology, health and disease. Options to study these functions with direct relevance to the human condition remain severely limited when using conventional cell cultures, microfluidic systems, organoids, animal surrogates or human studies. To replicate in vitro the tissue architecture and microenvironments of native intestine, we developed a 3D porous protein scaffolding system, containing a geometrically-engineered hollow lumen, with adaptability to both large and small intestines. These intestinal tissues demonstrated representative human responses by permitting continuous accumulation of mucous secretions on the epithelial surface, establishing low oxygen tension in the lumen, and interacting with gut-colonizing bacteria. The newly developed 3D intestine model enabled months-long sustained access to these intestinal functions in vitro, readily integrable with a multitude of different organ mimics and will therefore ensure a reliable ex vivo tissue system for studies in a broad context of human intestinal diseases and treatments.

  16. Comparative analysis of the composition of intestinal bacterial communities in Dastarcus helophoroides fed different diets.

    PubMed

    Wang, Wei-Wei; He, Cai; Cui, Jun; Wang, Hai-Dong; Li, Meng-Lou

    2014-01-01

    The diversity of the intestinal bacterial communities in Dastarcus helophoroides (Fairmaire) (Coleoptera: Bothrideridae) larvae and adults was assayed by PCR-DGGE to determine whether different artificial diets could influence these bacterial communities. Two diets were used for feeding the larvae and four for the adults. Escherichia, Desemzia, Staphylococcus, Asticcacaulis, Cellvibrio, Aurantimonas, and Planomicrobium were isolated from the gut of the adults, with Escherichia and Staphylococcus being the main bacterial communities, and the quantities of intestinal bacterial were different in the adults fed different diets. Specifically, the amount of intestinal bacteria from the adults fed different diets had the following ranking according to the major component of the diet: ant powder > darkling beetle pupa powder > cricket powder > silkworm pupa powder. Escherichia, Bacillus, Staphylococcus, Kurthia, Planococcaceae, Ralstonia, Leptothrix, Acinetobacter, and Pseudomonas were isolated from the gut of the larvae. The quantity of intestinal bacteria from the larvae fed the darkling beetle pupae was greater than that from the larvae fed other artificial diets. This study, for the first time, investigated the effect of artificial diets on the bacterial community and the intestinal microbial diversity of D. helophoroides.

  17. Comparative Analysis of the Composition of Intestinal Bacterial Communities in Dastarcus helophoroides Fed Different Diets

    PubMed Central

    Wang, Wei-Wei; He, Cai; Cui, Jun; Wang, Hai-Dong; Li, Meng-Lou

    2014-01-01

    The diversity of the intestinal bacterial communities in Dastarcus helophoroides (Fairmaire) (Coleoptera: Bothrideridae) larvae and adults was assayed by PCR-DGGE to determine whether different artificial diets could influence these bacterial communities. Two diets were used for feeding the larvae and four for the adults. Escherichia, Desemzia, Staphylococcus, Asticcacaulis, Cellvibrio, Aurantimonas, and Planomicrobium were isolated from the gut of the adults, with Escherichia and Staphylococcus being the main bacterial communities, and the quantities of intestinal bacterial were different in the adults fed different diets. Specifically, the amount of intestinal bacteria from the adults fed different diets had the following ranking according to the major component of the diet: ant powder > darkling beetle pupa powder > cricket powder > silkworm pupa powder. Escherichia, Bacillus, Staphylococcus, Kurthia, Planococcaceae, Ralstonia, Leptothrix, Acinetobacter, and Pseudomonas were isolated from the gut of the larvae. The quantity of intestinal bacteria from the larvae fed the darkling beetle pupae was greater than that from the larvae fed other artificial diets. This study, for the first time, investigated the effect of artificial diets on the bacterial community and the intestinal microbial diversity of D. helophoroides. PMID:25199878

  18. Intestinal bacterial flora of the household lizard, Gecko gecko.

    PubMed

    Tan, R J; Lim, E W; Ishak, B

    1978-03-01

    A total of 114 isolates was recovered from the intestines of 43 househould lizards, Gecko gecko. Among the important ones were Staphylococcus aureus, Salmonella typhimurium, Pseudomonas aeruginosa, Proteus mirabilis and Edwardsiella tarda.

  19. Intestinal permeability and bacterial translocation following small bowel transplantation in the rat

    SciTech Connect

    Grant, D.; Hurlbut, D.; Zhong, R.; Wang, P.Z.; Chen, H.F.; Garcia, B.; Behme, R.; Stiller, C.; Duff, J. )

    1991-08-01

    In addition to its role in absorbing nutrients, the intestinal mucosa provides an important barrier against toxins and bacteria in the bowel lumen. The present study evaluated gut barrier function following orthotopic (in continuity) intestinal grafting in rats. Graft histology, intestinal permeability, and bacterial translocation to the grafted mesenteric lymph nodes, the host's liver, and the host's spleen were assessed on the 3rd, 5th, and 7th postoperative days. The study group received no immunosuppression after allotransplantation. The two control groups included rats with isografts and rats with cyclosporine-treated allografts. On the 7th POD, the study animals had moderate transmural inflammation due to rejection, with normal histology in the isografts and CsA-treated allografts; increased intestinal permeability, measured by urinary excretion of oral 51Cr-EDTA (P less than 0.01); and increased number of bacteria in the MLN and spleen (P less than 0.05). The number of bacteria in the MLN and spleen of the study group positively correlated with the changes in intestinal permeability (P less than 0.05). Rejection of the orthotopic intestinal graft leads to increased intestinal permeability and bacterial translocation from the lumen of the graft to the host's reticuloendothelial system. Measures to improve gut barrier function and antibiotic therapy during rejection episodes may help reduce the incidence of septic complications after intestinal grafting.

  20. Establishment of novel prediction system of intestinal absorption in humans using human intestinal tissues.

    PubMed

    Miyake, Masateru; Toguchi, Hajime; Nishibayashi, Toru; Higaki, Kazutaka; Sugita, Akira; Koganei, Kazutaka; Kamada, Nobuhiko; Kitazume, Mina T; Hisamatsu, Tadakazu; Sato, Toshiro; Okamoto, Susumu; Kanai, Takanori; Hibi, Toshifumi

    2013-08-01

    The objective of this study was to establish a novel prediction system of drug absorption in humans by utilizing human intestinal tissues. Based on the transport index (TI), a newly defined parameter, calculated by taking account of the change in drug concentrations because of precipitation on the apical side and the amounts accumulated in the tissue and transported to the basal side, the absorbability of drugs in rank order as well as the fraction of dose absorbed (Fa) in humans were estimated. Human intestinal tissues taken from ulcerative colitis or Crohn's disease patients were mounted in a mini-Ussing chamber and transport studies were performed to evaluate the permeation of drugs, including FD-4, a very low permeable marker, atenolol, a low permeable marker, and metoprolol, a high permeable marker. Although apparent permeability coefficients calculated by the conventional equation did not reflect human Fa values for FD-4, atenolol, and metoprolol, TI values were well correlated with Fa values, which are described by 100 · [1 - e (- f · (TI - α)) ]. Based on this equation, Fa values in humans for other test drugs were predicted successfully, indicating that our new system utilizing human intestinal tissues would be valuable for predicting oral drug absorption in humans.

  1. The Role of Milk Sialyllactose in Intestinal Bacterial Colonization123

    PubMed Central

    Weiss, G. Adrienne; Hennet, Thierry

    2012-01-01

    Milk oligosaccharides influence the composition of intestinal microbiota and thereby mucosal inflammation. Some of the major milk oligosaccharides are α2,3-sialyllactose (3SL) and α2,6-sialyllactose, which are mainly produced by the sialyltransferases ST3GAL4 and ST6GAL1, respectively. Recently, we showed that mice fed milk deficient in 3SL were more resistant to dextran sulfate sodium-induced colitis. By contrast, the exposure to milk containing or deficient in 3SL had no impact on the development of mucosal leukocyte populations. Milk 3SL mainly affected the colonization of the intestine by clostridial cluster IV bacteria. PMID:22585928

  2. A General O-Glycosylation System Important to the Physiology of a Major Human Intestinal Symbiont

    PubMed Central

    Fletcher, C. Mark; Coyne, Michael J.; Villa, Otto F.; Chatzidaki-Livanis, Maria; Comstock, Laurie E.

    2009-01-01

    SUMMARY The Bacteroides are a numerically dominant genus of the human intestinal microbiota. These organisms harbor a rare bacterial pathway for incorporation of exogenous fucose into capsular polysaccharides and glycoproteins. The infrequency of glycoprotein synthesis by bacteria prompted a more detailed analysis of this process. Here, we demonstrate that Bacteroides fragilis has a general O-glycosylation system. The proteins targeted for glycosylation include those predicted to be involved in protein folding, protein-protein interactions, peptide degradation, as well as surface lipoproteins. Protein glycosylation is central to the physiology of B. fragilis and is necessary for the organism to competitively colonize the mammalian intestine. We provide evidence that general O-glycosylation systems are conserved among intestinal Bacteroides species and likely contribute to the predominance of Bacteroides in the human intestine. PMID:19379697

  3. Plant innate immunity against human bacterial pathogens

    PubMed Central

    Melotto, Maeli; Panchal, Shweta; Roy, Debanjana

    2014-01-01

    Certain human bacterial pathogens such as the enterohemorrhagic Escherichia coli and Salmonella enterica are not proven to be plant pathogens yet. Nonetheless, under certain conditions they can survive on, penetrate into, and colonize internal plant tissues causing serious food borne disease outbreaks. In this review, we highlight current understanding on the molecular mechanisms of plant responses against human bacterial pathogens and discuss salient common and contrasting themes of plant interactions with phytopathogens or human pathogens. PMID:25157245

  4. Human milk oligosaccharides: the novel modulator of intestinal microbiota.

    PubMed

    Jeong, Kyunghun; Nguyen, Vi; Kim, Jaehan

    2012-08-01

    Human milk, which nourishes the early infants, is a source of bioactive components for the infant growth, development and commensal formulation as well. Human milk oligosaccharide is a group of complex and diverse glycans that is apparently not absorbed in human gastrointestinal tract. Although most mammalian milk contains oligosaccharides, oligosaccharides in human milk exhibit unique features in terms of their types, amounts, sizes, and functionalities. In addition to the prevention of infectious bacteria and the development of early immune system, human milk oligosaccharides are able to facilitate the healthy intestinal microbiota. Bifidobacterial intestinal microbiota appears to be established by the unilateral interaction between milk oligosaccharides, human intestinal activity and commensals. Digestibility, membrane transportation and catabolic activity by bacteria and intestinal epithelial cells, all of which are linked to the structural of human milk oligosaccharides, are crucial in determining intestinal microbiota.

  5. Fermented liquid feed enhances bacterial diversity in piglet intestine.

    PubMed

    Tajima, Kiyoshi; Ohmori, Hideyuki; Aminov, Rustam I; Kobashi, Yuri; Kawashima, Tomoyuki

    2010-02-01

    Because of limitations imposed on the antibiotic use in animal industry, there is a need for alternatives to maintain the efficiency of production. One of them may be the use of fermented liquid feed (FLF) but how it affects gut ecology is poorly understood. We investigated the effect of three diets, standard dry feed (control), dry feed supplemented with antibiotics, and fermented liquid feed (FLF, fermented with Lactobacillus plantarum), on gut bacterial diversity in piglets. The structure of the ileal and caecal communities was estimated by sequencing the SSU rRNA gene libraries. Antibiotic-supplemented feed slightly increased bacterial diversity in the ileum but reduced it in the caecum while in FLF-fed animals bacterial diversity was elevated. The majority of bacterial sequences in the ileum of all three groups belonged to lactobacilli (92-98%). In the caecum the lactobacilli were still dominant in control and antibiotic-fed animals (59% and 64% of total bacterial sequences, respectively) but in FLF-fed animals they fell to 31% with the concomitant increase in the Firmicutes diversity represented by the Dorea, Coprococcus, Roseburia and Faecalibacterium genera. Thus FLF affects the gut ecology in a different way than antibiotics and contributes to the enhanced bacterial diversity in the gastrointestinal tract.

  6. The intestinal bacterial community in the food waste-reducing larvae of Hermetia illucens.

    PubMed

    Jeon, Hyunbum; Park, Soyoung; Choi, Jiyoung; Jeong, Gilsang; Lee, Sang-Beom; Choi, Youngcheol; Lee, Sung-Jae

    2011-05-01

    As it is known that food waste can be reduced by the larvae of Hermetia illucens (Black soldier fly, BSF), the scientific and commercial value of BSF larvae has increased recently. We hypothesised that the ability of catabolic degradation by BSF larvae might be due to intestinal microorganisms. Herein, we analysed the bacterial communities in the gut of BSF larvae by pyrosequencing of extracting intestinal metagenomic DNA from larvae that had been fed three different diets. The 16S rRNA sequencing results produced 9737, 9723 and 5985 PCR products from larval samples fed food waste, cooked rice and calf forage, respectively. A BLAST search using the EzTaxon program showed that the bacterial community in the gut of larvae fed three different diets was mainly composed of the four phyla with dissimilar proportions. Although the composition of the bacterial communities depended on the different nutrient sources, the identified bacterial strains in the gut of BSF larvae represented unique bacterial species that were unlike the intestinal microflora of other insects. Thus, our study analysed the structure of the bacterial communities in the gut of BSF larvae after three different feedings and assessed the application of particular bacteria for the efficient degradation of organic compounds.

  7. Giardia duodenalis induces pathogenic dysbiosis of human intestinal microbiota biofilms.

    PubMed

    Beatty, Jennifer K; Akierman, Sarah V; Motta, Jean-Paul; Muise, Stacy; Workentine, Matthew L; Harrison, Joe J; Bhargava, Amol; Beck, Paul L; Rioux, Kevin P; McKnight, Gordon Webb; Wallace, John L; Buret, Andre G

    2017-02-22

    Giardia duodenalis is a prevalent cause of acute diarrheal disease worldwide. However, recent outbreaks in Italy and Norway have revealed a link between giardiasis and the subsequent development of chronic post-infectious irritable bowel syndrome. While the mechanisms underlying the causation of post-infectious irritable bowel syndrome remain obscure, recent findings suggest that alterations in gut microbiota communities are linked to the pathophysiology of irritable bowel syndrome. In the present study, we use a laboratory biofilm system to culture and enrich mucosal microbiota from human intestinal biopsies. Subsequently, we show that co-culture with Giardia induces disturbances in biofilm species composition and biofilm structure resulting in microbiota communities that are intrinsically dysbiotic - even after the clearance of Giardia. These microbiota abnormalities were mediated in part by secretory-excretory Giardia cysteine proteases. Using in vitro cell culture and germ-free murine infection models, we show that Giardia-induced disruptions of microbiota promote bacterial invasion, resulting in epithelial apoptosis, tight junctional disruption, and bacterial translocation across an intestinal epithelial barrier. Additionally, these dysbiotic microbiota communities resulted in increased activation of the Toll-like receptor 4 signalling pathway, and overproduction of the pro-inflammatory cytokine IL-1beta in humanized germ-free mice. Previous studies that have sought explanations and risk factors for the development of post-infectious irritable bowel syndrome have focused on features of enteropathogens and attributes of the infected host. We propose that polymicrobial interactions involving Giardia and gut microbiota may cause persistent dysbiosis, offering a new interpretation of the reasons why those afflicted with giardiasis are predisposed to gastrointestinal disorders post-infection.

  8. Immunohistochemical detection of human intestinal spirochetosis.

    PubMed

    Ogata, Sho; Shimizu, Ken; Oda, Tomohiro; Tominaga, Susumu; Nakanishi, Kuniaki

    2016-12-01

    Human intestinal spirochetosis (HIS) is a colorectal infection by Brachyspira species of spiral bacteria. Immunohistochemical cross-reaction to an antibody for Treponema pallidum aids its histologic diagnosis. This study's aim was to analyze the immunohistochemical characteristics of HIS. In this analysis, on 223 specimens from 83 HIS cases, we focused on so-called fringe formation (a histologic hallmark of HIS), spiral organisms within mucus or within crypts, and strong immunopositive materials in the mucosa, together with their location and the types of lesions. Fringe formation was found in 81.6% of all specimens and spiral organisms within mucus or within crypts in 97.3% and 57.0%, respectively. Strong immunopositive materials were observed in the surface epithelial layer in 87.9%, in the subepithelial layer in 94.6%, and in deeper mucosa in 2.2% of all specimens. The positive rates in conventional adenomas (24.0%, n = 146) and hyperplastic nodules (100%, n = 17) were each different from that found in inflammation (70.8%, n = 24), and spiral organisms were seen more frequently in the right-side large intestine than in the left (within mucus, 100%, n = 104 versus 95.0%, n = 119; within crypts, 65.4%, n = 104 versus 49.6%, n = 119). Thus, immunohistochemistry was effective not only in supporting the diagnosis of HIS but also in highlighting spiral organisms within mucus or crypts that were invisible in routine histology. Possibly, these spiral organisms may spread throughout the entire large intestine, although there is a potential problem with antibody specificity.

  9. Bacterial diversity in the intestine of young farmed puffer fish Takifugu rubripes

    NASA Astrophysics Data System (ADS)

    Li, Yanyu; Zhang, Tao; Zhang, Congyao; Zhu, Ying; Ding, Jianfeng; Ma, Yuexin

    2015-07-01

    The aim of the study was to examine the bacterial community associated with the intestinal mucus of young farmed puffer fish Takifugu rubripes. Polymerase chain reaction and partial 16S rDNA sequencing was performed on DNA from bacteria cultivated on Zobell 2216E medium. All the isolates were classified into two phyla—Proteobacteria and Firmicutes. Proteobacteria were the dominant, culturable intestinal microbiota (68.3%). At the genus level, Vibrio, Enterobacter, Bacillus, Pseudomonas, Exiguobacterium, Staphylococcus, Acinetobacter, Pseudoalteromonas and Shewanella were isolated from the intestine, with representatives of the genera Vibrio, Enterobacter and Bacillus accounting for 70.7% of the total. This is the first report of Enterobacter, Bacillus, Exiguobacterium and Staphylococcus as part of the intestinal bacterial microflora in T. rubripes. The profile of the culturable bacterial community differed between samples collected from the same tank at 2-month intervals, as indicated by Bray-Curtis and Sorensen indices, and the impact on the intestinal physiology and health of puffer fish requires further investigation.

  10. Human intestinal lipoproteins. Studies in chyluric subjects.

    PubMed

    Green, P H; Glickman, R M; Saudek, C D; Blum, C B; Tall, A R

    1979-07-01

    To explore the role of the human intestine as a source of apolipoproteins, we have studied intestinal lipoproteins and apoprotein secretion in two subjects with chyluria (mesenteric lymphatic-urinary fistulae). After oral corn oil, apolipoprotein A-I (apoA-I) and apolipoprotein A-II (apoA-II) output in urine increased in parallel to urinary triglyceride. One subject, on two occasions, after 40 g of corn oil, excreted 8.4 and 8.6 g of triglyceride together with 196 and 199 mg apoA-I and on one occasion, 56 mg apoA-II. The other subject, after 40 g corn oil, excreted 0.3 g triglyceride and 17.5 mg apoA-I, and, after 100 g of corn oil, excreted 44.8 mg apoA-I and 5.8 mg apoA-II. 14.5+/-2.1% of apoA-I and 17.7+/-4.3% of apoA-II in chylous urine was in the d < 1.006 fraction (chylomicrons and very low density lipoprotein). Calculations based on the amount of apoA-I and apoA-II excreted on triglyceride-rich lipoproteins revealed that for these lipid loads, intestinal secretion could account for 50 and 33% of the calculated daily synthetic rate of apoA-I and apoA-II, respectively. Similarly, subject 2 excreted 48-70% and 14% of the calculated daily synthetic rate of apoA-I and apoA-II, respectively. Chylous urine contained chylomicrons, very low density lipoproteins and high density lipoproteins, all of which contained apoA-I. Chylomicrons and very low density lipoproteins contained a previously unreported human apoprotein of 46,000 mol wt. We have called this apoprotein apoA-IV because of the similarity of its molecular weight and amino acid composition to rat apoA-IV. In sodium dodecyl sulfate gels, chylomicron apoproteins consisted of apoB 3.4+/-0.7%, apoA-IV 10.0+/-3.3%, apoE 4.4+/-0.3%, apoA-I 15.0+/-1.8%, and apoC and apoA-II 43.3+/-11.3%. Very low density lipoprotein contained more apoB and apoA-IV and less apoC than chylomicrons. Ouchterlony immunodiffusion of chylomicron apoproteins revealed the presence of apoC-I, apoC-II, and apoC-III. In contrast, plasma

  11. The Human Vaginal Bacterial Biota and Bacterial Vaginosis

    PubMed Central

    Srinivasan, Sujatha; Fredricks, David N.

    2008-01-01

    The bacterial biota of the human vagina can have a profound impact on the health of women and their neonates. Changes in the vaginal microbiota have been associated with several adverse health outcomes including premature birth, pelvic inflammatory disease, and acquisition of HIV infection. Cultivation-independent molecular methods have provided new insights regarding bacterial diversity in this important niche, particularly in women with the common condition bacterial vaginosis (BV). PCR methods have shown that women with BV have complex communities of vaginal bacteria that include many fastidious species, particularly from the phyla Bacteroidetes and Actinobacteria. Healthy women are mostly colonized with lactobacilli such as Lactobacillus crispatus, Lactobacillus jensenii, and Lactobacillus iners, though a variety of other bacteria may be present. The microbiology of BV is heterogeneous. The presence of Gardnerella vaginalis and Atopobium vaginae coating the vaginal epithelium in some subjects with BV suggests that biofilms may contribute to this condition. PMID:19282975

  12. In vitro activity of rifaximin against isolates from patients with small intestinal bacterial overgrowth.

    PubMed

    Pistiki, Aikaterini; Galani, Irene; Pyleris, Emmanouel; Barbatzas, Charalambos; Pimentel, Mark; Giamarellos-Bourboulis, Evangelos J

    2014-03-01

    Rifaximin, a non-absorbable rifamycin derivative, has published clinical efficacy in the alleviation of symptoms in patients with irritable bowel syndrome (IBS). Small intestinal bacterial overgrowth (SIBO) is associated with the pathogenesis of IBS. This study describes for the first time the antimicrobial effect of rifaximin against SIBO micro-organisms from humans. Fluid was aspirated from the third part of the duodenum from 567 consecutive patients; quantitative cultures diagnosed SIBO in 117 patients (20.6%). A total of 170 aerobic micro-organisms were isolated and the in vitro efficacy of rifaximin was studied by (i) minimum inhibitory concentration (MIC) testing by a microdilution technique and (ii) time-kill assays using bile to simulate the small intestinal environment. At a breakpoint of 32 μg/mL, rifaximin inhibited in vitro 85.4% of Escherichia coli, 43.6% of Klebsiella spp., 34.8% of Enterobacter spp., 54.5% of other Enterobacteriaceae spp., 82.6% of non-Enterobacteriaceae Gram-negative spp., 100% of Enterococcus faecalis, 100% of Enterococcus faecium and 100% of Staphylococcus aureus. For the time-kill assays, 11 E. coli, 15 non-E. coli Gram-negative enterobacteria and three E. faecalis isolates were studied. Rifaximin produced a >3 log10 decrease in the starting inoculum against most of the tested isolates at 500 μg/mL after 24h of growth. The results indicate that rifaximin has a potent effect on specific small bowel flora associated with SIBO. This conclusion should be regarded in light of the considerable time-kill effect at concentrations lower than those achieved in the bowel lumen after administration of conventional doses in humans.

  13. Bacterial dynamics in intestines of the black tiger shrimp and the Pacific white shrimp during Vibrio harveyi exposure.

    PubMed

    Rungrassamee, Wanilada; Klanchui, Amornpan; Maibunkaew, Sawarot; Karoonuthaisiri, Nitsara

    2016-01-01

    The intestinal microbiota play important roles in health of their host, contributing to maintaining the balance and resilience against pathogen. To investigate effects of pathogen to intestinal microbiota, the bacterial dynamics upon a shrimp pathogen, Vibrio harveyi, exposures were determined in two economically important shrimp species; the black tiger shrimp (BT) and the Pacific white shrimp (PW). Both shrimp species were reared under the same diet and environmental conditions. Shrimp survival rates after the V. harveyi exposure revealed that the PW shrimp had a higher resistance to the pathogen than the BT shrimp. The intestinal bacterial profiles were determined by denaturing gradient gel electrophoresis (DGGE) and barcoded pyrosequencing of the 16S rRNA sequences under no pathogen challenge control and under pathogenic V. harveyi challenge. The DGGE profiles showed that the presence of V. harveyi altered the intestinal bacterial patterns in comparison to the control in BT and PW intestines. This implies that bacterial balance in shrimp intestines was disrupted in the presence of V. harveyi. The barcoded pyrosequencing analysis showed the similar bacterial community structures in intestines of BT and PW shrimp under a normal condition. However, during the time course exposure to V. harveyi, the relative abundance of bacteria belong to Vibrio genus was higher in the BT intestines at 12h after the exposure, whereas relative abundance of vibrios was more stable in PW intestines. The principle coordinates analysis based on weighted-UniFrac analysis showed that intestinal bacterial population in the BT shrimp lost their ability to restore their bacterial balance during the 72-h period of exposure to the pathogen, while the PW shrimp were able to reestablish their bacterial population to resemble those seen in the unexposed control group. This observation of bacterial disruption might correlate to different mortality rates observed between the two shrimp species

  14. Generation of tissue-engineered small intestine using embryonic stem cell-derived human intestinal organoids

    PubMed Central

    Finkbeiner, Stacy R.; Freeman, Jennifer J.; Wieck, Minna M.; El-Nachef, Wael; Altheim, Christopher H.; Tsai, Yu-Hwai; Huang, Sha; Dyal, Rachel; White, Eric S.; Grikscheit, Tracy C.; Teitelbaum, Daniel H.; Spence, Jason R.

    2015-01-01

    ABSTRACT Short bowel syndrome (SBS) is characterized by poor nutrient absorption due to a deficit of healthy intestine. Current treatment practices rely on providing supportive medical therapy with parenteral nutrition; while life saving, such interventions are not curative and are still associated with significant co-morbidities. As approaches to lengthen remaining intestinal tissue have been met with only limited success and intestinal transplants have poor survival outcomes, new approaches to treating SBS are necessary. Human intestine derived from embryonic stem cells (hESCs) or induced pluripotent stem cells (iPSCs), called human intestinal organoids (HIOs), have the potential to offer a personalized and scalable source of intestine for regenerative therapies. However, given that HIOs are small three-dimensional structures grown in vitro, methods to generate usable HIO-derived constructs are needed. We investigated the ability of hESCs or HIOs to populate acellular porcine intestinal matrices and artificial polyglycolic/poly L lactic acid (PGA/PLLA) scaffolds, and examined the ability of matrix/scaffolds to thrive when transplanted in vivo. Our results demonstrate that the acellular matrix alone is not sufficient to instruct hESC differentiation towards an endodermal or intestinal fate. We observed that while HIOs reseed acellular porcine matrices in vitro, the HIO-reseeded matrices do not thrive when transplanted in vivo. In contrast, HIO-seeded PGA/PLLA scaffolds thrive in vivo and develop into tissue that looks nearly identical to adult human intestinal tissue. Our results suggest that HIO-seeded PGA/PLLA scaffolds are a promising avenue for developing the mucosal component of tissue engineered human small intestine, which need to be explored further to develop them into fully functional tissue. PMID:26459240

  15. Generation of tissue-engineered small intestine using embryonic stem cell-derived human intestinal organoids.

    PubMed

    Finkbeiner, Stacy R; Freeman, Jennifer J; Wieck, Minna M; El-Nachef, Wael; Altheim, Christopher H; Tsai, Yu-Hwai; Huang, Sha; Dyal, Rachel; White, Eric S; Grikscheit, Tracy C; Teitelbaum, Daniel H; Spence, Jason R

    2015-10-12

    Short bowel syndrome (SBS) is characterized by poor nutrient absorption due to a deficit of healthy intestine. Current treatment practices rely on providing supportive medical therapy with parenteral nutrition; while life saving, such interventions are not curative and are still associated with significant co-morbidities. As approaches to lengthen remaining intestinal tissue have been met with only limited success and intestinal transplants have poor survival outcomes, new approaches to treating SBS are necessary. Human intestine derived from embryonic stem cells (hESCs) or induced pluripotent stem cells (iPSCs), called human intestinal organoids (HIOs), have the potential to offer a personalized and scalable source of intestine for regenerative therapies. However, given that HIOs are small three-dimensional structures grown in vitro, methods to generate usable HIO-derived constructs are needed. We investigated the ability of hESCs or HIOs to populate acellular porcine intestinal matrices and artificial polyglycolic/poly L lactic acid (PGA/PLLA) scaffolds, and examined the ability of matrix/scaffolds to thrive when transplanted in vivo. Our results demonstrate that the acellular matrix alone is not sufficient to instruct hESC differentiation towards an endodermal or intestinal fate. We observed that while HIOs reseed acellular porcine matrices in vitro, the HIO-reseeded matrices do not thrive when transplanted in vivo. In contrast, HIO-seeded PGA/PLLA scaffolds thrive in vivo and develop into tissue that looks nearly identical to adult human intestinal tissue. Our results suggest that HIO-seeded PGA/PLLA scaffolds are a promising avenue for developing the mucosal component of tissue engineered human small intestine, which need to be explored further to develop them into fully functional tissue.

  16. Intestinal bacterial community and growth performance of chickens fed diets containing antibiotics.

    PubMed

    Pedroso, A A; Menten, J F M; Lambais, M R; Racanicci, A M C; Longo, F A; Sorbara, J O B

    2006-04-01

    This study was conducted to relate the performance of broiler chickens fed diets containing growth-promoting antibiotics to changes in the intestinal microbiota. The technique of denaturing gradient gel electrophoresis (DGGE) of amplicons of the region V3 of 16S rDNA was used to characterize the microbiota. Two experiments were conducted, one with broilers raised in battery cages and the other with broilers raised in floor pens. Antibiotics improved the performance of the chickens raised in floor pens only. Avilamycin, bacitracin methylene disalicylate, and enramycin induced changes in the composition of the intestinal bacterial community of the birds in both experiments. The number of bacterial genotypes found in the intestinal tract of chickens was not reduced by the antibiotics supplemented in either environment. However, the changes in the composition of the intestinal bacterial community induced by antibiotics may be related to improvement in growth performance. This was indicated by the suppression of 6 amplicons and the presence of 4 amplicons exclusive to the treatment that had the best performance in the floor pen experiment.

  17. The effects of intestinal tract bacterial diversity on mortality following allogeneic hematopoietic stem cell transplantation

    PubMed Central

    Jenq, Robert R.; Perales, Miguel-Angel; Littmann, Eric R.; Morjaria, Sejal; Ling, Lilan; No, Daniel; Gobourne, Asia; Viale, Agnes; Dahi, Parastoo B.; Ponce, Doris M.; Barker, Juliet N.; Giralt, Sergio; van den Brink, Marcel; Pamer, Eric G.

    2014-01-01

    Highly diverse bacterial populations inhabit the gastrointestinal tract and modulate host inflammation and promote immune tolerance. In allogeneic hematopoietic stem cell transplantation (allo-HSCT), the gastrointestinal mucosa is damaged, and colonizing bacteria are impacted, leading to an impaired intestinal microbiota with reduced diversity. We examined the impact of intestinal diversity on subsequent mortality outcomes following transplantation. Fecal specimens were collected from 80 recipients of allo-HSCT at the time of stem cell engraftment. Bacterial 16S rRNA gene sequences were characterized, and microbial diversity was estimated using the inverse Simpson index. Subjects were classified into high, intermediate, and low diversity groups and assessed for differences in outcomes. Mortality outcomes were significantly worse in patients with lower intestinal diversity; overall survival at 3 years was 36%, 60%, and 67% for low, intermediate, and high diversity groups, respectively (P = .019, log-rank test). Low diversity showed a strong effect on mortality after multivariate adjustment for other clinical predictors (transplant related mortality: adjusted hazard ratio, 5.25; P = .014). In conclusion, the diversity of the intestinal microbiota at engraftment is an independent predictor of mortality in allo-HSCT recipients. These results indicate that the intestinal microbiota may be an important factor in the success or failure in allo-HSCT. PMID:24939656

  18. A Revised Model for Dosimetry in the Human Small Intestine

    SciTech Connect

    John Poston; Nasir U. Bhuiyan; R. Alex Redd; Neil Parham; Jennifer Watson

    2005-02-28

    A new model for an adult human gastrointestinal tract (GIT) has been developed for use in internal dose estimations to the wall of the GIT and to the other organs and tissues of the body from radionuclides deposited in the lumenal contents of the five sections of the GIT. These sections were the esophasgus, stomach, small intestine, upper large intestine, and the lower large intestine. The wall of each section was separated from its lumenal contents.

  19. Deoxycholic acid formation in gnotobiotic mice associated with human intestinal bacteria.

    PubMed

    Narushima, Seiko; Itoha, Kikuji; Miyamoto, Yukiko; Park, Sang-Hee; Nagata, Keiko; Kuruma, Kazuo; Uchida, Kiyohisa

    2006-09-01

    In humans and animals, intestinal flora is indispensable for bile acid transformation. The goal of our study was to establish gnotobiotic mice with intestinal bacteria of human origin in order to examine the role of intestinal bacteria in the transformation of bile acids in vivo using the technique of gnotobiology. Eight strains of bile acid-deconjugating bacteria were isolated from ex-germ-free mice inoculated with a human fecal dilution of 10(-6), and five strains of 7alpha-dehydroxylating bacteria were isolated from the intestine of limited human flora mice inoculated only with clostridia. The results of biochemical tests and 16S rDNA sequence analysis showed that seven out of eight bile acid-deconjugating strains belong to a bacteroides cluster (Bacteroides vulgatus, B. distasonis, and B. uniformis), and one strain had high similarity with Bilophila wadsworthia. All five strains that converted cholic acid to deoxycholic acid had greatest similarity with Clostridium hylemonae. A combination of 10 isolated strains converted taurocholic acid into deoxycholic acid both in vitro and in the mouse intestine. These results indicate that the predominant bacteria, mainly Bacteroides, in human feces comprise one of the main bacterial groups for the deconjugation of bile acids, and clostridia may play an important role in 7aplha-dehydroxylation of free-form primary bile acids in the intestine although these strains are not predominant. The gnotobiotic mouse with bacteria of human origin could be a useful model in studies of bile acid metabolism by human intestinal bacteria in vivo.

  20. Exploring food effects on indinavir absorption with human intestinal fluids in the mouse intestine.

    PubMed

    Holmstock, Nico; De Bruyn, Tom; Bevernage, Jan; Annaert, Pieter; Mols, Raf; Tack, Jan; Augustijns, Patrick

    2013-04-11

    Food can have a significant impact on the pharmacokinetics of orally administered drugs, as it may affect drug solubility as well as permeability. Since fed state conditions cannot easily be implemented in the presently available permeability tools, including the frequently used Caco-2 system, exploring food effects during drug development can be quite challenging. In this study, we investigated the effect of fasted and fed state conditions on the intestinal absorption of the HIV protease inhibitor indinavir using simulated and human intestinal fluids in the in situ intestinal perfusion technique in mice. Although the solubility of indinavir was 6-fold higher in fed state human intestinal fluids (FeHIF) as compared to fasted state HIF (FaHIF), the intestinal permeation of indinavir was 22-fold lower in FeHIF as compared to FaHIF. Dialysis experiments showed that only a small fraction of indinavir is accessible for absorption in FeHIF due to micellar entrapment, possibly explaining its low intestinal permeation. The presence of ritonavir, a known P-gp inhibitor, increased the intestinal permeation of indinavir by 2-fold in FaHIF, while there was no increase when using FeHIF. These data confirm that drug-food interactions form a complex interplay between solubility and permeability effects. The use of HIF in in situ intestinal perfusions holds great promise for biorelevant absorption evaluation as it allows to directly explore this complex solubility/permeability interplay on drug absorption.

  1. Identification of Population Bottlenecks and Colonization Factors during Assembly of Bacterial Communities within the Zebrafish Intestine

    PubMed Central

    Stephens, W. Zac; Wiles, Travis J.; Martinez, Emily S.; Jemielita, Matthew; Burns, Adam R.; Parthasarathy, Raghuveer; Bohannan, Brendan J. M.

    2015-01-01

    ABSTRACT The zebrafish, Danio rerio, is a powerful model for studying bacterial colonization of the vertebrate intestine, but the genes required by commensal bacteria to colonize the zebrafish gut have not yet been interrogated on a genome-wide level. Here we apply a high-throughput transposon mutagenesis screen to Aeromonas veronii Hm21 and Vibrio sp. strain ZWU0020 during their colonization of the zebrafish intestine alone and in competition with each other, as well as in different colonization orders. We use these transposon-tagged libraries to track bacterial population sizes in different colonization regimes and to identify gene functions required during these processes. We show that intraspecific, but not interspecific, competition with a previously established bacterial population greatly reduces the ability of these two bacterial species to colonize. Further, using a simple binomial sampling model, we show that under conditions of interspecific competition, genes required for colonization cannot be identified because of the population bottleneck experienced by the second colonizer. When bacteria colonize the intestine alone or at the same time as the other species, we find shared suites of functional requirements for colonization by the two species, including a prominent role for chemotaxis and motility, regardless of the presence of another species. PMID:26507229

  2. [Bacterial community structure in intestine of the white shrimp, Litopenaeus vannamei].

    PubMed

    Li, Ke; Zheng, Tian-ling; Tian, Yun; Yuan, Jian-jun

    2007-08-01

    The composition of bacterial community in the intestine of the white shrimp, Litopenaeus vannamei under laboratory culture condition was determined using the 16S rDNA clone library. 16s rRNA gene was amplified and a library was constructed by using the genomic DNA extracted from the bacteria in the shrimp intestine as template. 12 different RFLP patterns of the clones were obtained by restriction fragment length polymorphism analysis using Afa I and Msp I. Compared with the published sequences in GenBank database, sequencing results of cloned 16S rDNA amplicons revealed a diverse community including gamma-proteobacteria and Firmicutes in the intestine of artificial diet-fed shrimp. Results showed that the Firmicutes group can be a dominant component (75.4%) in the shrimp intestinal microflora and other clones belong to gamma-proteobacteria (24.6%) which were identified as Shewanella sp., Pantoea sp., Aranicola sp., Pseudomonas sp. and Vibrio sp., respectively. These results provide the first comprehensive description of microbial diversity of the white shrimp intestine and suggest that most of the bacteria associated with shrimp intestine are uncultured and novel species.

  3. R1: Immunohistochemical study of mucins in human intestinal spirochetosis.

    PubMed

    Ogata, Sho; Shimizu, Ken; Tominaga, Susumu; Nakanishi, Kuniaki

    2017-02-08

    Most patients with human intestinal spirochetosis (HIS; a colorectal bacterial infection caused by Brachyspira species) seem asymptomatic, and its pathogenicity remains unclear. Recently, alterations in mucin expression were reported in animal Brachyspira infection. The present question was "Is mucin expression altered in HIS?". Using antibodies for MUCs 1, 2, 4, 5 AC, and 6, we immunohistochemically compared 215 specimens from 83 histology-confirmed HIS cases with 106 specimens from 26 non-HIS cases. Positive staining (which included even focal positive staining) was rated "high (+)" or "low (+)". Results were analysed for four categories of lesions, and associations between MUC expression and spirochetal presence were also analysed. In the "specimens without polyps or adenocarcinoma" category: high (+) MUC2-positivity was more frequent in HIS than in control. In the hyperplasia/serrated polyp category: in HIS (vs. control), the MUC5AC-positivity rate was lower, while high (+) MUC4-positivity was more frequent. In the conventional adenoma category: in HIS (vs. control), the MUC1-positivity rate was lower, while both high (+) MUC2-positivity and high (+) MUC5AC-positivity were less frequent. In the adenocarcinoma category: high (+) MUC2-positivity was more frequent in HIS than in control. Among the above mucins, only MUC1-positivity was significantly associated with an absence of the so-called fringe formation, an absence of spiral organisms within mucus, and an absence of strong immunopositive materials within the epithelial layer and within the subepithelial layer. The results suggest that Brachyspira infection or a related change in the microbiome may alter the large intestine mucin-expression profile in humans.

  4. Transgenic milk containing recombinant human lactoferrin modulates the intestinal flora in piglets.

    PubMed

    Hu, Wenping; Zhao, Jie; Wang, Jianwu; Yu, Tian; Wang, Jing; Li, Ning

    2012-06-01

    Lactoferrin (LF) is a beneficial multifunctional protein in milk. The objective of this study was to determine whether bovine transgenic milk containing recombinant human lactoferrin (rhLF) can modulate intestinal flora in the neonatal pig as an animal model for the human infant. We fed 7-day-old piglets (i) ordinary whole milk (OM), (ii) a 1:1 mixture of OM and rhLF milk (MM), or (iii) rhLF milk (LFM). LFM provided better average daily mass gain than OM (P = 0.007). PCR-denaturing gradient gel electrophoresis and 16S rDNA sequencing analysis revealed that the LFM piglets exhibited more diversity of the intestinal flora than the OM group. Except for the colon in the LFM group, an increasing trend in microbial diversity occurred from the duodenum to the colon. Fecal flora was not different across different ages or different treatment groups, but a cluster analysis showed that the fecal flora of OM- and MM-fed piglets had a higher degree of similarity than that of LFM-fed piglets. Based on culture-based bacterial counts of intestinal content samples, concentrations of Salmonella spp. in the colon and of Escherichia coli throughout the intestine were reduced with LFM (P < 0.01). Concentrations of Bifidobacterium spp. in the ileum and of Lactobacillus spp. throughout the intestine were also increased with LFM (P ≤ 0.01). We suggest that rhLF can modulate the intestinal flora in piglets.

  5. Rye Affects Bacterial Translocation, Intestinal Viscosity, Microbiota Composition and Bone Mineralization in Turkey Poults

    PubMed Central

    Tellez, Guillermo; Latorre, Juan D.; Kuttappan, Vivek A.; Hargis, Billy M.; Hernandez-Velasco, Xochitl

    2015-01-01

    Previously, we have reported that rye significantly increased both viscosity and Clostridium perfringens proliferation when compared with corn in an in vitro digestive model. Two independent trials were conducted to evaluate the effect of rye as a source of energy on bacterial translocation, intestinal viscosity, gut microbiota composition, and bone mineralization, when compared with corn in turkey poults. In each experiment, day-of-hatch, turkey poults were randomly assigned to either a corn or a rye diet (n = 0 /group). At 10 d of age, in both experiments, 12 birds/group were given an oral gavage dose of fluorescein isothiocyanate dextran (FITC-d). After 2.5 h of oral gavage, blood and liver samples were collected to evaluate the passage of FITC-d and bacterial translocation (BT) respectively. Duodenum, ileum and cecum gut sections were collected to evaluate intestinal viscosity and to enumerate gut microbiota. Tibias were collected for observation of bone parameters. Broilers fed with a rye diet showed increased (p<0.05) intestinal viscosity, BT, and serum FITC-d. Bacterial enumeration revealed that turkey poults fed with rye had increased the number of total lactic acid bacteria (LAB) in all three sections of the gastrointestinal tract evaluated when compared to turkey poults fed with corn. Turkey poults fed with rye also had significantly higher coliforms in duodenum and ileum but not in the ceca, whereas the total number of anaerobes increased only in duodenum. A significant reduction in bone strength and bone mineralization was observed in turkey poults fed with rye when compared with corn fed turkey poults. In conclusion, rye evoked mucosal damage in turkey poults that increased intestinal viscosity, increased leakage through the intestinal tract, and altered the microbiota composition and bone mineralization. Studies to evaluate dietary inclusion of selected Direct-Fed Microbial (DFM) candidates that produce exogenous enzymes in rye fed turkey poults are

  6. Rye affects bacterial translocation, intestinal viscosity, microbiota composition and bone mineralization in Turkey poults.

    PubMed

    Tellez, Guillermo; Latorre, Juan D; Kuttappan, Vivek A; Hargis, Billy M; Hernandez-Velasco, Xochitl

    2015-01-01

    Previously, we have reported that rye significantly increased both viscosity and Clostridium perfringens proliferation when compared with corn in an in vitro digestive model. Two independent trials were conducted to evaluate the effect of rye as a source of energy on bacterial translocation, intestinal viscosity, gut microbiota composition, and bone mineralization, when compared with corn in turkey poults. In each experiment, day-of-hatch, turkey poults were randomly assigned to either a corn or a rye diet (n = 0 /group). At 10 d of age, in both experiments, 12 birds/group were given an oral gavage dose of fluorescein isothiocyanate dextran (FITC-d). After 2.5 h of oral gavage, blood and liver samples were collected to evaluate the passage of FITC-d and bacterial translocation (BT) respectively. Duodenum, ileum and cecum gut sections were collected to evaluate intestinal viscosity and to enumerate gut microbiota. Tibias were collected for observation of bone parameters. Broilers fed with a rye diet showed increased (p<0.05) intestinal viscosity, BT, and serum FITC-d. Bacterial enumeration revealed that turkey poults fed with rye had increased the number of total lactic acid bacteria (LAB) in all three sections of the gastrointestinal tract evaluated when compared to turkey poults fed with corn. Turkey poults fed with rye also had significantly higher coliforms in duodenum and ileum but not in the ceca, whereas the total number of anaerobes increased only in duodenum. A significant reduction in bone strength and bone mineralization was observed in turkey poults fed with rye when compared with corn fed turkey poults. In conclusion, rye evoked mucosal damage in turkey poults that increased intestinal viscosity, increased leakage through the intestinal tract, and altered the microbiota composition and bone mineralization. Studies to evaluate dietary inclusion of selected Direct-Fed Microbial (DFM) candidates that produce exogenous enzymes in rye fed turkey poults are

  7. Cultivable bacterial diversity from the human colon.

    PubMed

    Duncan, S H; Louis, P; Flint, H J

    2007-04-01

    Knowledge of the composition of the colonic microbiota is important for our understanding of how the balance of these microbes is influenced by diet and the environment, and which bacterial groups are important in maintaining gut health or promoting disease. Molecular methodologies have advanced our understanding of the composition and diversity of the colonic microbiota. Importantly, however, it is the continued isolation of bacterial representatives of key groups that offers the best opportunity to conduct detailed metabolic and functional studies. This also permits bacterial genome sequencing which will accelerate the linkage to functionality. Obtaining new human colonic bacterial isolates can be challenging, because most of these are strict anaerobes and many have rather exact nutritional and physical requirements. Despite this many new species are being isolated and described that occupy distinct niches in the colonic microbial community. This review focuses on these under-studied yet important gut anaerobes.

  8. Lipocalin 2 prevents intestinal inflammation by enhancing phagocytic bacterial clearance in macrophages.

    PubMed

    Toyonaga, Takahiko; Matsuura, Minoru; Mori, Kiyoshi; Honzawa, Yusuke; Minami, Naoki; Yamada, Satoshi; Kobayashi, Taku; Hibi, Toshifumi; Nakase, Hiroshi

    2016-10-13

    Lipocalin 2 (Lcn2), also called neutrophil gelatinase B-associated lipocalin (NGAL), is an anti-microbial peptide originally identified in neutrophil granules. Although Lcn2/NGAL expression is increased in the inflamed intestinal tissues of patients with inflammatory bowel disease, the role of Lcn2/NGAL in the development of intestinal inflammation remains unclear. Here we investigated the role of Lcn2/NGAL in intestinal inflammation using a spontaneous mouse colitis model, interleukin-10 knock out (IL-10 KO) mice. Lcn2 expression in the colonic tissues of IL-10 KO mice increased with the development of colitis. Lcn2/IL-10 double-KO mice showed a more rapid onset and development of colitis compared to IL-10 KO mice. Lcn2 enhanced phagocytic bacterial clearance in macrophages in vitro after infection with Escherichia coli. Transfer of Lcn2-repleted macrophages prevented the development of colitis in Lcn2/IL-10 double-KO mice in vivo. Our findings revealed that Lcn2 prevents the development of intestinal inflammation. One crucial factor seems to be the enhancement of phagocytic bacterial clearance in macrophages by Lcn2.

  9. Lipocalin 2 prevents intestinal inflammation by enhancing phagocytic bacterial clearance in macrophages

    PubMed Central

    Toyonaga, Takahiko; Matsuura, Minoru; Mori, Kiyoshi; Honzawa, Yusuke; Minami, Naoki; Yamada, Satoshi; Kobayashi, Taku; Hibi, Toshifumi; Nakase, Hiroshi

    2016-01-01

    Lipocalin 2 (Lcn2), also called neutrophil gelatinase B-associated lipocalin (NGAL), is an anti-microbial peptide originally identified in neutrophil granules. Although Lcn2/NGAL expression is increased in the inflamed intestinal tissues of patients with inflammatory bowel disease, the role of Lcn2/NGAL in the development of intestinal inflammation remains unclear. Here we investigated the role of Lcn2/NGAL in intestinal inflammation using a spontaneous mouse colitis model, interleukin-10 knock out (IL-10 KO) mice. Lcn2 expression in the colonic tissues of IL-10 KO mice increased with the development of colitis. Lcn2/IL-10 double-KO mice showed a more rapid onset and development of colitis compared to IL-10 KO mice. Lcn2 enhanced phagocytic bacterial clearance in macrophages in vitro after infection with Escherichia coli. Transfer of Lcn2-repleted macrophages prevented the development of colitis in Lcn2/IL-10 double-KO mice in vivo. Our findings revealed that Lcn2 prevents the development of intestinal inflammation. One crucial factor seems to be the enhancement of phagocytic bacterial clearance in macrophages by Lcn2. PMID:27734904

  10. Bacterial Population in Intestines of Litopenaeus vannamei Fed Different Probiotics or Probiotic Supernatant.

    PubMed

    Sha, Yujie; Liu, Mei; Wang, Baojie; Jiang, Keyong; Qi, Cancan; Wang, Lei

    2016-10-28

    The interactions of microbiota in the gut play an important role in promoting or maintaining the health of hosts. In this study, in order to investigate and compare the effects of dietary supplementation with Lactobacillus pentosus HC-2 (HC-2), Enterococcus faecium NRW-2, or the bacteria-free supernatant of a HC-2 culture on the bacterial composition of Litopenaeus vannamei, Illumina sequencing of the V1-V2 region of the 16S rRNA gene was used. The results showed that unique species exclusively existed in specific dietary groups, and the abundance of Actinobacteria was significantly increased in the intestinal bacterial community of shrimp fed with the bacteria-free supernatant of an HC-2 culture compared with the control. In addition, the histology of intestines of the shrimp from the four dietary groups was also described, but no obvious improvements in the intestinal histology were observed. The findings in this work will help to promote the understanding of the roles of intestinal bacteria in shrimps when fed with probiotics or probiotic supernatant.

  11. Efficient genetic engineering of human intestinal organoids using electroporation.

    PubMed

    Fujii, Masayuki; Matano, Mami; Nanki, Kosaku; Sato, Toshiro

    2015-10-01

    Gene modification in untransformed human intestinal cells is an attractive approach for studying gene function in intestinal diseases. However, because of the lack of practical tools, such studies have largely depended upon surrogates, such as gene-engineered mice or immortalized human cell lines. By taking advantage of the recently developed intestinal organoid culture method, we developed a methodology for modulating genes of interest in untransformed human colonic organoids via electroporation of gene vectors. Here we describe a detailed protocol for the generation of intestinal organoids by culture with essential growth factors in a basement membrane matrix. We also describe how to stably integrate genes via the piggyBac transposon, as well as precise genome editing using the CRISPR-Cas9 system. Beginning with crypt isolation from a human colon sample, genetically modified organoids can be obtained in 3 weeks.

  12. Intestinal bacterial overgrowth includes potential pathogens in the carbohydrate overload models of equine acute laminitis.

    PubMed

    Onishi, Janet C; Park, Joong-Wook; Prado, Julio; Eades, Susan C; Mirza, Mustajab H; Fugaro, Michael N; Häggblom, Max M; Reinemeyer, Craig R

    2012-10-12

    Carbohydrate overload models of equine acute laminitis are used to study the development of lameness. It is hypothesized that a diet-induced shift in cecal bacterial communities contributes to the development of the pro-inflammatory state that progresses to laminar failure. It is proposed that vasoactive amines, protease activators and endotoxin, all bacterial derived bioactive metabolites, play a role in disease development. Questions regarding the oral bioavailability of many of the bacterial derived bioactive metabolites remain. This study evaluates the possibility that a carbohydrate-induced overgrowth of potentially pathogenic cecal bacteria occurs and that bacterial translocation contributes toward the development of the pro-inflammatory state. Two groups of mixed-breed horses were used, those with laminitis induced by cornstarch (n=6) or oligofructan (n=6) and non-laminitic controls (n=8). Cecal fluid and tissue homogenates of extra-intestinal sites including the laminae were used to enumerate Gram-negative and -positive bacteria. Horses that developed Obel grade2 lameness, revealed a significant overgrowth of potentially pathogenic Gram-positive and Gram-negative intestinal bacteria within the cecal fluid. Although colonization of extra-intestinal sites with potentially pathogenic bacteria was not detected, results of this study indicate that cecal/colonic lymphadenopathy and eosinophilia develop in horses progressing to lameness. It is hypothesized that the pro-inflammatory state in carbohydrate overload models of equine acute laminitis is driven by an immune response to the rapid overgrowth of Gram-positive and Gram-negative cecal bacterial communities in the gut. Further equine research is indicated to study the immunological response, involving the lymphatic system that develops in the model.

  13. Milk sialyllactose influences colitis in mice through selective intestinal bacterial colonization

    PubMed Central

    Fuhrer, Andrea; Sprenger, Norbert; Kurakevich, Ekaterina; Borsig, Lubor; Chassard, Christophe

    2010-01-01

    Milk oligosaccharides contribute to the development of the intestinal environment by acting as decoy receptors for pathogens and as prebiotics, which promote the colonization of commensal bacteria. Here, using α2,3- and α2,6-sialyltransferase-deficient mice, we investigated the role of the sialylated milk oligosaccharides sialyl(α2,3)lactose and sialyl(α2,6)lactose on mucosal immunity. The exposure of newborn mice to milk containing or deficient in sialyllactose had no impact on the development of mucosal leukocyte populations. However, when challenged by dextran sulfate sodium (DSS) in drinking water, adult mice that had been fostered on sialyl(α2,3)lactose-deficient milk were more resistant to colitis compared with mice fostered on normal milk or sialyl(α2,6)lactose-deficient milk. Analysis of intestinal microbiota showed different colonization patterns depending on the presence or absence of sialyl(α2,3)lactose in the milk. Germ-free mice reconstituted with intestinal microbiota isolated from mice fed on sialyl(α2,3)lactose-deficient milk were more resistant to DSS-induced colitis than germ-free mice reconstituted with standard intestinal microbiota. Thus, exposure to sialyllactose during infancy affects bacterial colonization of the intestine, which influences the susceptibility to DSS-induced colitis in adult mice. PMID:21098096

  14. Methods to determine intestinal permeability and bacterial translocation during liver disease

    PubMed Central

    Wang, Lirui; Llorente, Cristina; Hartmann, Phillipp; Yang, An-Ming; Chen, Peng; Schnabl, Bernd

    2015-01-01

    Liver disease is often times associated with increased intestinal permeability. A disruption of the gut barrier allows microbial products and viable bacteria to translocate from the intestinal lumen to extraintestinal organs. The majority of the venous blood from the intestinal tract is drained into the portal circulation, which is part of the dual hepatic blood supply. The liver is therefore the first organ in the body to encounter not only absorbed nutrients, but also gut-derived bacteria and pathogen associated molecular patterns (PAMPs). Chronic exposure to increased levels of PAMPs has been linked to disease progression during early stages and to infectious complications during late stages of liver disease (cirrhosis). It is therefore important to assess and monitor gut barrier dysfunction during hepatic disease. We review methods to assess intestinal barrier disruption and discuss advantages and disadvantages. We will in particular focus on methods that we have used to measure increased intestinal permeability and bacterial translocation during experimental liver disease models. PMID:25595554

  15. Effect of Ceftaroline on Normal Human Intestinal Microflora▿

    PubMed Central

    Panagiotidis, Georgios; Bäckström, Tobias; Asker-Hagelberg, Charlotte; Jandourek, Alena; Weintraub, Andrej; Nord, Carl Erik

    2010-01-01

    Ceftaroline is a new broad-spectrum cephalosporin being developed for the treatment of serious bacterial infections, including those caused by aerobic Gram-positive and Gram-negative bacteria. The purpose of the present study was to investigate the effect of administration of ceftaroline on the intestinal flora of healthy subjects. Twelve healthy subjects (6 males and 6 females), 20 to 41 years of age, received ceftaroline (600 mg) by intravenous infusion every 12 h (q12h) for 7 days. Plasma and feces were collected for determination of ceftaroline concentration and analysis of fecal flora. Fecal specimens were cultured on nonselective and selective media. Different colony types were counted, isolated in pure culture, and identified to the genus level. All new strains of colonizing bacteria were tested for susceptibility to ceftaroline. The concentrations of ceftaroline in plasma were as follows: on day 2, 17.5 to 34.8 mg/liter; on day 5, 19.7 to 33.2 mg/liter; and on day 7, 18.0 to 29.8 mg/liter. No ceftaroline concentrations were found on day −1, 9, 14, or 21. No measurable concentrations in feces were found on day −1, 2, 5, 7, 9, 14, or 21. There was a minor impact on the numbers of Escherichia coli strains, while the numbers of enterococci and Candida albicans strains were not affected. There were moderate decreases in the numbers of bifidobacteria and lactobacilli during the first 7 days, while the numbers of clostridia increased during the same period. No impact on the numbers of Bacteroides bacteria was noticed. No new colonizing aerobic or anaerobic bacteria resistant to ceftaroline (MIC ≥ 4 mg/liter) were found. Ceftaroline had no significant ecological impact on the human intestinal microflora. PMID:20231399

  16. Active Transport of Phosphorylated Carbohydrates Promotes Intestinal Colonization and Transmission of a Bacterial Pathogen

    PubMed Central

    Sit, Brandon; Crowley, Shauna M.; Bhullar, Kirandeep; Lai, Christine Chieh-Lin; Tang, Calvin; Hooda, Yogesh; Calmettes, Charles; Khambati, Husain; Ma, Caixia; Brumell, John H.; Schryvers, Anthony B.; Vallance, Bruce A.; Moraes, Trevor F.

    2015-01-01

    Efficient acquisition of extracellular nutrients is essential for bacterial pathogenesis, however the identities and mechanisms for transport of many of these substrates remain unclear. Here, we investigate the predicted iron-binding transporter AfuABC and its role in bacterial pathogenesis in vivo. By crystallographic, biophysical and in vivo approaches, we show that AfuABC is in fact a cyclic hexose/heptose-phosphate transporter with high selectivity and specificity for a set of ubiquitous metabolites (glucose-6-phosphate, fructose-6-phosphate and sedoheptulose-7-phosphate). AfuABC is conserved across a wide range of bacterial genera, including the enteric pathogens EHEC O157:H7 and its murine-specific relative Citrobacter rodentium, where it lies adjacent to genes implicated in sugar sensing and acquisition. C. rodentium ΔafuA was significantly impaired in an in vivo murine competitive assay as well as its ability to transmit infection from an afflicted to a naïve murine host. Sugar-phosphates were present in normal and infected intestinal mucus and stool samples, indicating that these metabolites are available within the intestinal lumen for enteric bacteria to import during infection. Our study shows that AfuABC-dependent uptake of sugar-phosphates plays a critical role during enteric bacterial infection and uncovers previously unrecognized roles for these metabolites as important contributors to successful pathogenesis. PMID:26295949

  17. Intestinal microbiota in metabolic diseases: from bacterial community structure and functions to species of pathophysiological relevance.

    PubMed

    Clavel, Thomas; Desmarchelier, Charles; Haller, Dirk; Gérard, Philippe; Rohn, Sascha; Lepage, Patricia; Daniel, Hannelore

    2014-07-01

    The trillions of bacterial cells that colonize the mammalian digestive tract influence both host physiology and the fate of dietary compounds. Gnotobionts and fecal transplantation have been instrumental in revealing the causal role of intestinal bacteria in energy homeostasis and metabolic dysfunctions such as type-2 diabetes. However, the exact contribution of gut bacterial metabolism to host energy balance is still unclear and knowledge about underlying molecular mechanisms is scant. We have previously characterized cecal bacterial community functions and host responses in diet-induced obese mice using omics approaches. Based on these studies, we here discuss issues on the relevance of mouse models, give evidence that the metabolism of cholesterol-derived compounds by gut bacteria is of particular importance in the context of metabolic disorders and that dominant species of the family Coriobacteriaceae are good models to study these functions.

  18. Three-Dimensional Coculture Of Human Small-Intestine Cells

    NASA Technical Reports Server (NTRS)

    Wolf, David; Spaulding, Glen; Goodwin, Thomas J.; Prewett, Tracy

    1994-01-01

    Complex three-dimensional masses of normal human epithelial and mesenchymal small-intestine cells cocultured in process involving specially designed bioreactors. Useful as tissued models for studies of growth, regulatory, and differentiation processes in normal intestinal tissues; diseases of small intestine; and interactions between cells of small intestine and viruses causing disease both in small intestine and elsewhere in body. Process used to produce other tissue models, leading to advances in understanding of growth and differentiation in developing organisms, of renewal of tissue, and of treatment of myriad of clinical conditions. Prior articles describing design and use of rotating-wall culture vessels include "Growing And Assembling Cells Into Tissues" (MSC-21559), "High-Aspect-Ratio Rotating Cell-Culture Vessel" (MSC-21662), and "In Vitro, Matrix-Free Formation Of Solid Tumor Spheroids" (MSC-21843).

  19. Postnatal regulation of MAMDC4 in the porcine intestinal epithelium is influenced by bacterial colonization.

    PubMed

    Pasternak, Alex J; Hamonic, Glenn M; Van Kessel, Andrew; Wilson, Heather L

    2016-11-01

    The MAM domain-containing 4 (MAMDC4) protein is associated with the unique endocytotic mechanism observed in the intestine of mammals during the immediate postnatal period. Transcriptional expression of MAMDC4 was substantially upregulated at birth in both the piglet jejunum and ileum and its expression decreases after birth. The protein was found localized specifically to the apical region of the luminal epithelium, however, MAMDC4 protein expression was lost at day 10 and 15 in the jejunum and ileum, respectively, and was not associated with "fetal" enterocyte replacement. Although spatial variation in the subcellular localization of Claudin 1 (CLDN1) was noted at day 3, the loss of MAMDC4 at later stages of development did not appear to have any effect on the tight junction structure. Germ-free (GF) piglets and piglets whose gastrointestinal flora consists exclusively of Escherichia coli (EC) or Lactobacillus fermentum (LF) maintained MAMDC4 protein expression to 14 days of age in distal regions of the small intestine whereas those with conventionalized intestinal flora (CV) showed no MAMDC4 protein at this age. MAMDC4 protein expression was most pronounced in the LF and GF colonized piglets which showed staining in the epithelial cells at 75% and 95% of the length of the small intestine, respectively, which matched that of the newborn. In contrast, EC animals showed only a low abundance at these regions as well as a discontinuous staining pattern. Collectively these results suggest that maturation of MAMDC4 expression in the porcine epithelium occurs more rapidly than what is reported in previously studied rodent species. Furthermore, intestinal bacterial colonization is a major regulator of MAMDC4 in a manner specific to bacterial species and independent of enterocyte turnover.

  20. Characterization of human foetal intestinal alkaline phosphatase. Comparison with the isoenzymes from the adult intestine and human tumour cell lines.

    PubMed Central

    Behrens, C M; Enns, C A; Sussman, H H

    1983-01-01

    The molecular structure of human foetal intestinal alkaline phosphatase was defined by high-resolution two-dimensional polyacrylamide-gel electrophoresis and amino acid inhibition studies. Comparison was made with the adult form of intestinal alkaline phosphatase, as well as with alkaline phosphatases isolated from cultured foetal amnion cells (FL) and a human tumour cell line (KB). Two non-identical subunits were isolated from the foetal intestinal isoenzyme, one having same molecular weight and isoelectric point as placental alkaline phosphatase, and the other corresponding to a glycosylated subunit of the adult intestinal enzyme. The FL-cell and KB-cell alkaline phosphatases were also found to contain two subunits similar to those of the foetal intestinal isoenzyme. Characterization of neuraminidase digests of the non-placental subunit showed it to be indistinguishable from the subunits of the adult intestinal isoenzyme. This implies that no new phosphatase structural gene is involved in the transition from the expression of foetal to adult intestinal alkaline phosphatase, but that the molecular changes involve suppression of the placental subunit and loss of neuraminic acid from the non-placental subunit. Enzyme-inhibition studies demonstrated an intermediate response to the inhibitors tested for the foetal intestinal, FL-cell and KB-cell isoenzymes when compared with the placental, adult intestinal and liver forms. This result is consistent with the mixed-subunit structure observed for the former set of isoenzymes. In summary, this study has defined the molecular subunit structure of the foetal intestinal form of alkaline phosphatase and has demonstrated its expression in a human tumour cell line. Images Fig. 1. PMID:6882358

  1. Distinct human stem cell populations in small and large intestine.

    PubMed

    Cramer, Julie M; Thompson, Timothy; Geskin, Albert; LaFramboise, William; Lagasse, Eric

    2015-01-01

    The intestine is composed of an epithelial layer containing rapidly proliferating cells that mature into two regions, the small and the large intestine. Although previous studies have identified stem cells as the cell-of-origin for intestinal epithelial cells, no studies have directly compared stem cells derived from these anatomically distinct regions. Here, we examine intrinsic differences between primary epithelial cells isolated from human fetal small and large intestine, after in vitro expansion, using the Wnt agonist R-spondin 2. We utilized flow cytometry, fluorescence-activated cell sorting, gene expression analysis and a three-dimensional in vitro differentiation assay to characterize their stem cell properties. We identified stem cell markers that separate subpopulations of colony-forming cells in the small and large intestine and revealed important differences in differentiation, proliferation and disease pathways using gene expression analysis. Single cells from small and large intestine cultures formed organoids that reflect the distinct cellular hierarchy found in vivo and respond differently to identical exogenous cues. Our characterization identified numerous differences between small and large intestine epithelial stem cells suggesting possible connections to intestinal disease.

  2. Safety and risk assessment of the genetically modified Lactococci on rats intestinal bacterial flora.

    PubMed

    Lee, Kai-Chien; Liu, Chin-Feng; Lin, Tzu-Hsing; Pan, Tzu-Ming

    2010-08-15

    The interaction between Lactococcus lactis NZ9000/pNZPNK and intestinal microflora was evaluated as a method to assess safety of genetically modified microorganisms (GMMs). L. lactis NZ9000/pNZPNK is one kind of GMM and able to produce the intracellular subtilisin NAT (nattokinase) under induction with nisin. The host strain L. lactis NZ9000 was a generally recognized as safe (GRAS) microorganism. Six groups of Wistar rats were orally administered with L. lactis NZ9000/pNZPNK and L. lactis NZ9000 for 6 weeks. Fecal and cecal contents were collected to determine the number of L. lactis NZ9000, L. lactis NZ9000/pNZPNK, Lactobacillus, coliform bacteria, beneficial bacteria Bifidobacterium and harmful bacteria Clostridium perfringens. The liver, spleen, kidney and blood were evaluated for the bacterial translocation. After 6 weeks consumption with GM and non-GM Lactococcus, no adverse effects were observed on the rat's body weight, hematological or serum biochemical parameters, or intestinal microflora. The bacterial translocation test showed that L. lactis NZ9000/pNZPNK did not translocate to any organ or blood. Bifidobacterium was significantly increased in feces after administration of both Lactococcus strains (L. lactis NZ9000 and L. lactis NZ9000/pNZPNK), while C. perfringens remained undetectable during the experiment. These results suggested that L. lactis NZ9000/pNZPNK could be safe in animal experiments and monitoring of the interaction between test strains and intestinal microflora might be applied as a method for other GMM safety assessments.

  3. Glucagon effects on the human small intestine.

    PubMed

    Patel, G K; Whalen, G E; Soergel, K H; Wu, W C; Meade, R C

    1979-07-01

    In healthy volunteers, the effects of intravenously administered glucagon on small intestinal function was investigated. Bolus doses resulting in plasma glucagon concentrations of greater than 800 pg/ml (5 min after injection) abolished jejunal contractions for 4.4 +/- 0.4 (SEM) min after a latency period of 49 +/- 4 sec. During continuous intravenous glucagon infusion, jejunal dilatation and increase in mean transit time (MTT) occurred at plasma levels greater than 720 pg/ml, while inhibition of water and electrolyte absorption was observed only with plasma glucagon concentrations of 1760 +/- 114 pg/ml. Under these conditions, the propulsion of fasting intestinal contents was slowed without change in flow rate. The observed effects cannot be attributed to the simultaneously occurring rise in plasma insulin and glucose concentrations. Short-term increases in circulating glucagon concentration inhibit intestinal tone, contractions, and propulsion with only a minor effect on water and electrolyte absorption limited to a narrow concentration range of plasma glucagon. Neither effect occurs at glucagon levels likely to occur under physiologic concentrations. The latency period preceding the abolition of jejunal contractions suggests that glucagon does not act directly on intestinal smooth muscle cells.

  4. Prebiotic effects of almonds and almond skins on intestinal microbiota in healthy adult humans.

    PubMed

    Liu, Zhibin; Lin, Xiuchun; Huang, Guangwei; Zhang, Wen; Rao, Pingfan; Ni, Li

    2014-04-01

    Almonds and almond skins are rich in fiber and other components that have potential prebiotic properties. In this study we investigated the prebiotic effects of almond and almond skin intake in healthy humans. A total of 48 healthy adult volunteers consumed a daily dose of roasted almonds (56 g), almond skins (10 g), or commercial fructooligosaccharides (8 g) (as positive control) for 6 weeks. Fecal samples were collected at defined time points and analyzed for microbiota composition and selected indicators of microbial activity. Different strains of intestinal bacteria had varying degrees of growth sensitivity to almonds or almond skins. Significant increases in the populations of Bifidobacterium spp. and Lactobacillus spp. were observed in fecal samples as a consequence of almond or almond skin supplementation. However, the populations of Escherichia coli did not change significantly, while the growth of the pathogen Clostridum perfringens was significantly repressed. Modification of the intestinal microbiota composition induced changes in bacterial enzyme activities, specifically a significant increase in fecal β-galactosidase activity and decreases in fecal β-glucuronidase, nitroreductase and azoreductase activities. Our observations suggest that almond and almond skin ingestion may lead to an improvement in the intestinal microbiota profile and a modification of the intestinal bacterial activities, which would induce the promotion of health beneficial factors and the inhibition of harmful factors. Thus we believe that almonds and almond skins possess potential prebiotic properties.

  5. Hydrogen sulphide in exhaled breath: a potential biomarker for small intestinal bacterial overgrowth in IBS.

    PubMed

    Banik, Gourab Dutta; De, Anulekha; Som, Suman; Jana, Subhra; Daschakraborty, Sunil B; Chaudhuri, Sujit; Pradhan, Manik

    2016-05-10

    There is a pressing need to develop a novel early-detection strategy for the precise evolution of small intestinal bacterial overgrowth (SIBO) in irritable bowel syndrome (IBS) patients. The current method based on a hydrogen breath test (HBT) for the detection of SIBO is highly controversial. HBT has many limitations and drawbacks. It often fails to indentify SIBO when IBS individuals have 'non-hydrogen-producing' colonic bacteria. Here, we show that hydrogen sulphide (H2S) in exhaled breath is distinctly altered for diarrhea-predominant IBS individuals with positive and negative SIBO by the activity of intestinal sulphate-reducing bacteria. Subsequently, by analyzing the excretion kinetics of breath H2S, we found a missing link between breath H2S and SIBO when HBT often fails to diagnose SIBO. Moreover, breath H2S can track the precise evolution of SIBO, even after the eradication of bacterial overgrowth. Our findings suggest that the changes in H2S in the bacterial environment may contribute to the pathogenesis of SIBO and the breath H2S as a potential biomarker for non-invasive, rapid and precise assessment of SIBO without the endoscopy-based microbial culture of jejunal aspirates, and thus may open new perspectives into the pathophysiology of SIBO in IBS subjects.

  6. Understanding drug resistance in human intestinal protozoa.

    PubMed

    El-Taweel, Hend Aly

    2015-05-01

    Infections with intestinal protozoa continue to be a major health problem in many areas of the world. The widespread use of a limited number of therapeutic agents for their management and control raises concerns about development of drug resistance. Generally, the use of any antimicrobial agent should be accompanied by meticulous monitoring of its efficacy and measures to minimize resistance formation. Evidence for the occurrence of drug resistance in different intestinal protozoa comes from case studies and clinical trials, sometimes with a limited number of patients. Large-scale field-based assessment of drug resistance and drug sensitivity testing of clinical isolates are needed. Furthermore, the association of drug resistance with certain geographic isolates or genotypes deserves consideration. Drug resistance has been triggered in vitro and has been linked to modification of pyruvate:ferredoxin oxidoreductase, nitroreductases, antioxidant defense, or cytoskeletal system. Further mechanistic studies will have important implications in the development of second generation therapeutic agents.

  7. Investigation of the interactions between Chrysanthemum morifolium flowers extract and intestinal bacteria from human and rat.

    PubMed

    Tao, Jin-Hua; Duan, Jin-Ao; Qian, Yi-Yun; Qian, Da-Wei; Guo, Jian-Ming

    2016-11-01

    Flos Chrysanthemi, dried flower of Chrysanthemum morifolium Ramat, has drawn much attention recently owing to its potential beneficial health effects for human. Flos Chrysanthemi products are usually taken orally and metabolized by intestinal microflora. However, there has been no investigation of the comprehensive metabolic profile of the Flos Chrysanthemi extract by intestinal flora owing to its chemical complexity and the limitations of analytical methods. In this paper, a rapid, sensitive and automated analysis method, ultra-performance liquid chromatography/quadrupole time of flight mass spectrometry including MS(E) technology and automated data processing Metabolynx™ software, was developed and successfully applied for the biotransformation and metabolic profile of flavonoids in the Flos Chrysanthemi extract by intestinal flora from human and rat. A total of 32 metabolites were detected and tentatively identified in human and rat intestinal bacterial samples. These metabolites indicated that hydrolysis, hydroxylation, acetylation, methylation, hydrogenation and deoxygenation were the major conversion pathways of flavonoids in the Flos Chrysanthemi extract in vitro. Furthermore, the effects of the Flos Chrysanthemi extract on the growth of different intestinal bacteria were detected using an Emax precision microplate reader. Certain pathogenic bacteria such as Enterobacter, Enterococcus, Clostridium and Bacteroides were significantly inhibited by Flos Chrysanthemi, while commensal probiotics such as Lactobacillus and Bifidobacterium were moderately promoted. Our observation provided further evidence for the importance of intestinal bacteria in the metabolism and potential activity of the Flos Chrysanthemi extract. The results will also be helpful for the further pharmacokinetic study of Flos Chrysanthemi and to unravel how it works in vivo.

  8. Curcuminoid Demethylation as an Alternative Metabolism by Human Intestinal Microbiota.

    PubMed

    Burapan, Supawadee; Kim, Mihyang; Han, Jaehong

    2017-04-14

    Curcumin and other curcuminoids from Curcuma longa are important bioactive compounds exhibiting various pharmacological activities. In addition to the known reductive metabolism of curcuminoids, an alternative biotransformation of curcuminoids by human gut microbiota is reported herein. A curcuminoid mixture, composed of curcumin (1), demethoxycurcumin (2), and bisdemethoxycurcumin (3), was metabolized by the human intestinal bacterium Blautia sp. MRG-PMF1. 1 and 2 were converted to new metabolites by the methyl aryl ether cleavage reaction. Two metabolites, demethylcurcumin (4) and bisdemethylcurcumin (5), were sequentially produced from 1, and demethyldemethoxycurcumin (6) was produced from 2. Until now, sequential reduction of the heptadienone backbone of curcuminoids was the only known metabolism to occur in the human intestine. In this study, a new intestinal metabolism of curcuminoids was discovered. Demethylation of curcuminoids produced three new colonic metabolites that were already known as promising synthetic curcumin analogues. The results could explain the observed beneficial effects of turmeric.

  9. Intestine.

    PubMed

    Smith, J M; Skeans, M A; Horslen, S P; Edwards, E B; Harper, A M; Snyder, J J; Israni, A K; Kasiske, B L

    2016-01-01

    Intestine and intestine-liver transplant plays an important role in the treatment of intestinal failure, despite decreased morbidity associated with parenteral nutrition. In 2014, 210 new patients were added to the intestine transplant waiting list. Among prevalent patients on the list at the end of 2014, 65% were waiting for an intestine transplant and 35% were waiting for an intestine-liver transplant. The pretransplant mortality rate decreased dramatically over time for all age groups. Pretransplant mortality was highest for adult candidates, at 22.1 per 100 waitlist years compared with less than 3 per 100 waitlist years for pediatric candidates, and notably higher for candidates for intestine-liver transplant than for candidates for intestine transplant without a liver. Numbers of intestine transplants without a liver increased from a low of 51 in 2013 to 67 in 2014. Intestine-liver transplants increased from a low of 44 in 2012 to 72 in 2014. Short-gut syndrome (congenital and other) was the main cause of disease leading to both intestine and intestine-liver transplant. Graft survival improved over the past decade. Patient survival was lowest for adult intestine-liver recipients and highest for pediatric intestine recipients.

  10. Bacterial Respiratory Infections Complicating Human Immunodeficiency Virus.

    PubMed

    Feldman, Charles; Anderson, Ronald

    2016-04-01

    Opportunistic bacterial and fungal infections of the lower respiratory tract, most commonly those caused by Streptococcus pneumoniae (the pneumococcus), Mycobacterium tuberculosis, and Pneumocystis jirovecii, remain the major causes of mortality in those infected with human immunodeficiency virus (HIV). Bacterial respiratory pathogens most prevalent in those infected with HIV, other than M. tuberculosis, represent the primary focus of the current review with particular emphasis on the pneumococcus, the leading cause of mortality due to HIV infection in the developed world. Additional themes include (1) risk factors; (2) the predisposing effects of HIV-mediated suppression on pulmonary host defenses, possibly intensified by smoking; (3) clinical and laboratory diagnosis, encompassing assessment of disease severity and outcome; and (4) antibiotic therapy. The final section addresses current recommendations with respect to pneumococcal immunization in the context of HIV infection, including an overview of the rationale underpinning the current "prime-boost" immunization strategy based on sequential administration of pneumococcal conjugate vaccine 13 and pneumococcal polysaccharide vaccine 23.

  11. Probiotic yogurt in the elderly with intestinal bacterial overgrowth: endotoxaemia and innate immune functions.

    PubMed

    Schiffrin, Eduardo J; Parlesak, Alexandr; Bode, Christiane; Bode, J Christian; van't Hof, Martin A; Grathwohl, Dominik; Guigoz, Yves

    2009-04-01

    A study was conducted in healthy elderly living independently in senior housing to assess the impact of a probiotic yoghurt supplement on small intestinal bacterial overgrowth. Twenty-three participants with positive and thirteen participants with negative hydrogen breath test were studied before and after a period of 4 weeks of probiotic yoghurt administration. Intestinal permeability, plasma endotoxin levels, phagocytic activity of leucocytes, cytokine production by monocytes and free radical response of neutrophils were determined. Intestinal permeability was similar for the two groups and was unaffected by probiotic treatment. Both plasma endotoxin levels and the basal phagocytic activity of leucocytes decreased after yoghurt intake in the two groups. Exposure of monocytes and neutrophils ex vivo led to an increased cytokine response and free radical response, respectively. The normalisation of the various cytokine responses was more apparent in the group with positive breath test. In addition, the plasma levels of lipoplysaccharide binding protein and soluble CD14, lipoplysaccharide pattern recognition receptors and surrogate markers of lipoplysaccharide permeability were diminished by the end of the study. In conclusion, probiotic administration in the elderly normalises the response to endotoxin, and modulates activation markers in blood phagocytes, and therefore may help reduce low-grade chronic inflammation.

  12. Methane production and small intestinal bacterial overgrowth in children living in a slum

    PubMed Central

    Mello, Carolina Santos; Tahan, Soraia; Melli, Lígia Cristina FL; Rodrigues, Mirian Silva do Carmo; de Mello, Ricardo Martin Pereira; Scaletsky, Isabel Cristina Affonso; de Morais, Mauro Batista

    2012-01-01

    AIM: To analyze small intestinal bacterial overgrowth in school-aged children and the relationship between hydrogen and methane production in breath tests. METHODS: This transversal study included 85 children residing in a slum and 43 children from a private school, all aged between 6 and 10 years, in Osasco, Brazil. For characterization of the groups, data regarding the socioeconomic status and basic housing sanitary conditions were collected. Anthropometric data was obtained in children from both groups. All children completed the hydrogen (H2) and methane (CH4) breath test in order to assess small intestinal bacterial overgrowth (SIBO). SIBO was diagnosed when there was an increase in H2 ≥ 20 ppm or CH4 ≥ 10 ppm with regard to the fasting value until 60 min after lactulose ingestion. RESULTS: Children from the slum group had worse living conditions and lower nutritional indices than children from the private school. SIBO was found in 30.9% (26/84) of the children from the slum group and in 2.4% (1/41) from the private school group (P = 0.0007). Greater hydrogen production in the small intestine was observed in children from the slum group when compared to children from the private school (P = 0.007). A higher concentration of hydrogen in the small intestine (P < 0.001) and in the colon (P < 0.001) was observed among the children from the slum group with SIBO when compared to children from the slum group without SIBO. Methane production was observed in 63.1% (53/84) of the children from the slum group and in 19.5% (8/41) of the children from the private school group (P < 0.0001). Methane production was observed in 38/58 (65.5%) of the children without SIBO and in 15/26 (57.7%) of the children with SIBO from the slum. Colonic production of hydrogen was lower in methane-producing children (P = 0.017). CONCLUSION: Children who live in inadequate environmental conditions are at risk of bacterial overgrowth and methane production. Hydrogen is a substrate for

  13. The Bacterial Virulence Factor Lymphostatin Compromises Intestinal Epithelial Barrier Function by Modulating Rho GTPases

    PubMed Central

    Babbin, Brian A.; Sasaki, Maiko; Gerner-Schmidt, Kirsten W.; Nusrat, Asma; Klapproth, Jan-Michael A.

    2009-01-01

    Lymphocyte inhibitory factor A (lifA) in Citrobacter rodentium encodes the large toxin lymphostatin, which contains two enzymatic motifs associated with bacterial pathogenesis, a glucosyltransferase and a protease. Our aim was to determine the effects of each lymphostatin motif on intestinal epithelial-barrier function. In-frame mutations of C. rodentium lifA glucosyltransferase (CrGlM21) and protease (CrPrM5) were generated by homologous recombination. Infection of both model intestinal epithelial monolayers and mice with C. rodentium wild type resulted in compromised epithelial barrier function and mislocalization of key intercellular junction proteins in the tight junction and adherens junction. In contrast, CrGlM21 was impaired in its ability to reduce barrier function and influenced the tight junction proteins ZO-1 and occludin. CrPrM5 demonstrated decreased effects on the adherens junction proteins β-catenin and E-cadherin. Analysis of the mechanisms revealed that C. rodentium wild type differentially influenced Rho GTPase activation, suppressed Cdc42 activation, and induced Rho GTPase activation. CrGlM21 lost its suppressive effects on Cdc42 activation, whereas CrPrM5 was unable to activate Rho signaling. Rescue experiments using constitutively active Cdc42 or C3 exotoxin to inhibit Rho GTPase supported a role of Rho GTPases in the epithelial barrier compromise induced by C. rodentium. Taken together, our results suggest that lymphostatin is a bacterial virulence factor that contributes to the disruption of intestinal epithelial-barrier function via the modulation of Rho GTPase activities. PMID:19286565

  14. Transepithelial Transport of PAMAM Dendrimers Across Isolated Human Intestinal Tissue.

    PubMed

    Hubbard, Dallin; Enda, Michael; Bond, Tanner; Moghaddam, Seyyed Pouya Hadipour; Conarton, Josh; Scaife, Courtney; Volckmann, Eric; Ghandehari, Hamidreza

    2015-11-02

    Poly(amido amine) (PAMAM) dendrimers have shown transepithelial transport across intestinal epithelial barrier in rats and across Caco-2 cell monolayers. Caco-2 models innately lack mucous barriers, and rat isolated intestinal tissue has been shown to overestimate human permeability. This study is the first report of transport of PAMAM dendrimers across isolated human intestinal epithelium. It was observed that FITC labeled G4-NH2 and G3.5-COOH PAMAM dendrimers at 1 mM concentration do not have a statistically higher permeability compared to free FITC controls in isolated human jejunum and colonic tissues. Mannitol permeability was increased at 10 mM concentrations of G3.5-COOH and G4-NH2 dendrimers. Significant histological changes in human colonic and jejunal tissues were observed at G3.5-COOH and G4-NH2 concentrations of 10 mM implying that dose limiting toxicity may occur at similar concentrations in vivo. The permeability through human isolated intestinal tissue in this study was compared to previous rat and Caco-2 permeability data. This study implicates that PAMAM dendrimer oral drug delivery may be feasible, but it may be limited to highly potent drugs.

  15. Host-microbe interactions in the neonatal intestine: role of human milk oligosaccharides.

    PubMed

    Donovan, Sharon M; Wang, Mei; Li, Min; Friedberg, Iddo; Schwartz, Scott L; Chapkin, Robert S

    2012-05-01

    The infant intestinal microbiota is shaped by genetics and environment, including the route of delivery and early dietary intake. Data from germ-free rodents and piglets support a critical role for the microbiota in regulating gastrointestinal and immune development. Human milk oligosaccharides (HMO) both directly and indirectly influence intestinal development by regulating cell proliferation, acting as prebiotics for beneficial bacteria and modulating immune development. We have shown that the gut microbiota, the microbial metatranscriptome, and metabolome differ between porcine milk-fed and formula-fed (FF) piglets. Our goal is to define how early nutrition, specifically HMO, shapes host-microbe interactions in breast-fed (BF) and FF human infants. We an established noninvasive method that uses stool samples containing intact sloughed epithelial cells to quantify intestinal gene expression profiles in human infants. We hypothesized that a systems biology approach, combining i) HMO composition of the mother's milk with the infant's gut gene expression and fecal bacterial composition, ii) gene expression, and iii short-chain fatty acid profiles would identify important mechanistic pathways affecting intestinal development of BF and FF infants in the first few months of life. HMO composition was analyzed by HLPC Chip/time-of-flight MS and 3 HMO clusters were identified using principle component analysis. Initial findings indicated that both host epithelial cell mRNA expression and the microbial phylogenetic profiles provided strong feature sets that distinctly classified the BF and FF infants. Ongoing analyses are designed to integrate the host transcriptome, bacterial phylogenetic profiles, and functional metagenomic data using multivariate statistical analyses.

  16. Spatial organization of bacterial flora in normal and inflamed intestine: A fluorescence in situ hybridization study in mice

    PubMed Central

    Swidsinski, Alexander; Loening-Baucke, Vera; Lochs, Herbert; Hale, Laura P.

    2005-01-01

    AIM: To study the role of intestinal flora in inflammatory bowel disease (IBD). METHODS: The spatial organization of intestinal flora was investigated in normal mice and in two models of murine colitis using fluorescence in situ hybridization. RESULTS: The murine small intestine was nearly bacteria-free. The normal colonic flora was organized in three distinct compartments (crypt, interlaced, and fecal), each with different bacterial compositions. Crypt bacteria were present in the cecum and proximal colon. The fecal compartment was composed of homogeneously mixed bacterial groups that directly contacted the colonic wall in the cecum but were separated from the proximal colonic wall by a dense interlaced layer. Beginning in the middle colon, a mucus gap of growing thickness physically separated all intestinal bacteria from contact with the epithelium. Colonic inflammation was accompanied with a depletion of bacteria within the fecal compartment, a reduced surface area in which feces had direct contact with the colonic wall, increased thickness and spread of the mucus gap, and massive increases of bacterial concentrations in the crypt and interlaced compartments. Adhesive and infiltrative bacteria were observed in inflamed colon only, with dominant Bacteroides species. CONCLUSION: The proximal and distal colons are functionally different organs with respect to the intestinal flora, representing a bioreactor and a segregation device. The highly organized structure of the colonic flora, its specific arrangement in different colonic segments, and its specialized response to inflammatory stimuli indicate that the intestinal flora is an innate part of host immunity that is under complex control. PMID:15754393

  17. Diet and the development of the human intestinal microbiome

    PubMed Central

    Voreades, Noah; Kozil, Anne; Weir, Tiffany L.

    2014-01-01

    The important role of the gut microbiome in maintaining human health has necessitated a better understanding of the temporal dynamics of intestinal microbial communities as well as the host and environmental factors driving these dynamics. Genetics, mode of birth, infant feeding patterns, antibiotic usage, sanitary living conditions and long term dietary habits contribute to shaping the composition of the gut microbiome. This review focuses primarily on diet, as it is one of the most pivotal factors in the development of the human gut microbiome from infancy to the elderly. The infant gut microbiota is characterized by a high degree of instability, only reaching a state similar to that of adults by 2–3 years of age; consistent with the establishment of a varied solid food diet. The diet-related factors influencing the development of the infant gut microbiome include whether the child is breast or formula-fed as well as how and when solid foods are introduced. In contrast to the infant gut, the adult gut microbiome is resilient to large shifts in community structure. Several studies have shown that dietary changes induce transient fluctuations in the adult microbiome, sometimes in as little as 24 h; however, the microbial community rapidly returns to its stable state. Current knowledge of how long-term dietary habits shape the gut microbiome is limited by the lack of long-term feeding studies coupled with temporal gut microbiota characterization. However, long-term weight loss studies have been shown to alter the ratio of the Bacteroidetes and Firmicutes, the two major bacterial phyla residing in the human gastrointestinal tract. With aging, diet-related factors such as malnutrition are associated with microbiome shifts, although the cause and effect relationship between these factors has not been established. Increased pharmaceutical usage is also more prevalent in the elderly and can contribute to reduced gut microbiota stability and diversity. Foods containing

  18. Escherichia albertii, a novel human enteropathogen, colonizes rat enterocytes and translocates to extra-intestinal sites

    PubMed Central

    Yamamoto, Denise; Hernandes, Rodrigo T.; Liberatore, Ana Maria A.; Abe, Cecilia M.; de Souza, Rodrigo B.; Romão, Fabiano T.; Sperandio, Vanessa; Koh, Ivan H.

    2017-01-01

    Diarrhea is the second leading cause of death of children up to five years old in the developing countries. Among the etiological diarrheal agents are atypical enteropathogenic Escherichia coli (aEPEC), one of the diarrheagenic E. coli pathotypes that affects children and adults, even in developed countries. Currently, genotypic and biochemical approaches have helped to demonstrate that some strains classified as aEPEC are actually E. albertii, a recently recognized human enteropathogen. Studies on particular strains are necessary to explore their virulence potential in order to further understand the underlying mechanisms of E. albertii infections. Here we demonstrated for the first time that infection of fragments of rat intestinal mucosa is a useful tool to study the initial steps of E. albertii colonization. We also observed that an E. albertii strain can translocate from the intestinal lumen to Mesenteric Lymph Nodes and liver in a rat model. Based on our finding of bacterial translocation, we investigated how E. albertii might cross the intestinal epithelium by performing infections of M-like cells in vitro to identify the potential in vivo translocation route. Altogether, our approaches allowed us to draft a general E. albertii infection route from the colonization till the bacterial spreading in vivo. PMID:28178312

  19. Human milk oligosaccharide effects on intestinal function and inflammation after preterm birth in pigs.

    PubMed

    Rasmussen, Stine O; Martin, Lena; Østergaard, Mette V; Rudloff, Silvia; Roggenbuck, Michael; Nguyen, Duc Ninh; Sangild, Per T; Bering, Stine B

    2017-02-01

    Human milk oligosaccharides (HMOs) may mediate prebiotic and anti-inflammatory effects in newborns. This is particularly important for preterm infants who are highly susceptible to intestinal dysfunction and necrotizing enterocolitis (NEC). We hypothesized that HMO supplementation of infant formula (IF) improves intestinal function, bacterial colonization and NEC resistance immediately after preterm birth, as tested in a preterm pig model. Mixtures of HMOs were investigated in intestinal epithelial cells and in preterm pigs (n=112) fed IF supplemented without (CON) or with a mixture of four HMOs (4-HMO) or >25 HMOs (25-HMO, 5-10 g/L given for 5 or 11 days). The 25-HMO blend decreased cell proliferation and both HMO blends decreased lipopolysaccharide-induced interleukin-8 secretion in IPEC-J2 cells, relative to control (P<.05). All HMOs were found in urine and feces of HMO-treated pigs, and short-chain fatty acids in the colon were higher in HMO vs. CON pigs (P<.05). After 5 days, NEC lesions were similar between HMO and CON pigs and 25-HMO increased colon weights (P<.01). After 11 days, the 4-HMO diet did not affect NEC (56 vs. 79%, P=.2) but increased dehydration and diarrhea (P<.05) and expression of immune-related genes (IL10, IL12, TGFβ, TLR4; P<.05). Bacterial adherence and diversity was unchanged after HMO supplementation.

  20. Escherichia albertii, a novel human enteropathogen, colonizes rat enterocytes and translocates to extra-intestinal sites.

    PubMed

    Yamamoto, Denise; Hernandes, Rodrigo T; Liberatore, Ana Maria A; Abe, Cecilia M; Souza, Rodrigo B de; Romão, Fabiano T; Sperandio, Vanessa; Koh, Ivan H; Gomes, Tânia A T

    2017-01-01

    Diarrhea is the second leading cause of death of children up to five years old in the developing countries. Among the etiological diarrheal agents are atypical enteropathogenic Escherichia coli (aEPEC), one of the diarrheagenic E. coli pathotypes that affects children and adults, even in developed countries. Currently, genotypic and biochemical approaches have helped to demonstrate that some strains classified as aEPEC are actually E. albertii, a recently recognized human enteropathogen. Studies on particular strains are necessary to explore their virulence potential in order to further understand the underlying mechanisms of E. albertii infections. Here we demonstrated for the first time that infection of fragments of rat intestinal mucosa is a useful tool to study the initial steps of E. albertii colonization. We also observed that an E. albertii strain can translocate from the intestinal lumen to Mesenteric Lymph Nodes and liver in a rat model. Based on our finding of bacterial translocation, we investigated how E. albertii might cross the intestinal epithelium by performing infections of M-like cells in vitro to identify the potential in vivo translocation route. Altogether, our approaches allowed us to draft a general E. albertii infection route from the colonization till the bacterial spreading in vivo.

  1. Altered Virome and Bacterial Microbiome in Human Immunodeficiency Virus-Associated Acquired Immunodeficiency Syndrome.

    PubMed

    Monaco, Cynthia L; Gootenberg, David B; Zhao, Guoyan; Handley, Scott A; Ghebremichael, Musie S; Lim, Efrem S; Lankowski, Alex; Baldridge, Megan T; Wilen, Craig B; Flagg, Meaghan; Norman, Jason M; Keller, Brian C; Luévano, Jesús Mario; Wang, David; Boum, Yap; Martin, Jeffrey N; Hunt, Peter W; Bangsberg, David R; Siedner, Mark J; Kwon, Douglas S; Virgin, Herbert W

    2016-03-09

    Human immunodeficiency virus (HIV) infection is associated with increased intestinal translocation of microbial products and enteropathy as well as alterations in gut bacterial communities. However, whether the enteric virome contributes to this infection and resulting immunodeficiency remains unknown. We characterized the enteric virome and bacterial microbiome in a cohort of Ugandan patients, including HIV-uninfected or HIV-infected subjects and those either treated with anti-retroviral therapy (ART) or untreated. Low peripheral CD4 T cell counts were associated with an expansion of enteric adenovirus sequences and this increase was independent of ART treatment. Additionally, the enteric bacterial microbiome of patients with lower CD4 T counts exhibited reduced phylogenetic diversity and richness with specific bacteria showing differential abundance, including increases in Enterobacteriaceae, which have been associated with inflammation. Thus, immunodeficiency in progressive HIV infection is associated with alterations in the enteric virome and bacterial microbiome, which may contribute to AIDS-associated enteropathy and disease progression.

  2. Neutrophil priming by hypoxic preconditioning protects against epithelial barrier damage and enteric bacterial translocation in intestinal ischemia/reperfusion.

    PubMed

    Lu, Yen-Zhen; Wu, Chi-Chin; Huang, Yi-Chen; Huang, Ching-Ying; Yang, Chung-Yi; Lee, Tsung-Chun; Chen, Chau-Fong; Yu, Linda Chia-Hui

    2012-05-01

    Intestinal ischemia/reperfusion (I/R) induces mucosal barrier dysfunction and bacterial translocation (BT). Neutrophil-derived oxidative free radicals have been incriminated in the pathogenesis of ischemic injury in various organs, but their role in the bacteria-containing intestinal tract is debatable. Primed neutrophils are characterized by a faster and higher respiratory burst activity associated with more robust bactericidal effects on exposure to a second stimulus. Hypoxic preconditioning (HPC) attenuates ischemic injury in brain, heart, lung and kidney; no reports were found in the gut. Our aim is to investigate whether neutrophil priming by HPC protects against intestinal I/R-induced barrier damage and bacterial influx. Rats were raised in normoxia (NM) or kept in a hypobaric hypoxic chamber (380 Torr) 17 h/day for 3 weeks for HPC, followed by sham operation or intestinal I/R. Gut permeability was determined by using an ex vivo macromolecular flux assay and an in vivo magnetic resonance imaging-based method. Liver and spleen homogenates were plated for bacterial culturing. Rats raised in HPC showed diminished levels of BT, and partially improved mucosal histopathology and epithelial barrier function compared with the NM groups after intestinal I/R. Augmented cytokine-induced neutrophil chemoattractant (CINC)-1 and -3 levels and myeloperoxidase activity correlated with enhanced infiltration of neutrophils in intestines of HPC-I/R compared with NM-I/R rats. HPC alone caused blood neutrophil priming, as shown by elevated production of superoxide and hydrogen peroxide on stimulation, increased membrane translocation of cytosolic p47(phox) and p67(phox), as well as augmented bacterial-killing and phagocytotic activities. Neutrophil depletion reversed the mucosal protection by HPC, and aggravated intestinal leakiness and BT following I/R. In conclusion, neutrophil priming by HPC protects against I/R-induced BT via direct antimicrobial activity by oxidative

  3. Molecular Epidemiology of Human Intestinal Amoebas in Iran

    PubMed Central

    Hooshyar, H; Rostamkhani, P; Rezaian, M

    2012-01-01

    Many microscopic-based epidemiological surveys on the prevalence of human intestinal pathogenic and non-pathogenic protozoa including intestinal amoeba performed in Iran show a high prevalence of human intestinal amoeba in different parts of Iran. Such epidemiological studies on amoebiasis are confusing, mainly due to recently appreciated distinction between the Entamoeba histolytica, E. dispar and E. moshkovskii. Differential diagnosis can be done by some methods such as PCR-based methods, monoclonal antibodies and the analysis of isoenzyme typing, however the molecular study of these protozoa in Iran is low. Based on molecular studies, it seems that E. dispar is predominant species especially in the central and northern areas of Iran and amoebiasis due to E. histolytica is a rare infection in the country. It is suggested that infection with E. moshkovskii may be common among Iranians. Considering the importance of molecular epidemiology of amoeba in Iran and also the current data, the present study reviews the data currently available on the molecular distribution of intestinal human amoeba in Iran. PMID:23193500

  4. Identification of astilbin metabolites produced by human intestinal bacteria using UPLC-Q-TOF/MS.

    PubMed

    Zhao, Min; Xu, Jun; Qian, Dawei; Guo, Jianming; Jiang, Shu; Shang, Er-xin; Duan, Jin-ao

    2014-07-01

    Astilbin, mainly isolated from a commonly used herbal medicine, Smilax glabra Roxb (SGR), exhibits a variety of pharmacological activities and biological effects. It is metabolized by intestinal bacteria after oral administration which leads to the variation of ethnopharmacological profile of this traditional medicine. However, little is known on the interactions of this active compound with intestinal bacteria, which would be very helpful in unravelling how SGR works. In this study, different pure bacteria from human feces were isolated and were used to investigate their conversion capability of astilbin. Ultra-performance liquid chromatography/quadrupole-time-of-flight mass spectrometry (UPLC-Q-TOF/MS) technique combined with Metabolynx(TM) software was used to analyze astilbin and its metabolites. The parent compound and two metabolites (quercetin and eriodictyol) were detected in the isolated bacterial samples compared with blank samples. Quercetin was present in Enterococcus sp. 8B, 8-2 and 9-2 samples. Eriodictyol was only identified in Enterococcus sp. 8B sample. The metabolic routes and metabolites of astilbin produced by the different intestinal bacteria are reported for the first time. This will be useful for the investigation of the pharmacokinetic study of astilbin in vivo and the role of different intestinal bacteria in the metabolism of natural compounds.

  5. Human cytokine responses induced by Gram-positive cell walls of normal intestinal microbiota

    PubMed Central

    Chen, T; Isomäki, P; Rimpiläinen, M; Toivanen, P

    1999-01-01

    The normal microbiota plays an important role in the health of the host, but little is known of how the human immune system recognizes and responds to Gram-positive indigenous bacteria. We have investigated cytokine responses of peripheral blood mononuclear cells (PBMC) to Gram-positive cell walls (CW) derived from four common intestinal indigenous bacteria, Eubacterium aerofaciens (Eu.a.), Eubacterium limosum(Eu.l.), Lactobacillus casei(L.c.), and Lactobacillus fermentum (L.f.). Our results indicate that Gram-positive CW of the normal intestinal microbiota can induce cytokine responses of the human PBMC. The profile, level and kinetics of these responses are similar to those induced by lipopolysaccharide (LPS) or CW derived from a pathogen, Streptococcus pyogenes (S.p.). Bacterial CW are capable of inducing production of a proinflammatory cytokine, tumour necrosis factor-alpha (TNF-α), and an anti-inflammatory cytokine, IL-10, but not that of IL-4 or interferon-gamma (IFN-γ). Monocytes are the main cell population in PBMC to produce TNF-α and IL-10. Induction of cytokine secretion is serum-dependent; both CD14-dependent and -independent pathways are involved. These findings suggest that the human cytokine responses induced by Gram-positive CW of the normal intestinal microbiota are similar to those induced by LPS or Gram-positive CW of the pathogens. PMID:10540188

  6. Intestinal REG3 Lectins Protect Against Alcoholic Steatohepatitis by Reducing Mucosa-Associated Microbiota and Preventing Bacterial Translocation

    PubMed Central

    Wang, Lirui; Fouts, Derrick E.; Stärkel, Peter; Hartmann, Phillipp; Chen, Peng; Llorente, Cristina; DePew, Jessica; Moncera, Kelvin; Ho, Samuel B.; Brenner, David A.; Hooper, Lora V.; Schnabl, Bernd

    2016-01-01

    Summary Approximately half of all deaths from liver cirrhosis, the 10th leading cause of mortality in the United States, are related to alcohol use. Chronic alcohol consumption is accompanied by intestinal dysbiosis and bacterial overgrowth, yet little is known about the factors that alter the microbial composition or their contribution to liver disease. We previously associated chronic alcohol consumption with lower intestinal levels of the antimicrobial-regenerating islet-derived (REG)-3 lectins. Here, we demonstrate that intestinal deficiency in REG3B or REG3G increases numbers of mucosa-associated bacteria and enhances bacterial translocation to the mesenteric lymph nodes and liver, promoting the progression of ethanol-induced fatty liver disease toward steatohepatitis. Overexpression of Reg3g in intestinal epithelial cells restricts bacterial colonization of mucosal surfaces, reduces bacterial translocation, and protects mice from alcohol-induced steatohepatitis. Thus, alcohol appears to impair control of the mucosa-associated microbiota, and subsequent breach of the mucosal barrier facilitates progression of alcoholic liver disease. PMID:26867181

  7. Prevalence of Small Intestinal Bacterial Overgrowth among Chronic Pancreatitis Patients: A Case-Control Study

    PubMed Central

    Bouchard, Simon; Sidani, Sacha

    2016-01-01

    Background. Patients with chronic pancreatitis (CP) exhibit numerous risk factors for the development of small intestinal bacterial overgrowth (SIBO). Objective. To determine the prevalence of SIBO in patients with CP. Methods. Prospective, single-centre case-control study conducted between January and September 2013. Inclusion criteria were age 18 to 75 years and clinical and radiological diagnosis of CP. Exclusion criteria included history of gastric, pancreatic, or intestinal surgery or significant clinical gastroparesis. SIBO was detected using a standard lactulose breath test (LBT). A healthy control group also underwent LBT. Results. Thirty-one patients and 40 controls were included. The patient group was significantly older (53.8 versus 38.7 years; P < 0.01). The proportion of positive LBTs was significantly higher in CP patients (38.7 versus 2.5%: P < 0.01). A trend toward a higher proportion of positive LBTs in women compared with men was observed (66.6 versus 27.3%; P = 0.056). The subgroups with positive and negative LBTs were comparable in demographic and clinical characteristics, use of opiates, pancreatic enzymes replacement therapy (PERT), and severity of symptoms. Conclusion. The prevalence of SIBO detected using LBT was high among patients with CP. There was no association between clinical features and the risk for SIBO. PMID:27446865

  8. Selective intestinal decontamination for the prevention of early bacterial infections after liver transplantation

    PubMed Central

    Resino, Elena; San-Juan, Rafael; Aguado, Jose Maria

    2016-01-01

    Bacterial infection in the first month after liver transplantation is a frequent complication that poses a serious risk for liver transplant recipients as contributes substantially to increased length of hospitalization and hospital costs being a leading cause of death in this period. Most of these infections are caused by gram-negative bacilli, although gram-positive infections, especially Enterococcus sp. constitute an emerging infectious problem. This high rate of early postoperative infections after liver transplant has generated interest in exploring various prophylactic approaches to surmount this problem. One of these approaches is selective intestinal decontamination (SID). SID is a prophylactic strategy that consists of the administration of antimicrobials with limited anaerobicidal activity in order to reduce the burden of aerobic gram-negative bacteria and/or yeast in the intestinal tract and so prevent infections caused by these organisms. The majority of studies carried out to date have found SID to be effective in the reduction of gram-negative infection, but the effect on overall infection is limited due to a higher number of infection episodes by pathogenic enterococci and coagulase-negative staphylococci. However, difficulties in general extrapolation of the favorable results obtained in specific studies together with the potential risk of selection of multirresistant microorganisms has conditioned controversy about the routinely application of these strategies in liver transplant recipients. PMID:27468189

  9. Bacterial biofilms in the human gastrointestinal tract.

    PubMed

    Probert, H M; Gibson, G R

    2002-09-01

    Microbial biofilms were first described in 1936 and subsequent research has unveiled their ubiquity and physiological distinction from free-living (planktonic) microorganisms. In light of their emerging significance this review examines the bacterial biofilms within the human gastrointestinal tract. Attention is paid to the nature of these mucosally- associated populations, focusing on the protected environment afforded by the continual secretion of mucus by host epithelial cells. It also examines the attributes possessed by various bacterial species that facilitate habitation of this microenvironment. Additionally, contrasts are drawn between planktonic bacteria of the lumen and sessile (biofilm) bacteria growing in close association with host cells and food particles. In particular the different fermentation profiles exhibited by these two fractions are discussed. The potential role of these communities in host health and disease, as well as the stabilisation of the lumenal population, is also considered. Reference is made to the state of mutualism that exists between these little understood populations and the host epithelia, thus highlighting their ecological significance in terms of gastrointestinal health.

  10. Intermittent fasting promotes bacterial clearance and intestinal IgA production in Salmonella typhimurium-infected mice.

    PubMed

    Godínez-Victoria, M; Campos-Rodriguez, R; Rivera-Aguilar, V; Lara-Padilla, E; Pacheco-Yepez, J; Jarillo-Luna, R A; Drago-Serrano, M E

    2014-05-01

    The impact of intermittent fasting versus ad libitum feeding during Salmonella typhimurium infection was evaluated in terms of duodenum IgA levels, bacterial clearance and intestinal and extra-intestinal infection susceptibility. Mice that were intermittently fasted for 12 weeks or fed ad libitum were infected with S. typhimurium and assessed at 7 and 14 days post-infection. Next, we evaluated bacterial load in the faeces, Peyer's patches, spleen and liver by plate counting, as well as total and specific intestinal IgA and plasmatic corticosterone levels (by immunoenzymatic assay) and lamina propria IgA levels in plasma cells (by cytofluorometry). Polymeric immunoglobulin receptor, α- and J-chains, Pax-5 factor, pro-inflammatory cytokine (tumour necrosis factor-α and interferon-γ) and anti-inflammatory cytokine (transforming growth factor-β) mRNA levels were assessed in mucosal and liver samples (by real-time PCR). Compared with the infected ad libitum mice, the intermittently fasted infected animals had (1) lower intestinal and systemic bacterial loads; (2) higher SIgA and IgA plasma cell levels; (3) higher mRNA expression of most intestinal parameters; and (4) increased or decreased corticosterone levels on day 7 and 14 post-infection, respectively. No contribution of liver IgA was observed at the intestinal level. Apparently, the changes following metabolic stress induced by intermittent fasting during food deprivation days increased the resistance to S. typhimurium infection by triggering intestinal IgA production and presumably, pathogen elimination by phagocytic inflammatory cells.

  11. Associations between common intestinal parasites and bacteria in humans as revealed by qPCR.

    PubMed

    O'Brien Andersen, L; Karim, A B; Roager, H M; Vigsnæs, L K; Krogfelt, K A; Licht, T R; Stensvold, C R

    2016-09-01

    Several studies have shown associations between groups of intestinal bacterial or specific ratios between bacterial groups and various disease traits. Meanwhile, little is known about interactions and associations between eukaryotic and prokaryotic microorganisms in the human gut. In this work, we set out to investigate potential associations between common single-celled parasites such as Blastocystis spp. and Dientamoeba fragilis and intestinal bacteria. Stool DNA from patients with intestinal symptoms were selected based on being Blastocystis spp.-positive (B+)/negative (B-) and D. fragilis-positive (D+)/negative (D-), and split into four groups of 21 samples (B+ D+, B+ D-, B- D+, and B- D-). Quantitative PCR targeting the six bacterial taxa Bacteroides, Prevotella, the butyrate-producing clostridial clusters IV and XIVa, the mucin-degrading Akkermansia muciniphila, and the indigenous group of Bifidobacterium was subsequently performed, and the relative abundance of these bacteria across the four groups was compared. The relative abundance of Bacteroides in B- D- samples was significantly higher compared with B+ D- and B+ D+ samples (P < 0.05 and P < 0.01, respectively), and this association was even more significant when comparing all parasite-positive samples with parasite-negative samples (P < 0.001). Additionally, our data revealed that a low abundance of Prevotella and a higher abundance of Clostridial cluster XIVa was associated with parasite-negative samples (P < 0.05 and P < 0.01, respectively). Our data support the theory that Blastocystis alone or combined with D. fragilis is associated with gut microbiota characterized by low relative abundances of Bacteroides and Clostridial cluster XIVa and high levels of Prevotella.

  12. Intestinal Bacterial Infection Diagnosed by Histological Examination of Endoscopic Biopsy Specimens

    PubMed Central

    Yuki, Michiko; Emoto, Yuko; Yoshizawa, Katsuhiko; Yuri, Takashi; Tsubura, Airo

    2016-01-01

    Intestinal spirochetosis (IS) in humans is characterized by spirochetal microorganisms attached to the luminal surface of the colonic epithelium. In the present case, attached organisms appeared as 3- to 4 μm-thick (average thickness, 3.4 μm) basophilic fringes or haze in HE-stained endoscopic biopsy specimens. The basophilic fringes were clearly labeled by Treponema pallidum antiserum. Because IS is relatively rare in developed countries, thin basophilic fringes characteristic of IS are readily overlooked. Thus, the recognition of histological characteristics of this disease is important for its diagnosis. PMID:27920653

  13. Dual system of intestinal thiamine transport in humans

    SciTech Connect

    Hoyumpa, A.M. Jr.; Strickland, R.; Sheehan, J.J.; Yarborough, G.; Nichols, S.

    1982-05-01

    The transport of thiamine across the intestine has been characterized in rats but has not been adequately studied in humans. To determine the kinetics of thiamine intestinal transport directly in humans, mucosal tissues were obtained during routine endoscopy from normal-appearing sites at the second portion of the duodenum. With 3H-dextran as the marker of adherent volume, the uptake of 14C-thiamine hydrochloride by the excised mucosa was measured in vitro. By this method thiamine uptake was linear with tissue weight and with incubation time up to 5 min. Results showed that at low thiamine concentrations (0.2 to 2.0 microM), uptake was saturable whereas at high concentrations (5 to 50 microM), uptake was linear with thiamine concentrations. Pyrithiamine, anoxia, N-ethylmaleimide, and replacement of sodium chloride by mannitol reduced the uptake of 0.5 microM thiamine by 42%, 37%, 32% and 35%, respectively (p less than 0.05) but had no effect on the uptake of 20 microM thiamine. These data suggest that, as in the rat, the intestinal transport of thiamine in humans proceeds by a coexistent dual system. At physiologic concentrations, thiamine is transported primarily by an energy-requiring, sodium-dependent active process, whereas at higher pharmacologic concentrations thiamine uptake is predominantly a passive process.

  14. Evolution of Symbiotic Bacteria in the Distal Human Intestine

    PubMed Central

    Ley, Ruth E; Lozupone, Catherine A; Hamady, Micah; Martens, Eric C; Henrissat, Bernard; Coutinho, Pedro M; Minx, Patrick; Latreille, Philippe; Cordum, Holland; Van Brunt, Andrew; Kim, Kyung; Fulton, Robert S; Fulton, Lucinda A; Clifton, Sandra W; Wilson, Richard K; Knight, Robin D; Gordon, Jeffrey I

    2007-01-01

    The adult human intestine contains trillions of bacteria, representing hundreds of species and thousands of subspecies. Little is known about the selective pressures that have shaped and are shaping this community's component species, which are dominated by members of the Bacteroidetes and Firmicutes divisions. To examine how the intestinal environment affects microbial genome evolution, we have sequenced the genomes of two members of the normal distal human gut microbiota, Bacteroides vulgatus and Bacteroides distasonis, and by comparison with the few other sequenced gut and non-gut Bacteroidetes, analyzed their niche and habitat adaptations. The results show that lateral gene transfer, mobile elements, and gene amplification have played important roles in affecting the ability of gut-dwelling Bacteroidetes to vary their cell surface, sense their environment, and harvest nutrient resources present in the distal intestine. Our findings show that these processes have been a driving force in the adaptation of Bacteroidetes to the distal gut environment, and emphasize the importance of considering the evolution of humans from an additional perspective, namely the evolution of our microbiomes. PMID:17579514

  15. Bacterial species involved in the conversion of dietary flavonoids in the human gut

    PubMed Central

    Braune, Annett; Blaut, Michael

    2016-01-01

    ABSTRACT The gut microbiota plays a crucial role in the conversion of dietary flavonoids and thereby affects their health-promoting effects in the human host. The identification of the bacteria involved in intestinal flavonoid conversion has gained increasing interest. This review summarizes available information on the so far identified human intestinal flavonoid-converting bacterial species and strains as well as their enzymes catalyzing the underlying reactions. The majority of described species involved in flavonoid transformation are capable of carrying out the O-deglycosylation of flavonoids. Other bacteria cleave the less common flavonoid-C-glucosides and/or further degrade the aglycones of flavonols, flavanonols, flavones, flavanones, dihydrochalcones, isoflavones and monomeric flavan-3-ols. To increase the currently limited knowledge in this field, identification of flavonoid-converting bacteria should be continued using culture-dependent screening or isolation procedures and molecular approaches based on sequence information of the involved enzymes. PMID:26963713

  16. Subclinical atherosclerosis is linked to small intestinal bacterial overgrowth via vitamin K2-dependent mechanisms

    PubMed Central

    Ponziani, Francesca Romana; Pompili, Maurizio; Di Stasio, Enrico; Zocco, Maria Assunta; Gasbarrini, Antonio; Flore, Roberto

    2017-01-01

    AIM To assess the rate of matrix Gla-protein carboxylation in patients with small intestinal bacterial overgrowth (SIBO) and to decipher its association with subclinical atherosclerosis. METHODS Patients with suspected SIBO who presented with a low risk for cardiovascular disease and showed no evidence of atherosclerotic plaques were included in the study. A glucose breath test was performed in order to confirm the diagnosis of SIBO and vascular assessment was carried out by ultrasound examination. Plasma levels of the inactive form of MGP (dephosphorylated-uncarboxylated matrix Gla-protein) were quantified by ELISA and vitamin K2 intake was estimated using a food frequency questionnaire. RESULTS Thirty-nine patients were included in the study. SIBO was confirmed in 12/39 (30.8%) patients who also presented with a higher concentration of dephosphorylated-uncarboxylated matrix Gla-protein (9.5 μg/L vs 4.2 μg/L; P = 0.004). Arterial stiffness was elevated in the SIBO group (pulse-wave velocity 10.25 m/s vs 7.68 m/s; P = 0.002) and this phenomenon was observed to correlate linearly with the levels of dephosphorylated-uncarboxylated matrix Gla-protein (β = 0.220, R2 = 0.366, P = 0.03). Carotid intima-media thickness and arterial calcifications were not observed to be significantly elevated as compared to controls. CONCLUSION SIBO is associated with reduced matrix Gla-protein activation as well as arterial stiffening. Both these observations are regarded as important indicators of subclinical atherosclerosis. Hence, screening for SIBO, intestinal decontamination and supplementation with vitamin K2 has the potential to be incorporated into clinical practice as additional preventive measures. PMID:28275304

  17. Sequential cancer mutations in cultured human intestinal stem cells.

    PubMed

    Drost, Jarno; van Jaarsveld, Richard H; Ponsioen, Bas; Zimberlin, Cheryl; van Boxtel, Ruben; Buijs, Arjan; Sachs, Norman; Overmeer, René M; Offerhaus, G Johan; Begthel, Harry; Korving, Jeroen; van de Wetering, Marc; Schwank, Gerald; Logtenberg, Meike; Cuppen, Edwin; Snippert, Hugo J; Medema, Jan Paul; Kops, Geert J P L; Clevers, Hans

    2015-05-07

    Crypt stem cells represent the cells of origin for intestinal neoplasia. Both mouse and human intestinal stem cells can be cultured in medium containing the stem-cell-niche factors WNT, R-spondin, epidermal growth factor (EGF) and noggin over long time periods as epithelial organoids that remain genetically and phenotypically stable. Here we utilize CRISPR/Cas9 technology for targeted gene modification of four of the most commonly mutated colorectal cancer genes (APC, P53 (also known as TP53), KRAS and SMAD4) in cultured human intestinal stem cells. Mutant organoids can be selected by removing individual growth factors from the culture medium. Quadruple mutants grow independently of all stem-cell-niche factors and tolerate the presence of the P53 stabilizer nutlin-3. Upon xenotransplantation into mice, quadruple mutants grow as tumours with features of invasive carcinoma. Finally, combined loss of APC and P53 is sufficient for the appearance of extensive aneuploidy, a hallmark of tumour progression.

  18. Advancing the use of Lactobacillus acidophilus surface layer protein A for the treatment of intestinal disorders in humans

    PubMed Central

    Sahay, Bikash; Ge, Yong; Colliou, Natacha; Zadeh, Mojgan; Weiner, Chelsea; Mila, Ashley; Owen, Jennifer L; Mohamadzadeh, Mansour

    2015-01-01

    Intestinal immunity is subject to complex and fine-tuned regulation dictated by interactions of the resident microbial community and their gene products with host innate cells. Deterioration of this delicate process may result in devastating autoinflammatory diseases, including inflammatory bowel disease (IBD), which primarily comprises Crohn's disease (CD) and ulcerative colitis (UC). Efficacious interventions to regulate proinflammatory signals, which play critical roles in IBD, require further scientific investigation. We recently demonstrated that rebalancing intestinal immunity via the surface layer protein A (SlpA) from Lactobacillus acidophilus NCFM potentially represents a feasible therapeutic approach to restore intestinal homeostasis. To expand on these findings, we established a new method of purifying bacterial SlpA, a new SlpA-specific monoclonal antibody, and found no SlpA-associated toxicity in mice. Thus, these data may assist in our efforts to determine the immune regulatory efficacy of SlpA in humans. PMID:26647142

  19. Protective effects of terminal ileostomy against bacterial translocation in a rat model of intestinal ischemia/reperfusion injury

    PubMed Central

    Lin, Zhi-Liang; Yu, Wen-Kui; Tan, Shan-Jun; Duan, Kai-Peng; Dong, Yi; Bai, Xiao-Wu; Xu, Lin; Li, Ning

    2014-01-01

    AIM: To investigate the effects of terminal ileostomy on bacterial translocation (BT) and systemic inflammation after intestinal ischemia/reperfusion (I/R) injury in rats. METHODS: Thirty-two rats were assigned to either the sham-operated group, I/R group, I/R + resection and anastomosis group, or the I/R + ileostomy group. The superior mesenteric artery was occluded for 60 min. After 4 h, tissue samples were collected for analysis. BT was assessed by bacteriologic cultures, intestinal permeability and serum levels of endotoxin; systemic inflammation was assessed by serum levels of tumor necrosis factor (TNF)-α, interleukin (IL)-6 and IL-10, as well as by the activity of myeloperoxidase (MPO) and by intestinal histopathology. RESULTS: Intestinal I/R injury not only caused morphologic damage to ileal mucosa, but also induced BT, increased MPO activity and promoted the release of TNF-α, IL-6, and IL-10 in serum. BT and ileal mucosa injuries were significantly improved and levels of TNF-α and IL-6 in serum were decreased in the I/R + ileostomy group compared with the I/R + resection and anastomosis group. CONCLUSION: Terminal ileostomy can prevent the detrimental effects of intestinal I/R injury on BT, intestinal tissue, and inflammation. PMID:25548488

  20. Malabsorption, Orocecal Transit Time and Small Intestinal Bacterial Overgrowth in Type 2 Diabetic Patients: A Connection.

    PubMed

    Rana, S V; Malik, Aastha; Bhadada, Sanjay K; Sachdeva, Naresh; Morya, Rajesh Kumar; Sharma, Gaurav

    2017-03-01

    Type 2 diabetes mellitus consists of dysfunctions characterized by hyperglycemia and resulting from combination of resistance to insulin action and inadequate insulin secretion. Most of diabetic patients report significant gastrointestinal symptoms. Entire GI tract can be affected by diabetes from oral cavity to large bowel and anorectal region. Proteins, carbohydrates, fats, and most fluids are absorbed in small intestine. Malabsorption may occurs when proper absorption of nutrients does not take place due to bacterial overgrowth or altered gut motility. The present study was planned to measure various malabsorption parameters in type 2 diabetic patients. 175 patients and 175 age and sex matched healthy controls attending Endocrinology Clinic in PGI, Chandigarh were enrolled. Lactose intolerance was measured by using non-invasive lactose hydrogen breath test. Urinary d-xylose and fecal fat were estimated using standard methods. Orocecal transit time and small intestinal bacterial overgrowth were measured using non-invasive lactulose and glucose breath test respectively. Out of 175 diabetic patients enrolled, 87 were males while among 175 healthy subjects 88 were males. SIBO was observed in 14.8 % type 2 diabetic patients and in 2.8 % of controls. There was statistically significant increase (p < 0.002) in OCTT in type 2 diabetic patients compared with controls. OCTT was observed to be more delayed (p < 0.003) in patients who were found to have SIBO than in patients without SIBO. Lactose intolerance was observed in 60 % diabetic patients and 39.4 % in controls. Urinary d-xylose levels were also lower in case of diabetic patients but no significant difference was found in 72 h fecal fat excretion among diabetic patients and controls. Urinary d-xylose and lactose intolerance in SIBO positive type 2 diabetic patients was more severe as compared to SIBO negative diabetic patients. From this study we can conclude that delayed OCTT may have led to SIBO which may

  1. Commensal Streptococcus salivarius Modulates PPARγ Transcriptional Activity in Human Intestinal Epithelial Cells

    PubMed Central

    Couvigny, Benoît; de Wouters, Tomas; Kaci, Ghalia; Jacouton, Elsa; Delorme, Christine; Doré, Joël; Renault, Pierre; Blottière, Hervé M.

    2015-01-01

    The impact of commensal bacteria in eukaryotic transcriptional regulation has increasingly been demonstrated over the last decades. A multitude of studies have shown direct effects of commensal bacteria from local transcriptional activity to systemic impact. The commensal bacterium Streptococcus salivarius is one of the early bacteria colonizing the oral and gut mucosal surfaces. It has been shown to down-regulate nuclear transcription factor (NF-кB) in human intestinal cells, a central regulator of the host mucosal immune system response to the microbiota. In order to evaluate its impact on a further important transcription factor shown to link metabolism and inflammation in the intestine, namely PPARγ (peroxisome proliferator-activated receptor), we used human intestinal epithelial cell-lines engineered to monitor PPARγ transcriptional activity in response to a wide range of S. salivarius strains. We demonstrated that different strains from this bacterial group share the property to inhibit PPARγ activation independently of the ligand used. First attempts to identify the nature of the active compounds showed that it is a low-molecular-weight, DNase-, proteases- and heat-resistant metabolite secreted by S. salivarius strains. Among PPARγ-targeted metabolic genes, I-FABP and Angptl4 expression levels were dramatically reduced in intestinal epithelial cells exposed to S. salivarius supernatant. Both gene products modulate lipid accumulation in cells and down-regulating their expression might consequently affect host health. Our study shows that species belonging to the salivarius group of streptococci impact both host inflammatory and metabolic regulation suggesting a possible role in the host homeostasis and health. PMID:25946041

  2. [Bacterial proteases and bacterial resistance against human innate immunity factors].

    PubMed

    Tiurin, Iu A; Mustafin, I G; Fassakhov, R S

    2011-01-01

    The molecular and cell-mediated mechanisms that are developed by certain opportunistic and pathogenic bacteria and were obtained over the course of evolution to preserve resistance against principal components of human body innate immunity are summarized.

  3. Genome sequence of Victivallis vadensis ATCC BAA-548, an anaerobic bacterium from the phylum Lentisphaerae, isolated from the human gastro-intestinal tract

    SciTech Connect

    Van Passel, Mark W.J.; Kant, Ravi; Palva, Airi; Lucas, Susan; Copeland, A; Lapidus, Alla L.; Glavina Del Rio, Tijana; Dalin, Eileen; Tice, Hope; Bruce, David; Goodwin, Lynne A.; Pitluck, Sam; Davenport, Karen W.; Sims, David; Detter, J. Chris; Han, Cliff; Larimer, Frank W; Land, Miriam L; Hauser, Loren John; Kyrpides, Nikos C; Ovchinnikova, Galina; Richardson, Paul; De Vos, Willem M.; Smidt, Hauke; Zoetendal, Erwin G.

    2011-01-01

    Victivallis vadensis ATCC BAA-548 represents the first cultured representative from the novel phylum Lentisphaerae, a deep-branching bacterial lineage. Few cultured bacteria from this phylum are known, and V. vadensis therefore represents an important organism for evolutionary studies. V. vadensis is a strictly anaerobic sugar-fermenting isolate from the human gastro-intestinal tract.

  4. Loss of Sirt1 Function Improves Intestinal Anti-Bacterial Defense and Protects from Colitis-Induced Colorectal Cancer

    PubMed Central

    Lo Sasso, Giuseppe; Ryu, Dongryeol; Mouchiroud, Laurent; Fernando, Samodha C.; Anderson, Christopher L.; Katsyuba, Elena; Piersigilli, Alessandra; Hottiger, Michael O.; Schoonjans, Kristina; Auwerx, Johan

    2014-01-01

    Dysfunction of Paneth and goblet cells in the intestine contributes to inflammatory bowel disease (IBD) and colitis-associated colorectal cancer (CAC). Here, we report a role for the NAD+-dependent histone deacetylase SIRT1 in the control of anti-bacterial defense. Mice with an intestinal specific Sirt1 deficiency (Sirt1int−/−) have more Paneth and goblet cells with a consequent rearrangement of the gut microbiota. From a mechanistic point of view, the effects on mouse intestinal cell maturation are mediated by SIRT1-dependent changes in the acetylation status of SPDEF, a master regulator of Paneth and goblet cells. Our results suggest that targeting SIRT1 may be of interest in the management of IBD and CAC. PMID:25013930

  5. Human milk hyaluronan enhances innate defense of the intestinal epithelium.

    PubMed

    Hill, David R; Rho, Hyunjin K; Kessler, Sean P; Amin, Ripal; Homer, Craig R; McDonald, Christine; Cowman, Mary K; de la Motte, Carol A

    2013-10-04

    Breast-feeding is associated with enhanced protection from gastrointestinal disease in infants, mediated in part by an array of bioactive glycan components in milk that act through molecular mechanisms to inhibit enteric pathogen infection. Human milk contains hyaluronan (HA), a glycosaminoglycan polymer found in virtually all mammalian tissues. We have shown that synthetic HA of a specific size range promotes expression of antimicrobial peptides in intestinal epithelium. We hypothesize that hyaluronan from human milk also enhances innate antimicrobial defense. Here we define the concentration of HA in human milk during the first 6 months postpartum. Importantly, HA isolated from milk has a biological function. Treatment of HT-29 colonic epithelial cells with human milk HA at physiologic concentrations results in time- and dose-dependent induction of the antimicrobial peptide human β-defensin 2 and is abrogated by digestion of milk HA with a specific hyaluronidase. Milk HA induction of human β-defensin 2 expression is also reduced in the presence of a CD44-blocking antibody and is associated with a specific increase in ERK1/2 phosphorylation, suggesting a role for the HA receptor CD44. Furthermore, oral administration of human milk-derived HA to adult, wild-type mice results in induction of the murine Hβ D2 ortholog in intestinal mucosa and is dependent upon both TLR4 and CD44 in vivo. Finally, treatment of cultured colonic epithelial cells with human milk HA enhances resistance to infection by the enteric pathogen Salmonella typhimurium. Together, our observations suggest that maternally provided HA stimulates protective antimicrobial defense in the newborn.

  6. Smoking Cessation Induces Profound Changes in the Composition of the Intestinal Microbiota in Humans

    PubMed Central

    Biedermann, Luc; Zeitz, Jonas; Mwinyi, Jessica; Sutter-Minder, Eveline; Rehman, Ateequr; Ott, Stephan J.; Steurer-Stey, Claudia; Frei, Anja; Frei, Pascal; Scharl, Michael; Loessner, Martin J.; Vavricka, Stephan R.; Fried, Michael; Schreiber, Stefan; Schuppler, Markus; Rogler, Gerhard

    2013-01-01

    Background The human intestinal microbiota is a crucial factor in the pathogenesis of various diseases, such as metabolic syndrome or inflammatory bowel disease (IBD). Yet, knowledge about the role of environmental factors such as smoking (which is known to influence theses aforementioned disease states) on the complex microbial composition is sparse. We aimed to investigate the role of smoking cessation on intestinal microbial composition in 10 healthy smoking subjects undergoing controlled smoking cessation. Methods During the observational period of 9 weeks repetitive stool samples were collected. Based on abundance of 16S rRNA genes bacterial composition was analysed and compared to 10 control subjects (5 continuing smokers and 5 non-smokers) by means of Terminal Restriction Fragment Length Polymorphism analysis and high-throughput sequencing. Results Profound shifts in the microbial composition after smoking cessation were observed with an increase of Firmicutes and Actinobacteria and a lower proportion of Bacteroidetes and Proteobacteria on the phylum level. In addition, after smoking cessation there was an increase in microbial diversity. Conclusions These results indicate that smoking is an environmental factor modulating the composition of human gut microbiota. The observed changes after smoking cessation revealed to be similar to the previously reported differences in obese compared to lean humans and mice respectively, suggesting a potential pathogenetic link between weight gain and smoking cessation. In addition they give rise to a potential association of smoking status and the course of IBD. PMID:23516617

  7. Small Intestinal Bacterial Overgrowth and Irritable Bowel Syndrome: A Bridge between Functional Organic Dichotomy.

    PubMed

    Ghoshal, Uday C; Shukla, Ratnakar; Ghoshal, Ujjala

    2017-03-15

    The pathogenesis of irritable bowel syndrome (IBS), once thought to be largely psychogenic in origin, is now understood to be multifactorial. One of the reasons for this paradigm shift is the realization that gut dysbiosis, including small intestinal bacterial overgrowth (SIBO), causes IBS symptoms. Between 4% and 78% of patients with IBS and 1% and 40% of controls have SIBO; such wide variations in prevalence might result from population differences, IBS diagnostic criteria, and, most importantly, methods to diagnose SIBO. Although quantitative jejunal aspirate culture is considered the gold standard for the diagnosis of SIBO, noninvasive hydrogen breath tests have been popular. Although the glucose hydrogen breath test is highly specific, its sensitivity is low; in contrast, the early-peak criteria in the lactulose hydrogen breath test are highly nonspecific. Female gender, older age, diarrhea-predominant IBS, bloating and flatulence, proton pump inhibitor and narcotic intake, and low hemoglobin are associated with SIBO among IBS patients. Several therapeutic trials targeting gut microbes using antibiotics and probiotics have further demonstrated that not all symptoms in patients with IBS originate in the brain but rather in the gut, providing support for the micro-organic basis of IBS. A recent proof-of-concept study showing the high frequency of symptom improvement in patients with IBS with SIBO further supports this hypothesis.

  8. Small Intestinal Bacterial Overgrowth in Patients with Refractory Functional Gastrointestinal Disorders

    PubMed Central

    Shimura, Shino; Ishimura, Norihisa; Mikami, Hironobu; Okimoto, Eiko; Uno, Goichi; Tamagawa, Yuji; Aimi, Masahito; Oshima, Naoki; Sato, Shuichi; Ishihara, Shunji; Kinoshita, Yoshikazu

    2016-01-01

    Background/Aims Small intestinal bacterial overgrowth (SIBO) is considered to be involved in the pathogenesis of functional gastrointestinal disorders (FGID). However, the prevalence and clinical conditions of SIBO in patients with FGID remain to be fully elucidated. Here, we examined the frequency of SIBO in patients with refractory FGID. Methods We prospectively enrolled patients with refractory FGID based on Rome III criteria. A glucose hydrogen breath test (GHBT) was performed using a gas analyzer after an overnight fast, with breath hydrogen concentration measured at baseline and every 15 minutes after administration of glucose for a total of 3 hours. A peak hydrogen value ≥ 10 ppm above the basal value between 60 and 120 minutes after administration of glucose was diagnosed as SIBO. Results A total of 38 FGID patients, including 11 with functional dyspepsia (FD), 10 with irritable bowel syndrome (IBS), and 17 with overlapping with FD and IBS, were enrolled. Of those, 2 (5.3%) were diagnosed with SIBO (one patient diagnosed with FD; the other with overlapping FD and IBS). Their symptoms were clearly improved and breath hydrogen levels decreased to normal following levofloxacin administration for 7 days. Conclusions Two patients initially diagnosed with FD and IBS were also diagnosed with SIBO as assessed by GHBT. Although the frequency of SIBO is low among patients with FGID, it may be important to be aware of SIBO as differential diagnosis when examining patients with refractory gastrointestinal symptoms, especially bloating, as a part of routine clinical care. PMID:26554916

  9. Intestinal Dysbiosis, Gut Hyperpermeability and Bacterial Translocation: Missing Links Between Depression, Obesity and Type 2 Diabetes.

    PubMed

    Slyepchenko, Anastasiya; Maes, Michael; Machado-Vieira, Rodrigo; Anderson, George; Solmi, Marco; Sanz, Yolanda; Berk, Michael; Köhler, Cristiano A; Carvalho, André F

    2016-01-01

    The comorbid prevalence of major depressive disorder (MDD) with obesity and type II diabetes mellitus reflects the existence of a subset of individuals with a complex common pathophysiology and overlapping risk factors. Such comorbid disease presentations imply a number of difficulties, including: decreased treatment responsivity and adherence; altered glycemic control and increased risk of wider medical complications. A number of factors link MDD to metabolic-associated disorders, including: higher rates of shared risk factors such as poor diet and physical inactivity and biological elements including increased inflammation; insulin resistance; oxidative and nitrosative stress; and mitochondrial dysfunction. All of these biological factors have been extensively investigated in the pathophysiology of obesity and type 2 diabetes mellitus as well as MDD. In this review, we aim to: (1) overview the epidemiological links between MDD, obesity and type 2 diabetes mellitus; (2) discuss the role of synergistic neurotoxic effects in MDD comorbid with obesity, and type 2 diabetes mellitus; (3) review evidence of intestinal dysbiosis, leaky gut and increased bacterial translocation, in the pathophysiology of MDD, obesity and type 2 diabetes mellitus; and (4) propose a model in which the gut-brain axis could play a pivotal role in the comorbidity of these disorders.

  10. Small Intestinal Bacterial Overgrowth and Irritable Bowel Syndrome: A Bridge between Functional Organic Dichotomy

    PubMed Central

    Ghoshal, Uday C.; Shukla, Ratnakar; Ghoshal, Ujjala

    2017-01-01

    The pathogenesis of irritable bowel syndrome (IBS), once thought to be largely psychogenic in origin, is now understood to be multifactorial. One of the reasons for this paradigm shift is the realization that gut dysbiosis, including small intestinal bacterial overgrowth (SIBO), causes IBS symptoms. Between 4% and 78% of patients with IBS and 1% and 40% of controls have SIBO; such wide variations in prevalence might result from population differences, IBS diagnostic criteria, and, most importantly, methods to diagnose SIBO. Although quantitative jejunal aspirate culture is considered the gold standard for the diagnosis of SIBO, noninvasive hydrogen breath tests have been popular. Although the glucose hydrogen breath test is highly specific, its sensitivity is low; in contrast, the early-peak criteria in the lactulose hydrogen breath test are highly nonspecific. Female gender, older age, diarrhea-predominant IBS, bloating and flatulence, proton pump inhibitor and narcotic intake, and low hemoglobin are associated with SIBO among IBS patients. Several therapeutic trials targeting gut microbes using antibiotics and probiotics have further demonstrated that not all symptoms in patients with IBS originate in the brain but rather in the gut, providing support for the micro-organic basis of IBS. A recent proof-of-concept study showing the high frequency of symptom improvement in patients with IBS with SIBO further supports this hypothesis. PMID:28274108

  11. Nonsteroidal antiinflammatory drug-induced intestinal inflammation in humans

    SciTech Connect

    Bjarnason, I.; Zanelli, G.; Smith, T.; Prouse, P.; Williams, P.; Smethurst, P.; Delacey, G.; Gumpel, M.J.; Levi, A.J.

    1987-09-01

    This study examines the effects of nonsteroidal antiinflammatory drugs on the small intestine in humans. Using an /sup 111/In-leukocyte technique in patients with rheumatoid arthritis (n = 90) and osteoarthritis (n = 7), it appears that nonsteroidal antiinflammatory drugs cause small intestinal inflammation in two-thirds of patients on long-term treatment and on discontinuation, the inflammation may persist for up to 16 mo. The prevalence and magnitude of the intestinal inflammation was unrelated to the type and dose of nonsteroidal drugs and previous or concomitant second-line drug treatment. There was a significant inverse correlation (r = -0.29, p less than 0.05) between fecal /sup 111/In excretion and hemoglobin levels in patients treated with nonsteroidal antiinflammatory drugs. The kinetics of fecal indium 111 excretion in patients treated with nonsteroidal antiinflammatory drugs was almost identical to that of patients with small bowel Crohn's disease. Eighteen patients on nonsteroidal antiinflammatory drugs underwent a radiologic examination of the small bowel and 3 were found to have asymptomatic ileal disease with ulceration and strictures. Nineteen patients on nonsteroidal antiinflammatory drugs, 20 healthy controls, and 13 patients with Crohn's ileitis underwent a dual radioisotopic ileal function test with tauro 23 (/sup 75/Se) selena-25-homocholic acid and cobalt 58-labeled cyanocobalamine. On day 4, more than half of the patients with rheumatoid arthritis had evidence of bile acid malabsorption, but the ileal dysfunction was much milder than seen in patients with Crohn's ileitis.

  12. Intestinal microbiota modulates gluten-induced immunopathology in humanized mice.

    PubMed

    Galipeau, Heather J; McCarville, Justin L; Huebener, Sina; Litwin, Owen; Meisel, Marlies; Jabri, Bana; Sanz, Yolanda; Murray, Joseph A; Jordana, Manel; Alaedini, Armin; Chirdo, Fernando G; Verdu, Elena F

    2015-11-01

    Celiac disease (CD) is an immune-mediated enteropathy triggered by gluten in genetically susceptible individuals. The recent increase in CD incidence suggests that additional environmental factors, such as intestinal microbiota alterations, are involved in its pathogenesis. However, there is no direct evidence of modulation of gluten-induced immunopathology by the microbiota. We investigated whether specific microbiota compositions influence immune responses to gluten in mice expressing the human DQ8 gene, which confers moderate CD genetic susceptibility. Germ-free mice, clean specific-pathogen-free (SPF) mice colonized with a microbiota devoid of opportunistic pathogens and Proteobacteria, and conventional SPF mice that harbor a complex microbiota that includes opportunistic pathogens were used. Clean SPF mice had attenuated responses to gluten compared to germ-free and conventional SPF mice. Germ-free mice developed increased intraepithelial lymphocytes, markers of intraepithelial lymphocyte cytotoxicity, gliadin-specific antibodies, and a proinflammatory gliadin-specific T-cell response. Antibiotic treatment, leading to Proteobacteria expansion, further enhanced gluten-induced immunopathology in conventional SPF mice. Protection against gluten-induced immunopathology in clean SPF mice was reversed after supplementation with a member of the Proteobacteria phylum, an enteroadherent Escherichia coli isolated from a CD patient. The intestinal microbiota can both positively and negatively modulate gluten-induced immunopathology in mice. In subjects with moderate genetic susceptibility, intestinal microbiota changes may be a factor that increases CD risk.

  13. Diversity of human small intestinal Streptococcus and Veillonella populations.

    PubMed

    van den Bogert, Bartholomeus; Erkus, Oylum; Boekhorst, Jos; de Goffau, Marcus; Smid, Eddy J; Zoetendal, Erwin G; Kleerebezem, Michiel

    2013-08-01

    Molecular and cultivation approaches were employed to study the phylogenetic richness and temporal dynamics of Streptococcus and Veillonella populations in the small intestine. Microbial profiling of human small intestinal samples collected from four ileostomy subjects at four time points displayed abundant populations of Streptococcus spp. most affiliated with S. salivarius, S. thermophilus, and S. parasanguinis, as well as Veillonella spp. affiliated with V. atypica, V. parvula, V. dispar, and V. rogosae. Relative abundances varied per subject and time of sampling. Streptococcus and Veillonella isolates were cultured using selective media from ileostoma effluent samples collected at two time points from a single subject. The richness of the Streptococcus and Veillonella isolates was assessed at species and strain level by 16S rRNA gene sequencing and genetic fingerprinting, respectively. A total of 160 Streptococcus and 37 Veillonella isolates were obtained. Genetic fingerprinting differentiated seven Streptococcus lineages from ileostoma effluent, illustrating the strain richness within this ecosystem. The Veillonella isolates were represented by a single phylotype. Our study demonstrated that the small intestinal Streptococcus populations displayed considerable changes over time at the genetic lineage level because only representative strains of a single Streptococcus lineage could be cultivated from ileostoma effluent at both time points.

  14. Intestinal Microbiota Modulates Gluten-Induced Immunopathology in Humanized Mice

    PubMed Central

    Galipeau, Heather J.; McCarville, Justin L.; Huebener, Sina; Litwin, Owen; Meisel, Marlies; Jabri, Bana; Sanz, Yolanda; Murray, Joseph A.; Jordana, Manel; Alaedini, Armin; Chirdo, Fernando G.; Verdu, Elena F.

    2016-01-01

    Celiac disease (CD) is an immune-mediated enteropathy triggered by gluten in genetically susceptible individuals. The recent increase in CD incidence suggests that additional environmental factors, such as intestinal microbiota alterations, are involved in its pathogenesis. However, there is no direct evidence of modulation of gluten-induced immunopathology by the microbiota. We investigated whether specific microbiota compositions influence immune responses to gluten in mice expressing the human DQ8 gene, which confers moderate CD genetic susceptibility. Germ-free mice, clean specific-pathogen-free (SPF) mice colonized with a microbiota devoid of opportunistic pathogens and Proteobacteria, and conventional SPF mice that harbor a complex microbiota that includes opportunistic pathogens were used. Clean SPF mice had attenuated responses to gluten compared to germ-free and conventional SPF mice. Germ-free mice developed increased intraepithelial lymphocytes, markers of intraepithelial lymphocyte cytotoxicity, gliadin-specific antibodies, and a proinflammatory gliadin-specific T-cell response. Antibiotic treatment, leading to Proteobacteria expansion, further enhanced gluten-induced immunopathology in conventional SPF mice. Protection against gluten-induced immunopathology in clean SPF mice was reversed after supplementation with a member of the Proteobacteria phylum, an enteroadherent Escherichia coli isolated from a CD patient. The intestinal microbiota can both positively and negatively modulate gluten-induced immunopathology in mice. In subjects with moderate genetic susceptibility, intestinal microbiota changes may be a factor that increases CD risk. PMID:26456581

  15. [Influencing factors on infections of human intestinal helminthes in suburb of Shangyu City].

    PubMed

    Song-Lin, Hu

    2011-06-01

    The infections of human intestinal helminthes and socioeconomic status were investigated in suburb of Shangyu City in 1990 and 2005, respectively. The results showed that the economic status, the save drinking water and latrines, working environment, and health habits and consciousness of the residents improved obviously. The infection rate of intestinal helminthes decreased significantly and the prevalence of intestinal helminthosis was controlled effectively.

  16. Human milk and infant intestinal mucosal glycans guide succession of the neonatal intestinal microbiota.

    PubMed

    Newburg, David S; Morelli, Lorenzo

    2015-01-01

    Infants begin acquiring intestinal microbiota at parturition. Initial colonization by pioneer bacteria is followed by active succession toward a dynamic ecosystem. Keystone microbes engage in reciprocal transkingdom communication with the host, which is essential for human homeostasis and health; therefore, these bacteria should be considered mutualists rather than commensals. This review discusses the maternal role in providing infants with functional and stable microbiota. The initial fecal inoculum of microbiota results from the proximity of the birth canal and anus; the biological significance of this anatomic proximity could underlie observed differences in microbiota between vaginal and cesarean birth. Secondary sources of inocula include mouths and skin of kin, animals and objects, and the human milk microbiome, but guiding microbial succession may be a primary role of human milk. The unique glycans of human milk cannot be digested by the infant, but are utilized by mutualist bacteria. These prebiotic glycans support expansion of mutualist microbiota, which manifests as differences in microbiota among breastfed and artificially fed infants. Human milk glycans vary by maternal genotype. Milks of genetically distinct mothers and variations in infant mucosal glycan expression support discrete microbiota. Early colonization may permanently influence microbiota composition and function, with ramifications for health.

  17. Adenoviruses in Lymphocytes of the Human Gastro-Intestinal Tract

    PubMed Central

    Roy, Soumitra; Calcedo, Roberto; Medina-Jaszek, Angelica; Keough, Martin; Peng, Hui; Wilson, James M.

    2011-01-01

    Objective Persistent adenoviral shedding in stools is known to occur past convalescence following acute adenoviral infections. We wished to establish the frequency with which adenoviruses may colonize the gut in normal human subjects. Methods The presence of adenoviral DNA in intestinal specimens obtained at surgery or autopsy was tested using a nested PCR method. The amplified adenoviral DNA sequences were compared to each other and to known adenoviral species. Lamina propria lymphocytes (LPLs) were isolated from the specimens and the adenoviral copy numbers in the CD4+ and CD8+ fractions were determined by quantitative PCR. Adenoviral gene expression was tested by amplification of adenoviral mRNA. Results Intestinal tissue from 21 of 58 donors and LPLs from 21 of 24 donors were positive for the presence of adenoviral DNA. The majority of the sequences could be assigned to adenoviral species E, although species B and C sequences were also common. Multiple sequences were often present in the same sample. Forty-one non-identical sequences were identified from 39 different tissue donors. Quantitative PCR for adenoviral DNA in CD4+ and CD8+ fractions of LPLs showed adenoviral DNA to be present in both cell types and ranged from a few hundred to several million copies per million cells on average. Active adenoviral gene expression as evidenced by the presence of adenoviral messenger RNA in intestinal lymphocytes was demonstrated in 9 of the 11 donors tested. Conclusion Adenoviral DNA is highly prevalent in lymphocytes from the gastro-intestinal tract indicating that adenoviruses may be part of the normal gut flora. PMID:21980361

  18. Small intestinal bacterial overgrowth and thiamine deficiency after Roux-en-Y gastric bypass surgery in obese patients.

    PubMed

    Lakhani, Shilen V; Shah, Hiral N; Alexander, Kenneth; Finelli, Frederick C; Kirkpatrick, John R; Koch, Timothy R

    2008-05-01

    It has been proposed that thiamine deficiency after gastric bypass surgery in obese patients results from prolonged nausea and emesis. We hypothesized that thiamine deficiency is induced by altered gut ecology. This report includes 2 retrospective studies of obese patients who underwent Roux-en-Y gastric bypass surgery at our institution from 1999 to 2005. In the first study, 80 patients (52 women and 28 men) had measurement of whole-blood thiamine diphosphate level and serum folate level. In these 80 patients, 39 (49%) had thiamine diphosphate levels less than the lower limit of the reference range, and 28 (72%) of the 39 had folate levels higher than the upper limit of the reference range, an indicator of small intestinal bacterial overgrowth. In 41 patients with normal thiamine levels, only 14 (34%) had folate levels higher than the upper limit of the reference range (chi(2) test, P < .01). In the second study, 21 patients (17 women and 4 men) had thiamine diphosphate levels less than the lower limit of the reference range and abnormal glucose-hydrogen breath tests, consistent with small intestinal bacterial overgrowth. Fifteen patients received oral thiamine supplements, but repeated thiamine levels remained low in all 15. Nine of these patients then received oral antibiotic therapy; repeated thiamine levels were found to be normal in all 9 patients. These results support the hypothesis that small intestinal bacterial overgrowth results from altered gut ecology and induces thiamine deficiency after gastric bypass surgery in obese patients.

  19. Functional Characterization of Cholera Toxin Inhibitors Using Human Intestinal Organoids.

    PubMed

    Zomer-van Ommen, Domenique D; Pukin, Aliaksei V; Fu, Ou; Quarles van Ufford, Linda H C; Janssens, Hettie M; Beekman, Jeffrey M; Pieters, Roland J

    2016-07-28

    Preclinical drug testing in primary human cell models that recapitulate disease can significantly reduce animal experimentation and time-to-the-clinic. We used intestinal organoids to quantitatively study the potency of multivalent cholera toxin inhibitors. The method enabled the determination of IC50 values over a wide range of potencies (15 pM to 9 mM). The results indicate for the first time that an organoid-based swelling assay is a useful preclinical method to evaluate inhibitor potencies of drugs that target pathogen-derived toxins.

  20. [Progress in the knowledge of the intestinal human microbiota].

    PubMed

    Robles-Alonso, Virginia; Guarner, Francisco

    2013-01-01

    New sequencing technologies together with the development of bio-informatics allow a description of the full spectrum of the microbial communities that inhabit the human intestinal tract, as well as their functional contributions to host health. Most community members belong to the domain Bacteria, but Archaea, Eukaryotes (yeasts and protists), and Viruses are also present. Only 7 to 9 of the 55 known divisions or phyla of the domain Bacteria are detected in faecal or mucosal samples from the human gut. Most taxa belong to just two divisions: Bacteroidetes and Firmicutes, and the other divisions that have been consistently found are Proteobacteria, Actinobacteria, Fusobacteria, and Verrucomicrobia. Bacteroides, Faecalibacterium and Bifidobacterium are the most abundant genera but their relative proportion is highly variable across individuals. Full metagenomic analysis has identified more than 5 million non-redundant microbial genes encoding up to 20,000 biological functions related with life in the intestinal habitat. The overall structure of predominant genera in the human gut can be assigned into three robust clusters, which are known as "enterotypes". Each of the three enterotypes is identifiable by the levels of one of three genera: Bacteroides (enterotype 1), Prevotella (enterotype 2) and Ruminococcus (enterotype 3). This suggests that microbiota variations across individuals are stratified, not continuous. Next steps include the identification of changes that may play a role in certain disease states. A better knowledge of the contributions of microbial symbionts to host health will help in the design of interventions to improve symbiosis and combat disease.

  1. Derivation of Intestinal Organoids from Human Induced Pluripotent Stem Cells for Use as an Infection System.

    PubMed

    Forbester, Jessica L; Hannan, Nicholas; Vallier, Ludovic; Dougan, Gordon

    2016-08-31

    Intestinal human organoids (iHOs) provide an effective system for studying the intestinal epithelium and its interaction with various stimuli. By using combinations of different signaling factors, human induced pluripotent stem cells (hIPSCs) can be driven to differentiate down the intestinal lineage. Here, we describe the process for this differentiation, including the derivation of hindgut from hIPSCs, embedding hindgut into a pro-intestinal culture system and passaging the resulting iHOs. We then describe how to carry out microinjections to introduce bacteria to the apical side of the intestinal epithelial cells (IECs).

  2. A novel method for the culture and polarized stimulation of human intestinal mucosa explants.

    PubMed

    Tsilingiri, Katerina; Sonzogni, Angelica; Caprioli, Flavio; Rescigno, Maria

    2013-05-01

    Few models currently exist to realistically simulate the complex human intestine's micro-environment, where a variety of interactions take place. Proper homeostasis directly depends on these interactions, as they shape an entire immunological response inducing tolerance against food antigens while at the same time mounting effective immune responses against pathogenic microbes accidentally ingested with food. Intestinal homeostasis is preserved also through various complex interactions between the microbiota (including food-associated beneficial bacterial strains) and the host, that regulate the attachment/degradation of mucus, the production of antimicrobial peptides by the epithelial barrier, and the "education" of epithelial cells' that controls the tolerogenic or immunogenic phenotype of unique, gut-resident lymphoid cells' populations. These interactions have been so far very difficult to reproduce with in vitro assays using either cultured cell lines or peripheral blood mononuclear cells. In addition, mouse models differ substantially in components of the intestinal mucosa (mucus layer organization, commensal bacteria community) with respect to the human gut. Thus, studies of a variety of treatments to be brought in the clinics for important stress-related or pathological conditions such as irritable bowel syndrome, inflammatory bowel disease or colorectal cancer have been difficult to carry out. To address these issues, we developed a novel system that enables us to stimulate explants of human intestinal mucosa that retain their in situ conditioning by the host microbiota and immune response, in a polarized fashion. Polarized apical stimulation is of great importance for the outcome of the elicited immune response. It has been repeatedly shown that the same stimuli can produce completely different responses when they bypass the apical face of the intestinal epithelium, stimulating epithelial cells basolaterally or coming into direct contact with lamina

  3. Herbal Therapy Is Equivalent to Rifaximin for the Treatment of Small Intestinal Bacterial Overgrowth

    PubMed Central

    Chedid, Victor; Dhalla, Sameer; Clarke, John O.; Roland, Bani Chander; Dunbar, Kerry B.; Koh, Joyce; Justino, Edmundo; Tomakin, Eric

    2014-01-01

    Objective: Patients with small intestine bacterial overgrowth (SIBO) have chronic intestinal and extraintestinal symptomatology which adversely affects their quality of life. Present treatment of SIBO is limited to oral antibiotics with variable success. A growing number of patients are interested in using complementary and alternative therapies for their gastrointestinal health. The objective was to determine the remission rate of SIBO using either the antibiotic rifaximin or herbals in a tertiary care referral gastroenterology practice. Design: One hundred and four patients who tested positive for newly diagnosed SIBO by lactulose breath testing (LBT) were offered either rifaximin 1200 mg daily vs herbal therapy for 4 weeks with repeat LBT post-treatment. Results: Three hundred ninety-six patients underwent LBT for suspected SIBO, of which 251 (63.4%) were positive 165 underwent treatment and 104 had a follow-up LBT. Of the 37 patients who received herbal therapy, 17 (46%) had a negative follow-up LBT compared to 23/67 (34%) of rifaximin users (P=.24). The odds ratio of having a negative LBT after taking herbal therapy as compared to rifaximin was 1.85 (CI=0.77-4.41, P=.17) once adjusted for age, gender, SIBO risk factors and IBS status. Fourteen of the 44 (31.8%) rifaximin non-responders were offered herbal rescue therapy, with 8 of the 14 (57.1%) having a negative LBT after completing the rescue herbal therapy, while 10 non-responders were offered triple antibiotics with 6 responding (60%, P=.89). Adverse effects were reported among the rifaximin treated arm including 1 case of anaphylaxis, 2 cases of hives, 2 cases of diarrhea and 1 case of Clostridium difficile. Only one case of diarrhea was reported in the herbal therapy arm, which did not reach statistical significance (P=.22). Conclusion: SIBO is widely prevalent in a tertiary referral gastroenterology practice. Herbal therapies are at least as effective as rifaximin for resolution of SIBO by LBT. Herbals

  4. Human intestinal epithelial cells produce proinflammatory cytokines in response to infection in a SCID mouse-human intestinal xenograft model of amebiasis.

    PubMed Central

    Seydel, K B; Li, E; Swanson, P E; Stanley, S L

    1997-01-01

    The protozoan parasite Entamoeba histolytica causes amebic dysentery and amebic liver abscess, diseases associated with significant morbidity and mortality worldwide. E. histolytica infection appears to involve the initial attachment of amebic trophozoites to intestinal epithelial cells, followed by lysis of these cells and subsequent invasion into the submucosa. A recent in vitro study (L. Eckmann, S. L. Reed, J. R. Smith, and M. F. Kagnoff, J. Clin. Invest. 96:1269-1279, 1995) demonstrated that incubation of E. histolytica trophozoites with epithelial cell lines results in epithelial cell production of inflammatory cytokines, including interleukin-1 (IL-1) and IL-8, suggesting that intestinal epithelial cell production of cytokines might play a role in the inflammatory response and tissue damage seen in intestinal amebiasis. To determine whether intestinal epithelial cell production of IL-1 and IL-8 occurs in response to E. histolytica infection in vivo and as an approach to studying the specific interactions between amebic trophozoites and human intestine, we used a SCID mouse-human intestinal xenograft (SCID-HU-INT) model of disease, where human intestinal xenografts were infected with virulent E. histolytica trophozoites. Infection of xenografts with E. histolytica trophozoites resulted in extensive tissue damage, which was associated with the development of an early inflammatory response composed primarily of neutrophils. Using oligonucleotide primers that specifically amplify human IL-1beta and IL-8, we could demonstrate by reverse transcription PCR that mRNA for both IL-1beta and IL-8 is produced by human intestinal xenografts in response to amebic infection. The increase in human intestinal IL-1beta and IL-8 in response to invasive amebiasis was confirmed by enzyme-linked immunosorbent assays specific for human IL-1beta and IL-8. Using immunohistochemistry, we confirmed that human intestinal epithelial cells were the source of IL-8 in infected xenografts

  5. Ovine intestinal adenocarcinomas: histologic and phenotypic comparison with human colon cancer.

    PubMed

    Munday, John S; Brennan, Moira M; Jaber, Azhar M; Kiupel, Matti

    2006-04-01

    Approximately 7% of old, unthrifty sheep (Ovis aries) in New Zealand have intestinal adenocarcinomas. To investigate whether these sheep might be used as a model of human colonic neoplasia, the biologic behavior and histologic appearance of ovine intestinal adenocarcinomas were compared with those reported for human colonic adenocarcinomas. We collected 50 intestinal tracts with grossly visible intestinal neoplasia from slaughtered sheep. Neoplasms were assessed using World Health Organization guidelines for assessment of human colonic adenocarcinomas. All ovine adenocarcinomas developed in the small intestine. In contrast, only 4% of human intestinal tumors develop at this location, whereas the majority develop in the colon. A visible polyp is present within 89% of human colonic adenocarcinomas, whereas polyps were present in only 46% of the ovine neoplasms. Intestinal wall infiltration by the neoplastic cells and rates of lymph node (84% in sheep; 61% in humans) and distant (52% in sheep; 17% in humans) metastases were comparable between ovine and human adenocarcinomas. However, ovine adenocarcinomas developed more peritoneal and fewer hepatic metastases than human adenocarcinomas. Histologic grading of ovine tumors revealed cell differentiation similar to that reported within human colonic adenocarcinomas. In conclusion, ovine intestinal adenocarcinomas, like human colonic adenocarcinomas, typically arise spontaneously and consistently develop widespread metastases. In addition, tumors appear histologically similar between these species. Therefore, sheep may provide a model of advanced human colonic cancer, possibly allowing evaluation of novel therapeutics and surgical procedures.

  6. Alpha2 adrenoceptors regulate proliferation of human intestinal epithelial cells

    PubMed Central

    Schaak, S; Cussac, D; Cayla, C; Devedjian, J; Guyot, R; Paris, H; Denis, C

    2000-01-01

    BACKGROUND AND AIMS—Previous studies on rodents have suggested that catecholamines stimulate proliferation of the intestinal epithelium through activation of α2 adrenoceptors located on crypt cells. The occurrence of this effect awaits demonstration in humans and the molecular mechanisms involved have not yet been elucidated. Here, we examined the effect of α2 agonists on a clone of Caco2 cells expressing the human α2A adrenoceptor.
METHODS—Cells were transfected with a bicistronic plasmid containing the α2C10 and neomycin phosphotransferase genes. G418 resistant clones were assayed for receptor expression using radioligand binding. Receptor functionality was assessed by testing its ability to couple Gi proteins and to inhibit cAMP production. Mitogen activated protein kinase (MAPK) phosphorylation was followed by western blot, and cell proliferation was estimated by measuring protein and DNA content.
RESULTS—Permanent transfection of Caco2 cells allowed us to obtain a clone (Caco2-3B) expressing α2A adrenoceptors at a density similar to that found in normal human intestinal epithelium. Caco2-3B retained morphological features and brush border enzyme expression characteristic of enterocytic differentiation. The receptor was coupled to Gi2/Gi3 proteins and its stimulation caused marked diminution of forskolin induced cAMP production. Treatment of Caco2-3B with UK14304 (α2 agonist) induced a rapid increase in the phosphorylation state of MAPK, extracellular regulated protein kinase 1 (Erk1), and 2 (Erk2). This event was totally abolished in pertussis toxin treated cells and in the presence of kinase inhibitors (genistein or PD98059). It was unaffected by protein kinase C downregulation but correlated with a transient increase in Shc tyrosine phosphorylation. Finally, sustained exposure of Caco2-3B to UK14304 resulted in modest but significant acceleration of cell proliferation. None of these effects was observed in the parental cell line Caco2.

  7. Multiscale analysis of the murine intestine for modeling human diseases

    PubMed Central

    Lyons, Jesse; Herring, Charles A.; Banerjee, Amrita; Simmons, Alan J.

    2015-01-01

    When functioning properly, the intestine is one of the key interfaces between the human body and its environment. It is responsible for extracting nutrients from our food and excreting our waste products. It provides an environment for a host of healthful microbes and serves as a first defense against pathogenic ones. These processes require tight homeostatic controls, which are provided by the interactions of a complex mix of epithelial, stromal, neural and immune cells, as well as the resident microflora. This homeostasis can be disrupted by invasive microbes, genetic lesions, and carcinogens, resulting in diseases such Clostridium difficile infection, inflammatory bowel disease (IBD) and cancer. Enormous strides have been made in understanding how this important organ functions in health and disease using everything from cell culture systems to animal models to human tissue samples. This has resulted in better therapies for all of these diseases, but there is still significant room for improvement. In the United States alone, 14000 people per year die of C. difficile, up to 1.6 million people suffer from IBD, and more than 50000 people die every year from colon cancer. Because these and other intestinal diseases arise from complex interactions between the different components of the gut ecosystem, we propose that systems approaches that address this complexity in an integrative manner may eventually lead to improved therapeutics that deliver lasting cures. This review will discuss the use of systems biology for studying intestinal diseases in vivo with particular emphasis on mouse models. Additionally, it will focus on established experimental techniques that have been used to drive this systems-level analysis, and emerging techniques that will push this field forward in the future. PMID:26040649

  8. Binding of diarrheagenic Escherichia coli to 32- to 33-kilodalton human intestinal brush border proteins.

    PubMed Central

    Manjarrez-Hernandez, A; Gavilanes-Parra, S; Chavez-Berrocal, M E; Molina-Lopez, J; Cravioto, A

    1997-01-01

    We have detected human intestinal brush border proteins to which Escherichia coli strains adhere by means of a blotting-nitrocellulose method in which the binding of radiolabeled bacteria to sodium dodecyl sulfate-polyacrylamide gel electrophoresis-separated intestinal cell membranes was evaluated. The brush border fraction contained several polypeptides that bound only adherent E. coli strains. The most prominent and consistent of these proteins had apparent molecular masses of 32 to 33 kDa. Additional polypeptides ranging from 50 to 70, from 105 to 130, and from 180 to 200 kDa were also recognized by adherent E. coli strains, although with less intensity (in accordance with the number of bound bacteria to these polypeptides). Independently of the pattern of adherence (localized [LA], diffuse [DA], or aggregative [AggA]) all HEp-2-adhering strains recognized, with different intensities, the 32- to 33-kDa brush border proteins, whereas nonadhesive strains did not. The relative avidity of an LA strain to bind to the 32- to 33-kDa proteins was approximately seven- and sixfold higher than the binding of strains with aggregative and diffuse adherence, respectively. Thus, it is reasonable to think that LA, DA, and AggA strains have a common adhesin that mediates binding to the 32- to 33-kDa bands. Inhibition experiments using HEp-2 cells demonstrated that isolated 32- to 33-kDa proteins or specific antiserum blocked preferentially bacterial adherence of the LA pattern. Delipidization and protein digestion of the human brush borders confirmed that E. coli bound to structures of a proteinaceous nature. Deglycosylation studies and sodium meta-periodate oxidation of the intestinal cell membranes decreased bacterial binding activity significantly, indicating that E. coli bound to carbohydrate moieties in the glycoproteins. These results suggest that binding of E. coli strains, mainly of the LA phenotype, to the 32- to 33-kDa proteins could play a role in colonization through

  9. Differentiation of epithelial cells to M cells in response to bacterial colonization on the follicle-associated epithelium of Peyer's patch in rat small intestine.

    PubMed

    Chin, Keigi; Onishi, Sachiko; Yuji, Midori; Inamoto, Tetsurou; Qi, Wang-Mei; Warita, Katsuhiko; Yokoyama, Toshifumi; Hoshi, Nobuhiko; Kitagawa, Hiroshi

    2006-10-01

    To clarify the relationship between M cells and intestinal microflora, histoplanimetrical investigation into the bacterial colonization and the differentiation to M cells was carried out in rat Peyer's patch under physiological conditions. The follicle-associated epithelium (FAE), except for the narrow area of apical region, was closely covered with both neighboring intestinal villi and a thick mucous layer, the latter of which also filled the intervillous spaces as well as the space between the FAE and the neighboring intestinal villi. Indigenous bacteria adhered almost constantly to the narrow areas of apical regions of both intestinal villi and the FAE. Bacterial colonies were occasionally located on the basal to middle region of FAE, where M cells also appeared, forming large pockets. When bacterial colonies were located on the basal to middle region of FAE, bacteria with the same morphological characteristics also proliferated in the intervillous spaces neighboring the Peyer's patch. In cases with no bacterial colonies on the basal to middle region of FAE, however, M cells were rare in the FAE. Histoplanimetrical analysis showed the similar distribution pattern of bacterial colonies on the FAE and M cells in the FAE. M cells ultrastructurally engulfed indigenous bacteria, which were then transported to the pockets. These results suggest that indigenous bacterial colonization on the FAE stimulates the differentiation of M cells in the FAE under physiological conditions. The uptake of bacteria by M cells might contribute the regulation of the development of indigenous bacterial colonies in the small intestine.

  10. Utilization of rye as energy source affects bacterial translocation, intestinal viscosity, microbiota composition, and bone mineralization in broiler chickens

    PubMed Central

    Tellez, Guillermo; Latorre, Juan D.; Kuttappan, Vivek A.; Kogut, Michael H.; Wolfenden, Amanda; Hernandez-Velasco, Xochitl; Hargis, Billy M.; Bottje, Walter G.; Bielke, Lisa R.; Faulkner, Olivia B.

    2014-01-01

    Two independent trials were conducted to evaluate the utilization of rye as energy source on bacterial translocation (BT), intestinal viscosity, gut integrity, gut microbiota composition, and bone mineralization, when compared with a traditional cereal (corn) in broiler chickens. In each experiment, day-of-hatch, broiler chickens were randomly assigned to either a corn or a rye diet (n = 20 chickens/group). At 10 d of age, in both experiments, 12 chickens/group were randomly selected, and given an oral gavage dose of fluorescein isothiocyanate dextran (FITC-d). After 2.5 h of oral gavage, blood samples were collected to determine the passage of FITC-d. The liver was collected from each bird to evaluate BT. Duodenum, ileum, and cecum gut sections were collected to evaluate intestinal viscosity and to enumerate gut microbiota. Tibias were collected for observation of bone parameters. Broilers fed with rye showed increased (p < 0.05) intestinal viscosity, BT, and serum FITC-d. Bacterial enumeration revealed that chickens fed with rye had increased the number of total lactic acid bacteria in all three sections of the gastrointestinal tract evaluated when compared to chickens fed with corn. Chickens fed with rye also had significantly higher coliforms in duodenum and ileum, whereas the total number of anaerobes increased only in duodenum. A significant reduction in bone strength and bone mineralization was observed in chickens fed with rye when compared with corn fed chickens. In conclusion, rye evoked mucosal damage in chickens that alter the intestinal viscosity, increased leakage through the intestinal tract, and altered the microbiota composition as well as bone mineralization. Studies to evaluate dietary inclusion of selected DFM candidates that produce exogenous enzymes in rye fed chickens are currently being evaluated. PMID:25309584

  11. Effects of laxative and N-acetylcysteine on mucus accumulation, bacterial load, transit, and inflammation in the cystic fibrosis mouse small intestine.

    PubMed

    De Lisle, Robert C; Roach, Eileen; Jansson, Kyle

    2007-09-01

    The accumulation of mucus in affected organs is characteristic of cystic fibrosis (CF). The CF mouse small intestine has dramatic mucus accumulation and exhibits slower interdigestive intestinal transit. These factors are proposed to play cooperative roles that foster small intestinal bacterial overgrowth (SIBO) and contribute to the innate immune response of the CF intestine. It was hypothesized that decreasing the mucus accumulation would reduce SIBO and might improve other aspects of the CF intestinal phenotype. To test this, solid chow-fed CF mice were treated with an osmotic laxative to improve gut hydration or liquid-fed mice were treated orally with N-acetylcysteine (NAC) to break mucin disulfide bonds. Treatment with laxative or NAC reduced mucus accumulation by 43% and 50%, respectively, as measured histologically as dilation of the intestinal crypts. Laxative and NAC also reduced bacterial overgrowth in the CF intestine by 92% and 63%, respectively. Treatment with laxative normalized small intestinal transit in CF mice, whereas NAC did not. The expression of innate immune response-related genes was significantly reduced in laxative-treated CF mice, whereas there was no significant effect in NAC-treated CF mice. In summary, laxative and NAC treatments of CF mice reduced mucus accumulation to a similar extent, but laxative was more effective than NAC at reducing bacterial load. Eradication of bacterial overgrowth by laxative treatment was associated with normalized intestinal transit and a reduction in the innate immune response. These results suggest that both mucus accumulation and slowed interdigestive small intestinal transit contribute to SIBO in the CF intestine.

  12. Molecular diagnosis of bacterial vaginosis: Does adjustment for total bacterial load or human cellular content improve diagnostic performance?

    PubMed

    Plummer, E L; Garland, S M; Bradshaw, C S; Law, M G; Vodstrcil, L A; Hocking, J S; Fairley, C K; Tabrizi, S N

    2017-02-01

    We investigated the utility of quantitative PCR assays for diagnosis of bacterial vaginosis and found that while the best model utilized bacterial copy number adjusted for total bacterial load (sensitivity=98%, specificity=93%, AUC=0.95[95%CI=0.93,0.97]), adjusting for total bacterial or human cell load did not consistently increase the diagnostic performance of the assays.

  13. Prevalence and treatment of small intestinal bacterial overgrowth in postoperative patients with colorectal cancer

    PubMed Central

    DENG, LIHUA; LIU, YANG; ZHANG, DONGSHENG; LI, YUAN; XU, LIN

    2016-01-01

    To investigate the prevalence of small intestinal bacterial overgrowth (SIBO) in patients with colorectal cancer (CRC) after surgical treatment and observe whether gastrointestinal symptoms may improve with rifaximin, 43 postoperative CRC patients (CRC group) and 30 healthy individuals (normal group) were subjected to the glucose hydrogen breath test (GHBT). All the patients were asked to evaluate the gastrointestinal symptoms using the visual analogue scale (VAS). SIBO-positive patients were administered rifaximin for 10 days on the basis of the original treatment. After the treatment, the patients were asked to undergo GHBT and re-evaluate the gastrointestinal symptoms score (GISS). The results demonstrated that 18 of the 43 postoperative patients with CRC were SIBO-positive (41.86%), which was significantly higher compared with the incidence in normal controls (6.67%) (P<0.05). GISS was higher in SIBO-positive patients (P<0.05). Following rifaximin treatment, 6 of the 18 (33.33%) SIBO-positive patients had improved, as evaluated by GHBT and VAS. Additionally, the GISS in the SIBO-turned-negative group had improved significantly compared with that in the non-turned-negative group (P<0.05). The symptoms of all 18 SIBO-positive patients following rifaximin treatment improved notably, particularly diarrhea (P<0.05). In conclusion, postoperative CRC patients are more likely to develop SIBO compared with healthy individuals, and SIBO may aggravate digestive symptoms. The administration of rifaximin improved the overall gastrointestinal symptoms, particularly diarrhea, in SIBO-positive patients. PMID:27123301

  14. Carriage of λ Latent Virus Is Costly for Its Bacterial Host due to Frequent Reactivation in Monoxenic Mouse Intestine

    PubMed Central

    De Paepe, Marianne; Tournier, Laurent; Moncaut, Elisabeth; Son, Olivier; Langella, Philippe; Petit, Marie-Agnès

    2016-01-01

    Temperate phages, the bacterial viruses able to enter in a dormant prophage state in bacterial genomes, are present in the majority of bacterial strains for which the genome sequence is available. Although these prophages are generally considered to increase their hosts’ fitness by bringing beneficial genes, studies demonstrating such effects in ecologically relevant environments are relatively limited to few bacterial species. Here, we investigated the impact of prophage carriage in the gastrointestinal tract of monoxenic mice. Combined with mathematical modelling, these experimental results provided a quantitative estimation of key parameters governing phage-bacteria interactions within this model ecosystem. We used wild-type and mutant strains of the best known host/phage pair, Escherichia coli and phage λ. Unexpectedly, λ prophage caused a significant fitness cost for its carrier, due to an induction rate 50-fold higher than in vitro, with 1 to 2% of the prophage being induced. However, when prophage carriers were in competition with isogenic phage susceptible bacteria, the prophage indirectly benefited its carrier by killing competitors: infection of susceptible bacteria led to phage lytic development in about 80% of cases. The remaining infected bacteria were lysogenized, resulting overall in the rapid lysogenization of the susceptible lineage. Moreover, our setup enabled to demonstrate that rare events of phage gene capture by homologous recombination occurred in the intestine of monoxenic mice. To our knowledge, this study constitutes the first quantitative characterization of temperate phage-bacteria interactions in a simplified gut environment. The high prophage induction rate detected reveals DNA damage-mediated SOS response in monoxenic mouse intestine. We propose that the mammalian gut, the most densely populated bacterial ecosystem on earth, might foster bacterial evolution through high temperate phage activity. PMID:26871586

  15. An in vivo model of human small intestine using pluripotent stem cells.

    PubMed

    Watson, Carey L; Mahe, Maxime M; Múnera, Jorge; Howell, Jonathan C; Sundaram, Nambirajan; Poling, Holly M; Schweitzer, Jamie I; Vallance, Jefferson E; Mayhew, Christopher N; Sun, Ying; Grabowski, Gregory; Finkbeiner, Stacy R; Spence, Jason R; Shroyer, Noah F; Wells, James M; Helmrath, Michael A

    2014-11-01

    Differentiation of human pluripotent stem cells (hPSCs) into organ-specific subtypes offers an exciting avenue for the study of embryonic development and disease processes, for pharmacologic studies and as a potential resource for therapeutic transplant. To date, limited in vivo models exist for human intestine, all of which are dependent upon primary epithelial cultures or digested tissue from surgical biopsies that include mesenchymal cells transplanted on biodegradable scaffolds. Here, we generated human intestinal organoids (HIOs) produced in vitro from human embryonic stem cells (ESCs) or induced pluripotent stem cells (iPSCs) that can engraft in vivo. These HIOs form mature human intestinal epithelium with intestinal stem cells contributing to the crypt-villus architecture and a laminated human mesenchyme, both supported by mouse vasculature ingrowth. In vivo transplantation resulted in marked expansion and maturation of the epithelium and mesenchyme, as demonstrated by differentiated intestinal cell lineages (enterocytes, goblet cells, Paneth cells, tuft cells and enteroendocrine cells), presence of functional brush-border enzymes (lactase, sucrase-isomaltase and dipeptidyl peptidase 4) and visible subepithelial and smooth muscle layers when compared with HIOs in vitro. Transplanted intestinal tissues demonstrated digestive functions as shown by permeability and peptide uptake studies. Furthermore, transplanted HIO-derived tissue was responsive to systemic signals from the host mouse following ileocecal resection, suggesting a role for circulating factors in the intestinal adaptive response. This model of the human small intestine may pave the way for studies of intestinal physiology, disease and translational studies.

  16. Human and mouse tissue-engineered small intestine both demonstrate digestive and absorptive function.

    PubMed

    Grant, Christa N; Mojica, Salvador Garcia; Sala, Frederic G; Hill, J Ryan; Levin, Daniel E; Speer, Allison L; Barthel, Erik R; Shimada, Hiroyuki; Zachos, Nicholas C; Grikscheit, Tracy C

    2015-04-15

    Short bowel syndrome (SBS) is a devastating condition in which insufficient small intestinal surface area results in malnutrition and dependence on intravenous parenteral nutrition. There is an increasing incidence of SBS, particularly in premature babies and newborns with congenital intestinal anomalies. Tissue-engineered small intestine (TESI) offers a therapeutic alternative to the current standard treatment, intestinal transplantation, and has the potential to solve its biggest challenges, namely donor shortage and life-long immunosuppression. We have previously demonstrated that TESI can be generated from mouse and human small intestine and histologically replicates key components of native intestine. We hypothesized that TESI also recapitulates native small intestine function. Organoid units were generated from mouse or human donor intestine and implanted into genetically identical or immunodeficient host mice. After 4 wk, TESI was harvested and either fixed and paraffin embedded or immediately subjected to assays to illustrate function. We demonstrated that both mouse and human tissue-engineered small intestine grew into an appropriately polarized sphere of intact epithelium facing a lumen, contiguous with supporting mesenchyme, muscle, and stem/progenitor cells. The epithelium demonstrated major ultrastructural components, including tight junctions and microvilli, transporters, and functional brush-border and digestive enzymes. This study demonstrates that tissue-engineered small intestine possesses a well-differentiated epithelium with intact ion transporters/channels, functional brush-border enzymes, and similar ultrastructural components to native tissue, including progenitor cells, whether derived from mouse or human cells.

  17. Distribution of the IgG Fc Receptor, FcRn, in the Human Fetal Intestine

    PubMed Central

    Shah, Uzma; Dickinson, Bonny L.; Blumberg, Richard S.; Simister, Neil E.; Lencer, Wayne I.; Walker, W. Allan

    2010-01-01

    The intestinal Fc receptor, FcRn, functions in the maternofetal transfer of gamma globulin (IgG) in the neonatal rodent. In humans, most of this transfer is presumed to occur in utero via the placenta. Although the fetus swallows amniotic fluid that contains immunoglobulin, it is unknown whether this transfer also occurs via the fetal intestine. A human FcRn has been identified in the syncytiotrophoblast that mediates the maternofetal transfer of antibody. It has also been identified in human fetal intestine and is postulated to function in IgG transport. We hypothesize that the human fetal intestinal FcRn may play a role in IgG transport from the amniotic fluid into the fetal circulation. The aim of this study was to characterize the distribution of the FcRn along the human fetal intestine. Lysates prepared from human fetal intestine and from a nonmalignant human fetal intestinal epithelial cell line (H4) were subjected to Western blot analysis and probed using anti-FcRn antibodies. A 42-kD band, consistent with the known molecular weight of the FcRn, was detected along the human fetal intestine and in H4 cells. Expression of the human FcRn was confirmed with immunohistochemistry. Our study demonstrates the expression of FcRn along the human fetal intestine and in a human nonmalignant fetal intestinal epithelial cell line (H4), which by location indicates that FcRn could play a role in the uptake and transport of IgG in the human fetus. PMID:12538789

  18. A Novel Model of P-Glycoprotein Inhibitor Screening Using Human Small Intestinal Organoids.

    PubMed

    Zhao, Junfang; Zeng, Zhiyang; Sun, Jialiang; Zhang, Yuanjin; Li, Dali; Zhang, Xueli; Liu, Mingyao; Wang, Xin

    2017-03-01

    P-glycoprotein (P-gp), an important efflux transporter in intestine, regulates the bioavailability of orally taken drugs. To develop an in vitro model that preferably mimics the physiological microenvironment of human intestine, we employed the three-dimensionally (3D) cultured organoids from human normal small intestinal epithelium. It was observed that the intestinal crypts could efficiently form cystic organoid structure with the extension of culture time. Furthermore, the physiological expression of ABCB1 was detected at both mRNA and protein levels in cultured organoids. Rhodamine 123 (Rh123), a typical substrate of P-gp, was actively transported across 3D organoids and accumulated in the luminal space. This transport process was also inhibited by verapamil and mitotane. In summary, the above-mentioned model based on human small intestinal 3D organoids is suitable to imitate the small intestinal epithelium and could be used as a novel in vitro model especially for P-gp inhibitor screening.

  19. Intestinal Short Chain Fatty Acids and their Link with Diet and Human Health

    PubMed Central

    Ríos-Covián, David; Ruas-Madiedo, Patricia; Margolles, Abelardo; Gueimonde, Miguel; de los Reyes-Gavilán, Clara G.; Salazar, Nuria

    2016-01-01

    The colon is inhabited by a dense population of microorganisms, the so-called “gut microbiota,” able to ferment carbohydrates and proteins that escape absorption in the small intestine during digestion. This microbiota produces a wide range of metabolites, including short chain fatty acids (SCFA). These compounds are absorbed in the large bowel and are defined as 1-6 carbon volatile fatty acids which can present straight or branched-chain conformation. Their production is influenced by the pattern of food intake and diet-mediated changes in the gut microbiota. SCFA have distinct physiological effects: they contribute to shaping the gut environment, influence the physiology of the colon, they can be used as energy sources by host cells and the intestinal microbiota and they also participate in different host-signaling mechanisms. We summarize the current knowledge about the production of SCFA, including bacterial cross-feedings interactions, and the biological properties of these metabolites with impact on the human health. PMID:26925050

  20. Bacterial expression of human kynurenine 3-monooxygenase: solubility, activity, purification.

    PubMed

    Wilson, K; Mole, D J; Binnie, M; Homer, N Z M; Zheng, X; Yard, B A; Iredale, J P; Auer, M; Webster, S P

    2014-03-01

    Kynurenine 3-monooxygenase (KMO) is an enzyme central to the kynurenine pathway of tryptophan metabolism. KMO has been implicated as a therapeutic target in several disease states, including Huntington's disease. Recombinant human KMO protein production is challenging due to the presence of transmembrane domains, which localise KMO to the outer mitochondrial membrane and render KMO insoluble in many in vitro expression systems. Efficient bacterial expression of human KMO would accelerate drug development of KMO inhibitors but until now this has not been achieved. Here we report the first successful bacterial (Escherichia coli) expression of active FLAG™-tagged human KMO enzyme expressed in the soluble fraction and progress towards its purification.

  1. Helicobacter pylori infection but not small intestinal bacterial overgrowth may play a pathogenic role in rosacea

    PubMed Central

    Federico, A; Ruocco, E; Lo Schiavo, A; Masarone, M; Tuccillo, C; Peccerillo, F; Miranda, A; Romano, L; de Sio, C; de Sio, I; Persico, M; Ruocco, V; Riegler, G; Loguercio, C; Romano, M

    2015-01-01

    Background and aims Recent studies suggest a potential relationship between rosacea and Helicobacter pylori (H. pylori) infection or small intestinal bacterial overgrowth (SIBO), but there is no firm evidence of an association between rosacea and H. pylori infection or SIBO. We performed a prospective study to assess the prevalence of H. pylori infection and/or SIBO in patients with rosacea and evaluated the effect of H. pylori or SIBO eradication on rosacea. Methods We enrolled 90 patients with rosacea from January 2012 to January 2013 and a control group consisting of 90 patients referred to us because of mapping of nevi during the same period. We used the 13C Urea Breath Test and H. pylori stool antigen (HpSA) test to assess H. pylori infection and the glucose breath test to assess SIBO. Patients infected by H. pylori were treated with clarithromycin-containing sequential therapy. Patients positive for SIBO were treated with rifaximin. Results We found that 44/90 (48.9%) patients with rosacea and 24/90 (26.7%) control subjects were infected with H. pylori (p = 0.003). Moreover, 9/90 (10%) patients with rosacea and 7/90 (7.8%) subjects in the control group had SIBO (p = 0.6). Within 10 weeks from the end of antibiotic therapy, the skin lesions of rosacea disappeared or decreased markedly in 35/36 (97.2%) patients after eradication of H. pylori and in 3/8 (37.5%) patients who did not eradicate the infection (p < 0.0001). Rosacea skin lesions decreased markedly in 6/7 (85.7%) after eradication of SIBO whereas of the two patients who did not eradicate SIBO, one (50%) showed an improvement in rosacea (p = 0.284). Conclusions Prevalence of H. pylori infection was significantly higher in patients with rosacea than control group, whereas SIBO prevalence was comparable between the two groups. Eradication of H. pylori infection led to a significant improvement of skin symptoms in rosacea patients. PMID:25653855

  2. Induction of bacterial antigen-specific colitis by a simplified human microbiota consortium in gnotobiotic interleukin-10-/- mice.

    PubMed

    Eun, Chang Soo; Mishima, Yoshiyuki; Wohlgemuth, Steffen; Liu, Bo; Bower, Maureen; Carroll, Ian M; Sartor, R Balfour

    2014-06-01

    We evaluated whether a simplified human microbiota consortium (SIHUMI) induces colitis in germfree (GF) 129S6/SvEv (129) and C57BL/6 (B6) interleukin-10-deficient (IL-10(-/-)) mice, determined mouse strain effects on colitis and the microbiota, examined the effects of inflammation on relative bacterial composition, and identified immunodominant bacterial species in "humanized" IL-10(-/-) mice. GF wild-type (WT) and IL-10(-/-) 129 and B6 mice were colonized with 7 human-derived inflammatory bowel disease (IBD)-related intestinal bacteria and maintained under gnotobiotic conditions. Quantification of bacteria in feces, ileal and colonic contents, and tissues was performed using 16S rRNA gene selective quantitative PCR. Colonic segments were scored histologically, and gamma interferon (IFN-γ), IL-12p40, and IL-17 levels were measured in supernatants of unstimulated colonic tissue explants and of mesenteric lymph node (MLN) cells stimulated by lysates of individual or aggregate bacterial strains. Relative bacterial species abundances changed over time and differed between 129 and B6 mice, WT and IL-10(-/-) mice, luminal and mucosal samples, and ileal and colonic or fecal samples. SIHUMI induced colitis in all IL-10(-/-) mice, with more aggressive colitis and MLN cell activation in 129 mice. Escherichia coli LF82 and Ruminococcus gnavus lysates induced dominant effector ex vivo MLN TH1 and TH17 responses, although the bacterial mucosal concentrations were low. In summary, this study shows that a simplified human bacterial consortium induces colitis in ex-GF 129 and B6 IL-10(-/-) mice. Relative concentrations of individual SIHUMI species are determined by host genotype, the presence of inflammation, and anatomical location. A subset of IBD-relevant human enteric bacterial species preferentially stimulates bacterial antigen-specific TH1 and TH17 immune responses in this model, independent of luminal and mucosal bacterial concentrations.

  3. Human Intestinal Enteroids: a New Model To Study Human Rotavirus Infection, Host Restriction, and Pathophysiology

    PubMed Central

    Saxena, Kapil; Blutt, Sarah E.; Ettayebi, Khalil; Zeng, Xi-Lei; Broughman, James R.; Crawford, Sue E.; Karandikar, Umesh C.; Sastri, Narayan P.; Conner, Margaret E.; Opekun, Antone R.; Graham, David Y.; Qureshi, Waqar; Sherman, Vadim; Foulke-Abel, Jennifer; In, Julie; Kovbasnjuk, Olga; Zachos, Nicholas C.; Donowitz, Mark

    2015-01-01

    ABSTRACT Human gastrointestinal tract research is limited by the paucity of in vitro intestinal cell models that recapitulate the cellular diversity and complex functions of human physiology and disease pathology. Human intestinal enteroid (HIE) cultures contain multiple intestinal epithelial cell types that comprise the intestinal epithelium (enterocytes and goblet, enteroendocrine, and Paneth cells) and are physiologically active based on responses to agonists. We evaluated these nontransformed, three-dimensional HIE cultures as models for pathogenic infections in the small intestine by examining whether HIEs from different regions of the small intestine from different patients are susceptible to human rotavirus (HRV) infection. Little is known about HRVs, as they generally replicate poorly in transformed cell lines, and host range restriction prevents their replication in many animal models, whereas many animal rotaviruses (ARVs) exhibit a broader host range and replicate in mice. Using HRVs, including the Rotarix RV1 vaccine strain, and ARVs, we evaluated host susceptibility, virus production, and cellular responses of HIEs. HRVs infect at higher rates and grow to higher titers than do ARVs. HRVs infect differentiated enterocytes and enteroendocrine cells, and viroplasms and lipid droplets are induced. Heterogeneity in replication was seen in HIEs from different patients. HRV infection and RV enterotoxin treatment of HIEs caused physiological lumenal expansion detected by time-lapse microscopy, recapitulating one of the hallmarks of rotavirus-induced diarrhea. These results demonstrate that HIEs are a novel pathophysiological model that will allow the study of HRV biology, including host restriction, cell type restriction, and virus-induced fluid secretion. IMPORTANCE Our research establishes HIEs as nontransformed cell culture models to understand human intestinal physiology and pathophysiology and the epithelial response, including host restriction of

  4. Quantitative prediction of intestinal metabolism in humans from a simplified intestinal availability model and empirical scaling factor.

    PubMed

    Kadono, Keitaro; Akabane, Takafumi; Tabata, Kenji; Gato, Katsuhiko; Terashita, Shigeyuki; Teramura, Toshio

    2010-07-01

    This study aimed to establish a practical and convenient method of predicting intestinal availability (F(g)) in humans for highly permeable compounds at the drug discovery stage, with a focus on CYP3A4-mediated metabolism. We constructed a "simplified F(g) model," described using only metabolic parameters, assuming that passive diffusion is dominant when permeability is high and that the effect of transporters in epithelial cells is negligible. Five substrates for CYP3A4 (alprazolam, amlodipine, clonazepam, midazolam, and nifedipine) and four for both CYP3A4 and P-glycoprotein (P-gp) (nicardipine, quinidine, tacrolimus, and verapamil) were used as model compounds. Observed fraction of drug absorbed (F(a)F(g)) values for these compounds were calculated from in vivo pharmacokinetic (PK) parameters, whereas in vitro intestinal intrinsic clearance (CL(int,intestine)) was determined using human intestinal microsomes. The CL(int,intestine) for the model compounds corrected with that of midazolam was defined as CL(m,index) and incorporated into a simplified F(g) model with empirical scaling factor. Regardless of whether the compound was a P-gp substrate, the F(a)F(g) could be reasonably fitted by the simplified F(g) model, and the value of the empirical scaling factor was well estimated. These results suggest that the effects of P-gp on F(a) and F(g) are substantially minor, at least in the case of highly permeable compounds. Furthermore, liver intrinsic clearance (CL(int,liver)) can be used as a surrogate index of intestinal metabolism based on the relationship between CL(int,liver) and CL(m,index). F(g) can be easily predicted using a simplified F(g) model with the empirical scaling factor, enabling more confident selection of drug candidates with desirable PK profiles in humans.

  5. The Human Microbiome during Bacterial Vaginosis

    PubMed Central

    Delaney, Mary L.; Fichorova, Raina N.

    2016-01-01

    SUMMARY Bacterial vaginosis (BV) is the most commonly reported microbiological syndrome among women of childbearing age. BV is characterized by a shift in the vaginal flora from the dominant Lactobacillus to a polymicrobial flora. BV has been associated with a wide array of health issues, including preterm births, pelvic inflammatory disease, increased susceptibility to HIV infection, and other chronic health problems. A number of potential microbial pathogens, singly and in combinations, have been implicated in the disease process. The list of possible agents continues to expand and includes members of a number of genera, including Gardnerella, Atopobium, Prevotella, Peptostreptococcus, Mobiluncus, Sneathia, Leptotrichia, Mycoplasma, and BV-associated bacterium 1 (BVAB1) to BVAB3. Efforts to characterize BV using epidemiological, microscopic, microbiological culture, and sequenced-based methods have all failed to reveal an etiology that can be consistently documented in all women with BV. A careful analysis of the available data suggests that what we term BV is, in fact, a set of common clinical signs and symptoms that can be provoked by a plethora of bacterial species with proinflammatory characteristics, coupled to an immune response driven by variability in host immune function. PMID:26864580

  6. Fermentation in the human large intestine: its physiologic consequences and the potential contribution of prebiotics.

    PubMed

    Macfarlane, George T; Macfarlane, Sandra

    2011-11-01

    The human large intestine harbors a complex microbiota containing many hundreds of different bacterial species. Although structure/function relationships between different components of the microbiota are unclear, this complex multicellular entity plays an important role in maintaining homeostasis in the body. Many of the physiologic properties of the microbiota can be attributed to fermentation and the production of short-chain fatty acids (SCFAs), particularly acetate, propionate, and butyrate. In healthy people, fermentation processes are largely controlled by the amounts and different types of substrate, particularly complex carbohydrates that are accessible to bacteria in the colonic ecosystem. However, other factors impact on bacterial metabolism in the large gut, including large bowel transit time, the availability of inorganic terminal electron acceptors, such as nitrate and sulfate, and gut pH. They all affect the types and levels of SCFA that can be formed by the microbiota. This is important because to a large extent, acetate, propionate, and butyrate have varying physiologic effects in different body tissues. Prebiotics such as galactooligosaccharides together with inulins and their fructooligosaccharide derivatives have been shown to modify the species composition of the colonic microbiota, and in various degrees, to manifest several health-promoting properties related to enhanced mineral absorption, laxation, potential anticancer properties, lipid metabolism, and anti-inflammatory and other immune effects, including atopic disease. Many of these phenomena can be linked to their digestion and SCFA production by bacteria in the large gut.

  7. Antibiotics conspicuously affect community profiles and richness, but not the density of bacterial cells associated with mucosa in the large and small intestines of mice.

    PubMed

    Puhl, Nathan J; Uwiera, Richard R E; Yanke, L Jay; Selinger, L Brent; Inglis, G Douglas

    2012-02-01

    The influence of three antibiotics (bacitracin, enrofloxacin, and neomycin sulfate) on the mucosa-associated enteric microbiota and the intestines of mice was examined. Antibiotics caused conspicuous enlargement of ceca and an increase in overall length of the intestine. However, there were no pathologic changes associated with increased cecal size or length of the intestine. Conspicuous reductions in the richness of mucosa-associated bacteria and changes to community profiles within the small (duodenum, proximal jejunum, middle jejunum, distal jejunum, and ileum) and large (cecum, ascending colon, and descending colon) intestine occurred in mice administered antibiotics. Communities in antibiotic-treated mice were dominated by a limited number of Clostridium-like (i.e. clostridial cluster XIVa) and Bacteroides species. The richness of mucosa-associated communities within the small and large intestine increased during the 14-day recovery period. However, community profiles within the large intestine did not return to baseline (i.e. relative to the control). Although antibiotic administration greatly reduced bacterial richness, densities of mucosa-associated bacteria were not reduced correspondingly. These data showed that the antibiotics, bacitracin, enrofloxacin, and neomycin sulfate, administered for 21 days to mice did not sterilize the intestine, but did impart a tremendous and prolonged impact on mucosa-associated bacterial communities throughout the small and large intestine.

  8. Metabolism of green tea catechins by the human small intestine.

    PubMed

    Schantz, Markus; Erk, Thomas; Richling, Elke

    2010-10-01

    Numerous studies have shown that green tea polyphenols can be degraded in the colon, and there is abundant knowledge about the metabolites of these substances that appear in urine and plasma after green tea ingestion. However, there is very little information on the extent and nature of intestinal degradation of green tea catechins in humans. Therefore, the aim of this study was to examine in detail the microbial metabolism and chemical stability of these polyphenols in the small intestine using a well-established ex vivo model. For this purpose, fresh ileostomy fluids from two probands were incubated for 24 h under anaerobic conditions with (+)-catechin (C), (-)-epicatechin (EC), (-)-epicatechin 3-O-gallate (ECG), (-)-epigallocatechin (EGC), (-)-epigallocatchin 3-O-gallate (EGCG) and gallic acid (GA). After lyophilisation and extraction, metabolites were separated, identified and quantified by high performance liquid chromatography-photodiode array detection (HPLC-DAD) and HPLC-ESI-tandem mass spectrometry. Two metabolites of EC and C (3', 4', 5'-trihydroxyphenyl-γ-valerolactone and 3', 4'-dihydroxyphenyl-γ-valerolactone) were identified. In addition, 3', 4', 5'-trihydroxyphenyl-γ-valerolactone was detected as a metabolite of EGC, and (after 24-h incubation) pyrogallol as a degradation product of GA. Cleavage of the GA esters of EGCG and ECG was also observed, with variations dependent on the sources (probands) of the ileal fluids, which differed substantially microbiotically. The results provide new information about the degradation of green tea catechins in the gastrointestinal tract, notably that microbiota-dependent liberation of GA esters may occur before these compounds reach the colon.

  9. Cloning and expression of the human vasoactive intestinal peptide receptor.

    PubMed Central

    Sreedharan, S P; Robichon, A; Peterson, K E; Goetzl, E J

    1991-01-01

    Vasoactive intestinal peptide (VIP) is a neuroendocrine mediator found in the central and peripheral nervous system. Distinct subsets of neural, respiratory, gastrointestinal, and immune cells bear specific high-affinity receptors for VIP, which are associated with a guanine nucleotide-binding (G) protein capable of activating adenylate cyclase. A cDNA clone (GPRN1) encoding the human VIP receptor was identified in libraries prepared from the Nalm 6 line of leukemic pre-B lymphoblasts and the HT-29 line of colon carcinoma cells. The deduced 362-amino acid polypeptide sequence encoded by GPRN1 shares a seven-transmembrane-segment hydropathicity profile with other G protein-coupled receptors. Northern blot analyses identified a 2.7-kilobase transcript of the VIP receptor in Nalm 6 and HT-29 cells as well as in tissues from rat brain, colon, heart, lung, kidney, spleen, and small intestine. COS-6 cells transfected with GPRN1 bound 125I-labeled VIP specifically with a dissociation constant (Kd) of 2.5 nM. VIP--and less effectively secretin, peptide histidine isoleucine (PHI), and glucagon competitively displaced bound 125I-VIP from transfected COS-6 cells, with potencies in the order VIP greater than secretin = PHI much greater than glucagon. VIP stimulated adenylate cyclase activity in stably transfected Chinese hamster ovary K1 cells, inducing a 3-fold increase in the intracellular level of cAMP. When the antisense orientation of the VIP receptor clone was introduced into HT-29 cells, there was a 50% suppression of the specific binding of 125I-VIP and of the VIP-induced increase in cAMP level, relative to untransfected cells. The VIP receptor cloned exhibits less than or equal to 24% homology with other receptors in the same superfamily and thus represents a subset of G protein-coupled receptors for peptide ligands. Images PMID:1675791

  10. Initial insights into bacterial succession during human decomposition.

    PubMed

    Hyde, Embriette R; Haarmann, Daniel P; Petrosino, Joseph F; Lynne, Aaron M; Bucheli, Sibyl R

    2015-05-01

    Decomposition is a dynamic ecological process dependent upon many factors such as environment, climate, and bacterial, insect, and vertebrate activity in addition to intrinsic properties inherent to individual cadavers. Although largely attributed to microbial metabolism, very little is known about the bacterial basis of human decomposition. To assess the change in bacterial community structure through time, bacterial samples were collected from several sites across two cadavers placed outdoors to decompose and analyzed through 454 pyrosequencing and analysis of variable regions 3-5 of the bacterial 16S ribosomal RNA (16S rRNA) gene. Each cadaver was characterized by a change in bacterial community structure for all sites sampled as time, and decomposition, progressed. Bacteria community structure is variable at placement and before purge for all body sites. At bloat and purge and until tissues began to dehydrate or were removed, bacteria associated with flies, such as Ignatzschineria and Wohlfahrtimonas, were common. After dehydration and skeletonization, bacteria associated with soil, such as Acinetobacter, were common at most body sites sampled. However, more cadavers sampled through multiple seasons are necessary to assess major trends in bacterial succession.

  11. Exploring the bacterial assemblages along the human nasal passage.

    PubMed

    Wos-Oxley, Melissa L; Chaves-Moreno, Diego; Jáuregui, Ruy; Oxley, Andrew P A; Kaspar, Ursula; Plumeier, Iris; Kahl, Silke; Rudack, Claudia; Becker, Karsten; Pieper, Dietmar H

    2016-07-01

    The human nasal passage, from the anterior nares through the nasal vestibule to the nasal cavities, is an important habitat for opportunistic pathogens and commensals alike. This work sampled four different anatomical regions within the human nasal passage across a large cohort of individuals (n = 79) comprising individuals suffering from chronic nasal inflammation clinically known as chronic rhinosinusitis (CRS) and individuals not suffering from inflammation (CRS-free). While individuals had their own unique bacterial fingerprint that was consistent across the anatomical regions, these bacterial fingerprints formed into distinct delineated groups comprising core bacterial members, which were consistent across all four swabbed anatomical regions irrespective of health status. The most significant observed pattern was the difference between the global bacterial profiles of swabbed and tissue biopsy samples from the same individuals, being also consistent across different anatomical regions. Importantly, no statistically significant differences could be observed concerning the global bacterial communities, any of the bacterial species or the range of diversity indices used to compare between CRS and CRS-free individuals, and between two CRS phenotypes (without nasal polyps and with nasal polyps). Thus, the role of bacteria in the pathogenesis of sinusitis remains uncertain.

  12. Evolution of Bacterial Pathogens within the Human Host

    PubMed Central

    Bliven, Kimberly A.; Maurelli, Anthony T.

    2015-01-01

    Selective pressures within the human host, including interactions with innate and adaptive immune responses, exposure to medical interventions such as antibiotics, and competition with commensal microbiota all facilitate the evolution of bacterial pathogens. In this chapter, we present examples of pathogen strategies which emerged as a result of selective pressures within the human host niche, and discuss the resulting co-evolutionary ‘arms race’ between these organisms. In bacterial pathogens, many of the genes responsible for these strategies are encoded on mobile pathogenicity islands (PAIs) or plasmids, underscoring the importance of horizontal gene transfer (HGT) in the emergence of virulent microbial species. PMID:26999399

  13. Human colostrum oligosaccharides modulate major immunologic pathways of immature human intestine

    PubMed Central

    He, YingYing; Liu, ShuBai; Leone, Serena; Newburg, David S.

    2014-01-01

    The immature neonatal intestinal immune system hyperreacts to newly colonizing unfamiliar bacteria. The hypothesis that human milk oligosaccharides from colostrum (cHMOS) can directly modulate the signaling pathways of the immature mucosa was tested. Modulation of cytokine immune signaling by HMOS was measured ex vivo in intact immature (fetal) human intestinal mucosa. From the genes whose transcription was modulated by colostrum HMOS (cHMOS), Ingenuity Pathway Analysis identified networks controlling immune cell communication, intestinal mucosal immune system differentiation, and homeostasis. cHMOS attenuate pathogen-associated molecular pattern (PAMP)-stimulated acute phase inflammatory cytokine protein levels (IL-8, IL-6, MCP-1/2, IL-1β), while elevating cytokines involved in tissue repair and homeostasis. 3’-, 4-, and 6’-galactosyllactoses of cHMOS account for specific immunomodulation of PIC-induced IL-8 levels. cHMOS attenuate mucosal responses to surface inflammatory stimuli during early development, while enhancing signals that support maturation of the intestinal mucosal immune system. PMID:24691111

  14. Low dietary iron intake restrains the intestinal inflammatory response and pathology of enteric infection by food-borne bacterial pathogens

    PubMed Central

    Kortman, Guus AM; Mulder, Michelle LM; Richters, Thijs JW; Shanmugam, Nanda KN; Trebicka, Estela; Boekhorst, Jos; Timmerman, Harro M; Roelofs, Rian; Wiegerinck, Erwin T; Laarakkers, Coby M; Swinkels, Dorine W; Bolhuis, Albert; Cherayil, Bobby J; Tjalsma, Harold

    2015-01-01

    Orally administrated iron is suspected to increase susceptibility to enteric infections among children in infection endemic regions. Here we investigated the effect of dietary iron on the pathology and local immune responses in intestinal infection models. Mice were held on iron-deficient, normal-iron, or high-iron diets and after two weeks they were orally challenged with the pathogen Citrobacter rodentium. Microbiome analysis by pyrosequencing revealed profound iron- and infection-induced shifts in microbiota composition. Fecal levels of the innate defensive molecules and markers of inflammation lipocalin-2 and calprotectin were not influenced by dietary iron intervention alone, but were markedly lower in mice on the iron-deficient diet after infection. Next, mice on the iron-deficient diet tended to gain more weight and to have a lower grade of colon pathology. Furthermore, survival of the nematode Caenorhabditis elegans infected with Salmonella enterica serovar Typhimurium was prolonged after iron-deprivation. Together, these data show that iron limitation restricts disease pathology upon bacterial infection. However, our data also showed decreased intestinal inflammatory responses of mice fed on high-iron diets. Thus additionally, our study indicates that the effects of iron on processes at the intestinal host-pathogen interface may highly depend on host iron status, immune status and gut microbiota composition. PMID:26046550

  15. Protective Capacity of Resveratrol, a Natural Polyphenolic Compound, against Deoxynivalenol-Induced Intestinal Barrier Dysfunction and Bacterial Translocation.

    PubMed

    Ling, Ka-Ho; Wan, Murphy Lam Yim; El-Nezami, Hani; Wang, Mingfu

    2016-05-16

    Contamination of food/feedstuffs by mycotoxins is a serious problem worldwide, causing severe economic losses and serious health problems in animals/humans. Deoxynivalenol (DON) is a major mycotoxin contaminant and is known to impair intestinal barrier function. Grapes and red wine are rich in polyphenols, such as resveratrol (RES), which has striking antioxidant and anti-inflammatory activities. RES is a food-derived component; therefore, it may be simultaneously present with DON in the gastrointestinal tract. The aim of this study was to explore in vitro protective effects of RES against DON-induced intestinal damage. The results showed that RES could protect DON-induced bacteria translocation because of enhanced of intestinal barrier function by restoring the DON-induced decrease in transepithelial electrical resistance and increase in paracellular permeability. Further mechanistic studies demonstrated that RES protects against DON-induced barrier dysfunction by promoting the assembly of claudin-4 in the tight junction complex. This is probably mediated through modulation of IL-6 and IL-8 secretion via mitogen-activated protein kinase-dependent pathways. Our results imply that RES can protect against DON-induced intestinal damage and that RES may be used as a novel dietary intervention strategy to reduce DON toxicity in animals/humans.

  16. Frequency of occurrence and antimicrobial susceptibility of bacterial isolates from the intestinal and female genital tracts.

    PubMed

    Sutter, V L

    1983-01-01

    In the selection of empiric therapy for infections of the female genital tract and intraabdominal infections in humans, the requisite information includes (1) the frequency of isolation of bacterial species dominant in the normal gastrointestinal and female genital tracts and in intraabdominal and female genital tract infections and (2) the in vitro susceptibilities of the dominant species to drugs that may be used against them (e.g., penicillin G, cefoperazone, cefoxitin, chloramphenicol, clindamycin, and metronidazole). The predominant bacteria in fecal contents are not necessarily those most frequently found in infections. Intraabdominal and perirectal infections are usually polymicrobial, resulting from mixtures of facultative species (coliforms and streptococci) and anaerobes. The predominant bacteria of the normal vagina and cervix are lactobacilli, facultative streptococci, Peptococcus species, and Peptostreptococcus species. Most infections of the female genital tract are due to mixtures of facultative enteric bacilli, streptococci, and anaerobes (Peptococcus and Peptostreptococcus species, Bacteroides fragilis, Bacteroides disiens, Bacteroides melaninogenicus, Bacteroides bivius, and Bacteroides asaccharolyticus). Of the antibiotics tested, clindamycin appears the most active against many of the groups of bacteria isolated.

  17. Ileocecal valve dysfunction in small intestinal bacterial overgrowth: A pilot study

    PubMed Central

    Miller, Larry S; Vegesna, Anil K; Sampath, Aiswerya Madanam; Prabhu, Shital; Kotapati, Sesha Krishna; Makipour, Kian

    2012-01-01

    AIM: To explore whether patients with a defective ileocecal valve (ICV)/cecal distension reflex have small intestinal bacterial overgrowth. METHODS: Using a colonoscope, under conscious sedation, the ICV was intubated and the colonoscope was placed within the terminal ileum (TI). A manometry catheter with 4 pressure channels, spaced 1 cm apart, was passed through the biopsy channel of the colonoscope into the TI. The colonoscope was slowly withdrawn from the TI while the manometry catheter was advanced. The catheter was placed across the ICV so that at least one pressure port was within the TI, ICV and the cecum respectively. Pressures were continuously measured during air insufflation into the cecum, under direct endoscopic visualization, in 19 volunteers. Air was insufflated to a maximum of 40 mmHg to prevent barotrauma. All subjects underwent lactulose breath testing one month after the colonoscopy. The results of the breath tests were compared with the results of the pressures within the ICV during air insufflation. RESULTS: Nineteen subjects underwent colonoscopy with measurements of the ICV pressures after intubation of the ICV with a colonoscope. Initial baseline readings showed no statistical difference in the pressures of the TI and ICV, between subjects with positive lactulose breath tests and normal lactulose breath tests. The average peak ICV pressure during air insufflation into the cecum in subjects with normal lactulose breath tests was significantly higher than cecal pressures during air insufflation (49.33 ± 7.99 mmHg vs 16.40 ± 2.14 mmHg, P = 0.0011). The average percentage difference of the area under the pressure curve of the ICV from the cecum during air insufflations in subjects with normal lactulose breath tests was significantly higher (280.72% ± 43.29% vs 100% ± 0%, P = 0.0006). The average peak ICV pressure during air insufflation into the cecum in subjects with positive lactulose breath tests was not significantly different than cecal

  18. Human mini-guts: new insights into intestinal physiology and host–pathogen interactions

    PubMed Central

    In, Julie G.; Foulke-Abel, Jennifer; Estes, Mary K.; Zachos, Nicholas C.; Kovbasnjuk, Olga; Donowitz, Mark

    2016-01-01

    The development of indefinitely propagating human ‘mini-guts’ has led to a rapid advance in gastrointestinal research related to transport physiology, developmental biology, pharmacology, and pathophysiology. These mini-guts, also called enteroids or colonoids, are derived from LGR5+ intestinal stem cells isolated from the small intestine or colon. Addition of WNT3A and other growth factors promotes stemness and results in viable, physiologically functional human intestinal or colonic cultures that develop a crypt–villus axis and can be differentiated into all intestinal epithelial cell types. The success of research using human enteroids has highlighted the limitations of using animals or in vitro, cancer-derived cell lines to model transport physiology and pathophysiology. For example, curative or preventive therapies for acute enteric infections have been limited, mostly due to the lack of a physiological human intestinal model. However, the human enteroid model enables specific functional studies of secretion and absorption in each intestinal segment as well as observations of the earliest molecular events that occur during enteric infections. This Review describes studies characterizing these human mini-guts as a physiological model to investigate intestinal transport and host pathogen interactions. PMID:27677718

  19. Human mini-guts: new insights into intestinal physiology and host-pathogen interactions.

    PubMed

    In, Julie G; Foulke-Abel, Jennifer; Estes, Mary K; Zachos, Nicholas C; Kovbasnjuk, Olga; Donowitz, Mark

    2016-11-01

    The development of indefinitely propagating human 'mini-guts' has led to a rapid advance in gastrointestinal research related to transport physiology, developmental biology, pharmacology, and pathophysiology. These mini-guts, also called enteroids or colonoids, are derived from LGR5(+) intestinal stem cells isolated from the small intestine or colon. Addition of WNT3A and other growth factors promotes stemness and results in viable, physiologically functional human intestinal or colonic cultures that develop a crypt-villus axis and can be differentiated into all intestinal epithelial cell types. The success of research using human enteroids has highlighted the limitations of using animals or in vitro, cancer-derived cell lines to model transport physiology and pathophysiology. For example, curative or preventive therapies for acute enteric infections have been limited, mostly due to the lack of a physiological human intestinal model. However, the human enteroid model enables specific functional studies of secretion and absorption in each intestinal segment as well as observations of the earliest molecular events that occur during enteric infections. This Review describes studies characterizing these human mini-guts as a physiological model to investigate intestinal transport and host-pathogen interactions.

  20. Levothyroxine therapy and impaired clearance are the strongest contributors to small intestinal bacterial overgrowth: Results of a retrospective cohort study

    PubMed Central

    Brechmann, Thorsten; Sperlbaum, Andre; Schmiegel, Wolff

    2017-01-01

    AIM To identify a set of contributors, and weight and rank them on a pathophysiological basis. METHODS Patients who have undergone a lactulose or glucose hydrogen breath test to rule out small intestinal bacterial overgrowth (SIBO) for various clinical symptoms, including diarrhoea, weight loss, abdominal pain, cramping or bloating, were seen as eligible for inclusion in a retrospective single-centre study. Clinical data such as co-morbidities, medication, laboratory parameters and other possible risk factors have been identified from the electronic data system. Cases lacking or with substantially incomplete clinical data were excluded from the analysis. Suspected contributors were summarised under four different pathophysiological pathways (impaired gastric acid barrier, impaired intestinal clearance, immunosuppression and miscellaneous factors including thyroid gland variables) and investigated using the χ2 test, Student’s t-test and logistic regression models. RESULTS A total of 1809 patients who had undergone hydrogen breath testing were analysed. Impairment of the gastric acid barrier (gastrectomy, odds ratio: OR = 3.5, PPI therapy OR = 1.4), impairment of intestinal clearance (any resecting gastric surgery OR = 2.6, any colonic resection OR = 1.9, stenosis OR = 3.4, gastroparesis OR = 3.4, neuropathy 2.2), immunological factors (any drug-induced immunosuppression OR = 1.8), altered thyroid gland metabolism (hypothyroidism OR = 2.6, levothyroxine therapy OR = 3.0) and diabetes mellitus (OR = 1.9) were associated significantly to SIBO. Any abdominal surgery, ileocecal resection, vagotomy or IgA-deficiency did not have any influence, and a history of appendectomy decreased the risk of SIBO. Multivariate analysis revealed gastric surgery, stenoses, medical immunosuppression and levothyroxine to be the strongest predictors. Levothyroxine therapy was the strongest contributor in a simplified model (OR = 3.0). CONCLUSION The most important contributors for the

  1. The action of berry phenolics against human intestinal pathogens.

    PubMed

    Puupponen-Pimiä, Riitta; Nohynek, Liisa; Alakomi, Hanna-Leena; Oksman-Caldentey, Kirsi-Marja

    2005-01-01

    Phenolic compounds present in berries selectively inhibit the growth of human gastrointestinal pathogens. Especially cranberry, cloudberry, raspberry, strawberry and bilberry possess clear antimicrobial effects against e.g. salmonella and staphylococcus. Complex phenolic polymers, such as ellagitannins, are strong antibacterial agents present in cloudberry, raspberry and strawberry. Berry phenolics seem to affect the growth of different bacterial species with different mechanisms. Adherence of bacteria to epithelial surfaces is a prerequisite for colonization and infection of many pathogens. Antimicrobial activity of berries may also be related to anti-adherence activity of the berries. Utilization of enzymes in berry processing increases the amount of phenolics and antimicrobial activity of the berry products. Antimicrobial berry compounds are likely to have many important applications in the future as natural antimicrobial agents for food industry as well as for medicine.

  2. Looking in ticks for human bacterial pathogens.

    PubMed

    Mediannikov, O; Fenollar, F

    2014-12-01

    Ticks are considered to be second worldwide to mosquitoes as vectors of human diseases and the most important vectors of disease-causing pathogens in domestic and wild animals. A number of emerging tick-borne pathogens are already discovered; however, the proportion of undiagnosed infectious diseases, especially in tropical regions, may suggest that there are still more pathogens associated with ticks. Moreover, the identification of bacteria associated with ticks may provide new tool for the control of ticks and tick-borne diseases. Described here molecular methods of screening of ticks, extensive use of modern culturomics approach, newly developed artificial media and different cell line cultures may significantly improve our knowledge about the ticks as the agents of human and animal pathology.

  3. Involvement of Concentrative Nucleoside Transporter 1 in Intestinal Absorption of Trifluridine Using Human Small Intestinal Epithelial Cells.

    PubMed

    Takahashi, Koichi; Yoshisue, Kunihiro; Chiba, Masato; Nakanishi, Takeo; Tamai, Ikumi

    2015-09-01

    TAS-102, which is effective for refractory metastatic colorectal cancer, is a combination drug of anticancer trifluridine (FTD; which is derived from pyrimidine nucleoside) and FTD-metabolizing enzyme inhibitor tipiracil hydrochloride (TPI) at a molecular ratio of 1:0.5. To evaluate the intestinal absorption mechanism of FTD, the uptake and transcellular transport of FTD by human small intestinal epithelial cell (HIEC) monolayer as a model of human intestinal epithelial cells was investigated. The uptake and membrane permeability of FTD by HIEC monolayers were saturable, Na(+) -dependent, and inhibited by nucleosides. These transport characteristics are mostly comparable with those of concentrative nucleoside transporters (CNTs). Moreover, the uptake of FTD by CNT1-expressing Xenopus oocytes was the highest among human CNT transporters. The obtained Km and Vmax values of FTD by CNT1 were 69.0 μM and 516 pmol/oocyte/30 min, respectively. The transcellular transport of FTD by Caco-2 cells, where CNT1 is heterologously expressed, from apical to basolateral side was greater than that by Mock cells. In conclusion, these results demonstrated that FTD exhibits high oral absorption by the contribution of human CNT1.

  4. The Metabolic Profiling of Isorhamnetin-3-O-Neohesperidoside Produced by Human Intestinal Flora Employing UPLC-Q-TOF/MS.

    PubMed

    Du, Le-Yue; Zhao, Min; Tao, Jin-Hua; Qian, Da-Wei; Jiang, Shu; Shang, Er-Xin; Guo, Jian-Ming; Liu, Pei; Su, Shu-Lan; Duan, Jin-Ao

    2016-11-23

    Isorhamnetin-3-O-neohesperidoside is the major active substance of Puhuang, a traditional herb medicine widely used in clinical practice to tackle many chronic diseases. However, little is known about the interactions between this ingredient and intestinal flora. In this study, ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry together with automated data analysis software (Metabolynx™) was used for analysis of the metabolic profile of isorhamnetin-3-O-neohesperidoside by the isolated human intestinal bacteria. The parent and three metabolites isorhamnetin-3-O-glucoside, isorhamnetin and quercetin were detected and identified based on the characteristics of their deprotonated molecules. These metabolites indicated that isorhamnetin-3-O-neohesperidoside was firstly deglycosylated to isorhamnetin-3-O-glucoside and subsequently to the aglycone isorhamnetin, and the latter was demethylated to quercetin. The majority of bacteria such as Escherichia sp. 23 were capable of converting isorhamnetin-3-O-neohesperidoside to considerable amounts of aglycone isorhamnetin and further to minor amounts of quercetin, while minor amounts of isorhamnetin-3-O-glucoside were detected in minority of bacterial samples such as Enterococcus sp. 30. The metabolic pathway and metabolites of isorhamnetin-3-O-neohesperidoside by the different human intestinal bacteria were firstly investigated. Furthermore, the metabolites of isorhamnetin-3-O-neohesperidoside might influence the effects of traditional herb medicines. Thus, our study is helpful to further unravel how isorhamnetin-3-O-neohesperidoside and Puhuang work in vivo.

  5. Insights into bacterial protein glycosylation in human microbiota

    PubMed Central

    Zhu, Fan; Wu, Hui

    2017-01-01

    The study of human microbiota is an emerging research topic. The past efforts have mainly centered on studying the composition and genomic landscape of bacterial species within the targeted communities. The interaction between bacteria and hosts is the pivotal event in the initiation and progression of infectious diseases. There is a great need to identify and characterize the molecules that mediate the bacteria-host interaction. Bacterial surface exposed proteins play an important role in the bacteria-host interaction. Numerous surface proteins are glycosylated, and the glycosylation is crucial for their function in mediating the bacterial interaction with hosts. Here we present an overview of surface glycoproteins from bacteria that inhabit three major mucosal environments across human body: oral, gut and skin. We describe the important enzymes involved in the process of protein glycosylation, and discuss how the process impacts the bacteria-host interaction. Emerging molecular details underlying glycosylation of bacterial surface proteins may lead to new opportunities for designing anti-infective small molecules, and developing novel vaccines in order to treat or prevent bacterial infection. PMID:26712033

  6. Insights into bacterial protein glycosylation in human microbiota.

    PubMed

    Zhu, Fan; Wu, Hui

    2016-01-01

    The study of human microbiota is an emerging research topic. The past efforts have mainly centered on studying the composition and genomic landscape of bacterial species within the targeted communities. The interaction between bacteria and hosts is the pivotal event in the initiation and progression of infectious diseases. There is a great need to identify and characterize the molecules that mediate the bacteria-host interaction. Bacterial surface exposed proteins play an important role in the bacteria- host interaction. Numerous surface proteins are glycosylated, and the glycosylation is crucial for their function in mediating the bacterial interaction with hosts. Here we present an overview of surface glycoproteins from bacteria that inhabit three major mucosal environments across human body: oral, gut and skin. We describe the important enzymes involved in the process of protein glycosylation, and discuss how the process impacts the bacteria-host interaction. Emerging molecular details underlying glycosylation of bacterial surface proteins may lead to new opportunities for designing anti-infective small molecules, and developing novel vaccines in order to treat or prevent bacterial infection.

  7. An in vivo model of human small intestine using pluripotent stem cells

    PubMed Central

    Watson, Carey L; Mahe, Maxime M; Múnera, Jorge; Howell, Jonathan C; Sundaram, Nambirajan; Poling, Holly M; Schweitzer, Jamie I; Vallance, Jefferson E; Mayhew, Christopher N; Sun, Ying; Grabowski, Gregory; Finkbeiner, Stacy R; Spence, Jason R; Shroyer, Noah F; Wells, James M; Helmrath, Michael A

    2015-01-01

    Differentiation of human pluripotent stem cells (hPSCs) into organ-specific subtypes offers an exciting avenue for the study of embryonic development and disease processes, for pharmacologic studies and as a potential resource for therapeutic transplant1,2. To date, limited in vivo models exist for human intestine, all of which are dependent upon primary epithelial cultures or digested tissue from surgical biopsies that include mesenchymal cells transplanted on biodegradable scaffolds3,4. Here, we generated human intestinal organoids (HIOs) produced in vitro from human embryonic stem cells (ESCs) or induced pluripotent stem cells (iPSCs)5,6 that can engraft in vivo. These HIOs form mature human intestinal epithelium with intestinal stem cells contributing to the cryptvillus architecture and a laminated human mesenchyme, both supported by mouse vasculature ingrowth. In vivo transplantation resulted in marked expansion and maturation of the epithelium and mesenchyme, as demonstrated by differentiated intestinal cell lineages (enterocytes, goblet cells, Paneth cells, tuft cells and enteroendocrine cells), presence of functional brush-border enzymes (lactase, sucrase-isomaltase and dipeptidyl peptidase 4) and visible subepithelial and smooth muscle layers when compared with HIOs in vitro. Transplanted intestinal tissues demonstrated digestive functions as shown by permeability and peptide uptake studies. Furthermore, transplanted HIO-derived tissue was responsive to systemic signals from the host mouse following ileocecal resection, suggesting a role for circulating factors in the intestinal adaptive response7–9. This model of the human small intestine may pave the way for studies of intestinal physiology, disease and translational studies. PMID:25326803

  8. Human intestinal absorption--neutral molecules and ionic species.

    PubMed

    Abraham, Michael H

    2014-07-01

    Analysis of percentage human intestinal absorption (%HIA) for 280 drugs shows that an excellent fit can be obtained using only three descriptors for neutral molecules with a SD of 13.9%. Use of descriptors for individual cations and anions does not lead to any better goodness-of-fit. It is noted that diffusion coefficients in water for ionized molecules are almost identical to those for the corresponding neutral molecules. Comparison of equation coefficients for HIA with those for other processes shows that HIA resembles diffusion in water but does not resemble permeation through biological bilayers. It is shown that compound substituent effects on HIA are near those for diffusion but are far away from substituent effects on permeation through a typical bilayer. Calculations indicate that rates of permeation through an unstirred mucosal layer are of the same order as experimental rates of permeation in HIA. It is concluded that for the 280 compound set, diffusion through the unstirred mucosal layer is the rate determining step. The effect on pK(a) in transfer of acids and bases from water to another solvent, and of diffusion past a negative charge in a phase/bilayer is also considered.

  9. Protective effect of glutamine on intestinal injury and bacterial community in rats exposed to hypobaric hypoxia environment

    PubMed Central

    Xu, Chun-Lan; Sun, Rui; Qiao, Xiang-Jin; Xu, Cui-Cui; Shang, Xiao-Ya; Niu, Wei-Ning

    2014-01-01

    AIM: To investigate the protective effect of glutamine (Gln) on intestinal injury and the bacterial community in rats exposed to hypobaric hypoxia environment. METHODS: Sprague-Dawley rats were divided into control, hypobaric hypoxia (HH), and hypobaric hypoxia + Gln (5.0 g/kg BW·d) (HG) groups. On the first 3 d, all rats were placed in a normal environment. After the third day, the HH and HG groups were transferred into a hypobaric chamber at a simulated elevation of 7000 m for 5 d. The rats in the HG group were given Gln by gavage daily for 8 d. The rats in the control and HH groups were treated with the same volume of saline. The intestinal morphology, serum levels of malondialdehyde (MDA), superoxide dismutase (SOD), interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), interferon-gamma (IFN-γ) and diamino oxidase (DAO) were examined. We also evaluated the expression levels of occludin, toll-like receptor 4 (TLR4), nuclear factor-κB p65 (NF-κB p65) and myeloid differentiation factor 88 (MyD88), and examined the bacterial community in caecal contents. RESULTS: Hypobaric hypoxia induced the enlargement of the heart, liver, lung and kidney, and caused spleen atrophy. Intestinal villi damage was also observed in the HH group. Supplementation with Gln significantly alleviated hypobaric-induced damage to main organs including the intestine, increased serum SOD (1.14 ± 0.03 vs 0.88 ± 0.04, P < 0.05) and MDA (8.35 ± 1.60, P < 0.01) levels and decreased serum IL-6 (1172.13±30.49 vs 1407.05 ± 34.36, P < 0.05), TNF-α (77.46 ± 0.78 vs 123.70 ± 3.03, P < 0.001), IFN-γ (1355.42 ± 72.80 vs 1830.16 ± 42.07, P < 0.01) and DAO (629.30 ± 9.15 vs 524.10 ± 13.34, P < 0.001) levels. Moreover, Gln significantly increased occludin (0.72 ± 0.05 vs 0.09 ± 0.01, P < 0.001), TLR4 (0.15 ± 0.05 vs 0.30 ±0.09, P < 0.05), MyD88 (0.32 ± 0.08 vs 0.71 ± 0.06, P < 0.01), and NF-κB p65 (0.16 ± 0.04 vs 0.44 ± 0.03, P < 0.01) expression levels and improved the intestinal

  10. Focused specificity of intestinal TH17 cells towards commensal bacterial antigens.

    PubMed

    Yang, Yi; Torchinsky, Miriam B; Gobert, Michael; Xiong, Huizhong; Xu, Mo; Linehan, Jonathan L; Alonzo, Francis; Ng, Charles; Chen, Alessandra; Lin, Xiyao; Sczesnak, Andrew; Liao, Jia-Jun; Torres, Victor J; Jenkins, Marc K; Lafaille, Juan J; Littman, Dan R

    2014-06-05

    T-helper-17 (TH17) cells have critical roles in mucosal defence and in autoimmune disease pathogenesis. They are most abundant in the small intestine lamina propria, where their presence requires colonization of mice with microbiota. Segmented filamentous bacteria (SFB) are sufficient to induce TH17 cells and to promote TH17-dependent autoimmune disease in animal models. However, the specificity of TH17 cells, the mechanism of their induction by distinct bacteria, and the means by which they foster tissue-specific inflammation remain unknown. Here we show that the T-cell antigen receptor (TCR) repertoire of intestinal TH17 cells in SFB-colonized mice has minimal overlap with that of other intestinal CD4(+) T cells and that most TH17 cells, but not other T cells, recognize antigens encoded by SFB. T cells with antigen receptors specific for SFB-encoded peptides differentiated into RORγt-expressing TH17 cells, even if SFB-colonized mice also harboured a strong TH1 cell inducer, Listeria monocytogenes, in their intestine. The match of T-cell effector function with antigen specificity is thus determined by the type of bacteria that produce the antigen. These findings have significant implications for understanding how commensal microbiota contribute to organ-specific autoimmunity and for developing novel mucosal vaccines.

  11. Study of the adhesion of Bifidobacterium bifidum MIMBb75 to human intestinal cell lines.

    PubMed

    Guglielmetti, Simone; Tamagnini, Isabella; Minuzzo, Mario; Arioli, Stefania; Parini, Carlo; Comelli, Elena; Mora, Diego

    2009-08-01

    The aim of this study was to investigate the adhesive phenotype of the human intestinal isolate Bifidobacterium bifidum MIMBb75 to human colon carcinoma cell lines. We have previously shown that the adhesion of this strain to Caco-2 cells is mediated by an abundant surface lipoprotein named BopA. In this study, we found that this strain adheres to Caco-2 and HT-29 cells, and that its adhesion strongly depends on the environmental conditions, including the presence of sugars and bile salts and the pH. Considerably more adhesion to a Caco-2 monolayer occurred in the presence of fucose and mannose and less when MIMBb75 grew in Oxgall bile salts compared to standard environmental conditions. In particular, growth in Oxgall bile salts reduced the adhesion ability of MIMBb75 and modified the SDS-PAGE profile of the cell wall associated proteins of the strain. The pH markedly affected both adhesion to Caco-2 and bacterial autoaggregation. Finally, experiments with sodium metaperiodate suggested that not only proteinaceous determinants are involved in the adhesion process of B. bifidum. In conclusion, it seems that the colonization strategy of this bacterium can be influenced by factors varying along the gastrointestinal tract, such as the presence of specific sugars and bile salts and the pH, possibly limiting the adhesion of B. bifidum to only restricted distal sites of the gut.

  12. Characterization of intracellular pteroylpolyglutamate hydrolase (PPH) from human intestinal mucosa

    SciTech Connect

    Wang, T.T.Y.; Chandler, C.J.; Halsted, C.H.

    1986-03-01

    There are two forms of pteroylpolyglutamate hydrolase (PPH) in the human intestinal mucosa, one in the brush border membrane and the other intracellular; brush border PPH is an exopeptidase with optimal activity at pH 6.5 and a requirement for zinc. The presence study characterized human intracellular PPH and compared its properties to those of brush border PPH. Intracellular PPH was purified 30-fold. The enzyme had a MW of 75,000 by gel filtration, was optimally active at pH 4.5, and had an isoelectric point at pH 8.0. In contrast to brush border PPH, intracellular PPH was unstable at increasing temperatures, was unaffected by dialysis against chelating agents and showed no requirement for Zn/sup 2 +/. Using PteGlu/sub 2/(/sup 14/C)Glu as substrate, they demonstrated a K/sub m/ of 1.2 ..mu..M and increasing affinity for folates with longer glutamate chains. Intracellular PPH required the complete folic acid (PteGlu) moiety and a ..gamma..-glutamyl linkage for activity. Using ion exchange chromatography and an HPLC method to determine the hydrolytic products of the reaction, they found intracellular PPH could cleave both internal and terminal ..gamma..-glutamyl linkages, with PteGlu as an end product. After subcellular fractionation of the mucosa, PPH was found in the lysosomes. In summary, the distinct characteristics of brush border and intracellular PPH suggest that the two hydrolases serve different roles in folate metabolism.

  13. Identification of Human Intestinal Bacteria that Promote or Inhibit Inflammation

    DTIC Science & Technology

    2012-11-01

    microbiota  composition  is  critical  in  intestinal  inflammation  and  their  perturbations  are...Evidence   suggests   that   intestinal   microbiota  is  a  very  important  factor   in   IBD   (Kaser   et   al.,   2009...treatment   of   intestinal   inflammation.   Pertinent   adjustments   in   the   microbiota   of   patients  

  14. Human intestinal parasites in primary school children in Kampala, Uganda.

    PubMed

    Kabatereine, N B; Kemijumbi, J; Kazibwe, F; Onapa, A W

    1997-05-01

    A cross sectional survey on intestinal parasite infections was carried out in 5,313 pupils between the ages of ten and fifteen years in 98 primary schools in Kampala. The aim was to identify the types and distribution of intestinal parasites and to estimate the prevalence in school children. Trichuris trichiura (28%), Ascaris lumbricoides (17%) and hookworms (12.9%) were common infections among the children. Other less commonly found parasites were S.mansoni, Strongyloides stercolaris, Taenia sp, Enterobius vermicularis, Giardia lamblia, Entamoeba coli and E. histolytica. Refuse dumps are probably a significant source of transmission of intestinal helminthic infections in Kampala.

  15. The metabolic profile of acteoside produced by human or rat intestinal bacteria or intestinal enzyme in vitro employed UPLC-Q-TOF-MS.

    PubMed

    Cui, Qingling; Pan, Yingni; Xu, Xiaotong; Zhang, Wenjie; Wu, Xiao; Qu, Shouhe; Liu, Xiaoqiu

    2016-03-01

    Acteoside, the main and representative phenylethanoid glycosides of Herba Cistanches, possesses wide bioactivities but low oral bioavailability. It may serve as the prodrug and be converted into the active forms in gastrointestinal tract, which mainly occurred in intestinal tract composed of intestinal bacteria and intestinal enzyme. Intestinal bacteria, a new drug target, take a significant role on exerting pharmacological effects of drugs by oral administration. In this paper, acteoside was incubated with human or rat intestinal bacteria or rat intestinal enzyme for 36 h to seek metabolites responsible for pharmacodynamics. The samples were analyzed by ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry. Besides the parent compound, 14 metabolites were detected and identified based on their retention times and fragmentation patterns in their MS spectra including 8 degradation metabolites, 2 isomers in intestinal bacteria and intestinal enzyme samples and 4 parent metabolites only found in intestinal enzymes. The metabolic pathway of acteoside was thus proposed. Identification of these metabolites of acteoside by the intestinal bacteria or intestinal enzyme gave an insight to clarify pharmacological mechanism of traditional Chinese medicines and identify the real active molecules.

  16. Topographic diversity of fungal and bacterial communities in human skin.

    PubMed

    Findley, Keisha; Oh, Julia; Yang, Joy; Conlan, Sean; Deming, Clayton; Meyer, Jennifer A; Schoenfeld, Deborah; Nomicos, Effie; Park, Morgan; Kong, Heidi H; Segre, Julia A

    2013-06-20

    Traditional culture-based methods have incompletely defined the microbial landscape of common recalcitrant human fungal skin diseases, including athlete's foot and toenail infections. Skin protects humans from invasion by pathogenic microorganisms and provides a home for diverse commensal microbiota. Bacterial genomic sequence data have generated novel hypotheses about species and community structures underlying human disorders. However, microbial diversity is not limited to bacteria; microorganisms such as fungi also have major roles in microbial community stability, human health and disease. Genomic methodologies to identify fungal species and communities have been limited compared with those that are available for bacteria. Fungal evolution can be reconstructed with phylogenetic markers, including ribosomal RNA gene regions and other highly conserved genes. Here we sequenced and analysed fungal communities of 14 skin sites in 10 healthy adults. Eleven core-body and arm sites were dominated by fungi of the genus Malassezia, with only species-level classifications revealing fungal-community composition differences between sites. By contrast, three foot sites--plantar heel, toenail and toe web--showed high fungal diversity. Concurrent analysis of bacterial and fungal communities demonstrated that physiologic attributes and topography of skin differentially shape these two microbial communities. These results provide a framework for future investigation of the contribution of interactions between pathogenic and commensal fungal and bacterial communities to the maintainenace of human health and to disease pathogenesis.

  17. In vitro patterning of pluripotent stem cell-derived intestine recapitulates in vivo human development

    PubMed Central

    Tsai, Yu-Hwai; Nattiv, Roy; Dedhia, Priya H.; Nagy, Melinda S.; Chin, Alana M.; Thomson, Matthew; Klein, Ophir D.

    2017-01-01

    ABSTRACT The intestine plays a central role in digestion, nutrient absorption and metabolism, with individual regions of the intestine having distinct functional roles. Many examples of region-specific gene expression in the adult intestine are known, but how intestinal regional identity is established during development is a largely unresolved issue. Here, we have identified several genes that are expressed in a region-specific manner in the developing human intestine. Using human embryonic stem cell-derived intestinal organoids, we demonstrate that the duration of exposure to active FGF and WNT signaling controls regional identity. Short-term exposure to FGF4 and CHIR99021 (a GSK3β inhibitor that stabilizes β-catenin) resulted in organoids with gene expression patterns similar to developing human duodenum, whereas longer exposure resulted in organoids similar to ileum. When region-specific organoids were transplanted into immunocompromised mice, duodenum-like organoids and ileum-like organoids retained their regional identity, demonstrating that regional identity of organoids is stable after initial patterning occurs. This work provides insights into the mechanisms that control regional specification of the developing human intestine and provides new tools for basic and translational research. PMID:27927684

  18. Type 1 diabetes: role of intestinal microbiome in humans and mice.

    PubMed

    Boerner, Brian P; Sarvetnick, Nora E

    2011-12-01

    Type 1 diabetes is a disease involving autoimmune destruction of pancreatic beta cells in genetically predisposed individuals. Identifying factors that trigger initiation and progression of autoimmunity may provide opportunities for directed prophylactic and therapeutic measures to prevent and/or treat type 1 diabetes. The human intestinal microbiome is a complex, symbiotic ecological community that influences human health and development, including the development and maintenance of the human immune system. The role of the intestinal microbiome in autoimmunity has garnered significant attention, and evidence suggests a particular role for intestinal microbiome alterations in autoimmune disease development, including type 1 diabetes. This review will examine the role of the intestinal microbiome in the development and function of the immune system and how this relates to the development of autoimmunity. Data from animal and human studies linking alterations in the intestinal microbiome and intestinal integrity with type 1 diabetes will be closely examined. Finally, we will examine the interactions between the intestinal microbiome and dietary exposures and how these interactions may further influence autoimmunity and type 1 diabetes development.

  19. In vitro patterning of pluripotent stem cell-derived intestine recapitulates in vivo human development.

    PubMed

    Tsai, Yu-Hwai; Nattiv, Roy; Dedhia, Priya H; Nagy, Melinda S; Chin, Alana M; Thomson, Matthew; Klein, Ophir D; Spence, Jason R

    2017-03-15

    The intestine plays a central role in digestion, nutrient absorption and metabolism, with individual regions of the intestine having distinct functional roles. Many examples of region-specific gene expression in the adult intestine are known, but how intestinal regional identity is established during development is a largely unresolved issue. Here, we have identified several genes that are expressed in a region-specific manner in the developing human intestine. Using human embryonic stem cell-derived intestinal organoids, we demonstrate that the duration of exposure to active FGF and WNT signaling controls regional identity. Short-term exposure to FGF4 and CHIR99021 (a GSK3β inhibitor that stabilizes β-catenin) resulted in organoids with gene expression patterns similar to developing human duodenum, whereas longer exposure resulted in organoids similar to ileum. When region-specific organoids were transplanted into immunocompromised mice, duodenum-like organoids and ileum-like organoids retained their regional identity, demonstrating that regional identity of organoids is stable after initial patterning occurs. This work provides insights into the mechanisms that control regional specification of the developing human intestine and provides new tools for basic and translational research.

  20. Metabolism of puerarin and daidzin by human intestinal bacteria and their relation to in vitro cytotoxicity.

    PubMed

    Kim, D H; Yu, K U; Bae, E A; Han, M J

    1998-06-01

    When puerarin or daidzin were incubated for 24 h with human intestinal bacteria, two metabolites, daidzein and calycosin, were produced from them, respectively. The metabolic time course of puerarin was as follows: at an early time, puerarin was converted to daidzin, and then calycosin. The metabolic time course of daidzin by human intestinal bacteria was also similar to that of puerarin. The in vitro cytotoxicities of these metabolites, calycosin and daidzein, were superior to those of puerarin and daidzein.

  1. Human Intestinal Barrier Function in Health and Disease

    PubMed Central

    König, Julia; Wells, Jerry; Cani, Patrice D; García-Ródenas, Clara L; MacDonald, Tom; Mercenier, Annick; Whyte, Jacqueline; Troost, Freddy; Brummer, Robert-Jan

    2016-01-01

    The gastrointestinal tract consists of an enormous surface area that is optimized to efficiently absorb nutrients, water, and electrolytes from food. At the same time, it needs to provide a tight barrier against the ingress of harmful substances, and protect against a reaction to omnipresent harmless compounds. A dysfunctional intestinal barrier is associated with various diseases and disorders. In this review, the role of intestinal permeability in common disorders such as infections with intestinal pathogens, inflammatory bowel disease, irritable bowel syndrome, obesity, celiac disease, non-celiac gluten sensitivity, and food allergies will be discussed. In addition, the effect of the frequently prescribed drugs proton pump inhibitors and non-steroidal anti-inflammatory drugs on intestinal permeability, as well as commonly used methods to assess barrier function will be reviewed. PMID:27763627

  2. [Intestinal microbiota].

    PubMed

    Debré, Patrice; Le Gall, Jean-Yves

    2014-12-01

    The human body normally lives in symbiosis with a considerable microscopic environment present on all interfaces with the external environment; it hosts ten times more microbes (microbiota) that it has somatic or germ cells, representing a gene diversity (microbiome) 100-150 times higher than the human genome. These germs are located mainly in the gut, where they represent a mass of about one kilogram. The primary colonization of the gastrointestinal tract depends on the delivery route, the bacterial flora rewarding then depending on the environment, food hygiene, medical treatments. The intestinal microbiota plays an important role in the maturation of the immune system and in different physiological functions: digestion of polysaccharides, glycosaminoglycans and glycoproteins, vitamins biosynthesis, bile salt metabolism of some amino acids and xenobiotics. Quantitative and qualitative changes in the microbiota are observed in a wide range of diseases: obesity, colorectal cancer, liver cancer, inflammatory bowel disease, autoimmune diseases, allergies... pharmacobiotics aim to modify the intestinal microbiota in a therapeutic goal and this by various means: prebiotics, probiotics, antibiotics or fecal transplants. Intestinal flora also plays a direct role in the metabolism of certain drugs and the microbiota should be considered as a predictive parameter of response to some chemotherapies.

  3. Role of Bacterial Endotoxins of Intestinal Origin in Rat Heat Stress Mortality

    DTIC Science & Technology

    1982-01-19

    Intestinal Origin in M -4/81 litle U H a B. CONTRACT OR GRANT NUMBER(’) D. A. DuBose, K. Basamania, L. Maglione , and J. Rowlands 9. PERFORMING...mortality D. A. DuBOSE, K. BASAMANIA, L. MAGLIONE , AND J. ROWLANDS Department of the Army, US Army Research Institute of Environmental Medicine...Natick, Massachusetts 01760 DX3BOSE, D. A., K. BASAMANIA, L. MAGLIONE , AND J. Row- mined the incidence of duodenal and extraintestinal in- LANDS. Role of

  4. Pseudomonas reactans, a bacterial strain isolated from the intestinal flora of Blattella germanica with anti-Beauveria bassiana activity.

    PubMed

    Zhang, Fan; Huang, Yan Hong; Liu, Shu Zhen; Zhang, Lei; Li, Bo Tai; Zhao, Xiao Xu; Fu, Ying; Liu, Jian Jun; Zhang, Xue Xia

    2013-06-01

    Anti-Beauveria bassiana activity of aqueous fecal extracts from conventional German cockroaches [Blattella germanica (L.)] was detected, but was not detected in samples from germ-free German cockroaches. Subsequently, bacterial strain BGI-14 was isolated from the gut of conventional German cockroaches and was identified as Pseudomonas reactans based on 16S rDNA sequence. The strain BGI-14 not only inhibited the germination of conidia, but also inhibited the growth of B. bassiana hyphae. Further studies demonstrated that B. bassiana infections in German cockroaches orally treated with the extracts of BGI-14 fermentation were significantly weakened. Compared with the control group, the cumulative mortality rate of treatment group was reduced by 10.3% at 20 d postinoculation. These studies imply that intestinal flora with anti-B. bassiana activity might contribute to resistance of infection by entomopathogenic fungi.

  5. Prostaglandin E1 maintains structural integrity of intestinal mucosa and prevents bacterial translocation during experimental obstructive jaundice.

    PubMed

    Gurleyik, Emin; Coskun, Ozgur; Ustundag, Nil; Ozturk, Elif

    2006-01-01

    The absence of bile in the gut lumen induces mucosal injury and promotes bacterial translocation (BT). Prostaglandin E (PGE) has a protective effect on the mucosal layer of the alimentary tract. We hypothesize that PGE1 may prevent BT by its beneficial action on the mucosa of the small bowel. Thirty Wistar albino rats were divided equally into 3 groups; Group 1 (control) underwent sham laparotomy, group 2 obstructive jaundice (OJ) and group 3 (OJ + PGE1) underwent common bile duct (CBD) ligation and transection. Groups 1 and 2 received; 1 mL normal saline and group 3 received 40 mg of the PGE1 analogue misoprostol dissolved in 1 mL normal saline administered by orogastric tube once daily. After 7 days, laparotomy and collection of samples for laboratory analyses were performed, including bacteriological analysis of intestine, mesenteric lymph nodes (MLNs), and blood, and histopathologic examination of intestinal mucosa to determine mucosal thickness and structural damage. Serum bilirubin and alkaline phosphatase levels confirmed OJ in all animals with CBD transection. The mucosal damage score was significantly reduced in jaundiced animals receiving PGE1 compared to jaundiced controls (2.15 +/- 0.74 vs 5.3 +/- 0.59; p < .00001) and mucosal thickness was greater (607 +/- 59.1 microm vs. 393 +/- 40.3 microm; p < .00001). The incidence of BT to MLNs decreased from 90% to 30% (p < .02) when jaundiced rats received PGE1. PGE1 treatment reduced the detection rate of viable enteric bacteria in the blood from 60% to 10% (p < .057). We conclude that administration of PGE1 provides protection against OJ-induced atrophy and damage of intestinal mucosa, and thereby prevents translocation of enteric bacteria to underlying tissues.

  6. Long-term changes of bacterial and viral compositions in the intestine of a recovered Clostridium difficile patient after fecal microbiota transplantation

    PubMed Central

    Broecker, Felix; Klumpp, Jochen; Schuppler, Markus; Russo, Giancarlo; Biedermann, Luc; Hombach, Michael; Rogler, Gerhard; Moelling, Karin

    2016-01-01

    Fecal microbiota transplantation (FMT) is an effective treatment for recurrent Clostridium difficile infections (RCDIs). However, long-term effects on the patients’ gut microbiota and the role of viruses remain to be elucidated. Here, we characterized bacterial and viral microbiota in the feces of a cured RCDI patient at various time points until 4.5 yr post-FMT compared with the stool donor. Feces were subjected to DNA sequencing to characterize bacteria and double-stranded DNA (dsDNA) viruses including phages. The patient's microbial communities varied over time and showed little overall similarity to the donor until 7 mo post-FMT, indicating ongoing gut microbiota adaption in this time period. After 4.5 yr, the patient's bacteria attained donor-like compositions at phylum, class, and order levels with similar bacterial diversity. Differences in the bacterial communities between donor and patient after 4.5 yr were seen at lower taxonomic levels. C. difficile remained undetectable throughout the entire timespan. This demonstrated sustainable donor feces engraftment and verified long-term therapeutic success of FMT on the molecular level. Full engraftment apparently required longer than previously acknowledged, suggesting the implementation of year-long patient follow-up periods into clinical practice. The identified dsDNA viruses were mainly Caudovirales phages. Unexpectedly, sequences related to giant algae–infecting Chlorella viruses were also detected. Our findings indicate that intestinal viruses may be implicated in the establishment of gut microbiota. Therefore, virome analyses should be included in gut microbiota studies to determine the roles of phages and other viruses—such as Chlorella viruses—in human health and disease, particularly during RCDI. PMID:27148577

  7. Human intestine luminal ACE2 and amino acid transporter expression increased by ACE-inhibitors.

    PubMed

    Vuille-dit-Bille, Raphael N; Camargo, Simone M; Emmenegger, Luca; Sasse, Tom; Kummer, Eva; Jando, Julia; Hamie, Qeumars M; Meier, Chantal F; Hunziker, Schirin; Forras-Kaufmann, Zsofia; Kuyumcu, Sena; Fox, Mark; Schwizer, Werner; Fried, Michael; Lindenmeyer, Maja; Götze, Oliver; Verrey, François

    2015-04-01

    Sodium-dependent neutral amino acid transporter B(0)AT1 (SLC6A19) and imino acid (proline) transporter SIT1 (SLC6A20) are expressed at the luminal membrane of small intestine enterocytes and proximal tubule kidney cells where they exert key functions for amino acid (re)absorption as documented by their role in Hartnup disorder and iminoglycinuria, respectively. Expression of B(0)AT1 was shown in rodent intestine to depend on the presence of the carboxypeptidase angiotensin-converting enzyme 2 (ACE2). This enzyme belongs to the renin-angiotensin system and its expression is induced by treatment with ACE-inhibitors (ACEIs) or angiotensin II AT1 receptor blockers (ARBs) in many rodent tissues. We show here in the Xenopus laevis oocyte expression system that human ACE2 also functionally interacts with SIT1. To investigate in human intestine the potential effect of ACEIs or ARBs on ACE2, we analysed intestinal biopsies taken during routine gastroduodenoscopy and ileocolonoscopy from 46 patients of which 9 were under ACEI and 13 ARB treatment. Analysis of transcript expression by real-time PCR and of proteins by immunofluorescence showed a co-localization of SIT1 and B(0)AT1 with ACE2 in the brush-border membrane of human small intestine enterocytes and a distinct axial expression pattern of the tested gene products along the intestine. Patients treated with ACEIs displayed in comparison with untreated controls increased intestinal mRNA levels of ACE2, peptide transporter PEPT1 (SLC15A1) and AA transporters B(0)AT1 and PAT1 (SLC36A1). This study unravels in human intestine the localization and distribution of intestinal transporters involved in amino acid absorption and suggests that ACEIs impact on their expression.

  8. The human intestinal microbiota of constipated-predominant irritable bowel syndrome patients exhibits anti-inflammatory properties

    PubMed Central

    Gobert, Alain P.; Sagrestani, Giulia; Delmas, Eve; Wilson, Keith T.; Verriere, Thomas G.; Dapoigny, Michel; Del’homme, Christophe; Bernalier-Donadille, Annick

    2016-01-01

    The intestinal microbiota of patients with constipated-predominant irritable bowel syndrome (C-IBS) displays chronic dysbiosis. Our aim was to determine whether this microbial imbalance instigates perturbation of the host intestinal mucosal immune response, using a model of human microbiota-associated rats (HMAR) and dextran sulfate sodium (DSS)-induced experimental colitis. The analysis of the microbiota composition revealed a decrease of the relative abundance of Bacteroides, Roseburia-Eubacterium rectale and Bifidobacterium and an increase of Enterobacteriaceae, Desulfovibrio sp., and mainly Akkermansia muciniphila in C-IBS patients compared to healthy individuals. The bacterial diversity of the gut microbiota of healthy individuals or C-IBS patients was maintained in corresponding HMAR. Animals harboring a C-IBS microbiota had reduced DSS colitis with a decreased expression of pro-inflammatory cytokines from innate, Th1, and Th17 responses. The pre-treatment of conventional C57BL/6 mice or HMAR with A. muciniphila, but not with Escherichia coli, prior exposure to DSS also resulted in a reduction of colitis severity, highlighting that the anti-inflammatory effect of the gut microbiota of C-IBS patients is mediated, in part, by A. muciniphila. This work highlights a novel aspect of the crosstalk between the gut microbiota of C-IBS patients and host intestinal homeostasis. PMID:27982124

  9. Contribution of gut bacterial metabolism to human metabolic disease.

    PubMed

    Bain, M D; Jones, M; Borriello, S P; Reed, P J; Tracey, B M; Chalmers, R A; Stacey, T E

    1988-05-14

    Metronidazole, an antibiotic with specific activity against anaerobic bacteria, was of clinical and biochemical benefit in two patients with methylmalonic aciduria. The virtual elimination of propionic acid from the stool suggested that propionic acid derived from faecal bacterial metabolism contributes substantially to methylmalonate production. These findings point to a novel avenue of treatment for these disorders of intermediary metabolism, and indicate the importance of microbial gut flora in normal human metabolism.

  10. Assessment of the prebiotic effect of quinoa and amaranth in the human intestinal ecosystem.

    PubMed

    Gullón, Beatriz; Gullón, Patricia; Tavaria, Freni K; Yáñez, Remedios

    2016-09-14

    Quinoa and amaranth belong to the group of the so called "superfoods" and have a nutritional composition that confers multiple benefits. In this work, we explored the possibility of these foods exhibiting a prebiotic effect. These pseudocereals were subjected to an in vitro digestion and used as carbon sources in batch cultures with faecal human inocula. The effects on the microbiota composition and their metabolic products were determined by assessment of variations in pH, short-chain fatty acid (SCFA) production and changes in the dynamic bacterial populations by fluorescence in situ hybridization (FISH). After 48 h of incubation, the total SCFAs were 106.5 mM for quinoa and 108.83 mM for amaranth, in line with the decrease in pH. Considerable differences (p < 0.05) were found in certain microbial groups, including Bifidobacterium spp., Lactobacillus-Enterococcus, Atopobium, Bacteroides-Prevotella, Clostridium coccoides-Eubacterium rectale, Faecalibacterium prausnitzii and Roseburia intestinalis. Our research suggests that these pseudocereals can have the prebiotic potential and that their intake may improve dysbiosis or maintain the gastrointestinal health through a balanced intestinal microbiota, although additional studies are necessary.

  11. Practical techniques for detection of Toll-like receptor-4 in the human intestine.

    PubMed

    Ungaro, Ryan; Abreu, Maria T; Fukata, Masayuki

    2009-01-01

    The human intestine has evolved in the presence of a diverse array of luminal microorganisms. In order to maintain intestinal homeostasis, mucosal immune responses to theses microorganisms must be tightly regulated. The intestine needs to be able to respond to pathogenic organisms while at the same time maintain tolerance to normal commensal flora. Toll-like receptors (TLRs) play an important role in this delicate balance. TLRs are transmembrane noncatalytic receptor proteins that induce activation of innate and adaptive immune responses to microorganisms by recognizing structurally conserved molecular patterns of microbes. Expression of TLRs by intestinal epithelial cell is normally down-regulated to maintain immune tolerance to the luminal microorganisms.One of the challenges of TLR research in the human intestine is that it is difficult for many experimental methods to detect very low expression of TLRs within the intestinal mucosa. Quantitative methods such as PCR are limited in their ability to detect TLR expression by specific cell types within a tissue sample, which can be important when studying the contribution of TLR signaling to pathological conditions. In this regard, immunohistochemistry (IHC) is advantageous in that one can visualize the distribution and localization of target proteins within both normal and pathologic parts of a given tissue sample. We found that a subset of human colorectal cancers over-express TLR4 by means of immunofluorescence (IF) and IHC methods. Localization of TLR4 within cancer tissue often appears to be patchy, making IHC an appropriate way to examine these changes. We will describe our current techniques to detect TLR4 in paraffin-embedded human large intestine sections. Establishing a practical IHC technique that may provide consistent results between laboratories will significantly enhance understanding of the role of TLRs in human intestinal health and disease.

  12. Immunological quantitation and localization of ACAT-1 and ACAT-2 in human liver and small intestine.

    PubMed

    Chang, C C; Sakashita, N; Ornvold, K; Lee, O; Chang, E T; Dong, R; Lin, S; Lee, C Y; Strom, S C; Kashyap, R; Fung, J J; Farese, R V; Patoiseau, J F; Delhon, A; Chang, T Y

    2000-09-08

    By using specific anti-ACAT-1 antibodies in immunodepletion studies, we previously found that ACAT-1, a 50-kDa protein, plays a major catalytic role in the adult human liver, adrenal glands, macrophages, and kidneys but not in the intestine. Acyl-coenzyme A:cholesterol acyltransferase (ACAT) activity in the intestine may be largely derived from a different ACAT protein. To test this hypothesis, we produced specific polyclonal anti-ACAT-2 antibodies that quantitatively immunodepleted human ACAT-2, a 46-kDa protein expressed in Chinese hamster ovary cells. In hepatocyte-like HepG2 cells, ACAT-1 comprises 85-90% of the total ACAT activity, with the remainder attributed to ACAT-2. In adult intestines, most of the ACAT activity can be immunodepleted by anti-ACAT-2. ACAT-1 and ACAT-2 do not form hetero-oligomeric complexes. In differentiating intestinal enterocyte-like Caco-2 cells, ACAT-2 protein content increases by 5-10-fold in 6 days, whereas ACAT-1 protein content remains relatively constant. In the small intestine, ACAT-2 is concentrated at the apices of the villi, whereas ACAT-1 is uniformly distributed along the villus-crypt axis. In the human liver, ACAT-1 is present in both fetal and adult hepatocytes. In contrast, ACAT-2 is evident in fetal but not adult hepatocytes. Our results collectively suggest that in humans, ACAT-2 performs significant catalytic roles in the fetal liver and in intestinal enterocytes.

  13. Evolution of antibiotic resistance by human and bacterial niche construction.

    PubMed

    Boni, Maciej F; Feldman, Marcus W

    2005-03-01

    Antibiotic treatment by humans generates strong viability selection for antibiotic-resistant bacterial strains. The frequency of host antibiotic use often determines the strength of this selection, and changing patterns of antibiotic use can generate many types of behaviors in the population dynamics of resistant and sensitive bacterial populations. In this paper, we present a simple model of hosts dimorphic for their tendency to use/avoid antibiotics and bacterial pathogens dimorphic in their resistance/sensitivity to antibiotic treatment. When a constant fraction of hosts uses antibiotics, the two bacterial strain populations can coexist unless host use-frequency is above a critical value; this critical value is derived as the ratio of the fitness cost of resistance to the fitness cost of undergoing treatment. When strain frequencies can affect host behavior, the dynamics may be analyzed in the light of niche construction. We consider three models underlying changing host behavior: conformism, the avoidance of long infections, and adherence to the advice of public health officials. In the latter two, we find that the pathogen can have quite a strong effect on host behavior. In particular, if antibiotic use is discouraged when resistance levels are high, we observe a classic niche-construction phenomenon of maintaining strain polymorphism even in parameter regions where it would not be expected.

  14. Characterization of two cysteine proteases secreted by Blastocystis ST7, a human intestinal parasite.

    PubMed

    Wawrzyniak, Ivan; Texier, Catherine; Poirier, Philippe; Viscogliosi, Eric; Tan, Kevin S W; Delbac, Frédéric; El Alaoui, Hicham

    2012-09-01

    Blastocystis spp. are unicellular anaerobic intestinal parasites of both humans and animals and the most prevalent ones found in human stool samples. Their association with various gastrointestinal disorders raises the questions of its pathogenicity and of the molecular mechanisms involved. Since secreted proteases are well-known to be implicated in intestinal parasite virulence, we intended to determine whether Blastocystis spp. possess such pathogenic factors. In silico analysis of the Blastocystis subtype 7 (ST7) genome sequence highlighted 22 genes coding proteases which were predicted to be secreted. We characterized the proteolytic activities in the secretory products of Blastocystis ST7 using specific protease inhibitors. Two cysteine proteases, a cathepsin B and a legumain, were identified in the parasite culture supernatant by gelatin zymographic SDS-PAGE gel and MS/MS analysis. These proteases might act on intestinal cells and disturb gut function. This work provides serious molecular candidates to link Blastocystis spp. and intestinal disorders.

  15. Isolation and identification of intestinal CYP3A inhibitors from cranberry (Vaccinium macrocarpon) using human intestinal microsomes.

    PubMed

    Kim, Eunkyung; Sy-Cordero, Arlene; Graf, Tyler N; Brantley, Scott J; Paine, Mary F; Oberlies, Nicholas H

    2011-02-01

    Cranberry juice is used routinely, especially among women and the elderly, to prevent and treat urinary tract infections. These individuals are likely to be taking medications concomitantly with cranberry juice, leading to concern about potential drug-dietary substance interactions, particularly in the intestine, which, along with the liver, is rich in expression of the prominent drug metabolizing enzyme, cytochrome P450 3A (CYP3A). Using a systematic in vitro-in vivo approach, a cranberry juice product was identified recently that elicited a pharmacokinetic interaction with the CYP3A probe substrate midazolam in 16 healthy volunteers. Relative to water, cranberry juice inhibited intestinal first-pass midazolam metabolism. In vitro studies were initiated to identify potential enteric CYP3A inhibitors from cranberry via a bioactivity-directed fractionation approach involving dried whole cranberry [Vaccinium macrocarpon Ait. (Ericaceae)], midazolam, and human intestinal microsomes (HIM). Three triterpenes (maslinic acid, corosolic acid, and ursolic acid) were isolated. The inhibitory potency (IC(50)) of maslinic acid, corosolic acid, and ursolic acid was 7.4, 8.8, and < 10 µM, respectively, using HIM as the enzyme source and 2.8, 4.3, and < 10 µM, respectively, using recombinant CYP3A4 as the enzyme source. These in vitro inhibitory potencies, which are within the range of those reported for two CYP3A inhibitory components in grapefruit juice, suggest that these triterpenes may have contributed to the midazolam-cranberry juice interaction observed in the clinical study.

  16. Antimicrobial peptide LL-37 promotes bacterial phagocytosis by human macrophages.

    PubMed

    Wan, Min; van der Does, Anne M; Tang, Xiao; Lindbom, Lennart; Agerberth, Birgitta; Haeggström, Jesper Z

    2014-06-01

    LL-37/hCAP-18 is the only human member of the cathelicidin family and plays an important role in killing various pathogens, as well as in immune modulation. In this study, we investigated the effect of LL-37 on bacterial phagocytosis by macrophages and demonstrate that LL-37 enhances phagocytosis of IgG-opsonized Gram-negative and Gram-positive bacteria in a dose- and time-dependent manner by dTHP-1 cells. In addition, LL-37 enhanced phagocytosis of nonopsonized Escherichia coli by human macrophages. Consistently, LL-37 elevated the expression of FcγRs on macrophages but not the complement receptors CD11b and -c. Further studies revealed that the expression of TLR4 and CD14 is also increased on LL-37-treated macrophages. Several lines of evidence indicated that the FPR2/ALX receptor mediated LL-37-induced phagocytosis. However, TLR4 signaling was also coupled to the phagocytic response, as a specific TLR4 antibody significantly suppressed phagocytosis of IgG-opsonized E. coli and nonopsonized E. coli by dTHP-1 cells. Finally, macrophages from Cnlp(-/-) mice exhibited diminished bacterial phagocytosis compared with macrophages from their WT littermates. In conclusion, we demonstrate a novel, immune-modulatory mechanism of LL-37, which may contribute to bacterial clearance.

  17. Diversity of human vaginal bacterial communities and associations with clinically defined bacterial vaginosis.

    PubMed

    Oakley, Brian B; Fiedler, Tina L; Marrazzo, Jeanne M; Fredricks, David N

    2008-08-01

    Bacterial vaginosis (BV) is a common syndrome associated with numerous adverse health outcomes in women. Despite its medical importance, the etiology and microbial ecology of BV remain poorly understood. We used broad-range PCR to census the community structure of the healthy and BV-affected vaginal microbial ecosystems and synthesized current publicly available bacterial 16S rRNA gene sequence data from this environment. The community of vaginal bacteria detected in subjects with BV was much more taxon rich and diverse than in subjects without BV. At a 97% sequence similarity cutoff, the number of operational taxonomic units (OTUs) per patient in 28 subjects with BV was nearly three times greater than in 13 subjects without BV: 14.8 +/- 0.7 versus 5.2 +/- 0.75 (mean +/- standard error). OTU-based analyses revealed previously hidden diversity for many vaginal bacteria that are currently poorly represented in GenBank. Our sequencing efforts yielded many novel phylotypes (123 of our sequences represented 38 OTUs not previously found in the vaginal ecosystem), including several novel BV-associated OTUs, such as those belonging to the Prevotella species complex, which remain severely underrepresented in the current NCBI database. Community composition was highly variable among subjects at a fine taxonomic scale, but at the phylum level, Actinobacteria and Bacteroidetes were strongly associated with BV. Our data describe a previously unrecognized extent of bacterial diversity in the vaginal ecosystem. The human vagina hosts many bacteria that are only distantly related to known species, and subjects with BV harbor particularly taxon-rich and diverse bacterial communities.

  18. Diversity of Human Vaginal Bacterial Communities and Associations with Clinically Defined Bacterial Vaginosis▿ †

    PubMed Central

    Oakley, Brian B.; Fiedler, Tina L.; Marrazzo, Jeanne M.; Fredricks, David N.

    2008-01-01

    Bacterial vaginosis (BV) is a common syndrome associated with numerous adverse health outcomes in women. Despite its medical importance, the etiology and microbial ecology of BV remain poorly understood. We used broad-range PCR to census the community structure of the healthy and BV-affected vaginal microbial ecosystems and synthesized current publicly available bacterial 16S rRNA gene sequence data from this environment. The community of vaginal bacteria detected in subjects with BV was much more taxon rich and diverse than in subjects without BV. At a 97% sequence similarity cutoff, the number of operational taxonomic units (OTUs) per patient in 28 subjects with BV was nearly three times greater than in 13 subjects without BV: 14.8 ± 0.7 versus 5.2 ± 0.75 (mean ± standard error). OTU-based analyses revealed previously hidden diversity for many vaginal bacteria that are currently poorly represented in GenBank. Our sequencing efforts yielded many novel phylotypes (123 of our sequences represented 38 OTUs not previously found in the vaginal ecosystem), including several novel BV-associated OTUs, such as those belonging to the Prevotella species complex, which remain severely underrepresented in the current NCBI database. Community composition was highly variable among subjects at a fine taxonomic scale, but at the phylum level, Actinobacteria and Bacteroidetes were strongly associated with BV. Our data describe a previously unrecognized extent of bacterial diversity in the vaginal ecosystem. The human vagina hosts many bacteria that are only distantly related to known species, and subjects with BV harbor particularly taxon-rich and diverse bacterial communities. PMID:18487399

  19. Translocation of coagulase-negative bacterial staphylococci in rats following intestinal ischemia-reperfusion injury.

    PubMed

    Luo, Chih-Cheng; Shih, Hsiang-Hung; Chiu, Cheng-Hsun; Lin, Jer-Nan

    2004-01-01

    Many patients with sepsis have bacteremia for which no septic focus is identified either clinically or by autopsy. This study was designed to determine the relationship between the ischemia-reperfusion injury (IRI) and bacterial translocation that might be involved in the pathogenesis of necrotizing enterocolitis. In the first experiment, a total of 32 Sprague-Dawley rats weighing 150-200 g were divided into four groups. The mesentery to isolated loop was occluded for 30, 60, and 90 min following 30-min reperfusion in the three groups of experimental animals with a micro-bulldog clamp. A control group involved the same technique and exposure, without occlusion of the mesentery. Two sets of blood culture were taken through a catheter in the portal vein immediately and 15 min after the reperfusion, respectively. In another experiment, bacteria isolated were fed in different doses to control rats and those after 30- or 60-min ischemia and 30-min reperfusion. Two sets of blood culture were taken following the procedure. Invasion and transcytosis of the bacteria through epithelial cells were studied in vitro using a Madin-Derby canine kidney (MDCK) cell monolayer model. PCR for delta toxin gene was performed on all bacteria isolated, using Staphylococcus epidermidis as the control. Coagulase-negative staphylococci (CoNS) were invariably isolated from mice with prolonged ischemia (90 min) and reperfusion. When bacteria were fed into mice with only 30-min ischemia, an inoculum as low as 5 x 10(5) CFU/ml could induce bacteremia. No bacterial translocation was found in control mice even fed with a higher dose of bacteria (5 x 10(8) CFU/ml). In vitro experiments showed that CoNS failed to transcytose MDCK monolayer. These isolates were not cytotoxic to MDCK cells and contained no delta toxin gene. Bacterial translocation of CoNS occurred following severe bowel ischemia and reperfusion injury. Intact mucosa integrity readily prevented bacterial translocation; however

  20. The Mouse Intestinal Bacterial Collection (miBC) provides host-specific insight into cultured diversity and functional potential of the gut microbiota.

    PubMed

    Lagkouvardos, Ilias; Pukall, Rüdiger; Abt, Birte; Foesel, Bärbel U; Meier-Kolthoff, Jan P; Kumar, Neeraj; Bresciani, Anne; Martínez, Inés; Just, Sarah; Ziegler, Caroline; Brugiroux, Sandrine; Garzetti, Debora; Wenning, Mareike; Bui, Thi P N; Wang, Jun; Hugenholtz, Floor; Plugge, Caroline M; Peterson, Daniel A; Hornef, Mathias W; Baines, John F; Smidt, Hauke; Walter, Jens; Kristiansen, Karsten; Nielsen, Henrik B; Haller, Dirk; Overmann, Jörg; Stecher, Bärbel; Clavel, Thomas

    2016-08-08

    Intestinal bacteria influence mammalian physiology, but many types of bacteria are still uncharacterized. Moreover, reference strains of mouse gut bacteria are not easily available, although mouse models are extensively used in medical research. These are major limitations for the investigation of intestinal microbiomes and their interactions with diet and host. It is thus important to study in detail the diversity and functions of gut microbiota members, including those colonizing the mouse intestine. To address these issues, we aimed at establishing the Mouse Intestinal Bacterial Collection (miBC), a public repository of bacterial strains and associated genomes from the mouse gut, and studied host-specificity of colonization and sequence-based relevance of the resource. The collection includes several strains representing novel species, genera and even one family. Genomic analyses showed that certain species are specific to the mouse intestine and that a minimal consortium of 18 strains covered 50-75% of the known functional potential of metagenomes. The present work will sustain future research on microbiota-host interactions in health and disease, as it will facilitate targeted colonization and molecular studies. The resource is available at www.dsmz.de/miBC.

  1. Recognition of human milk oligosaccharides by bacterial exotoxins.

    PubMed

    El-Hawiet, Amr; Kitova, Elena N; Klassen, John S

    2015-08-01

    The affinities of the most abundant oligosaccharides found in human milk for four bacterial exotoxins (from Vibrio cholerae and pathogenic Escherichia coli) were quantified for the first time. Association constants (Ka) for a library of 20 human milk oligosaccharides (HMOs) binding to Shiga toxin type 2 holotoxin (Stx2) and the B subunit homopentamers of cholera toxin, heat-labile toxin and Shiga toxin type 1 (CTB5, HLTB5 and Stx1B5) were measured at 25°C and pH 7 using the direct electrospray ionization mass spectrometry assay. Notably, all four bacterial toxins bind to a majority of the HMOs tested and five of the HMOs (2'-fucosyllactose, lacto-N-tetraose, lacto-N-fucopentaose I, lacto-N-fucopentaose II and lacto-N-fucopentaose III) are ligands for all four toxins. These five HMOs are also reported to bind to other bacterial toxins (e.g. toxin A and toxin B of Clostridium difficile). In all cases, the HMO affinities (apparent Ka) are relatively modest (≤15,000 M(-1)). However, at the high concentrations of HMOs typically ingested by infants, a significant fraction of these toxins, if present, is expected to be bound to HMOs. Binding measurements carried out with 2'-fucosyllactose or lacto-N-fucopentaose I, together with a high-affinity ligand based on the native carbohydrate receptor, revealed that all four toxins possess HMO-binding sites that are distinct from those of the native receptors, although evidence of competitive binding was found for lacto-N-fucopentaose I with Stx2 and 2'-fucosyllactose and lacto-N-fucopentaose I with HLTB5. Taken together, the results of this study suggest that, while HMOs are expected to bind extensively to these bacterial toxins, it is unlikely that HMO binding will effectively inhibit their interactions with their cellular receptors.

  2. Expression, Distribution and Role of Aquaporin Water Channels in Human and Animal Stomach and Intestines.

    PubMed

    Zhu, Cui; Chen, Zhuang; Jiang, Zongyong

    2016-08-29

    Stomach and intestines are involved in the secretion of gastrointestinal fluids and the absorption of nutrients and fluids, which ensure normal gut functions. Aquaporin water channels (AQPs) represent a major transcellular route for water transport in the gastrointestinal tract. Until now, at least 11 AQPs (AQP1-11) have been found to be present in the stomach, small and large intestines. These AQPs are distributed in different cell types in the stomach and intestines, including gastric epithelial cells, gastric glands cells, absorptive epithelial cells (enterocytes), goblet cells and Paneth cells. AQP1 is abundantly distributed in the endothelial cells of the gastrointestinal tract. AQP3 and AQP4 are mainly distributed in the basolateral membrane of epithelial cells in the stomach and intestines. AQP7, AQP8, AQP10 and AQP11 are distributed in the apical of enterocytes in the small and large intestines. Although AQP-null mice displayed almost no phenotypes in gastrointestinal tracts, the alterations of the expression and localization of these AQPs have been shown to be associated with the pathology of gastrointestinal disorders, which suggests that AQPs play important roles serving as potential therapeutic targets. Therefore, this review provides an overview of the expression, localization and distribution of AQPs in the stomach, small and large intestine of human and animals. Furthermore, this review emphasizes the potential roles of AQPs in the physiology and pathophysiology of stomach and intestines.

  3. Expression, Distribution and Role of Aquaporin Water Channels in Human and Animal Stomach and Intestines

    PubMed Central

    Zhu, Cui; Chen, Zhuang; Jiang, Zongyong

    2016-01-01

    Stomach and intestines are involved in the secretion of gastrointestinal fluids and the absorption of nutrients and fluids, which ensure normal gut functions. Aquaporin water channels (AQPs) represent a major transcellular route for water transport in the gastrointestinal tract. Until now, at least 11 AQPs (AQP1–11) have been found to be present in the stomach, small and large intestines. These AQPs are distributed in different cell types in the stomach and intestines, including gastric epithelial cells, gastric glands cells, absorptive epithelial cells (enterocytes), goblet cells and Paneth cells. AQP1 is abundantly distributed in the endothelial cells of the gastrointestinal tract. AQP3 and AQP4 are mainly distributed in the basolateral membrane of epithelial cells in the stomach and intestines. AQP7, AQP8, AQP10 and AQP11 are distributed in the apical of enterocytes in the small and large intestines. Although AQP-null mice displayed almost no phenotypes in gastrointestinal tracts, the alterations of the expression and localization of these AQPs have been shown to be associated with the pathology of gastrointestinal disorders, which suggests that AQPs play important roles serving as potential therapeutic targets. Therefore, this review provides an overview of the expression, localization and distribution of AQPs in the stomach, small and large intestine of human and animals. Furthermore, this review emphasizes the potential roles of AQPs in the physiology and pathophysiology of stomach and intestines. PMID:27589719

  4. Composition and Metabolic Activities of Bacterial Biofilms Colonizing Food Residues in the Human Gut

    PubMed Central

    Macfarlane, Sandra; Macfarlane, George T.

    2006-01-01

    Bacteria growing in the human large intestine live in intimate association with the host and play an important role in host digestive processes, gut physiology, and metabolism. Fecal bacteria have been investigated extensively, but few studies have been done on biofilms that form on digestive wastes in the large bowel. The aims of this investigation were to investigate the composition and metabolic activities of bacterial communities that colonize the surfaces of food residues in fecal material, with respect to their role in the fermentation of complex carbohydrates. Fresh stools were obtained from 15 healthy donors, and food residues were separated by filtration. Adherent bacteria were removed by surfactant treatment for microbiological analysis and fermentation studies. Scanning electron microscopy and fluorescent in situ hybridization in conjunction with confocal laser scanning microscopy (CLSM) were used to visualize intact biofilms. Results showed that bacterial populations strongly adhering to particulate matter were phenotypically similar in composition to unattached communities, with bacteroides and bifidobacteria predominating. Biofilms comprised a mixture of living and dead bacteria, and CLSM showed that bifidobacteria in the biofilms occurred as isolated dispersed cells and in microcolonies near the interface with the substratum. Fermentation experiments with a variety of complex carbohydrates demonstrated that biofilm populations were more efficient in digesting polysaccharides, while nonadhering communities fermented oligosaccharides most rapidly. Acetate was the principal fermentation product formed by biofilm bacteria, whereas higher levels of butyrate were produced by nonadherent populations, showing that the two communities were metabolically distinct. PMID:16957247

  5. Bacterial Community Associated with the Intestinal Tract of Chinese Mitten Crab (Eriocheir sinensis) Farmed in Lake Tai, China

    PubMed Central

    Chen, Xiaobing; Di, Panpan; Wang, Hongming; Li, Bailin; Pan, Yingjie; Yan, Shuling; Wang, Yongjie

    2015-01-01

    Chinese mitten crab (CMC, Eriocheir sinensis) is an economically valuable species in South-East Asia that has been widely farmed in China. Characterization of the intestinal bacterial diversity of CMC will provide insights into the aquaculturing of CMCs. Based on the analysis of cloned 16S rRNA genes from culture-independent CMC gut bacteria, 124 out of 128 different clones reveal >95% nucleotide similarity to the species belonging to the four phyla of Tenericutes, Bacteroidetes, Firmicutes and Proteobacteria; one clone shows 91% sequence similarity to the member of TM7 (a candidate phylum without cultured representatives). Fluorescent in situ hybridization also reveals the abundance of Bacteroidetes in crab intestine. Electron micrographs show that spherical and filamentous bacteria are closely associated with the microvillus brush border of the midgut epithelium and are often inserted into the space between the microvilli using a stalk-like cell appendage. In contrast, the predominant rod-shaped bacteria in the hindgut are tightly attached to the epithelium surface by an unusual pili-like structure. Both 16S rRNA gene denaturing gel gradient electrophoresis and metagenome library indicate that the CMC Mollicutes group 2 appears to be present in both the midgut and hindgut with no significant difference in abundance. The CMC Mollicutes group 1, however, was found mostly in the midgut of CMCs. The CMC gut Mollicutes phylotypes appear to be most closely related to Mollicutes symbionts detected in the gut of isopods (Crustacea: Isopoda). Overall, the results suggest that CMCs harbor diverse, novel and specific gut bacteria, which are likely to live in close relationships with the CMC host. PMID:25875449

  6. The human milk oligosaccharide 2′-fucosyllactose augments the adaptive response to extensive intestinal

    PubMed Central

    Hawkins, Jennifer A.; Ollberding, Nicholas J.; Karns, Rebekah; Morrow, Ardythe L.; Helmrath, Michael A.

    2015-01-01

    Intestinal resection resulting in short bowel syndrome (SBS) carries a heavy burden of long-term morbidity, mortality, and cost of care, which can be attenuated with strategies that improve intestinal adaptation. SBS infants fed human milk, compared with formula, have more rapid intestinal adaptation. We tested the hypothesis that the major noncaloric human milk oligosaccharide 2′-fucosyllactose (2′-FL) contributes to the adaptive response after intestinal resection. Using a previously described murine model of intestinal adaptation, we demonstrated increased weight gain from 21 to 56 days (P < 0.001) and crypt depth at 56 days (P < 0.0095) with 2′-FL supplementation after ileocecal resection. Furthermore, 2′-FL increased small bowel luminal content microbial alpha diversity following resection (P < 0.005) and stimulated a bloom in organisms of the genus Parabacteroides (log2-fold = 4.1, P = 0.035). Finally, transcriptional analysis of the intestine revealed enriched ontologies and pathways related to antimicrobial peptides, metabolism, and energy processing. We conclude that 2′-FL supplementation following ileocecal resection increases weight gain, energy availability through microbial community modulation, and histological changes consistent with improved adaptation. PMID:26702137

  7. Circulating intestine-derived exosomal miR-328 in plasma, a possible biomarker for estimating BCRP function in the human intestines.

    PubMed

    Gotanda, Keisuke; Hirota, Takeshi; Saito, Jumpei; Fukae, Masato; Egashira, Yu; Izumi, Noritomo; Deguchi, Mariko; Kimura, Miyuki; Matsuki, Shunji; Irie, Shin; Ieiri, Ichiro

    2016-08-30

    A variant in the breast cancer resistance protein (BCRP) gene, 421C> A is a useful biomarker for describing large inter-individual differences in the pharmacokinetics of sulfasalazine (SASP), a BCRP substrate. However, large intra-genotypic variability still exists in spite of the incorporation of this variant into the pharmacokinetics of SASP. Since miR-328 negatively regulates BCRP expression in human tissues, we hypothesized that exosomal miR-328 in plasma, which leaks from the intestines, is a possible biomarker for estimating BCRP activity in the human intestines. We established an immunoprecipitation-based quantitative method for circulating intestine-derived miR-328 in plasma using an anti-glycoprotein A33 antibody. A clinical study was conducted with an open-label, non-randomized, and single-arm design involving 33 healthy participants. Intestine-derived exosomal miR-328 levels positively correlated (P < 0.05) with SASP AUC0-48, suggesting that subjects with high miR-328 levels have low intestinal BCRP activity, resulting in the high AUC of SASP. Circulating intestine-derived exosomal miR-328 in plasma has potential as a possible biomarker for estimating BCRP function in the human intestines.

  8. Study of the Biotransformation of Tongmai Formula by Human Intestinal Flora and Its Intestinal Permeability across the Caco-2 Cell Monolayer.

    PubMed

    Wu, Shuai; Xu, Wei; Wang, Fu-Rong; Yang, Xiu-Wei

    2015-10-15

    Tongmai formula (TMF) is a well-known Chinese medicinal preparation that contains isoflavones as its major bioactive constituents. As traditional Chinese medicines (TCMs) are usually used by oral administration, their fate inside the intestinal lumen, including their biotransformation by human intestinal flora (HIF) and intestinal absorption deserves study. In this work TMF extract was incubated with human intestinal bacteria under anaerobic conditions and the changes in the twelve main constituents of TMF were then investigated. Their intestinal permeabilities, i.e., the transport capability across the intestinal brush border were investigated with a human colon carcinoma cell line (Caco-2) cell monolayer model to predict the absorption mechanism. Meanwhile, rapid HPLC-DAD methods were established for the assay. According to the biotransformation curves of the twelve constituents and the permeability coefficients, the intestinal absorption capacity of the typical compounds was elevated from the levels of 10(-7) cm/s to 10(-5) cm/s from those of the original compounds in TMF. Among them the main isoflavone glycosides puerarin (4), mirificin (6) and daidzin (7) were transformed into the same aglycone, daidzein (10). Therefore it was predicted that the aglycone compounds might be the real active ingredients in TMF. The models used can represent a novel path for the TCM studies.

  9. Saccharomyces cerevisiae strain UFMG 905 protects against bacterial translocation, preserves gut barrier integrity and stimulates the immune system in a murine intestinal obstruction model.

    PubMed

    Generoso, Simone V; Viana, Mirelle; Santos, Rosana; Martins, Flaviano S; Machado, José A N; Arantes, Rosa M E; Nicoli, Jacques R; Correia, Maria I T D; Cardoso, Valbert N

    2010-06-01

    Probiotic is a preparation containing microorganisms that confers beneficial effect to the host. This work assessed whether oral treatment with viable or heat-killed yeast Saccharomyces cerevisiae strain UFMG 905 prevents bacterial translocation (BT), intestinal barrier integrity, and stimulates the immunity, in a murine intestinal obstruction (IO) model. Four groups of mice were used: mice undergoing only laparotomy (CTL), undergoing intestinal obstruction (IO) and undergoing intestinal obstruction after previous treatment with viable or heat-killed yeast. BT, determined as uptake of (99m)Tc-E. coli in blood, mesenteric lymph nodes, liver, spleen and lungs, was significantly higher in IO group than in CTL group. Treatments with both yeasts reduced BT in blood and all organs investigated. The treatment with both yeasts also reduced intestinal permeability as determined by blood uptake of (99m)Tc-DTPA. Immunological data demonstrated that both treatments were able to significantly increase IL-10 levels, but only viable yeast had the same effect on sIgA levels. Intestinal lesions were more severe in IO group when compared to CTL and yeasts groups. Concluding, both viable and heat-killed cells of yeast prevent BT, probably by immunomodulation and by maintaining gut barrier integrity. Only the stimulation of IgA production seems to depend on the yeast viability.

  10. Cyclical DNA Methylation and Histone Changes Are Induced by LPS to Activate COX-2 in Human Intestinal Epithelial Cells

    PubMed Central

    Brancaccio, Mariarita; Coretti, Lorena; Florio, Ermanno; Pezone, Antonio; Calabrò, Viola; Falco, Geppino; Keller, Simona; Lembo, Francesca; Avvedimento, Vittorio Enrico; Chiariotti, Lorenzo

    2016-01-01

    Bacterial lipopolysaccharide (LPS) induces release of inflammatory mediators both in immune and epithelial cells. We investigated whether changes of epigenetic marks, including selected histone modification and DNA methylation, may drive or accompany the activation of COX-2 gene in HT-29 human intestinal epithelial cells upon exposure to LPS. Here we describe cyclical histone acetylation (H3), methylation (H3K4, H3K9, H3K27) and DNA methylation changes occurring at COX-2 gene promoter overtime after LPS stimulation. Histone K27 methylation changes are carried out by the H3 demethylase JMJD3 and are essential for COX-2 induction by LPS. The changes of the histone code are associated with cyclical methylation signatures at the promoter and gene body of COX-2 gene. PMID:27253528

  11. From bacterial to human dihydrouridine synthase: automated structure determination

    SciTech Connect

    Whelan, Fiona Jenkins, Huw T.; Griffiths, Samuel C.; Byrne, Robert T.; Dodson, Eleanor J.; Antson, Alfred A.

    2015-06-30

    The crystal structure of a human dihydrouridine synthase, an enzyme associated with lung cancer, with 18% sequence identity to a T. maritima enzyme, has been determined at 1.9 Å resolution by molecular replacement after extensive molecular remodelling of the template. The reduction of uridine to dihydrouridine at specific positions in tRNA is catalysed by dihydrouridine synthase (Dus) enzymes. Increased expression of human dihydrouridine synthase 2 (hDus2) has been linked to pulmonary carcinogenesis, while its knockdown decreased cancer cell line viability, suggesting that it may serve as a valuable target for therapeutic intervention. Here, the X-ray crystal structure of a construct of hDus2 encompassing the catalytic and tRNA-recognition domains (residues 1–340) determined at 1.9 Å resolution is presented. It is shown that the structure can be determined automatically by phenix.mr-rosetta starting from a bacterial Dus enzyme with only 18% sequence identity and a significantly divergent structure. The overall fold of the human Dus2 is similar to that of bacterial enzymes, but has a larger recognition domain and a unique three-stranded antiparallel β-sheet insertion into the catalytic domain that packs next to the recognition domain, contributing to domain–domain interactions. The structure may inform the development of novel therapeutic approaches in the fight against lung cancer.

  12. P-gp activity and inhibition in the different regions of human intestine ex vivo.

    PubMed

    Li, Ming; de Graaf, Inge A M; de Jager, Marina H; Groothuis, Geny M M

    2017-03-01

    Although intestinal P-glycoprotein (P-gp) has been extensively studied in vitro and in animals, its activity and the consequences of P-gp inhibition for drug disposition and toxicity in humans are still difficult to accurately extrapolate from these studies. Moreover, existing in vitro models do not take into consideration that the intestine is heterogeneous with respect to P-gp expression. Recently, we reported rat precision-cut intestinal slices (PCIS) as a physiological ex vivo model to study the regional gradient of P-gp activity and inhibition. Here we extended the application of PCIS to the human intestine. For this purpose rhodamine 123 (R123) accumulation in the presence or absence of the P-gp inhibitors verapamil, cyclosporine A, quinidine, ketoconazole, PSC833 and CP100356 was measured in PCIS of human duodenum, jejunum, ileum and colon. R123 accumulation in the presence of the P-gp inhibitors appeared to be most enhanced in the ileum compared to the other regions. Moreover, the regional differences in accumulation are in line with published differences in abundance of P-gp. The rank order of the potency of the P-gp inhibitors, reflected by their IC50 , was comparable to that in rat PCIS. However, the increase in accumulation of the P-gp substrate R123 by the inhibitors was larger in human ileum PCIS than in rat PCIS, indicating species difference in P-gp abundance. These data show that human PCIS are an appropriate ex vivo model to study the activity of intestinal P-gp and predict the inhibitory effect of drugs and of transporter-mediated drug-drug interactions in the human intestine. Copyright © 2016 John Wiley & Sons, Ltd.

  13. Anti-Infective Activities of Lactobacillus Strains in the Human Intestinal Microbiota: from Probiotics to Gastrointestinal Anti-Infectious Biotherapeutic Agents

    PubMed Central

    Liévin-Le Moal, Vanessa

    2014-01-01

    SUMMARY A vast and diverse array of microbial species displaying great phylogenic, genomic, and metabolic diversity have colonized the gastrointestinal tract. Resident microbes play a beneficial role by regulating the intestinal immune system, stimulating the maturation of host tissues, and playing a variety of roles in nutrition and in host resistance to gastric and enteric bacterial pathogens. The mechanisms by which the resident microbial species combat gastrointestinal pathogens are complex and include competitive metabolic interactions and the production of antimicrobial molecules. The human intestinal microbiota is a source from which Lactobacillus probiotic strains have often been isolated. Only six probiotic Lactobacillus strains isolated from human intestinal microbiota, i.e., L. rhamnosus GG, L. casei Shirota YIT9029, L. casei DN-114 001, L. johnsonii NCC 533, L. acidophilus LB, and L. reuteri DSM 17938, have been well characterized with regard to their potential antimicrobial effects against the major gastric and enteric bacterial pathogens and rotavirus. In this review, we describe the current knowledge concerning the experimental antibacterial activities, including antibiotic-like and cell-regulating activities, and therapeutic effects demonstrated in well-conducted, placebo-controlled, randomized clinical trials of these probiotic Lactobacillus strains. What is known about the antimicrobial activities supported by the molecules secreted by such probiotic Lactobacillus strains suggests that they constitute a promising new source for the development of innovative anti-infectious agents that act luminally and intracellularly in the gastrointestinal tract. PMID:24696432

  14. Association of germ-free mice with a simplified human intestinal microbiota results in a shortened intestine.

    PubMed

    Slezak, Kathleen; Krupova, Zuzana; Rabot, Sylvie; Loh, Gunnar; Levenez, Florence; Descamps, Amandine; Lepage, Patricia; Doré, Joël; Bellier, Sylvain; Blaut, Michael

    2014-01-01

    Genetic, nutritional, and gut microbiota-derived factors have been proposed to play a role in the development of the whole intestine that is around 40% longer in PRM/Alf mice compared with other mouse strains. The PRM/Alf genotype explains 60% of this length difference. The remaining 40% are due to a maternal effect that could depend on the gut microbiota transmitted by the mother to their pups. Germ-free PRM/Alf mice and C3H/He mice were associated with a simplified human microbiota (SIHUMI) to study its impact on gut length. The small intestines of the SIHUMI-associated mice were 16.4% (PRM/Alf) and 9.7% (C3H/He) shorter than those of the corresponding germ-free counterparts. Temporal temperature gradient gel electrophoresis and quantitative real-time PCR revealed differences in microbiota composition between both SIHUMI-associated mouse strains. Anaerostipes caccae was one log lower in PRM/Alf mice than in C3H/He mice. Since polyamines and short-chain fatty acids (SCFAs) are important intestinal growth factors, their concentrations were explored. Cecal concentrations of putrescine, spermine, spermidine, and N-acetylspermine were 1.5-fold, 3.7-fold, 2.2-fold, and 1.4-fold higher, respectively, in the SIHUMI-C3H/He mice compared with the SIHUMI-PRM/Alf mice. In addition, cecal acetate, propionate, and butyrate concentrations in SIHUMI-C3H/He mice were 1.4-fold, 1.1-fold, and 2.1-fold higher, respectively, than in SIHUMI-PRM/Alf mice. These results indicate that polyamines and SCFAs did not promote gut lengthening in any of the two mouse strains. This suggests that as yet unknown factors provided by the SIHUMI prevented gut lengthening in the SIHUMI-associated mice compared with the germfree mice.

  15. The influence of the immunostimulation by bacterial cell components derived from altered large intestinal microbiota on probiotic anti-inflammatory benefits.

    PubMed

    Matsumoto, Mitsuharu; Hara, Kurt; Benno, Yoshimi

    2007-04-01

    Using murine macrophage-like J774.1 cells and fecal precipitates prepared from the feces of elderly volunteers whose acute inflammation had been inhibited by LKM512 yogurt consumption, we investigated the likelihood that immunostimulation by altered intestinal bacterial cell components contribute to the anti-inflammatory benefits of this yogurt. Tumor necrosis factor-alpha production due to stimulation by fecal precipitates obtained during LKM512 yogurt consumption tended to be higher than due to stimulation by precipitates obtained from preconsumption (P=0.0827), although acute phase response was suppressed by LKM512 yogurt consumption. We suggest that the anti-inflammatory benefits of LKM512 yogurt on elderly volunteers are independent of direct immunostimulation by the bacterial cell components derived from altered intestinal microbiota.

  16. Role of TLR4/NF-κB in Damage to Intestinal Mucosa Barrier Function and Bacterial Translocation in Rats Exposed to Hypoxia

    PubMed Central

    Luo, Han; Guo, Ping; Zhou, Qiquan

    2012-01-01

    The role of Toll-like receptor 4 (TLR4)/nuclear factor-kappa-B (NF-κB) in intestinal mucosal barrier damage and bacterial translocation under hypoxic exposure is unclear. Here, we investigated their role using an acute hypobaric hypoxia model. Adult Sprague-Dawley rats were divided into control (C), hypoxia (H), hypoxia+NF-κB inhibitor pyrrolidinedithiocarbamic acid (PDTC) (100 mg. kg) (HP), hypoxia+0.5 mg/kg lipopolysaccharide (HPL), and hypoxia+PDTC+LPS (HPL) group. Except control group, other four groups were placed in a hypobaric chamber set at 7000 m. Samples were collected at 72 h after pressure reduction. Damage in ultrastructure of the intestinal tract was examined by transmission electron microscopy and bacterial translocation was detected by cultivation. Kinetic turbidimetric assay was used to measure the serum LPS. ELISA was performed to detect TNF-α and IL-6 serum concentrations. Fluorescent quantitative RT-PCR was used to measure TLR4 mRNA levels was measured using quantitative RT-PCR and protein of NF-κB p65 was measured by western blotting. Different degrees of intestinal mucosa damage were observed in groups H and HL. The damage was significantly alleviated after blockage of the TLR4/NF-κB signaling pathway. PDTC- treatment also reversed hyoxia- and LPS-induced bacterial translocation rate and increased serum levels of LPS, TNF-α, and IL-6. TLR4 mRNA levels and NF-κB p65 expression were consistent with the serum factor results. This study suggested that TLR4 and NF-κB expression increased in rat intestinal tissues after acute hypoxia exposure. PDTC-treatment reversed TLR4 and NF-κB upregulation and alleviated damage to the intestinal tract and bacterial translocation. Thus, the TLR4/NF-κB signaling pathway may be critical to the mechanism underlying hypoxia-induced damage to intestinal barrier function and bacterial translocation. PMID:23082119

  17. Generating human intestinal tissues from pluripotent stem cells to study development and disease

    PubMed Central

    Sinagoga, Katie L; Wells, James M

    2015-01-01

    As one of the largest and most functionally complex organs of the human body, the intestines are primarily responsible for the breakdown and uptake of macromolecules from the lumen and the subsequent excretion of waste from the body. However, the intestine is also an endocrine organ, regulating digestion, metabolism, and feeding behavior. Intricate neuronal, lymphatic, immune, and vascular systems are integrated into the intestine and are required for its digestive and endocrine functions. In addition, the gut houses an extensive population of microbes that play roles in digestion, global metabolism, barrier function, and host–parasite interactions. With such an extensive array of cell types working and performing in one essential organ, derivation of functional intestinal tissues from human pluripotent stem cells (PSCs) represents a significant challenge. Here we will discuss the intricate developmental processes and cell types that are required for assembly of this highly complex organ and how embryonic processes, particularly morphogenesis, have been harnessed to direct differentiation of PSCs into 3-dimensional human intestinal organoids (HIOs) in vitro. We will further describe current uses of HIOs in development and disease research and how additional tissue complexity might be engineered into HIOs for better functionality and disease modeling. PMID:25792515

  18. Generating human intestinal tissues from pluripotent stem cells to study development and disease.

    PubMed

    Sinagoga, Katie L; Wells, James M

    2015-05-05

    As one of the largest and most functionally complex organs of the human body, the intestines are primarily responsible for the breakdown and uptake of macromolecules from the lumen and the subsequent excretion of waste from the body. However, the intestine is also an endocrine organ, regulating digestion, metabolism, and feeding behavior. Intricate neuronal, lymphatic, immune, and vascular systems are integrated into the intestine and are required for its digestive and endocrine functions. In addition, the gut houses an extensive population of microbes that play roles in digestion, global metabolism, barrier function, and host-parasite interactions. With such an extensive array of cell types working and performing in one essential organ, derivation of functional intestinal tissues from human pluripotent stem cells (PSCs) represents a significant challenge. Here we will discuss the intricate developmental processes and cell types that are required for assembly of this highly complex organ and how embryonic processes, particularly morphogenesis, have been harnessed to direct differentiation of PSCs into 3-dimensional human intestinal organoids (HIOs) in vitro. We will further describe current uses of HIOs in development and disease research and how additional tissue complexity might be engineered into HIOs for better functionality and disease modeling.

  19. Small Intestinal Bacterial Overgrowth in Patients with Irritable Bowel Syndrome: Clinical Characteristics, Psychological Factors, and Peripheral Cytokines

    PubMed Central

    Chu, Hua; Fox, Mark; Zheng, Xia; Deng, Yanyong; Long, Yanqin; Huang, Zhihui; Du, Lijun; Xu, Fei; Dai, Ning

    2016-01-01

    Small intestinal bacterial overgrowth (SIBO) has been implicated in the pathogenesis of irritable bowel syndrome (IBS). Psychosocial factors and low-grade colonic mucosal immune activation have been suggested to play important roles in the pathophysiology of IBS. In total, 94 patients with IBS and 13 healthy volunteers underwent a 10 g lactulose hydrogen breath test (HBT) with concurrent 99mTc scintigraphy. All participants also completed a face-to-face questionnaire survey, including the Hospital Anxiety and Depression Scale, Life Event Stress (LES), and general information. Serum tumour necrosis factor-α, interleukin- (IL-) 6, IL-8, and IL-10 levels were measured. The 89 enrolled patients with IBS and 13 healthy controls had no differences in baseline characteristics. The prevalence of SIBO in patients with IBS was higher than that in healthy controls (39% versus 8%, resp.; p = 0.026). Patients with IBS had higher anxiety, depression, and LES scores, but anxiety, depression, and LES scores were similar between the SIBO-positive and SIBO-negative groups. Psychological disorders were not associated with SIBO in patients with IBS. The serum IL-10 level was significantly lower in SIBO-positive than SIBO-negative patients with IBS. PMID:27379166

  20. Identification of intestinal bacterial flora in Rhipicephalus microplus ticks by conventional methods and PCR-DGGE analysis.

    PubMed

    Xu, Xing-Li; Cheng, Tian-Yin; Yang, Hu; Yan, Fen

    2015-06-01

    In this study, we have analyzed the intestinal microbial flora associated with Rhipicephalus microplus ticks using both culture-dependent and independent methods based on PCR and denaturing gradient gel electrophoresis (PCR-DGGE). The R. microplus ticks were collected from cattle and goats in Jiangxi, Hunan and Guizhou Provinces of China. Three distinct strains of bacteria were isolated using culture-dependent methods: Staphylococcus simulans, Bacillus subtilis and Bacillus flexus strain. Nineteen distinct DGGE bands were found using PCR-DGGE analysis, and their search for identity shows that they belonged to Rickettsiaceae, Xanthomonadaceae, Coxiella sp., Ehrlichia sp., Pseudomonas sp., Ehrlichia sp., Orphnebius sp., Rickettsia peacockii, Bacillus flexus. Rickettsia peacockii and Coxiella genus were the dominant strain of the R. microplus ticks from cattle, Pseudomonas sp. and B. flexus strain were the most common species in all tick samples from goats. Ehrlichia canis were detected only in R. microplus ticks from Yongshun area in Hunan Province. The results indicate that the intestinal microbial diversity of R. microplus ticks was influenced by tick hosts and local differences in the sampling location and these two aspects may affect transmission of pathogen to humans and animals.

  1. Brucella invasion of human intestinal epithelial cells elicits a weak proinflammatory response but a significant CCL20 secretion.

    PubMed

    Ferrero, Mariana C; Fossati, Carlos A; Rumbo, Martín; Baldi, Pablo C

    2012-10-01

    In spite of the frequent acquisition of Brucella infection by the oral route in humans, the interaction of the bacterium with cells of the intestinal mucosa has been poorly studied. Here, we show that different Brucella species can invade human colonic epithelial cell lines (Caco-2 and HT-29), in which only smooth species can replicate efficiently. Infection with smooth strains did not produce a significant cytotoxicity, while the rough strain RB51 was more cytotoxic. Infection of Caco-2 cells or HT-29 cells with either smooth or rough strains of Brucella did not result in an increased secretion of TNF-α, IL-1β, MCP-1, IL-10 or TGF-β as compared with uninfected controls, whereas all the infections induced the secretion of IL-8 and CCL20 by both cell types. The MCP-1 response to flagellin from Salmonella typhimurium was similar in Brucella-infected or uninfected cells, ruling out a bacterial inhibitory mechanism as a reason for the weak proinflammatory response. Infection did not modify ICAM-1 expression levels in Caco-2 cells, but increased them in HT-29 cells. These results suggest that Brucella induces only a weak proinflammatory response in gut epithelial cells, but produces a significant CCL20 secretion. The latter may be important for bacterial dissemination given the known ability of Brucella to survive in dendritic cells.

  2. From bacterial to human dihydrouridine synthase: automated structure determination

    PubMed Central

    Whelan, Fiona; Jenkins, Huw T.; Griffiths, Samuel C.; Byrne, Robert T.; Dodson, Eleanor J.; Antson, Alfred A.

    2015-01-01

    The reduction of uridine to dihydrouridine at specific positions in tRNA is catalysed by dihydrouridine synthase (Dus) enzymes. Increased expression of human dihydrouridine synthase 2 (hDus2) has been linked to pulmonary carcinogenesis, while its knockdown decreased cancer cell line viability, suggesting that it may serve as a valuable target for therapeutic intervention. Here, the X-ray crystal structure of a construct of hDus2 encompassing the catalytic and tRNA-recognition domains (residues 1–340) determined at 1.9 Å resolution is presented. It is shown that the structure can be determined automatically by phenix.mr_rosetta starting from a bacterial Dus enzyme with only 18% sequence identity and a significantly divergent structure. The overall fold of the human Dus2 is similar to that of bacterial enzymes, but has a larger recognition domain and a unique three-stranded antiparallel β-sheet insertion into the catalytic domain that packs next to the recognition domain, contributing to domain–domain interactions. The structure may inform the development of novel therapeutic approaches in the fight against lung cancer. PMID:26143927

  3. A review of drug solubility in human intestinal fluids: implications for the prediction of oral absorption.

    PubMed

    Augustijns, Patrick; Wuyts, Benjamin; Hens, Bart; Annaert, Pieter; Butler, James; Brouwers, Joachim

    2014-06-16

    The purpose of this paper is to collate all recently published solubility data of orally administered drugs in human intestinal fluids (HIF) that were aspirated from the upper small intestine (duodenum and jejunum). The data set comprises in total 102 solubility values in fasted state HIF and 37 solubility values in fed state HIF, covering 59 different drugs. Despite differences in the protocol for HIF sampling and subsequent handling, this summary of HIF solubilities provides a critical reference data set to judge the value of simulated media for intestinal solubility estimation. In this regard, the review includes correlations between the reported solubilizing capacity of HIF and fasted or fed state simulated intestinal fluid (FaSSIF/FeSSIF). Correlating with HIF solubilities enables the optimal use of solubility measurements in simulated biorelevant media to obtain accurate estimates of intestinal solubility during drug development. Considering the fraction of poorly soluble new molecular entities in contemporary drug discovery, adequate prediction of intestinal solubility is critical for efficient lead optimization, early candidate profiling, and further development.

  4. Heparan sulfate and syndecan-1 are essential in maintaining murine and human intestinal epithelial barrier function

    PubMed Central

    Bode, Lars; Salvestrini, Camilla; Park, Pyong Woo; Li, Jin-Ping; Esko, Jeffrey D.; Yamaguchi, Yu; Murch, Simon; Freeze, Hudson H.

    2007-01-01

    Patients with protein-losing enteropathy (PLE) fail to maintain intestinal epithelial barrier function and develop an excessive and potentially fatal efflux of plasma proteins. PLE occurs in ostensibly unrelated diseases, but emerging commonalities in clinical observations recently led us to identify key players in PLE pathogenesis. These include elevated IFN-γ, TNF-α, venous hypertension, and the specific loss of heparan sulfate proteoglycans from the basolateral surface of intestinal epithelial cells during PLE episodes. Here we show that heparan sulfate and syndecan-1, the predominant intestinal epithelial heparan sulfate proteoglycan, are essential in maintaining intestinal epithelial barrier function. Heparan sulfate– or syndecan-1–deficient mice and mice with intestinal-specific loss of heparan sulfate had increased basal protein leakage and were far more susceptible to protein loss induced by combinations of IFN-γ, TNF-α, and increased venous pressure. Similarly, knockdown of syndecan-1 in human epithelial cells resulted in increased basal and cytokine-induced protein leakage. Clinical application of heparin has been known to alleviate PLE in some patients but its unknown mechanism and severe side effects due to its anticoagulant activity limit its usefulness. We demonstrate here that non-anticoagulant 2,3-de-O-sulfated heparin could prevent intestinal protein leakage in syndecan-deficient mice, suggesting that this may be a safe and effective therapy for PLE patients. PMID:18064305

  5. Insights from human congenital disorders of intestinal lipid metabolism

    PubMed Central

    Levy, Emile

    2015-01-01

    The intestine must challenge the profuse daily flux of dietary fat that serves as a vital source of energy and as an essential component of cell membranes. The fat absorption process takes place in a series of orderly and interrelated steps, including the uptake and translocation of lipolytic products from the brush border membrane to the endoplasmic reticulum, lipid esterification, Apo synthesis, and ultimately the packaging of lipid and Apo components into chylomicrons (CMs). Deciphering inherited disorders of intracellular CM elaboration afforded new insight into the key functions of crucial intracellular proteins, such as Apo B, microsomal TG transfer protein, and Sar1b GTPase, the defects of which lead to hypobetalipoproteinemia, abetalipoproteinemia, and CM retention disease, respectively. These “experiments of nature” are characterized by fat malabsorption, steatorrhea, failure to thrive, low plasma levels of TGs and cholesterol, and deficiency of liposoluble vitamins and essential FAs. After summarizing and discussing the functions and regulation of these proteins for reader’s comprehension, the current review focuses on their specific roles in malabsorptions and dyslipidemia-related intestinal fat hyperabsorption while dissecting the spectrum of clinical manifestations and managements. The influence of newly discovered proteins (proprotein convertase subtilisin/kexin type 9 and angiopoietin-like 3 protein) on fat absorption has also been provided. Finally, it is stressed how the overexpression or polymorphism status of the critical intracellular proteins promotes dyslipidemia and cardiometabolic disorders. PMID:25387865

  6. Intestinal Microbiota Distinguish Gout Patients from Healthy Humans

    PubMed Central

    Guo, Zhuang; Zhang, Jiachao; Wang, Zhanli; Ang, Kay Ying; Huang, Shi; Hou, Qiangchuan; Su, Xiaoquan; Qiao, Jianmin; Zheng, Yi; Wang, Lifeng; Koh, Eileen; Danliang, Ho; Xu, Jian; Lee, Yuan Kun; Zhang, Heping

    2016-01-01

    Current blood-based approach for gout diagnosis can be of low sensitivity and hysteretic. Here via a 68-member cohort of 33 healthy and 35 diseased individuals, we reported that the intestinal microbiota of gout patients are highly distinct from healthy individuals in both organismal and functional structures. In gout, Bacteroides caccae and Bacteroides xylanisolvens are enriched yet Faecalibacterium prausnitzii and Bifidobacterium pseudocatenulatum depleted. The established reference microbial gene catalogue for gout revealed disorder in purine degradation and butyric acid biosynthesis in gout patients. In an additional 15-member validation-group, a diagnosis model via 17 gout-associated bacteria reached 88.9% accuracy, higher than the blood-uric-acid based approach. Intestinal microbiota of gout are more similar to those of type-2 diabetes than to liver cirrhosis, whereas depletion of Faecalibacterium prausnitzii and reduced butyrate biosynthesis are shared in each of the metabolic syndromes. Thus the Microbial Index of Gout was proposed as a novel, sensitive and non-invasive strategy for diagnosing gout via fecal microbiota. PMID:26852926

  7. Intestinal Microbiota Distinguish Gout Patients from Healthy Humans.

    PubMed

    Guo, Zhuang; Zhang, Jiachao; Wang, Zhanli; Ang, Kay Ying; Huang, Shi; Hou, Qiangchuan; Su, Xiaoquan; Qiao, Jianmin; Zheng, Yi; Wang, Lifeng; Koh, Eileen; Danliang, Ho; Xu, Jian; Lee, Yuan Kun; Zhang, Heping

    2016-02-08

    Current blood-based approach for gout diagnosis can be of low sensitivity and hysteretic. Here via a 68-member cohort of 33 healthy and 35 diseased individuals, we reported that the intestinal microbiota of gout patients are highly distinct from healthy individuals in both organismal and functional structures. In gout, Bacteroides caccae and Bacteroides xylanisolvens are enriched yet Faecalibacterium prausnitzii and Bifidobacterium pseudocatenulatum depleted. The established reference microbial gene catalogue for gout revealed disorder in purine degradation and butyric acid biosynthesis in gout patients. In an additional 15-member validation-group, a diagnosis model via 17 gout-associated bacteria reached 88.9% accuracy, higher than the blood-uric-acid based approach. Intestinal microbiota of gout are more similar to those of type-2 diabetes than to liver cirrhosis, whereas depletion of Faecalibacterium prausnitzii and reduced butyrate biosynthesis are shared in each of the metabolic syndromes. Thus the Microbial Index of Gout was proposed as a novel, sensitive and non-invasive strategy for diagnosing gout via fecal microbiota.

  8. Chemical form of selenium affects its uptake, transport and glutathione peroxidase activity in the human intestinal Caco-2 cell model

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Determining the effect of selenium (Se) chemical form on uptake and transport in human intestinal cells is critical to assess Se bioavailability. In the present study, we measured the uptake and transport of various Se compounds in the human intestinal Caco-2 cell model. We found that two sources...

  9. Commensal gut bacteria modulate phosphorylation-dependent PPARγ transcriptional activity in human intestinal epithelial cells

    PubMed Central

    Nepelska, Malgorzata; de Wouters, Tomas; Jacouton, Elsa; Béguet-Crespel, Fabienne; Lapaque, Nicolas; Doré, Joël; Arulampalam, Velmurugesan; Blottière, Hervé M.

    2017-01-01

    In healthy subjects, the intestinal microbiota interacts with the host’s epithelium, regulating gene expression to the benefit of both, host and microbiota. The underlying mechanisms remain poorly understood, however. Although many gut bacteria are not yet cultured, constantly growing culture collections have been established. We selected 57 representative commensal bacterial strains to study bacteria-host interactions, focusing on PPARγ, a key nuclear receptor in colonocytes linking metabolism and inflammation to the microbiota. Conditioned media (CM) were harvested from anaerobic cultures and assessed for their ability to modulate PPARγ using a reporter cell line. Activation of PPARγ transcriptional activity was linked to the presence of butyrate and propionate, two of the main metabolites of intestinal bacteria. Interestingly, some stimulatory CMs were devoid of these metabolites. A Prevotella and an Atopobium strain were chosen for further study, and shown to up-regulate two PPARγ-target genes, ANGPTL4 and ADRP. The molecular mechanisms of these activations involved the phosphorylation of PPARγ through ERK1/2. The responsible metabolites were shown to be heat sensitive but markedly diverged in size, emphasizing the diversity of bioactive compounds found in the intestine. Here we describe different mechanisms by which single intestinal bacteria can directly impact their host’s health through transcriptional regulation. PMID:28266623

  10. Commensal gut bacteria modulate phosphorylation-dependent PPARγ transcriptional activity in human intestinal epithelial cells.

    PubMed

    Nepelska, Malgorzata; de Wouters, Tomas; Jacouton, Elsa; Béguet-Crespel, Fabienne; Lapaque, Nicolas; Doré, Joël; Arulampalam, Velmurugesan; Blottière, Hervé M

    2017-03-07

    In healthy subjects, the intestinal microbiota interacts with the host's epithelium, regulating gene expression to the benefit of both, host and microbiota. The underlying mechanisms remain poorly understood, however. Although many gut bacteria are not yet cultured, constantly growing culture collections have been established. We selected 57 representative commensal bacterial strains to study bacteria-host interactions, focusing on PPARγ, a key nuclear receptor in colonocytes linking metabolism and inflammation to the microbiota. Conditioned media (CM) were harvested from anaerobic cultures and assessed for their ability to modulate PPARγ using a reporter cell line. Activation of PPARγ transcriptional activity was linked to the presence of butyrate and propionate, two of the main metabolites of intestinal bacteria. Interestingly, some stimulatory CMs were devoid of these metabolites. A Prevotella and an Atopobium strain were chosen for further study, and shown to up-regulate two PPARγ-target genes, ANGPTL4 and ADRP. The molecular mechanisms of these activations involved the phosphorylation of PPARγ through ERK1/2. The responsible metabolites were shown to be heat sensitive but markedly diverged in size, emphasizing the diversity of bioactive compounds found in the intestine. Here we describe different mechanisms by which single intestinal bacteria can directly impact their host's health through transcriptional regulation.

  11. Expression and membrane localization of MCT isoforms along the length of the human intestine.

    PubMed

    Gill, Ravinder K; Saksena, Seema; Alrefai, Waddah A; Sarwar, Zaheer; Goldstein, Jay L; Carroll, Robert E; Ramaswamy, Krishnamurthy; Dudeja, Pradeep K

    2005-10-01

    Recent studies from our laboratory and others have demonstrated the involvement of monocarboxylate transporter (MCT)1 in the luminal uptake of short-chain fatty acids (SCFAs) in the human intestine. Functional studies from our laboratory previously demonstrated kinetically distinct SCFA transporters on the apical and basolateral membranes of human colonocytes. Although apical SCFA uptake is mediated by the MCT1 isoform, the molecular identity of the basolateral membrane SCFA transporter(s) and whether this transporter is encoded by another MCT isoform is not known. The present studies were designed to assess the expression and membrane localization of different MCT isoforms in human small intestine and colon. Immunoblotting was performed with the purified apical and basolateral membranes from human intestinal mucosa obtained from organ donor intestine. Immunohistochemistry studies were done on paraffin-embedded sections of human colonic biopsy samples. Immunoblotting studies detected a protein band of approximately 39 kDa for MCT1, predominantly in the apical membranes. The relative abundance of MCT1 mRNA and protein increased along the length of the human intestine. MCT4 (54 kDa) and MCT5 (54 kDa) isoforms showed basolateral localization and were highly expressed in the distal colon. Immunohistochemical studies confirmed that human MCT1 antibody labeling was confined to the apical membranes, whereas MCT5 antibody staining was restricted to the basolateral membranes of the colonocytes. We speculate that distinct MCT isoforms may be involved in SCFA transport across the apical or basolateral membranes in polarized colonic epithelial cells.

  12. Effects of the Probiotic Enterococcus faecium and Pathogenic Escherichia coli Strains in a Pig and Human Epithelial Intestinal Cell Model

    PubMed Central

    Lodemann, Ulrike; Strahlendorf, Julia; Schierack, Peter; Klingspor, Shanti; Aschenbach, Jörg R.

    2015-01-01

    The aim of this study has been to elucidate the effect of the probiotic Enterococcus faecium NCIMB 10415 on epithelial integrity in intestinal epithelial cells and whether pre- and coincubation with this strain can reproducibly prevent damage induced by enterotoxigenic (ETEC) and enteropathogenic Escherichia coli (EPEC). Porcine (IPEC-J2) and human (Caco-2) intestinal epithelial cells were incubated with bacterial strains and epithelial integrity was assessed by measuring transepithelial electrical resistance (TEER) and mannitol flux rates. E. faecium alone increased TEER of Caco-2 cells without affecting mannitol fluxes whereas the E. coli strains decreased TEER and concomitantly increased mannitol flux rates in both cell lines. Preincubation with E. faecium had no effect on the TEER decrease induced by E. coli in preliminary experiments. However, in a second set of experiments using a slightly different protocol, E. faecium ameliorated the TEER decrease induced by ETEC at 4 h in IPEC-J2 and at 2, 4, and 6 h in Caco-2 cells. We conclude that E. faecium positively affected epithelial integrity in monoinfected Caco-2 cells and could ameliorate the damage on TEER induced by an ETEC strain. Reproducibility of the results is, however, limited when experiments are performed with living bacteria over longer periods. PMID:25883829

  13. Identification of the transcriptional response of human intestinal mucosa to Lactobacillus plantarum WCFS1 in vivo

    PubMed Central

    Troost, Freddy J; van Baarlen, Peter; Lindsey, Patrick; Kodde, Andrea; de Vos, Willem M; Kleerebezem, Michiel; Brummer, Robert-Jan M

    2008-01-01

    Background There is limited knowledge on the extent and dynamics of the mucosal response to commensal and probiotic species in the human intestinal lumen. This study aimed to identify the acute, time-dependent responses of intestinal mucosa to commensal Lactobacillus plantarum WCFS1 in vivo in two placebo-controlled human intervention studies in healthy volunteers. Transcriptional changes in duodenal mucosa upon continuous intraduodenal infusion of L. plantarum WCFS1 for one- and six h, respectively, were studied using oro- and nasogastric intubations with dedicated orogastric catheters and tissue sampling by standard flexible gastroduodenoscopy. Results One- and six-h exposure of small intestinal mucosa to L. plantarum WCFS1 induced differential expression of 669 and 424 gene reporters, respectively. While short-term exposure to L. plantarum WCFS1 inhibited fatty acid metabolism and cell cycle progression, cells switched to a more proliferative phase after prolonged exposure with an overall expression profile characterized by upregulation of genes involved in lipid metabolism, cellular growth and development. Cell death and immune responses were triggered, but cell death-executing genes or inflammatory signals were not expressed. Proteome analysis showed differential expression of several proteins. Only the microsomal protein 'microsomal triglyceride transfer protein' was regulated on both the transcriptional and the protein level in all subjects. Conclusion Overall, this study showed that intestinal exposure to L. plantarum WCFS1 induced consistent, time-dependent transcriptional responses in healthy intestinal mucosa. This extensive exploration of the human response to L. plantarum WCFS1 could eventually provide molecular support for specific or probiotic activity of this strain or species, and exemplifies the strength of the applied technology to identify the potential bio-activity of microbes in the human intestine. PMID:18681965

  14. hPSC-derived lung and intestinal organoids as models of human fetal tissue.

    PubMed

    Aurora, Megan; Spence, Jason R

    2016-12-15

    In vitro human pluripotent stem cell (hPSC) derived tissues are excellent models to study certain aspects of normal human development. Current research in the field of hPSC derived tissues reveals these models to be inherently fetal-like on both a morphological and gene expression level. In this review we briefly discuss current methods for differentiating lung and intestinal tissue from hPSCs into individual 3-dimensional units called organoids. We discuss how these methods mirror what is known about in vivo signaling pathways of the developing embryo. Additionally, we will review how the inherent immaturity of these models lends them to be particularly valuable in the study of immature human tissues in the clinical setting of premature birth. Human lung organoids (HLOs) and human intestinal organoids (HIOs) not only model normal development, but can also be utilized to study several important diseases of prematurity such as respiratory distress syndrome (RDS), bronchopulmonary dysplasia (BPD), and necrotizing enterocolitis (NEC).

  15. Modulation of Intestinal Barrier and Bacterial Endotoxin Production Contributes to the Beneficial Effect of Nicotinic Acid on Alcohol-Induced Endotoxemia and Hepatic Inflammation in Rats

    PubMed Central

    Zhong, Wei; Li, Qiong; Zhang, Wenliang; Sun, Qian; Sun, Xinguo; Zhou, Zhanxiang

    2015-01-01

    Alcohol consumption causes nicotinic acid deficiency. The present study was undertaken to determine whether dietary nicotinic acid supplementation provides beneficial effects on alcohol-induced endotoxin signaling and the possible mechanisms at the gut-liver axis. Male Sprague-Dawley rats were pair-fed the Lieber-DeCarli liquid diets containing ethanol or isocaloric maltose dextrin for eight weeks, with or without dietary supplementation with 750 mg/liter nicotinic acid. Chronic alcohol feeding elevated the plasma endotoxin level and activated hepatic endotoxin signaling cascade, which were attenuated by nicotinic acid supplementation. Alcohol consumption remarkably decreased the mRNA levels of claudin-1, claudin-5, and ZO-1 in the distal intestine, whereas nicotinic acid significantly up-regulated these genes. The concentrations of endotoxin, ethanol, and acetaldehyde in the intestinal contents were increased by alcohol exposure, and niacin supplementation reduced the intestinal endotoxin and acetaldehyde levels. Nicotinic acid supplementation upregulated the intestinal genes involved in aldehyde detoxification via transcriptional regulation. These results demonstrate that modulation of the intestinal barrier function and bacterial endotoxin production accounts for the inhibitory effects of nicotinic acid on alcohol-induced endotoxemia and hepatic inflammation. PMID:26501337

  16. Dietary interactions with the bacterial sensing machinery in the intestine: the plant polyphenol case

    PubMed Central

    Ahmed Nasef, Noha; Mehta, Sunali; Ferguson, Lynnette R.

    2014-01-01

    There are millions of microbes that live in the human gut. These are important in digestion as well as defense. The host immune system needs to be able to distinguish between the harmless bacteria and pathogens. The initial interaction between bacteria and the host happen through the pattern recognition receptors (PRRs). As these receptors are in direct contact with the external environment, this makes them important candidates for regulation by dietary components and therefore potential targets for therapy. In this review, we introduce some of the main PRRs including a cellular process known as autophagy, and how they function. Additionally we review dietary phytochemicals from plants which are believed to be beneficial for humans. The purpose of this review was to give a better understanding of how these components work in order to create better awareness on how they could be explored in the future. PMID:24772116

  17. Human Intestinal Raf Kinase Inhibitor Protein (RKIP) Catalyzes Prasugrel as a Bioactivation Hydrolase.

    PubMed

    Kazui, Miho; Ogura, Yuji; Hagihara, Katsunobu; Kubota, Kazuishi; Kurihara, Atsushi

    2016-01-01

    Prasugrel is a thienopyridine antiplatelet prodrug that undergoes rapid hydrolysis in vivo to a thiolactone metabolite by human carboxylesterase-2 (hCE2) during gastrointestinal absorption. The thiolactone metabolite is further converted to a pharmacologically active metabolite by cytochrome P450 isoforms. The aim of the current study was to elucidate hydrolases other than hCE2 involved in the bioactivation step of prasugrel in human intestine. Using size-exclusion column chromatography of a human small intestinal S9 fraction, another peak besides the hCE2 peak was observed to have prasugrel hydrolyzing activity, and this protein was found to have a molecular weight of about 20 kDa. This prasugrel hydrolyzing protein was successfully purified from a monkey small intestinal cytosolic fraction by successive four-step column chromatography and identified as Raf-1 kinase inhibitor protein (RKIP) by liquid chromatography-tandem mass spectrometry. Second, we evaluated the enzymatic kinetic parameters for prasugrel hydrolysis using recombinant human RKIP and hCE2 and estimated the contributions of these two hydrolyzing enzymes to the prasugrel hydrolysis reaction in human intestine, which were approximately 40% for hRKIP and 60% for hCE2. Moreover, prasugrel hydrolysis was inhibited by anti-hRKIP antibody and carboxylesterase-specific chemical inhibitor (bis p-nitrophenyl phosphate) by 30% and 60%, respectively. In conclusion, another protein capable of hydrolyzing prasugrel to its thiolactone metabolite was identified as RKIP, and this protein may play a significant role with hCE2 in prasugrel bioactivation in human intestine. RKIP is known to have diverse functions in many intracellular signaling cascades, but this is the first report describing RKIP as a hydrolase involved in drug metabolism.

  18. Attaching and effacing activities of rabbit and human enteropathogenic Escherichia coli in pig and rabbit intestines.

    PubMed Central

    Moon, H W; Whipp, S C; Argenzio, R A; Levine, M M; Giannella, R A

    1983-01-01

    Three strains of enteropathogenic Escherichia coli (EPEC), originally isolated from humans and previously shown to cause diarrhea in human volunteers by unknown mechanisms, and one rabbit EPEC strain were shown to attach intimately to and efface microvilli and cytoplasm from intestinal epithelial cells in both the pig and rabbit intestine. The attaching and effacing activities of these EPEC were demonstrable by light microscopic examination of routine histological sections and by transmission electron microscopy. It was suggested that intact colostrum-deprived newborn pigs and ligated intestinal loops in pigs and rabbits may be useful systems to detect EPEC that have attaching and effacing activities and for studying the pathogenesis of such infections. The lesions (attachment and effacement) produced by EPEC in these systems were multifocal, with considerable animal-to-animal variation in response to the same strain of EPEC. The EPEC strains also varied in the frequency and extent of lesion production. For example, three human EPEC strains usually caused extensive lesions in rabbit intestinal loops, whereas two other human EPEC strains usually did not produce lesions in this system. Images PMID:6350186

  19. Consensus hologram QSAR modeling for the prediction of human intestinal absorption.

    PubMed

    Moda, Tiago L; Andricopulo, Adriano D

    2012-04-15

    Consistent in silico models for ADME properties are useful tools in early drug discovery. Here, we report the hologram QSAR modeling of human intestinal absorption using a dataset of 638 compounds with experimental data associated. The final validated models are consistent and robust for the consensus prediction of this important pharmacokinetic property and are suitable for virtual screening applications.

  20. Poly-beta-hydroxybutyrate (PHB) increases growth performance and intestinal bacterial range-weighted richness in juvenile European sea bass, Dicentrarchus labrax.

    PubMed

    De Schryver, Peter; Sinha, Amit Kumar; Kunwar, Prabesh Singh; Baruah, Kartik; Verstraete, Willy; Boon, Nico; De Boeck, Gudrun; Bossier, Peter

    2010-05-01

    The bacterial storage polymer poly-beta-hydroxybutyrate (PHB) has the potential to be used as an alternative anti-infective strategy for aquaculture rearing. In this research, the effects of (partially) replacing the feed of European sea bass juveniles with PHB were investigated. During a 6-week trial period, the PHB showed the ability to act as an energy source for the fish. This indicated that PHB was degraded and used during gastrointestinal passage. The gut pH decreased from 7.7 to 7.2 suggesting that the presence of PHB in the gut led to the increased production of (short-chain fatty) acids. The diets supplemented with 2% and 5% PHB (w/w) induced a gain of the initial fish weight with a factor 2.4 and 2.7, respectively, relative to a factor 2.2 in the normal feed treatment. Simultaneously, these treatments showed the highest bacterial range-weighted richness in the fish intestine. Based on molecular analysis, higher dietary PHB levels induced larger changes in the bacterial community composition. From our results, it seems that PHB can have a beneficial effect on fish growth performance and that the intestinal bacterial community structure may be closely related to this phenomenon.

  1. Human ecology and behavior and sexually transmitted bacterial infections.

    PubMed Central

    Holmes, K K

    1994-01-01

    The three direct determinants of the rate of spread of sexually transmitted diseases (STDs) are sexual behaviors, the mean duration of infectiousness, and the mean efficiency of sexual transmission of each STD. Underlying ecological and behavioral factors that operate through one or more of these direct determinants lie on a continuum, ranging from those most proximate back to those more remote (in time or mechanism) from the direct determinants. Most remote and least modifiable are the historical stages of economic development that even today conspicuously influence patterns of sexual behavior. Next are the distribution and changing patterns of climate, hygiene, and population density; the global population explosion and stages of the demographic transition; and ongoing changes in human physiology (e.g., menarche at younger age) and culture (e.g., later marriage). More proximate on the continuum are war, migration, and travel; and current policies for economic development and social welfare. Most recent or modifiable are technologic and commercial product development (e.g., oral contraceptives); circumcision, condom, spermicide, and contraception practices; patterns of illicit drug use that influence sexual behaviors; and the accessibility, quality, and use of STD health care. These underlying factors help explain why the curable bacterial STDs are epidemic in developing countries and why the United States is the only industrialized country that has failed to control bacterial STDs during the AIDS era. Images PMID:8146138

  2. Binding of Candida albicans to Human CEACAM1 and CEACAM6 Modulates the Inflammatory Response of Intestinal Epithelial Cells

    PubMed Central

    Müller, Mario M.; Schäfer, Miriam R.; Clauder, Ann-Katrin; Feer, Sabina; Heyl, Kerstin A.; Stock, Magdalena; Klassert, Tilman E.; Zipfel, Peter F.; Singer, Bernhard B.

    2017-01-01

    ABSTRACT Candida albicans colonizes human mucosa, including the gastrointestinal tract, as a commensal. In immunocompromised patients, C. albicans can breach the intestinal epithelial barrier and cause fatal invasive infections. Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1; CD66a), CEACAM5 (CEA), and CEACAM6 (CD66c) are immunomodulatory receptors expressed on human mucosa and are recruited by bacterial and viral pathogens. Here we show for the first time that a fungal pathogen (i.e., C. albicans) also binds directly to the extracellular domain of human CEACAM1, CEACAM3, CEACAM5, and CEACAM6. Binding was specific for human CEACAMs and mediated by the N-terminal IgV-like domain. In enterocytic C2BBe1 cells, C. albicans caused a transient tyrosine phosphorylation of CEACAM1 and induced higher expression of membrane-bound CEACAM1 and soluble CEACAM6. Lack of the CEACAM1 receptor after short hairpin RNA (shRNA) knockdown abolished CXCL8 (interleukin-8) secretion by C2BBe1 cells in response to C. albicans. In CEACAM1-competent cells, the addition of recombinant soluble CEACAM6 reduced the C. albicans-induced CXCL8 secretion. PMID:28292985

  3. Preterm infant gut microbiota affects intestinal epithelial development in a humanized microbiome gnotobiotic mouse model.

    PubMed

    Yu, Yueyue; Lu, Lei; Sun, Jun; Petrof, Elaine O; Claud, Erika C

    2016-09-01

    Development of the infant small intestine is influenced by bacterial colonization. To promote establishment of optimal microbial communities in preterm infants, knowledge of the beneficial functions of the early gut microbiota on intestinal development is needed. The purpose of this study was to investigate the impact of early preterm infant microbiota on host gut development using a gnotobiotic mouse model. Histological assessment of intestinal development was performed. The differentiation of four epithelial cell lineages (enterocytes, goblet cells, Paneth cells, enteroendocrine cells) and tight junction (TJ) formation was examined. Using weight gain as a surrogate marker for health, we found that early microbiota from a preterm infant with normal weight gain (MPI-H) induced increased villus height and crypt depth, increased cell proliferation, increased numbers of goblet cells and Paneth cells, and enhanced TJs compared with the changes induced by early microbiota from a poor weight gain preterm infant (MPI-L). Laser capture microdissection (LCM) plus qRT-PCR further revealed, in MPI-H mice, a higher expression of stem cell marker Lgr5 and Paneth cell markers Lyz1 and Cryptdin5 in crypt populations, along with higher expression of the goblet cell and mature enterocyte marker Muc3 in villus populations. In contrast, MPI-L microbiota failed to induce the aforementioned changes and presented intestinal characteristics comparable to a germ-free host. Our data demonstrate that microbial communities have differential effects on intestinal development. Future studies to identify pioneer settlers in neonatal microbial communities necessary to induce maturation may provide new insights for preterm infant microbial ecosystem therapeutics.

  4. Consumption of a Bifidobacterium bifidum Strain for 4 Weeks Modulates Dominant Intestinal Bacterial Taxa and Fecal Butyrate in Healthy Adults

    PubMed Central

    Gargari, Giorgio; Taverniti, Valentina; Balzaretti, Silvia; Ferrario, Chiara; Gardana, Claudio; Simonetti, Paolo

    2016-01-01

    the intestinal microbiota, resulting in the modulation of short-chain fatty acid concentrations in the gut. The overall changes witnessed in the probiotic intervention indicate a mechanism of microbiota modulation that could have potential effects on human health. PMID:27451450

  5. Human-derived probiotic Lactobacillus reuteri strains differentially reduce intestinal inflammation.

    PubMed

    Liu, Yuying; Fatheree, Nicole Y; Mangalat, Nisha; Rhoads, Jon Marc

    2010-11-01

    Lactobacillus reuteri (L. reuteri) is a probiotic that inhibits the severity of enteric infections and modulates the immune system. Human-derived L. reuteri strains DSM17938, ATCC PTA4659, ATCC PTA 5289, and ATCC PTA 6475 have demonstrated strain-specific immunomodulation in cultured monocytoid cells, but information about how these strains affect inflammation in intestinal epithelium is limited. We determined the effects of the four different L. reuteri strains on lipopolysaccharide (LPS)-induced inflammation in small intestinal epithelial cells and in the ileum of newborn rats. IPEC-J2 cells (derived from the jejunal epithelium of a neonatal piglet) and IEC-6 cells (derived from the rat crypt) were treated with L. reuteri. Newborn rat pups were gavaged cow milk formula supplemented with L. reuteri strains in the presence or absence of LPS. Protein and mRNA levels of cytokines and histological changes were measured. We demonstrate that even though one L. reuteri strain (DSM 17938) did not inhibit LPS-induced IL-8 production in cultured intestinal cells, all strains significantly reduced intestinal mucosal levels of KC/GRO (∼IL-8) and IFN-γ when newborn rat pups were fed formula containing LPS ± L. reuteri. Intestinal histological damage produced by LPS plus cow milk formula was also significantly reduced by all four strains. Cow milk formula feeding (without LPS) produced mild gut inflammation, evidenced by elevated mucosal IFN-γ and IL-13 levels, a process that could be suppressed by strain 17938. Other cytokines and chemokines were variably affected by the different strains, and there was no toxic effect of L. reuteri on intestinal cells or mucosa. In conclusion, L. reuteri strains differentially modulate LPS-induced inflammation. Probiotic interactions with both epithelial and nonepithelial cells in vivo must be instrumental in modulating intrinsic anti-inflammatory effects in the intestine. We suggest that the terms anti- and proinflammatory be used only

  6. CFTR is a tumor suppressor gene in murine and human intestinal cancer

    PubMed Central

    Than, BLN; Linnekamp, JF; Starr, TK; Largaespada, DA; Rod, A; Zhang, Y; Bruner, V; Abrahante, J; Schumann, A; Luczak, T; Niemczyk, A; O’Sullivan, MG; Medema, JP; Fijneman, RJA; Meijer, GA; Van den Broek, E; Hodges, CA; Scott, PM; Vermeulen, L; Cormier, RT

    2016-01-01

    CFTR, the cystic fibrosis (CF) gene, encodes for the CFTR protein that plays an essential role in anion regulation and tissue homeostasis of various epithelia. In the gastrointestinal (GI) tract CFTR promotes chloride and bicarbonate secretion, playing an essential role in ion and acid–base homeostasis. Cftr has been identified as a candidate driver gene for colorectal cancer (CRC) in several Sleeping Beauty DNA transposon-based forward genetic screens in mice. Further, recent epidemiological and clinical studies indicate that CF patients are at high risk for developing tumors in the colon. To investigate the effects of CFTR dysregulation on GI cancer, we generated ApcMin mice that carried an intestinal-specific knockout of Cftr. Our results indicate that Cftr is a tumor suppressor gene in the intestinal tract as Cftr mutant mice developed significantly more tumors in the colon and the entire small intestine. In Apc+/+ mice aged to ~ 1 year, Cftr deficiency alone caused the development of intestinal tumors in >60% of mice. Colon organoid formation was significantly increased in organoids created from Cftr mutant mice compared with wild-type controls, suggesting a potential role of Cftr in regulating the intestinal stem cell compartment. Microarray data from the Cftr-deficient colon and the small intestine identified dysregulated genes that belong to groups of immune response, ion channel, intestinal stem cell and other growth signaling regulators. These associated clusters of genes were confirmed by pathway analysis using Ingenuity Pathway Analysis and gene set enrichment analysis (GSEA). We also conducted RNA Seq analysis of tumors from Apc+/+ Cftr knockout mice and identified sets of genes dysregulated in tumors including altered Wnt β-catenin target genes. Finally we analyzed expression of CFTR in early stage human CRC patients stratified by risk of recurrence and found that loss of expression of CFTR was significantly associated with poor disease

  7. CFTR is a tumor suppressor gene in murine and human intestinal cancer.

    PubMed

    Than, B L N; Linnekamp, J F; Starr, T K; Largaespada, D A; Rod, A; Zhang, Y; Bruner, V; Abrahante, J; Schumann, A; Luczak, T; Niemczyk, A; O'Sullivan, M G; Medema, J P; Fijneman, R J A; Meijer, G A; Van den Broek, E; Hodges, C A; Scott, P M; Vermeulen, L; Cormier, R T

    2016-08-11

    CFTR, the cystic fibrosis (CF) gene, encodes for the CFTR protein that plays an essential role in anion regulation and tissue homeostasis of various epithelia. In the gastrointestinal (GI) tract CFTR promotes chloride and bicarbonate secretion, playing an essential role in ion and acid-base homeostasis. Cftr has been identified as a candidate driver gene for colorectal cancer (CRC) in several Sleeping Beauty DNA transposon-based forward genetic screens in mice. Further, recent epidemiological and clinical studies indicate that CF patients are at high risk for developing tumors in the colon. To investigate the effects of CFTR dysregulation on GI cancer, we generated Apc(Min) mice that carried an intestinal-specific knockout of Cftr. Our results indicate that Cftr is a tumor suppressor gene in the intestinal tract as Cftr mutant mice developed significantly more tumors in the colon and the entire small intestine. In Apc(+/+) mice aged to ~1 year, Cftr deficiency alone caused the development of intestinal tumors in >60% of mice. Colon organoid formation was significantly increased in organoids created from Cftr mutant mice compared with wild-type controls, suggesting a potential role of Cftr in regulating the intestinal stem cell compartment. Microarray data from the Cftr-deficient colon and the small intestine identified dysregulated genes that belong to groups of immune response, ion channel, intestinal stem cell and other growth signaling regulators. These associated clusters of genes were confirmed by pathway analysis using Ingenuity Pathway Analysis and gene set enrichment analysis (GSEA). We also conducted RNA Seq analysis of tumors from Apc(+/+) Cftr knockout mice and identified sets of genes dysregulated in tumors including altered Wnt β-catenin target genes. Finally we analyzed expression of CFTR in early stage human CRC patients stratified by risk of recurrence and found that loss of expression of CFTR was significantly associated with poor disease

  8. Probiotics for Preventing and Treating Small Intestinal Bacterial Overgrowth: A Meta-Analysis and Systematic Review of Current Evidence.

    PubMed

    Zhong, Changqing; Qu, Changmin; Wang, Baoyan; Liang, Shuwen; Zeng, Bolun

    2017-04-01

    The present study conducted a meta-analysis and systematic review of current evidence to assess the efficacy of probiotics in preventing or treating small intestinal bacterial overgrowth (SIBO). Relevant studies from PubMed, Embase, and the Cochrane Central Register of Controlled Trials, until May 2016, were assimilated. The prevention efficacy was assessed by the incidence of SIBO in the probiotic group, and the treatment efficacy by the SIBO decontamination rate, reduction in H2 concentration, and symptom improvement. The relative risk (RR) and weighted mean difference (WMD) were used as effect measures and the random-effects model used for meta-analysis. A total of 14 full-text articles and 8 abstracts were included for the systematic review, and 18 studies were eligible for data synthesis. Patients on probiotic usage showed an insignificant trend toward low SIBO incidence [RR=0.54; 95% confidence intervals (CI), 0.19-1.52; P=0.24]. The pooled SIBO decontamination rate was 62.8% (51.5% to 72.8%). The probiotics group showed a significantly higher SIBO decontamination rate than the nonprobiotic group (RR=1.61; 95% CI, 1.19-2.17; P<0.05). Also, the H2 concentration was significantly reduced among probiotic users (WMD=-36.35 ppm; 95% CI, -44.23 to -28.47 ppm; P<0.05). Although probiotics produced a marked decrease in the abdominal pain scores (WMD=-1.17; 95% CI, -2.30 to -0.04; P<0.05), it did not significantly reduce the daily stool frequency (WMD=-0.09; 95% CI, -0.47 to 0.29). Therefore, the present findings indicated that probiotics supplementation could effectively decontaminate SIBO, decrease H2 concentration, and relieve abdominal pain, but were ineffective in preventing SIBO.

  9. Identification of a human intestinal myeloid cell subset that regulates gut homeostasis.

    PubMed

    Barman, Soumik; Kayama, Hisako; Okuzaki, Daisuke; Ogino, Takayuki; Osawa, Hideki; Matsuno, Hiroshi; Mizushima, Tsunekazu; Mori, Masaki; Nishimura, Junichi; Takeda, Kiyoshi

    2016-11-01

    Inappropriate activation of T helper (Th) cells, such as Th1 and Th17 cells, is implicated in the pathogenesis of chronic inflammatory disorders including ulcerative colitis (UC). CX3CR1(high) macrophages contribute to intestinal homeostasis through various mechanisms in mice. However, whether mononuclear phagocytes with regulatory functions are present in the human colon is not clearly defined. We investigated whether innate myeloid cells that suppress activation of effector T cells exist in the human intestinal mucosa. Among intestinal lamina propria cells, Lin(-) HLA-DR(high) CD14(+) CD163(high) cells were subdivided into CD160(low) and CD160(high) cells. Both subsets produced high levels of IL-10. CD163(high) CD160(high) cells suppressed effector T cell proliferation, whereas CD163(high) CD160(low) cells induced Th17 differentiation. Patients with UC exhibited increased numbers of CD163(high) CD160(low) cells, while showing profoundly decreased numbers of CD163(high) CD160(high) cells. In this context, CD163(high) CD160(high) cells had higher CD80/CD86 expression and lower IL10RB expression, and these cells did not suppress effector T cell proliferation. The CD163(high) CD160(high) subset in normal intestinal mucosa inhibits inappropriate Th1/Th17 responses through suppression of their proliferation, and its number and suppressive activity are impaired in patients with UC. These findings indicate how human innate immune cells might prevent UC development.

  10. Reductive dechlorination of methoxychlor and DDT by human intestinal bacterium Eubacterium limosum under anaerobic conditions.

    PubMed

    Yim, You-Jin; Seo, Jiyoung; Kang, Su-Il; Ahn, Joong-Hoon; Hur, Hor-Gil

    2008-04-01

    Methoxychlor [1,1,1-trichloro-2,2-bis(p-methoxyphenyl)ethane], a substitute for 1,1,1-trichloro-2,2-bis(p-chlorophenyl)ethane (DDT), is a compound of environmental concern because of potential long-term health risks related to its endocrine-disrupting and carcinogenic potency. In order to determine the metabolic fate of methoxychlor and DDT in the human intestinal gut, Eubacterium limosum (ATCC 8486), a strict anaerobe isolated from the human intestine that is capable of O-demethylation toward O-methylated isoflavones, was used as a model intestinal microbial organism. Under anaerobic incubation conditions, E. limosum completely transformed methoxychlor and DDT in 16 days. Based on gas chromatography-mass chromatography analyses, the metabolites produced from methoxychlor and DDT by E. limosum were confirmed to be 1,1-dichloro-2,2-bis(p-methoxyphenyl)ethane (methoxydichlor) and 1,1-dichloro-2,2-bis(p-chlorophenyl)ethane (DDD), respectively. This study suggests that E. limosum in the human intestinal gut might be a participant in the reductive dechlorination of methoxychlor to the more antiandrogenic active methoxydichlor.

  11. Metabolism of Isoflavones Found in the Pueraria thomsonii Flower by Human Intestinal Microbiota.

    PubMed

    Hirayama, Kazuhiro; Matsuzuka, Yuki; Kamiya, Tomoyasu; Ikeguchi, Motoya; Takagaki, Kinya; Itoh, Kikuji

    2011-01-01

    Isoflavones contained in the root and flower of Kudzu (Pueraria lobata and related species) are suggested to be the critical component for its effects. Although metabolism of soy isoflavones has been well studied, the composition of isoflavones found in Kudzu is completely different from that of soy isoflavones. In the present study, we investigated whether isoflavones found in the flower of Pueraria thomsonii, a species of Kudzu, were metabolized by human fecal microbiota and murine small intestinal enzymes. Among 5 glycosidic isoflavones of the Pueraria thomsonii flower, tectorigenin 7-O-xylosylglucoside, tectoridin, genistin and glycitin were completely hydrolyzed by a homogenate of germfree mouse small intestine without contribution of bacteria. Released aglycones were not further metabolized, except that up to half of glycitein disappeared. Mouse small intestinal enzymes did not metabolize 6-hydroxygenistein 6,7-di-O-glucoside. Isoflavone aglycones as well as 6-hydroxygenistein 6,7-di-O-glucoside were highly metabolized by most of the human fecal suspensions. Metabolites were not detected with the present analytical methods in most cases. Although further investigations of the pharmacokinetics of Pueraria thomsonii flower isoflavones are needed, the results of the present study indicate active metabolism of Pueraria thomsonii flower isoflavones in the human intestine.

  12. Detection of Intracellular Bacterial Communities in Human Urinary Tract Infection

    PubMed Central

    Rosen, David A; Hooton, Thomas M; Stamm, Walter E; Humphrey, Peter A; Hultgren, Scott J

    2007-01-01

    Background Urinary tract infections (UTIs) are one of the most common bacterial infections and are predominantly caused by uropathogenic Escherichia coli (UPEC). While UTIs are typically considered extracellular infections, it has been recently demonstrated that UPEC bind to, invade, and replicate within the murine bladder urothelium to form intracellular bacterial communities (IBCs). These IBCs dissociate and bacteria flux out of bladder facet cells, some with filamentous morphology, and ultimately establish quiescent intracellular reservoirs that can seed recurrent infection. This IBC pathogenic cycle has not yet been investigated in humans. In this study we sought to determine whether evidence of an IBC pathway could be found in urine specimens from women with acute UTI. Methods and Findings We collected midstream, clean-catch urine specimens from 80 young healthy women with acute uncomplicated cystitis and 20 asymptomatic women with a history of UTI. Investigators were blinded to culture results and clinical history. Samples were analyzed by light microscopy, immunofluorescence, and electron microscopy for evidence of exfoliated IBCs and filamentous bacteria. Evidence of IBCs was found in 14 of 80 (18%) urines from women with UTI. Filamentous bacteria were found in 33 of 80 (41%) urines from women with UTI. None of the 20 urines from the asymptomatic comparative group showed evidence of IBCs or filaments. Filamentous bacteria were present in all 14 of the urines with IBCs compared to 19 (29%) of 66 samples with no evidence of IBCs (p < 0.001). Of 65 urines from patients with E. coli infections, 14 (22%) had evidence of IBCs and 29 (45%) had filamentous bacteria, while none of the gram-positive infections had IBCs or filamentous bacteria. Conclusions The presence of exfoliated IBCs and filamentous bacteria in the urines of women with acute cystitis suggests that the IBC pathogenic pathway characterized in the murine model may occur in humans. The findings

  13. Metabolism of heme and bilirubin in rat and human small intestinal mucosa.

    PubMed Central

    Hartmann, F; Bissell, D M

    1982-01-01

    Formation of heme, bilirubin, and bilirubin conjugates has been examined in mucosal cells isolated from the rat upper small intestine. Intact, viable cells were prepared by enzymatic dissociation using a combined vascular and luminal perfusion and incubated with an isotopically labeled precursor, delta-amino-[2,3-3H]levulinic acid. Labeled heme and bile pigment were formed with kinetics similar to those exhibited by hepatocytes. Moreover, the newly formed bilirubin was converted rapidly to both mono- and diglucuronide conjugates. In addition, cell-free extracts of small intestinal mucosa from rats or humans exhibited a bilirubin-UDP-glucuronyl transferase activity that was qualitatively similar to that present in liver. The data suggest that the small intestinal mucosa normally contributes to bilirubin metabolism. PMID:6806320

  14. Interaction of Salmonella enterica Serovar Typhimurium with Intestinal Organoids Derived from Human Induced Pluripotent Stem Cells.

    PubMed

    Forbester, Jessica L; Goulding, David; Vallier, Ludovic; Hannan, Nicholas; Hale, Christine; Pickard, Derek; Mukhopadhyay, Subhankar; Dougan, Gordon

    2015-07-01

    The intestinal mucosa forms the first line of defense against infections mediated by enteric pathogens such as salmonellae. Here we exploited intestinal "organoids" (iHOs) generated from human induced pluripotent stem cells (hIPSCs) to explore the interaction of Salmonella enterica serovar Typhimurium with iHOs. Imaging and RNA sequencing were used to analyze these interactions, and clear changes in transcriptional signatures were detected, including altered patterns of cytokine expression after the exposure of iHOs to bacteria. S. Typhimurium microinjected into the lumen of iHOs was able to invade the epithelial barrier, with many bacteria residing within Salmonella-containing vacuoles. An S. Typhimurium invA mutant defective in the Salmonella pathogenicity island 1 invasion apparatus was less capable of invading the iHO epithelium. Hence, we provide evidence that hIPSC-derived organoids are a promising model of the intestinal epithelium for assessing interactions with enteric pathogens.

  15. Detection of fastidious mycobacteria in human intestines by the polymerase chain reaction.

    PubMed

    Dumonceau, J M; Van Gossum, A; Adler, M; Van Vooren, J P; Fonteyne, P A; De Beenhouwer, H; Portaels, F

    1997-05-01

    The aim of this study was to determine whether difficult-to-grow mycobacteria are present in human intestines. Intestinal tissue samples were subjected to both mycobacterial culture and a polymerase chain reaction (PCR) assay. After detection by PCR, species identity was determined by hybridizing the amplified 16S rRNA gene fragments with species-specific oligonucleotides. Intestinal biopsies from 63 patients with noninflammatory bowel diseases (n = 22), Crohn's disease (n = 31), or ulcerative colitis (n = 10) were analyzed. Culture and PCR revealed mycobacteria in four (6%) and 25 (40%) samples, respectively. Samples positive by PCR were negative with all probes specific to nine common cultivable species but were positive with Mycobacterium genavense-specific probe in 68% of cases. Mycobacterial isolates were identified as Mycobacterium gordonae and Mycobacterium chelonae. Findings were similar in Crohn's disease samples compared to non-Chron's disease samples. This study shows that difficult-to-grow mycobacteria can be detected by PCR in large and similar proportions of inflamed intestinal tissue from patients with inflammatory bowel disease and intestinal tissue that appears normal from patients with noninflammatory bowel disease.

  16. Recombinant Human Epidermal Growth Factor Accelerates Recovery of Mouse Small Intestinal Mucosa After Radiation Damage

    SciTech Connect

    Lee, Kang Kyoo; Jo, Hyang Jeong; Hong, Joon Pio; Lee, Sang-wook Sohn, Jung Sook; Moon, Soo Young; Yang, Sei Hoon; Shim, Hyeok; Lee, Sang Ho; Ryu, Seung-Hee; Moon, Sun Rock

    2008-07-15

    Purpose: To determine whether systemically administered recombinant human epidermal growth factor (rhEGF) accelerates the recovery of mouse small intestinal mucosa after irradiation. Methods and Materials: A mouse mucosal damage model was established by administering radiation to male BALB/c mice with a single dose of 15 Gy applied to the abdomen. After irradiation, rhEGF was administered subcutaneously at various doses (0.04, 0.2, 1.0, and 5.0 mg/kg/day) eight times at 2- to 3-day intervals. The evaluation methods included histologic changes of small intestinal mucosa, change in body weight, frequency of diarrhea, and survival rate. Results: The recovery of small intestinal mucosa after irradiation was significantly improved in the mice treated with a high dose of rhEGF. In the mice that underwent irradiation without rhEGF treatment, intestinal mucosal ulceration, mucosal layer damage, and severe inflammation occurred. The regeneration of villi was noticeable in mice treated with more than 0.2 mg/kg rhEGF, and the villi recovered fully in mice given more than 1 mg/kg rhEGF. The frequency of diarrhea persisting for more than 3 days was significantly greater in the radiation control group than in the rhEGF-treated groups. Conclusions: Systemic administration of rhEGF accelerates recovery from mucosal damage induced by irradiation. We suggest that rhEGF treatment shows promise for the reduction of small intestinal damage after irradiation.

  17. Protein abundance of clinically relevant multidrug transporters along the entire length of the human intestine.

    PubMed

    Drozdzik, Marek; Gröer, Christian; Penski, Jette; Lapczuk, Joanna; Ostrowski, Marek; Lai, Yurong; Prasad, Bhagwat; Unadkat, Jashvant D; Siegmund, Werner; Oswald, Stefan

    2014-10-06

    Intestinal transporters are crucial determinants in the oral absorption of many drugs. We therefore studied the mRNA expression (N = 33) and absolute protein content (N = 10) of clinically relevant transporters in healthy epithelium of the duodenum, the proximal and distal jejunum and ileum, and the ascending, transversal, descending, and sigmoidal colon of six organ donors (24-54 years). In the small intestine, the abundance of nearly all studied proteins ranged between 0.2 and 1.6 pmol/mg with the exception of those of OCT3 (<0.1 pmol/mg) and PEPT1 (2.6-4.9 pmol/mg) that accounted for ∼50% of all measured transporters. OATP1A2 was not detected in any intestinal segment. ABCB1, ABCG2, PEPT1, and ASBT were significantly more abundant in jejunum and ileum than in colon. In contrast to this, the level of expression of ABCC2, ABCC3, and OCT3 was found to be highest in colon. Site-dependent differences in the levels of gene and protein expression were observed for ABCB1 and ASBT. Significant correlations between mRNA and protein levels have been found for ABCG2, ASBT, OCT3, and PEPT1 in the small intestine. Our data provide further physiological pieces of the puzzle required to predict intestinal drug absorption in humans.

  18. Human, rat and chicken small intestinal Na+-Cl−-creatine transporter: functional, molecular characterization and localization

    PubMed Central

    Peral, M J; García-Delgado, M; Calonge, M L; Durán, J M; De La Horra, M C; Wallimann, T; Speer, O; Ilundáin, A A

    2002-01-01

    In spite of all the fascinating properties of oral creatine supplementation, the mechanism(s) mediating its intestinal absorption has(have) not been investigated. The purpose of this study was to characterize intestinal creatine transport. [14C]Creatine uptake was measured in chicken enterocytes and rat ileum, and expression of the creatine transporter CRT was examined in human, rat and chicken small intestine by reverse transcription-polymerase chain reaction, Northern blot, in situ hybridization, immunoblotting and immunohistochemistry. Results show that enterocytes accumulate creatine against its concentration gradient. This accumulation was electrogenic, Na+- and Cl−-dependent, with a probable stoichiometry of 2 Na+: 1 Cl−: 1 creatine, and inhibited by ouabain and iodoacetic acid. The kinetic study revealed a Km for creatine of 29 μm. [14C]Creatine uptake was efficiently antagonized by non-labelled creatine, guanidinopropionic acid and cyclocreatine. More distant structural analogues of creatine, such as GABA, choline, glycine, β-alanine, taurine and betaine, had no effect on intestinal creatine uptake, indicating a high substrate specificity of the creatine transporter. Consistent with these functional data, messenger RNA for CRT was detected only in the cells lining the intestinal villus. The sequences of partial clones, and of the full-length cDNA clone, isolated from human and rat small intestine were identical to previously cloned CRT cDNAs. Immunological analysis revealed that CRT protein was mainly associated with the apical membrane of the enterocytes. This study reports for the first time that mammalian and avian enterocytes express CRT along the villus, where it mediates high-affinity, Na+- and Cl−-dependent, apical creatine uptake. PMID:12433955

  19. Human milk mucin 1 and mucin 4 inhibit Salmonella enterica serovar Typhimurium invasion of human intestinal epithelial cells in vitro.

    PubMed

    Liu, Bo; Yu, Zhuoteng; Chen, Ceng; Kling, David E; Newburg, David S

    2012-08-01

    Many human milk glycans inhibit pathogen binding to host receptors and their consumption by infants is associated with reduced risk of disease. Salmonella infection is more frequent among infants than among the general population, but the incidence is lower in breast-fed babies, suggesting that human milk could contain components that inhibit Salmonella. This study aimed to test whether human milk per se inhibits Salmonella invasion of human intestinal epithelial cells in vitro and, if so, to identify the milk components responsible for inhibition. Salmonella enterica serovar Typhimurium SL1344 (SL1344) invasion of FHs 74 Int and Caco-2 cells were the models of human intestinal epithelium infection. Internalization of fluorescein-5-isothiocyanate-labeled SL1344 into intestinal cells was measured by flow cytometry to quantify infection. Human milk and its fractions inhibited infection; the inhibitory activity localized to the high molecular weight glycans. Mucin 1 and mucin 4 were isolated to homogeneity. At 150 μg/L, a typical concentration in milk, human milk mucin 1 and mucin 4 inhibited SL1344 invasion of both target cell types. These mucins inhibited SL1344 invasion of epithelial cells in a dose-dependent manner. Thus, mucins may prove useful as a basis for developing novel oral prophylactic and therapeutic agents that inhibit infant diseases caused by Salmonella and related pathogens.

  20. Biotransformation on the flavonolignan constituents of Silybi Fructus by an intestinal bacterial strain Eubacterium limosum ZL-II.

    PubMed

    Zhang, Ying; Yang, Dong-Hui; Zhang, Ying-Tao; Chen, Xiu-Min; Li, Li-Li; Cai, Shao-Qing

    2014-01-01

    Eubacterium limosum ZL-II is an anaerobic bacterium with demethylated activity, which was isolated from human intestinal bacteria in our previous work. In this study, the flavonolignan constituents of Silybi Fructus were biotransformed by E. limosum(1) ZL-II, producing four new transformation products - demethylisosilybin B (T1), demethylisosilybin A (T2), demethylsilybin B (T3) and demethylsilybin A (T4), among which T1 and T2 were new compounds. Their chemical structures were identified by ESI-TOF/MS, (1)H NMR, (13)C NMR, HMBC and CD spectroscopic data. The bioassay results showed that the transformation products T1-T4 exhibited significant inhibitory activities on Alzheimer's amyloid-β 42 (Aβ42(2)) aggregation with IC50 values at 7.49 μM-10.46 μM, which were comparable with that of the positive control (epigallocatechin gallate, EGCG(3), at 9.01 μM) and much lower than those of their parent compounds (at not less than 145.10 μM). The method of biotransformation by E. limosum ZL-II explored a way to develop the new and active lead compounds in Alzheimer's disease from Silybi Fructus. However, the transformation products T1-T4 exhibited decreased inhibitory activities against human tumor cell lines comparing with their parent compounds.

  1. Human small intestinal epithelial cells differentiated from adult intestinal stem cells as a novel system for predicting oral drug absorption in humans.

    PubMed

    Takenaka, Toru; Harada, Naomoto; Kuze, Jiro; Chiba, Masato; Iwao, Takahiro; Matsunaga, Tamihide

    2014-11-01

    Adult intestinal stem cells (ISCs) possess both a long-term proliferation ability and differentiation capability into enterocytes. As a novel in vitro system for the evaluation of drug absorption, we characterized a human small intestinal epithelial cell (HIEC) monolayer that differentiated from adult ISCs. Continuous proliferation/differentiation from ISCs consistently conferred the capability of maturation of enterocytes to HIECs over 25 passages. The morphologically matured HIEC monolayer consisted of polarized columnar epithelia with dense microvilli, tight junctions, and desmosomes 8 days after seeding onto culture inserts. Transepithelial electrical resistance across the monolayer was 9-fold lower in HIECs (98.9 Ω × cm(2)) than in Caco-2 cells (900 Ω × cm(2)), which indicated that the looseness of the tight junctions in the HIEC monolayer was similar to that in the human small intestine (approximately 40 Ω × cm(2)). No significant differences were observed in the overall gene expression patterns of the major drug-metabolizing enzymes and transporters between the HIEC and Caco-2 cell monolayers. Furthermore, the functions of P-glycoprotein and breast cancer resistance protein in the HIEC monolayer were confirmed by the vectorial transport of marker substrates and their disappearance in the presence of specific inhibitors. The apparent drug permeability values of paracellularly transported compounds (fluorescein isothiocyanate-dextran 4000, atenolol, and terbutaline) and nucleoside transporter substrates (didanosine, ribavirin, and doxifluridine) in the HIEC monolayer were markedly higher than those of Caco-2 cells, whereas transcellularly transported drugs (pindolol and midazolam) were equally well permeated. In conclusion, the HIEC monolayer can serve as a novel and superior alternative to the conventional Caco-2 cell monolayer for predicting oral absorption in humans.

  2. Diversity of halophilic archaea in fermented foods and human intestines and their application.

    PubMed

    Lee, Han-Seung

    2013-12-01

    Archaea are prokaryotic organisms distinct from bacteria in the structural and molecular biological sense, and these microorganisms are known to thrive mostly at extreme environments. In particular, most studies on halophilic archaea have been focused on environmental and ecological researches. However, new species of halophilic archaea are being isolated and identified from high salt-fermented foods consumed by humans, and it has been found that various types of halophilic archaea exist in food products by culture-independent molecular biological methods. In addition, even if the numbers are not quite high, DNAs of various halophilic archaea are being detected in human intestines and much interest is given to their possible roles. This review aims to summarize the types and characteristics of halophilic archaea reported to be present in foods and human intestines and to discuss their application as well.

  3. Hyperforin Exhibits Antigenotoxic Activity on Human and Bacterial Cells.

    PubMed

    Imreova, Petronela; Feruszova, Jana; Kyzek, Stanislav; Bodnarova, Kristina; Zduriencikova, Martina; Kozics, Katarina; Mucaji, Pavel; Galova, Eliska; Sevcovicova, Andrea; Miadokova, Eva; Chalupa, Ivan

    2017-01-21

    Hyperforin (HF), a substance that accumulates in the leaves and flowers of Hypericum perforatum L. (St. John's wort), consists of a phloroglucinol skeleton with lipophilic isoprene chains. HF exhibits several medicinal properties and is mainly used as an antidepressant. So far, the antigenotoxicity of HF has not been investigated at the level of primary genetic damage, gene mutations, and chromosome aberrations, simultaneously. The present work is designed to investigate the potential antigenotoxic effects of HF using three different experimental test systems. The antigenotoxic effect of HF leading to the decrease of primary/transient promutagenic genetic changes was detected by the alkaline comet assay on human lymphocytes. The HF antimutagenic effect leading to the reduction of gene mutations was assessed using the Ames test on the standard Salmonella typhimurium (TA97, TA98, and TA100) bacterial strains, and the anticlastogenic effect of HF leading to the reduction of chromosome aberrations was evaluated by the in vitro mammalian chromosome aberration test on the human tumor cell line HepG2 and the non-carcinogenic cell line VH10. Our findings provided evidence that HF showed antigenotoxic effects towards oxidative mutagen zeocin in the comet assay and diagnostic mutagen (4-nitroquinoline-1-oxide) in the Ames test. Moreover, HF exhibited an anticlastogenic effect towards benzo(a)pyrene and cisplatin in the chromosome aberration test.

  4. Doublecortin-like kinase 1-positive enterocyte - a new cell type in human intestine.

    PubMed

    Leppänen, Joni; Helminen, Olli; Huhta, Heikki; Kauppila, Joonas H; Miinalainen, Ilkka; Ronkainen, Veli-Pekka; Saarnio, Juha; Lehenkari, Petri P; Karttunen, Tuomo J

    2016-11-01

    Doublecortin-like kinase 1 (DCLK1) is a microtubule-associated kinase. In murine intestine, DCLK1 marks tuft cells with characteristic microvilli, features of neuroendocrine cells and also quiescent stem cell-like properties. The occurrence and pathological role of DCLK1-positive cells in human intestinal mucosa is unknown. We analysed DCLK1 expression in healthy duodenal, jejunal and colorectal mucosa samples (n = 35), and in duodenal specimens from patients with coeliac disease (n = 20). The samples were immunohistochemically double-stained with DCLK1, and synaptophysin, chromogranin A and Ki-67. Ultrastructure of DCLK1-expressing duodenal cells was assessed using correlative light and electron microscopy. DCLK1 expression was seen in about 1% of epithelial cells diffusely scattered through the intestinal epithelium. Electron microscopy showed that the duodenal DCLK1-positive cells had short apical microvilli similar to neighbouring enterocytes and cytoplasmic granules on the basal side. DCLK1-positive cells were stained with synaptophysin. The number of DCLK1-positive cells was decreased in villus atrophy in coeliac disease. Our findings indicate that in human intestinal epithelium, DLCK1-positive cells form a subpopulation of non-proliferating neuroendocrine cells with apical brush border similar to that in enterocytes, and their number is decreased in untreated coeliac disease.

  5. Lactobacillus reuteri Inhibition of Enteropathogenic Escherichia coli Adherence to Human Intestinal Epithelium

    PubMed Central

    Walsham, Alistair D. S.; MacKenzie, Donald A.; Cook, Vivienne; Wemyss-Holden, Simon; Hews, Claire L.; Juge, Nathalie; Schüller, Stephanie

    2016-01-01

    Enteropathogenic Escherichia coli (EPEC) is a major cause of diarrheal infant death in developing countries, and probiotic bacteria have been shown to provide health benefits in gastrointestinal infections. In this study, we have investigated the influence of the gut symbiont Lactobacillus reuteri on EPEC adherence to the human intestinal epithelium. Different host cell model systems including non-mucus-producing HT-29 and mucus-producing LS174T intestinal epithelial cell lines as well as human small intestinal biopsies were used. Adherence of L. reuteri to HT-29 cells was strain-specific, and the mucus-binding proteins CmbA and MUB increased binding to both HT-29 and LS174T cells. L. reuteri ATCC PTA 6475 and ATCC 53608 significantly inhibited EPEC binding to HT-29 but not LS174T cells. While pre-incubation of LS174T cells with ATCC PTA 6475 did not affect EPEC attaching/effacing (A/E) lesion formation, it increased the size of EPEC microcolonies. ATCC PTA 6475 and ATCC 53608 binding to the mucus layer resulted in decreased EPEC adherence to small intestinal biopsy epithelium. Our findings show that L. reuteri reduction of EPEC adhesion is strain-specific and has the potential to target either the epithelium or the mucus layer, providing further rationale for the selection of probiotic strains. PMID:26973622

  6. Short- and long-term effects of oral vancomycin on the human intestinal microbiota

    PubMed Central

    Isaac, Sandrine; Scher, Jose U.; Djukovic, Ana; Jiménez, Nuria; Littman, Dan R.; Abramson, Steven B.; Pamer, Eric G.; Ubeda, Carles

    2017-01-01

    Background Oral vancomycin remains the mainstay of therapy for severe infections produced by Clostridium difficile, the most prevalent cause of healthcare-associated infectious diarrhoea in developed countries. However, its short- and long-term effects on the human intestinal microbiota remain largely unknown. Methods We utilized high-throughput sequencing to analyse the effects of vancomycin on the faecal human microbiota up to 22 weeks post-antibiotic cessation. The clinical relevance of the observed microbiota perturbations was studied in mice. Results During vancomycin therapy, most intestinal microbiota genera and operational taxonomic units (OTUs) were depleted in all analysed subjects, including all baseline OTUs from the phylum Bacteroidetes. This was accompanied by a vast expansion of genera associated with infections, including Klebsiella and Escherichia/Shigella. Following antibiotic cessation, marked differences in microbiota resilience were observed among subjects. While some individuals recovered a microbiota close to baseline composition, in others, up to 89% of abundant OTUs could no longer be detected. The clinical relevance of the observed microbiota changes was further demonstrated in mice, which developed analogous microbiota alterations. During vancomycin treatment, mice were highly susceptible to intestinal colonization by an antibiotic-resistant pathogen and, upon antibiotic cessation, a less-resilient microbiota allowed higher levels of pathogen colonization. Conclusions Oral vancomycin induces drastic and consistent changes in the human intestinal microbiota. Upon vancomycin cessation, the microbiota recovery rate varied considerably among subjects, which could influence, as validated in mice, the level of susceptibility to pathogen intestinal colonization. Our results demonstrate the negative long-term effects of vancomycin, which should be considered as a fundamental aspect of the cost–benefit equation for antibiotic prescription. PMID

  7. Megaselia scalaris causing human intestinal myiasis in Egypt.

    PubMed

    Mazayad, Said A M; Rifaat, Manal M A

    2005-04-01

    Megaselia scalaris is a worldwide distributed insect of medical importance. In a laboratory-based study, stool samples with undefined maggot infestation were examined and the presence of M. scalaris maggots was confirmed. Binocular stereo-microscopy was used for identification of the maggots. Larvae were allowed to develop into adults onto a human stool culture. The larvae and the emerged flies were identified using standard keys. This may be the first report of M. scalaris as a causative agent of human myiasis in Egypt. Details of the third instar larva, pupa and adults were given.

  8. Modeling colorectal cancer using CRISPR-Cas9-mediated engineering of human intestinal organoids.

    PubMed

    Matano, Mami; Date, Shoichi; Shimokawa, Mariko; Takano, Ai; Fujii, Masayuki; Ohta, Yuki; Watanabe, Toshiaki; Kanai, Takanori; Sato, Toshiro

    2015-03-01

    Human colorectal tumors bear recurrent mutations in genes encoding proteins operative in the WNT, MAPK, TGF-β, TP53 and PI3K pathways. Although these pathways influence intestinal stem cell niche signaling, the extent to which mutations in these pathways contribute to human colorectal carcinogenesis remains unclear. Here we use the CRISPR-Cas9 genome-editing system to introduce multiple such mutations into organoids derived from normal human intestinal epithelium. By modulating the culture conditions to mimic that of the intestinal niche, we selected isogenic organoids harboring mutations in the tumor suppressor genes APC, SMAD4 and TP53, and in the oncogenes KRAS and/or PIK3CA. Organoids engineered to express all five mutations grew independently of niche factors in vitro, and they formed tumors after implantation under the kidney subcapsule in mice. Although they formed micrometastases containing dormant tumor-initiating cells after injection into the spleen of mice, they failed to colonize in the liver. In contrast, engineered organoids derived from chromosome-instable human adenomas formed macrometastatic colonies. These results suggest that 'driver' pathway mutations enable stem cell maintenance in the hostile tumor microenvironment, but that additional molecular lesions are required for invasive behavior.

  9. Hydrolysis of pyrethroids by human and rat tissues: Examination of intestinal, liver and serum carboxylesterases

    SciTech Connect

    Crow, J. Allen; Borazjani, Abdolsamad; Potter, Philip M.; Ross, Matthew K. . E-mail: mross@cvm.msstate.edu

    2007-05-15

    Hydrolytic metabolism of pyrethroid insecticides in humans is one of the major catabolic pathways that clear these compounds from the body. Rodent models are often used to determine the disposition and clearance rates of these esterified compounds. In this study the distribution and activities of esterases that catalyze pyrethroid metabolism have been investigated in vitro using several human and rat tissues, including small intestine, liver and serum. The major esterase in human intestine is carboxylesterase 2 (hCE2). We found that the pyrethroid trans-permethrin is effectively hydrolyzed by a sample of pooled human intestinal microsomes (5 individuals), while deltamethrin and bioresmethrin are not. This result correlates well with the substrate specificity of recombinant hCE2 enzyme. In contrast, a sample of pooled rat intestinal microsomes (5 animals) hydrolyze trans-permethrin 4.5-fold slower than the sample of human intestinal microsomes. Furthermore, it is demonstrated that pooled samples of cytosol from human or rat liver are {approx} 2-fold less hydrolytically active (normalized per mg protein) than the corresponding microsomal fraction toward pyrethroid substrates; however, the cytosolic fractions do have significant amounts ({approx} 40%) of the total esteratic activity. Moreover, a 6-fold interindividual variation in carboxylesterase 1 protein expression in human hepatic cytosols was observed. Human serum was shown to lack pyrethroid hydrolytic activity, but rat serum has hydrolytic activity that is attributed to a single CE isozyme. We purified the serum CE enzyme to homogeneity to determine its contribution to pyrethroid metabolism in the rat. Both trans-permethrin and bioresmethrin were effectively cleaved by this serum CE, but deltamethrin, esfenvalerate, alpha-cypermethrin and cis-permethrin were slowly hydrolyzed. Lastly, two model lipase enzymes were examined for their ability to hydrolyze pyrethroids. However, no hydrolysis products could be

  10. Comparative genomics analysis of Streptococcus isolates from the human small intestine reveals their adaptation to a highly dynamic ecosystem.

    PubMed

    Van den Bogert, Bartholomeus; Boekhorst, Jos; Herrmann, Ruth; Smid, Eddy J; Zoetendal, Erwin G; Kleerebezem, Michiel

    2013-01-01

    The human small-intestinal microbiota is characterised by relatively large and dynamic Streptococcus populations. In this study, genome sequences of small-intestinal streptococci from S. mitis, S. bovis, and S. salivarius species-groups were determined and compared with those from 58 Streptococcus strains in public databases. The Streptococcus pangenome consists of 12,403 orthologous groups of which 574 are shared among all sequenced streptococci and are defined as the Streptococcus core genome. Genome mining of the small-intestinal streptococci focused on functions playing an important role in the interaction of these streptococci in the small-intestinal ecosystem, including natural competence and nutrient-transport and metabolism. Analysis of the small-intestinal Streptococcus genomes predicts a high capacity to synthesize amino acids and various vitamins as well as substantial divergence in their carbohydrate transport and metabolic capacities, which is in agreement with observed physiological differences between these Streptococcus strains. Gene-specific PCR-strategies enabled evaluation of conservation of Streptococcus populations in intestinal samples from different human individuals, revealing that the S. salivarius strains were frequently detected in the small-intestine microbiota, supporting the representative value of the genomes provided in this study. Finally, the Streptococcus genomes allow prediction of the effect of dietary substances on Streptococcus population dynamics in the human small-intestine.

  11. Refining small intestinal bacterial overgrowth diagnosis by means of carbohydrate specificity: a proof-of-concept study

    PubMed Central

    Enko, Dietmar; Halwachs-Baumann, Gabriele; Stolba, Robert; Mangge, Harald; Kriegshäuser, Gernot

    2015-01-01

    Background: Diagnosis of small intestinal bacterial overgrowth (SIBO) remains challenging. This study aimed at proving the diagnostic concept of carbohydrate-specific SIBO (cs-SIBO) using glucose, fructose and sorbitol hydrogen (H2)/methane (CH4) breath testing (HMBT). Methods: In this study 125 patients referred to our outpatient clinic for SIBO testing were included. All individuals underwent glucose (50 g), fructose (25 g) and sorbitol (12.5 g) HMBT at 3 consecutive days. Patients with cs-SIBO (i.e. early H2/CH4 peak) were given rifaximin (600 mg/day) in a 10-day treatment. HMBT results were reassessed in a subset of patients 3–6 months after antibiotic therapy. In view of cs-SIBO diagnosis, agreements between HMBT results obtained for different sugars were calculated using Cohen’s kappa (κ) with 95% confidence intervals (CIs). Results: A total of 59 (47.2%) patients presented an early H2/CH4 peak with one or more sugars. Among these, 21 (16.8%), 10 (8.0%) and 7 (5.6%) individuals had a positive HMBT result with either glucose, fructose or sorbitol, respectively. Another 21 (16.8%) patients with a positive glucose HMBT result were also found positive with an early H2/CH4 peak obtained after ingestion of fructose and/or sorbitol. Fair agreement was observed between glucose and fructose (κ = 0.26, p = 0.0018) and between glucose and sorbitol (κ = 0.18, p = 0.0178) HMBT results. Slight agreement was observed between fructose and sorbitol (κ = 0.03, p = 0.6955) HMBT results only. Successful antibiotic therapy with rifaximin could be demonstrated in 26/30 (86.7%) of patients as indicated by normal HMBT results and symptom remission. Conclusions: Combined glucose, fructose and sorbitol HMBT has the potential to optimize cs-SIBO diagnosis. Furthermore, the majority of patients with cs-SIBO seem to benefit from rifaximin therapy regardless of its carbohydrate specificity. PMID:27134657

  12. Metabolomics analysis of Cistus monspeliensis leaf extract on energy metabolism activation in human intestinal cells.

    PubMed

    Shimoda, Yoichi; Han, Junkyu; Kawada, Kiyokazu; Smaoui, Abderrazak; Isoda, Hiroko

    2012-01-01

    Energy metabolism is a very important process to improve and maintain health from the point of view of physiology. It is well known that the intracellular ATP production is contributed to energy metabolism in cells. Cistus monspeliensis is widely used as tea, spices, and medical herb; however, it has not been focusing on the activation of energy metabolism. In this study, C. monspeliensis was investigated as the food resources by activation of energy metabolism in human intestinal epithelial cells. C. monspeliensis extract showed high antioxidant ability. In addition, the promotion of metabolites of glycolysis and TCA cycle was induced by C. monspeliensis treatment. These results suggest that C. monspeliensis extract has an ability to enhance the energy metabolism in human intestinal cells.

  13. Diagnosis of edema and inflammation in human intestines using ultrawideband radar

    NASA Astrophysics Data System (ADS)

    Smith, Sonny; Narayanan, Ram M.; Messaris, Evangelos

    2015-05-01

    Human intestines are vital organs, which are often subjected to chronic issues. In particular, Crohn's disease is a bowel aliment resulting in inflammation along the lining of one's digestive tract. Moreover, such an inflammatory condition causes changes in the thickness of the intestines; and we posit induce changes in the dielectric properties detectable by radar. This detection hinges on the increase in fluid content in the afflicted area, which is described by effective medium approximations (EMA). In this paper, we consider one of the constitutive parameters (i.e. relative permittivity) of different human tissues and introduce a simple numerical, electromagnetic multilayer model. We observe how the increase in water content in one layer can be approximated to predict the effective permittivity of that layer. Moreover, we note trends in how such an accumulation can influence the total effective reflection coefficient of the multiple layers.

  14. Metabolomics Analysis of Cistus monspeliensis Leaf Extract on Energy Metabolism Activation in Human Intestinal Cells

    PubMed Central

    Shimoda, Yoichi; Han, Junkyu; Kawada, Kiyokazu; Smaoui, Abderrazak; Isoda, Hiroko

    2012-01-01

    Energy metabolism is a very important process to improve and maintain health from the point of view of physiology. It is well known that the intracellular ATP production is contributed to energy metabolism in cells. Cistus monspeliensis is widely used as tea, spices, and medical herb; however, it has not been focusing on the activation of energy metabolism. In this study, C. monspeliensis was investigated as the food resources by activation of energy metabolism in human intestinal epithelial cells. C. monspeliensis extract showed high antioxidant ability. In addition, the promotion of metabolites of glycolysis and TCA cycle was induced by C. monspeliensis treatment. These results suggest that C. monspeliensis extract has an ability to enhance the energy metabolism in human intestinal cells. PMID:22523469

  15. Human intestinal gas measurement systems: in vitro fermentation and gas capsules.

    PubMed

    Ou, Jian Zhen; Yao, C K; Rotbart, Asaf; Muir, Jane G; Gibson, Peter R; Kalantar-zadeh, Kourosh

    2015-04-01

    The biological and clinical significance of the human gut microbiome is currently attracting worldwide attention. While rRNA and DNA technologies led to a quantum leap in our understanding of the numbers and types of gut microorganisms, much less is known about these microorganisms' activity in situ and in real time. Accurately measuring their byproducts, including intestinal gases, may offer unique biomarkers for specific gut microbiota, accelerating our understanding of the relationships among intestinal gases, the metabolic activity of the gut microbiome, and human health states. Here we present two novel techniques, namely in vitro fermentation and gas capsule systems, for measuring and assessing selected gas species. We discuss new developments with these technologies and the methods of their implementation and provide an overall review of their operation.

  16. Generation of L cells in mouse and human small intestine organoids.

    PubMed

    Petersen, Natalia; Reimann, Frank; Bartfeld, Sina; Farin, Henner F; Ringnalda, Femke C; Vries, Robert G J; van den Brink, Stieneke; Clevers, Hans; Gribble, Fiona M; de Koning, Eelco J P

    2014-02-01

    Upon a nutrient challenge, L cells produce glucagon-like peptide 1 (GLP-1), a powerful stimulant of insulin release. Strategies to augment endogenous GLP-1 production include promoting L-cell differentiation and increasing L-cell number. Here we present a novel in vitro platform to generate functional L cells from three-dimensional cultures of mouse and human intestinal crypts. We show that short-chain fatty acids selectively increase the number of L cells, resulting in an elevation of GLP-1 release. This is accompanied by the upregulation of transcription factors associated with the endocrine lineage of intestinal stem cell development. Thus, our platform allows us to study and modulate the development of L cells in mouse and human crypts as a potential basis for novel therapeutic strategies in patients with type 2 diabetes.

  17. Safety assessment of genetically modified rice expressing human serum albumin from urine metabonomics and fecal bacterial profile.

    PubMed

    Qi, Xiaozhe; Chen, Siyuan; Sheng, Yao; Guo, Mingzhang; Liu, Yifei; He, Xiaoyun; Huang, Kunlun; Xu, Wentao

    2015-02-01

    The genetically modified (GM) rice expressing human serum albumin (HSA) is used for non-food purposes; however, its food safety assessment should be conducted due to the probability of accidental mixture with conventional food. In this research, Sprague Dawley rats were fed diets containing 50% (wt/wt) GM rice expressing HSA or non-GM rice for 90 days. Urine metabolites were detected by (1)H NMR to examine the changes of the metabolites in the dynamic process of metabolism. Fecal bacterial profiles were detected by denaturing gradient gel electrophoresis to reflect intestinal health. Additionally, short chain fatty acids and fecal enzymes were investigated. The results showed that compared with rats fed the non-GM rice, some significant differences were observed in rats fed with the GM rice; however, these changes were not significantly different from the control diet group. Additionally, the gut microbiota was associated with blood indexes and urine metabolites. In conclusion, the GM rice diet is as safe as the traditional daily diet. Furthermore, urine metabonomics and fecal bacterial profiles provide a non-invasive food safety assessment rat model for genetically modified crops that are used for non-food/feed purposes. Fecal bacterial profiles have the potential for predicting the change of blood indexes in future.

  18. Metabolism of liriodendrin and syringin by human intestinal bacteria and their relation to in vitro cytotoxicity.

    PubMed

    Kim, D H; Lee, K T; Bae, E A; Han, M J; Park, H J

    1999-02-01

    When liriodendrin or syringin was incubated for 24 h with human intestinal bacteria, two metabolites, (+)-syringaresinol-beta-D-glucopyranoside and (+)-syringaresinol, from liriodendrin and one metabolite, synapyl alcohol, from syringin were produced. The metabolic time course of liriodendrin was as follows: at early time, liriodendrin was converted to (+)-syringaresinol-beta-D-glucopyranoside, and then (+)-syringaresinol. The in vitro cytotoxicities of these metabolites, (+)-syringaresinol and synapyl alcohol, were superior to those of liriodendrin and syringin.

  19. A bioassay using intestinal organoids to measure CFTR modulators in human plasma.

    PubMed

    Dekkers, R; Vijftigschild, L A W; Vonk, A M; Kruisselbrink, E; de Winter-de Groot, K M; Janssens, H M; van der Ent, C K; Beekman, J M

    2015-03-01

    Treatment efficacies of drugs depend on patient-specific pharmacokinetic and pharmacodynamic properties. Here, we developed an assay to measure functional levels of the CFTR potentiator VX-770 in human plasma and observed that VX-770 in plasma from different donors induced variable CFTR function in intestinal organoids. This assay can help to understand variability in treatment response to CFTR potentiators by functionally modeling individual pharmacokinetics.

  20. Rapid and Accurate Diagnosis of Human Intestinal Spirochetosis by Fluorescence In Situ Hybridization▿

    PubMed Central

    Schmiedel, Dinah; Epple, Hans-Jörg; Loddenkemper, Christoph; Ignatius, Ralf; Wagner, Jutta; Hammer, Bettina; Petrich, Annett; Stein, Harald; Göbel, Ulf B.; Schneider, Thomas; Moter, Annette

    2009-01-01

    Human intestinal spirochetosis (HIS) is associated with overgrowth of the large intestine by spirochetes of the genus Brachyspira. The microbiological diagnosis of HIS is hampered by the fastidious nature and slow growth of Brachyspira spp. In clinical practice, HIS is diagnosed histopathologically, and a significant portion of cases may be missed. Fluorescence in situ hybridization (FISH) is a molecular method that allows the visualization and identification of single bacteria within tissue sections. In this study, we analyzed intestinal biopsy samples from five patients with possible HIS. All specimens yielded positive results by histopathological techniques. PCR amplification and sequencing of the 16S rRNA gene were performed. Sequences of two isolates clustered in the group of Brachyspira aalborgi, whereas in three cases, the sequences were highly similar to that of Brachyspira pilosicoli. Three phylotypes showed mismatches at distinct nucleotide positions with Brachyspira sp. sequences published previously. In addition, culture for Brachyspira was successful in three cases. On the basis of these data, we designed and evaluated a Brachyspira genus-specific 16S rRNA-directed FISH probe that detects all of the Brachyspira spp. published to date. FISH of biopsy samples resulted in strong, unequivocal signals of brush-like formations at the crypt surfaces. This technique allowed simultaneous visualization of single spirochetes and their identification as Brachyspira spp. In conclusion, FISH provides a fast and accurate technique for the visualization and identification of intestinal spirochetes in tissue sections. It therefore represents a valuable tool for routine diagnosis of HIS. PMID:19279178

  1. Evidence of native starch degradation with human small intestinal maltase-glucoamylase (recombinant).

    PubMed

    Ao, Zihua; Quezada-Calvillo, Roberto; Sim, Lyann; Nichols, Buford L; Rose, David R; Sterchi, Erwin E; Hamaker, Bruce R

    2007-05-29

    Action of human small intestinal brush border carbohydrate digesting enzymes is thought to involve only final hydrolysis reactions of oligosaccharides to monosaccharides. In vitro starch digestibility assays use fungal amyloglucosidase to provide this function. In this study, recombinant N-terminal subunit enzyme of human small intestinal maltase-glucoamylase (rhMGAM-N) was used to explore digestion of native starches from different botanical sources. The susceptibilities to enzyme hydrolysis varied among the starches. The rate and extent of hydrolysis of amylomaize-5 and amylomaize-7 into glucose were greater than for other starches. Such was not observed with fungal amyloglucosidase or pancreatic alpha-amylase. The degradation of native starch granules showed a surface furrowed pattern in random, radial, or tree-like arrangements that differed substantially from the erosion patterns of amyloglucosidase or alpha-amylase. The evidence of raw starch granule degradation with rhMGAM-N indicates that pancreatic alpha-amylase hydrolysis is not a requirement for native starch digestion in the human small intestine.

  2. Effects of human fecal flora on intestinal morphology and mucosal immunity in human flora-associated piglet.

    PubMed

    Che, C; Pang, X; Hua, X; Zhang, B; Shen, J; Zhu, J; Wei, H; Sun, L; Chen, P; Cui, L; Zhao, L; Yang, Q

    2009-03-01

    Human flora-associated (HFA) piglet model was established to examine the effects of gut microbes from a different donor species on the intestinal morphology and mucosal immunity. Newborn germ-free piglets, obtained by caesarean section, were orally inoculated with a human and a porcine faecal suspension, and artificially fed to establish a HFA group (n = 7) and pig flora-associated (PFA) group (n = 7), respectively. All pigs were killed 6 weeks later. Tissue samples from duodenum, jejunum, ileum and colon were collected and studied by histochemistry and immunohistochemistry methods for intestinal morphological analyses and detection of immunocompetent cells. In summary, both groups of pigs performed well but HFA pigs had a somewhat better daily weight gain, and their jejunal villus height and crypt depth were significantly higher. In comparison with PFA pigs, the number of intraepithelial lymphocytes in jejunum was lower but the number of goblet cells containing neutral mucins was significantly increased in HFA pigs. No difference was observed in the number of mast cells. The areas of IgA producing cells and CD4(+) T cells in the jejunum and IgG producing cells in the small intestine were significantly higher in HFA pigs. However, the areas of MHC class II expressing cells were significantly increased in the duodenum and colon. Additionally, the amount of Bifidobacteria spp. was significantly higher in HFA pigs. This study confirms that the composition of gut microbes differentially affects the host intestinal mucosal immunity and suggests that commensal bacteria have great effects on intestinal health and development.

  3. Prediction of human drug clearance by multiple metabolic pathways: integration of hepatic and intestinal microsomal and cytosolic data.

    PubMed

    Cubitt, Helen E; Houston, J Brian; Galetin, Aleksandra

    2011-05-01

    The current study assesses hepatic and intestinal glucuronidation, sulfation, and cytochrome P450 (P450) metabolism of raloxifene, quercetin, salbutamol, and troglitazone using different in vitro systems. The fraction metabolized by conjugation and P450 metabolism was estimated in liver and intestine, and the importance of multiple metabolic pathways on accuracy of clearance prediction was assessed. In vitro intrinsic sulfation clearance (CL(int, SULT)) was determined in human intestinal and hepatic cytosol and compared with hepatic and intestinal microsomal glucuronidation (CL(int, UGT)) and P450 clearance (CL(int, CYP)) expressed per gram of tissue. Hepatic and intestinal cytosolic scaling factors of 80.7 mg/g liver and 18 mg/g intestine were estimated from published data. Scaled CL(int, SULT) ranged between 0.7 and 11.4 ml · min(-1) · g(-1) liver and 0.1 and 3.3 ml · min(-1) · g(-1) intestine (salbutamol and quercetin were the extremes). Salbutamol was the only compound with a high extent of sulfation (51 and 28% of total CL(int) for liver and intestine, respectively) and also significant renal clearance (26-57% of observed plasma clearance). In contrast, the clearance of quercetin was largely accounted for by glucuronidation. Drugs metabolized by multiple pathways (raloxifene and troglitazone) demonstrated improved prediction of intravenous clearance using data from all hepatic pathways (44-86% of observed clearance) compared with predictions based only on the primary pathway (22-36%). The assumption of no intestinal first pass resulted in underprediction of oral clearance for raloxifene, troglitazone, and quercetin (3-22% of observed, respectively). Accounting for the intestinal contribution to oral clearance via estimated intestinal availability improved prediction accuracy for raloxifene and troglitazone (within 2.5-fold of observed). Current findings emphasize the importance of both hepatic and intestinal conjugation for in vitro-in vivo extrapolation

  4. Ex vivo permeability experiments in excised rat intestinal tissue and in vitro solubility measurements in aspirated human intestinal fluids support age-dependent oral drug absorption.

    PubMed

    Annaert, Pieter; Brouwers, Joachim; Bijnens, Ann; Lammert, Frank; Tack, Jan; Augustijns, Patrick

    2010-01-31

    The possible influence of advanced age on intestinal drug absorption was investigated by determining the effects of aging on (i) solubility of model drugs in human intestinal fluids (HIF) obtained from two age groups (18-25 years; 62-72 years); and (ii) transepithelial permeation of model drugs across intestinal tissue excised from young, adult and old rats. Average equilibrium solubility values for 10 poorly soluble compounds in HIF aspirated from both age groups showed high interindividual variability, but did not reveal significant differences. Characterization of the HIF from both age groups demonstrated comparable pH profiles, while concentrations of individual bile salts showed pronounced variability between individuals, however without statistical differences between age groups. Transepithelial permeation of the transcellular probe metoprolol was significantly increased in old rats (38 weeks) compared to the younger age groups, while the modulatory role of P-glycoprotein in transepithelial talinolol transport was observed in adult and old rats but not in young rats. In conclusion, age-dependent permeability of intestinal tissue (rather than age-dependent luminal drug solubility) may contribute to altered intestinal drug absorption in older patients compared to young adults.

  5. Species differences in hepatic and intestinal metabolic activities for 43 human cytochrome P450 substrates between humans and rats or dogs.

    PubMed

    Nishimuta, Haruka; Nakagawa, Tetsuya; Nomura, Naruaki; Yabuki, Masashi

    2013-11-01

    1. Prediction of human pharmacokinetics might be made more precise by using species with similar metabolic activities to humans. We had previously reported the species differences in intestinal and hepatic metabolic activities of 43 cytochrome P450 (CYP) substrates between cynomolgus monkeys and humans. However, the species differences between humans and rats or dogs had not yet been determined using comparable data sets with sufficient number of compounds. 2. Here, we investigated metabolic stabilities in intestinal and liver microsomes obtained from rats, dogs and humans using 43 substrates of human CYP1A2, CYP2J2, CYP2C, CYP2D6 and CYP3A. 3. Hepatic intrinsic clearance (CLint) values for most compounds in dogs were comparable to those in humans (within 10-fold), whereas in rats, those for the human CYP2D6 substrates were much higher and showed low correlation with humans. In dog intestine, as with human intestine, CLint values for almost all human CYP1A2, CYP2C, CYP2D6 substrates were not determined because they were very low. Intestinal CLint values for human CYP3A substrates in rats and dogs appeared to be lower for most of the compounds and showed moderate correlation with those in humans. 4. In conclusion, dogs showed the most similar metabolic activity to humans.

  6. Ricin crosses polarized human intestinal cells and intestines of ricin-gavaged mice without evident damage and then disseminates to mouse kidneys.

    PubMed

    Flora, Alyssa D; Teel, Louise D; Smith, Mark A; Sinclair, James F; Melton-Celsa, Angela R; O'Brien, Alison D

    2013-01-01

    Ricin is a potent toxin found in the beans of Ricinus communis and is often lethal for animals and humans when aerosolized or injected and causes significant morbidity and occasional death when ingested. Ricin has been proposed as a bioweapon because of its lethal properties, environmental stability, and accessibility. In oral intoxication, the process by which the toxin transits across intestinal mucosa is not completely understood. To address this question, we assessed the impact of ricin on the gastrointestinal tract and organs of mice after dissemination of toxin from the gut. We first showed that ricin adhered in a specific pattern to human small bowel intestinal sections, the site within the mouse gut in which a variable degree of damage has been reported by others. We then monitored the movement of ricin across polarized human HCT-8 intestinal monolayers grown in transwell inserts and in HCT-8 cell organoids. We observed that, in both systems, ricin trafficked through the cells without apparent damage until 24 hours post intoxication. We delivered a lethal dose of purified fluorescently-labeled ricin to mice by oral gavage and followed transit of the toxin from the gastrointestinal tracts to the internal organs by in vivo imaging of whole animals over time and ex vivo imaging of organs at various time points. In addition, we harvested organs from unlabeled ricin-gavaged mice and assessed them for the presence of ricin and for histological damage. Finally, we compared serum chemistry values from buffer-treated versus ricin-intoxicated animals. We conclude that ricin transverses human intestinal cells and mouse intestinal cells in situ prior to any indication of enterocyte damage and that ricin rapidly reaches the kidneys of intoxicated mice. We also propose that mice intoxicated orally with ricin likely die from distributive shock.

  7. Anthocyanin Absorption and Metabolism by Human Intestinal Caco-2 Cells--A Review.

    PubMed

    Kamiloglu, Senem; Capanoglu, Esra; Grootaert, Charlotte; Van Camp, John

    2015-09-08

    Anthocyanins from different plant sources have been shown to possess health beneficial effects against a number of chronic diseases. To obtain any influence in a specific tissue or organ, these bioactive compounds must be bioavailable, i.e., effectively absorbed from the gut into the circulation and transferred to the appropriate location within the body while still maintaining their bioactivity. One of the key factors affecting the bioavailability of anthocyanins is their transport through the gut epithelium. The Caco-2 cell line, a human intestinal epithelial cell model derived from a colon carcinoma, has been proven to be a good alternative to animal studies for predicting intestinal absorption of anthocyanins. Studies investigating anthocyanin absorption by Caco-2 cells report very low absorption of these compounds. However, the bioavailability of anthocyanins may be underestimated since the metabolites formed in the course of digestion could be responsible for the health benefits associated with anthocyanins. In this review, we critically discuss recent findings reported on the anthocyanin absorption and metabolism by human intestinal Caco-2 cells.

  8. Comprehensive Survey of Intestinal Microbiota Changes in Offspring of Human Microbiota-Associated Mice.

    PubMed

    von Klitzing, Eliane; Öz, Fulya; Ekmekciu, Ira; Escher, Ulrike; Bereswill, Stefan; Heimesaat, Markus M

    2017-03-01

    Secondary abiotic mice generated by broad-spectrum antibiotic treatment provide a valuable tool for association studies with microbiota derived from different vertebrate hosts. We here generated human microbiota-associated (hma) mice by human fecal microbiota transplantation of secondary abiotic mice and performed a comprehensive survey of the intestinal microbiota dynamics in offspring of hma mice over 18 weeks following weaning as compared to their mothers applying both cultural and molecular methods. Mice were maintained under standard hygienic conditions with open cages, handled under aseptic conditions, and fed autoclaved chow and water. Within 1 week post weaning, fecal loads of commensal enterobacteria and enterococci had decreased, whereas obligate anaerobic bacteria such as Bacteroides/Prevotella species and clostridia were stably colonizing the intestines of hma offspring at high loads. Lactobacilli numbers were successively increasing until 18 weeks post weaning in both hma offspring and mothers, whereas by then, bifidobacteria were virtually undetectable in the former only. Interestingly, fecal lactobacilli and bifidobacteria were higher in mothers as compared to their offspring at 5 and 18 weeks post weaning. We conclude that the intestinal microbiota composition changes in offspring of hma mice, but also their mothers over time particularly affecting aerobic and microaerobic species.

  9. Comprehensive Survey of Intestinal Microbiota Changes in Offspring of Human Microbiota-Associated Mice

    PubMed Central

    von Klitzing, Eliane; Öz, Fulya; Ekmekciu, Ira; Escher, Ulrike; Bereswill, Stefan; Heimesaat, Markus M.

    2017-01-01

    Secondary abiotic mice generated by broad-spectrum antibiotic treatment provide a valuable tool for association studies with microbiota derived from different vertebrate hosts. We here generated human microbiota-associated (hma) mice by human fecal microbiota transplantation of secondary abiotic mice and performed a comprehensive survey of the intestinal microbiota dynamics in offspring of hma mice over 18 weeks following weaning as compared to their mothers applying both cultural and molecular methods. Mice were maintained under standard hygienic conditions with open cages, handled under aseptic conditions, and fed autoclaved chow and water. Within 1 week post weaning, fecal loads of commensal enterobacteria and enterococci had decreased, whereas obligate anaerobic bacteria such as Bacteroides/Prevotella species and clostridia were stably colonizing the intestines of hma offspring at high loads. Lactobacilli numbers were successively increasing until 18 weeks post weaning in both hma offspring and mothers, whereas by then, bifidobacteria were virtually undetectable in the former only. Interestingly, fecal lactobacilli and bifidobacteria were higher in mothers as compared to their offspring at 5 and 18 weeks post weaning. We conclude that the intestinal microbiota composition changes in offspring of hma mice, but also their mothers over time particularly affecting aerobic and microaerobic species. PMID:28386472

  10. Anthocyanin Absorption and Metabolism by Human Intestinal Caco-2 Cells—A Review

    PubMed Central

    Kamiloglu, Senem; Capanoglu, Esra; Grootaert, Charlotte; Van Camp, John

    2015-01-01

    Anthocyanins from different plant sources have been shown to possess health beneficial effects against a number of chronic diseases. To obtain any influence in a specific tissue or organ, these bioactive compounds must be bioavailable, i.e., effectively absorbed from the gut into the circulation and transferred to the appropriate location within the body while still maintaining their bioactivity. One of the key factors affecting the bioavailability of anthocyanins is their transport through the gut epithelium. The Caco-2 cell line, a human intestinal epithelial cell model derived from a colon carcinoma, has been proven to be a good alternative to animal studies for predicting intestinal absorption of anthocyanins. Studies investigating anthocyanin absorption by Caco-2 cells report very low absorption of these compounds. However, the bioavailability of anthocyanins may be underestimated since the metabolites formed in the course of digestion could be responsible for the health benefits associated with anthocyanins. In this review, we critically discuss recent findings reported on the anthocyanin absorption and metabolism by human intestinal Caco-2 cells. PMID:26370977

  11. Subversion of human intestinal mucosa innate immunity by a Crohn's disease-associated E. coli.

    PubMed

    Jarry, A; Crémet, L; Caroff, N; Bou-Hanna, C; Mussini, J M; Reynaud, A; Servin, A L; Mosnier, J F; Liévin-Le Moal, V; Laboisse, C L

    2015-05-01

    Adherent-invasive Escherichia coli (AIEC), associated with Crohn's disease, are likely candidate contributory factors in the disease. However, signaling pathways involved in human intestinal mucosa innate host response to AIEC remain unknown. Here we use a 3D model of human intestinal mucosa explant culture to explore the effects of the AIEC strain LF82 on two innate immunity platforms, i.e., the inflammasome through evaluation of caspase-1 status, and NFκB signaling. We showed that LF82 bacteria enter and survive within a few intestinal epithelial cells and macrophages, without altering the mucosa overall architecture. Although 4-h infection with a Salmonella strain caused crypt disorganization, caspase-1 activation, and mature IL-18 production, LF82 bacteria were unable to activate caspase-1 and induce IL-18 production. In parallel, LF82 bacteria activated NFκB signaling in epithelial cells through IκBα phosphorylation, NFκBp65 nuclear translocation, and TNFα secretion. In addition, NFκB activation was crucial for the maintenance of epithelial homeostasis upon LF82 infection. In conclusion, here we decipher at the whole-mucosa level the mechanisms of the LF82-induced subversion of innate immunity that, by maintaining host cell integrity, ensure intracellular bacteria survival.

  12. Drug resistance and adherence to human intestines of enteroaggregative Escherichia coli.

    PubMed

    Yamamoto, T; Echeverria, P; Yokota, T

    1992-04-01

    Clinical isolates of enteroaggregative Escherichia coli (EAggEC) were tested for their in vitro susceptibilities to 27 antimicrobial agents. Marked drug resistance was observed with sulfamethoxazole, ampicillin, and chloramphenicol in contrast to such antimicrobial agents as cefixime, sparfloxacin, and ciprofloxacin. One of the EAggEC strains carried a plasmid that conferred on its host resistance to ampicillin, tetracycline, sulfamethoxazole, streptomycin, and spectinomycin and an ability to adhere to child ileal villi or HeLa cells in the characteristic aggregative pattern. This plasmid also mediated D-mannose-resistant hemagglutinin production and bacterial clump formation (autoagglutination). The data demonstrate appearance of marked drug resistance and an intestine-adherence and drug-resistance plasmid in the newest category of diarrheagenic E. coli.

  13. Vasoactive Intestinal Peptide Inhibits Human Small-Cell Lung Cancer Proliferation in vitro and in vivo

    NASA Astrophysics Data System (ADS)

    Maruno, Kaname; Absood, Afaf; Said, Sami I.

    1998-11-01

    Small-cell lung carcinoma (SCLC) is an aggressive, rapidly growing and metastasizing, and highly fatal neoplasm. We report that vasoactive intestinal peptide inhibits the proliferation of SCLC cells in culture and dramatically suppresses the growth of SCLC tumor-cell implants in athymic nude mice. In both cases, the inhibition was mediated apparently by a cAMP-dependent mechanism, because the inhibition was enhanced by the adenylate cyclase activator forskolin and the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine in proportion to increases in intracellular cAMP levels, and the inhibition was abolished by selective inhibition of cAMP-dependent protein kinase. If confirmed in clinical trials, this antiproliferative action of vasoactive intestinal peptide may offer a new and promising means of suppressing SCLC in human subjects, without the toxic side effects of chemotherapeutic agents.

  14. hPSC-derived lung and intestinal organoids as models of human fetal tissue

    PubMed Central

    Aurora, Megan; Spence, Jason R.

    2016-01-01

    In vitro human pluripotent stem cell (hPSC) derived tissues are excellent models to study certain aspects of normal human development. Current research in the field of hPSC derived tissues reveals these models to be inherently fetal-like on both a morphological and gene expression level. In this review we briefly discuss current methods for differentiating lung and intestinal tissue from hPSCs into individual 3-dimensional units called organoids. We discuss how these methods mirror what is known about in vivo signaling pathways of the developing embryo. Additionally, we will review how the inherent immaturity of these models lends them to be particularly valuable in the study of immature human tissues in the clinical setting of premature birth. Human lung organoids (HLOs) and human intestinal organoids (HIOs) not only model normal development, but can also be utilized to study several important diseases of prematurity such as respiratory distress syndrome (RDS), bronchopulmonary dysplasia (BPD), and necrotizing enterocolitis (NEC). PMID:27287882

  15. Intestinal epithelial culture under an air-liquid interface: a tool for studying human and mouse esophagi.

    PubMed

    Yokobori, T; Suzuki, S; Miyazaki, T; Sohda, M; Sakai, M; Tanaka, N; Ozawa, D; Hara, K; Honjo, H; Altan, B; Fukuchi, M; Ishii, H; Iwatsuki, M; Sugimachi, K; Sudo, T; Iwaya, T; Nishida, N; Mimori, K; Kuwano, H; Mori, M

    2016-10-01

    This study investigated whether an intestinal epithelial culture method can be applied to mouse and human esophageal cultures. The esophagi harvested from 1-day-old mice and adult humans were maintained in collagen gels. A commercially available culture medium for human embryonic stem cells was used for the human esophageal culture. We discovered that the intestinal epithelial culture method can be successfully applied to both mouse and human esophageal cultures. The long-term cultured esophageal organoids were rod-like luminal structures lined with myofibroblasts. We discovered that regeneration of the esophageal mucosal surface can be almost completely achieved in vitro, and the advantage of this method is that organoid cultures may be generated using host-derived fibroblasts as a niche. This method is a promising tool for mouse and human research in intestinal biology, carcinogenesis, and regenerative medicine.

  16. Natural Pig Plasma Immunoglobulins Have Anti-Bacterial Effects: Potential for Use as Feed Supplement for Treatment of Intestinal Infections in Pigs

    PubMed Central

    Hedegaard, Chris J.; Strube, Mikael L.; Hansen, Marie B.; Lindved, Bodil K.; Lihme, Allan; Boye, Mette; Heegaard, Peter M. H.

    2016-01-01

    There is an increasing demand for non-antibiotics solutions to control infectious disease in intensive pig production. Here, one such alternative, namely pig antibodies purified from slaughterhouse blood was investigated in order to elucidate its potential usability to control post-weaning diarrhoea (PWD), which is one of the top indications for antibiotics usage in the pig production. A very cost-efficient and rapid one-step expanded bed adsorption (EBA) chromatography procedure was used to purify pig immunoglobulin G from slaughterhouse pig plasma (more than 100 litres), resulting in >85% pure pig IgG (ppIgG). The ppIgG thus comprised natural pig immunoglobulins and was subsequently shown to contain activity towards four pig-relevant bacterial strains (three different types of Escherichia coli and one type of Salmonella enterica) but not towards a fish pathogen (Yersinia ruckeri), and was demonstrated to inhibit the binding of the four pig relevant bacteria to a pig intestinal cell line (IPEC-J2). Finally it was demonstrated in an in vivo weaning piglet model for intestinal colonization with an E. coli F4+ challenge strain that ppIgG given in the feed significantly reduced shedding of the challenge strain, reduced the proportion of the bacterial family Enterobacteriaceae, increased the proportion of families Enterococcoceae and Streptococcaceae and generally increased ileal microbiota diversity. Conclusively, our data support the idea that natural IgG directly purified from pig plasma and given as a feed supplement can be used in modern swine production as an efficient and cost-effective means for reducing both occurrence of PWD and antibiotics usage and with a potential for the prevention and treatment of other intestinal infectious diseases even if the causative agent might not be known. PMID:26824607

  17. Natural Pig Plasma Immunoglobulins Have Anti-Bacterial Effects: Potential for Use as Feed Supplement for Treatment of Intestinal Infections in Pigs.

    PubMed

    Hedegaard, Chris J; Strube, Mikael L; Hansen, Marie B; Lindved, Bodil K; Lihme, Allan; Boye, Mette; Heegaard, Peter M H

    2016-01-01

    There is an increasing demand for non-antibiotics solutions to control infectious disease in intensive pig production. Here, one such alternative, namely pig antibodies purified from slaughterhouse blood was investigated in order to elucidate its potential usability to control post-weaning diarrhoea (PWD), which is one of the top indications for antibiotics usage in the pig production. A very cost-efficient and rapid one-step expanded bed adsorption (EBA) chromatography procedure was used to purify pig immunoglobulin G from slaughterhouse pig plasma (more than 100 litres), resulting in >85% pure pig IgG (ppIgG). The ppIgG thus comprised natural pig immunoglobulins and was subsequently shown to contain activity towards four pig-relevant bacterial strains (three different types of Escherichia coli and one type of Salmonella enterica) but not towards a fish pathogen (Yersinia ruckeri), and was demonstrated to inhibit the binding of the four pig relevant bacteria to a pig intestinal cell line (IPEC-J2). Finally it was demonstrated in an in vivo weaning piglet model for intestinal colonization with an E. coli F4+ challenge strain that ppIgG given in the feed significantly reduced shedding of the challenge strain, reduced the proportion of the bacterial family Enterobacteriaceae, increased the proportion of families Enterococcoceae and Streptococcaceae and generally increased ileal microbiota diversity. Conclusively, our data support the idea that natural IgG directly purified from pig plasma and given as a feed supplement can be used in modern swine production as an efficient and cost-effective means for reducing both occurrence of PWD and antibiotics usage and with a potential for the prevention and treatment of other intestinal infectious diseases even if the causative agent might not be known.

  18. Interleukin 2 modulates ion secretion and cell proliferation in cultured human small intestinal enterocytes

    PubMed Central

    O'Loughlin, E; Pang, G; Noltorp, R; Koina, C; Batey, R; Clancy, R

    2001-01-01

    AIMS—To determine if interleukin 2 (IL-2) alters epithelial transport and barrier function in cultured human small intestinal enterocytes.
METHODS—Confluent monolayers of small intestinal cells derived from duodenal biopsies were treated with IL-2 0.2-50 U/ml for 24 hours prior to study. Transport measurements were performed under short circuited conditions in Ussing chambers, with and without the secretagogues forskolin and 3-isobutyl-1-methyl xanthine (IBMX). Serosal to mucosal flux of 3[H] mannitol (permeability) and 3[H] thymidine uptake (proliferation) were measured. IL-2 receptor and cystic fibrosis transmembrane conductance regulator (CFTR) mRNA were identified using reverse transcription-polymerase chain reaction (RT-PCR).
RESULTS—IL-2 did not alter baseline electrical parameters but caused a significant increase in cAMP dependent chloride secretion. The effect was mediated by the IL-2 receptor and paralleled a rapid increase in tyrosine phosphorylation, janus kinase 1, and signal transducers and activators of transcription (STATs) 1, 3, and 5. IL-2 significantly increased proliferation but at a lower dose than observed for enhanced secretion but did not alter permeability. IL-2 receptor β and γc chains and CFTR mRNA were identified by RT-PCR.
CONCLUSIONS—IL-2 treatment enhances cAMP stimulated chloride secretion and cellular proliferation in a human small intestinal cell line expressing a functional IL-2 receptor.


Keywords: interleukin 2; ion secretion; cell proliferation; enterocytes; small intestine PMID:11600465

  19. Lysophosphatidylcholine enhances carotenoid uptake from mixed micelles by Caco-2 human intestinal cells.

    PubMed

    Sugawara, T; Kushiro, M; Zhang, H; Nara, E; Ono, H; Nagao, A

    2001-11-01

    Despite the interest in the beneficial roles of dietary carotenoids in human health, little is known about their solubilization from foods to mixed bile micelles during digestion and the intestinal uptake from the micelles. We investigated the absorption of carotenoids solubilized in mixed micelles by differentiated Caco-2 human intestinal cells, which is a useful model for studying the absorption of dietary compounds by intestinal cells. The micelles were composed of 1 micromol/L carotenoids, 2 mmol/L sodium taurocholate, 100 micromol/L monoacylglycerol, 33.3 micromol/L fatty acid and phospholipid (0-200 micromol/L). The phospholipid content of micelles had profound effects on the cellular uptake of carotenoids. Uptake of micellar beta-carotene and lutein was greatly suppressed by phosphatidylcholine (PC) in a dose-dependent manner, whereas lysophosphatidylcholine (lysoPC), the lipolysis product of PC by phospholipase A2 (PLA2), markedly enhanced both beta-carotene and lutein uptake. The addition of PLA2 from porcine pancreas to the medium also enhanced the uptake of carotenoids from micelles containing PC. Caco-2 cells could take up 15 dietary carotenoids, including epoxy carotenoids, such as violaxanthin, neoxanthin and fucoxanthin, from micellar carotenoids, and the uptakes showed a linear correlation with their lipophilicity, defined as the distribution coefficient in 1-octanol/water (log P(ow)). These results suggest that pancreatic PLA2 and lysoPC are important in regulating the absorption of carotenoids in the digestive tract and support a simple diffusion mechanism for carotenoid absorption by the intestinal epithelium.

  20. Tea Catechin Auto-oxidation Dimers are Accumulated and Retained by Caco-2 Human Intestinal Cells

    PubMed Central

    Neilson, Andrew P.; Song, Brian J.; Sapper, Teryn N.; Bomser, Joshua A.; Ferruzzi, Mario G.

    2010-01-01

    Despite the presence of bioactive catechin B-ring auto-oxidation dimers in tea, little is known regarding their absorption in humans. Our hypothesis for this research is that catechin auto-oxidation dimers are present in teas and are absorbable by human intestinal epithelial cells. Dimers [theasinensins (THSNs) and P-2 analogs) were quantified in commercial teas by HPLC-MS. (−)-Epigallocatechin (EGC) and (−)-epigallocatechin gallate (EGCG) homodimers were present at 10–43 and 0–62 µmol/g leaf, respectively. EGC-EGCG heterodimers were present at 0–79 µmol/g. The potential intestinal absorption of these dimers was assessed using Caco-2 intestinal cells. Catechin monomers and dimers were detected in cells exposed to media containing monomers and preformed dimers. Accumulation of dimers was significantly greater than monomers from test media. Three h accumulation of EGC and EGCG was 0.19– 0.55% and 1.24–1.35% respectively. Comparatively, 3h accumulation of the EGC P-2 analog, and THSNs C/E was 0.89 ± 0.28% and 1.53 ± 0.36%. Accumulation of P-2, and THSNs A/D was 6.93 ± 2.1%, and 10.1 ± 3.6%. EGCG-EGC heterodimer P-2 analog, and THSN B 3h accumulation was 4.87 ± 2.2%, and 4.65 ± 2.8% respectively. One h retention of P-2, and THSNs A/D was 171 ± 22%, and 29.6 ± 9.3% of accumulated amount suggesting intracellular oxidative conversion of THSNs to P-2. These data suggest that catechin dimers present in the gut lumen may be readily absorbed by intestinal epithelium. PMID:20579525

  1. Biotransformation and metabolic profile of buddleoside with human intestinal microflora by ultrahigh-performance liquid chromatography coupled to hybrid linear ion trap/orbitrap mass spectrometer.

    PubMed

    Tao, Jin-Hua; Duan, Jin-Ao; Jiang, Shu; Qian, Yi-Yun; Qian, Da-Wei

    2016-07-01

    Buddleoside (also known as linarin) as the major flavonoid in Chrysanthemum morifolium Ramat., has been reported to possess a wide range of pharmacological activities. The human intestinal microbiota might have an important impact on drug metabolism and ultimately on the drug oral bioavailability. However, the interaction of the buddleoside with human intestinal bacteria remains unknown. In this study, the conversion of buddleoside by different bacteria from human feces was firstly investigated. A reliable, sensitive and rapid analytical method, ultra performance liquid chromatography was established and successfully applied to investigate the metabolites and metabolic profile of buddleoside by human intestinal bacteria. Among the isolated bacteria, four strains including Escherichia sp. 4, Escherichia sp. 34, Enterococcus sp. 45 and Bacillus sp. 46 showed more powerful conversion capability. Based on the accurate mass data and the characteristic MS(n) product ions, the parent and six metabolites were detected and tentatively identified compared with blank samples. The metabolites were produced by four main metabolic pathways including deglycosylation, acetylation, methylation and hydroxylation. Buddleoside could be firstly converted to its aglycon acacetin (M2) by the majority of the isolated intestinal bacteria. Subsequently, M2 was further metabolize to its methylated (M3), acetylated (M4), hydroxylated (M5) and hydrogenated product (M6). However, acacetin-7-glucosid (M1) was obtained only from the minor bacterial samples like Bacillus sp. 46. To further explain the metabolism of buddleoside, the β-d-glucosidase and α-l-rhamnosidase activities of four strains were analyzed. Bacillus sp. 46 could only produce α-l-rhamnosidase, while the other three strains showed two kinds of enzyme activities. Furthermore, the activities of α-l-rhamnosidase and β-d-glucosidase reached the highest level at 12-18h and 10-12h, respectively. The metabolic routes and metabolites

  2. Intestinal Commitment and Maturation of Human Pluripotent Stem Cells Is Independent of Exogenous FGF4 and R-spondin1

    PubMed Central

    Tamminen, Kaisa; Balboa, Diego; Toivonen, Sanna; Pakarinen, Mikko P.; Wiener, Zoltan; Alitalo, Kari; Otonkoski, Timo

    2015-01-01

    Wnt/beta-catenin signaling plays a central role in guiding the differentiation of the posterior parts of the primitive gut tube into intestinal structures in vivo and some studies suggest that FGF4 is another crucial factor for intestinal development. The aim of this study was to define the effects of Wnt and FGF4 on intestinal commitment in vitro by establishing conditions for differentiation of human pluripotent stem cells (hPSC) into posterior endoderm (hindgut) and further to self-renewing intestinal-like organoids. The most prominent induction of the well-established intestinal marker gene CDX2 was achieved when hPSC-derived definitive endoderm cells were treated with Wnt agonist molecule CHIR99021 during differentiation to hindgut. FGF4 was found to be dispensable during intestinal commitment, but it had an early role in repressing development towards the hepatic lineage. When hindgut stage cells were further cultured in 3D, they formed self-renewing organoid structures containing all major intestinal cell types even without exogenous R-spondin1 (RSPO1), a crucial factor for the culture of epithelial organoids derived from adult intestine. This may be explained by the presence of a mesenchymal compartment in the hPSC-derived organoids. Addition of WNT3A increased the expression of the Paneth cell marker Lysozyme in hPSC-derived organoid cultures, whereas FGF4 inhibited both the formation and maturation of intestinal-like organoids. Similar hindgut and organoid cultures were established from human induced pluripotent stem cells, implying that this approach can be used to create patient-specific intestinal tissue models for disease modeling in vitro. PMID:26230325

  3. Intestinal Commitment and Maturation of Human Pluripotent Stem Cells Is Independent of Exogenous FGF4 and R-spondin1.

    PubMed

    Tamminen, Kaisa; Balboa, Diego; Toivonen, Sanna; Pakarinen, Mikko P; Wiener, Zoltan; Alitalo, Kari; Otonkoski, Timo

    2015-01-01

    Wnt/beta-catenin signaling plays a central role in guiding the differentiation of the posterior parts of the primitive gut tube into intestinal structures in vivo and some studies suggest that FGF4 is another crucial factor for intestinal development. The aim of this study was to define the effects of Wnt and FGF4 on intestinal commitment in vitro by establishing conditions for differentiation of human pluripotent stem cells (hPSC) into posterior endoderm (hindgut) and further to self-renewing intestinal-like organoids. The most prominent induction of the well-established intestinal marker gene CDX2 was achieved when hPSC-derived definitive endoderm cells were treated with Wnt agonist molecule CHIR99021 during differentiation to hindgut. FGF4 was found to be dispensable during intestinal commitment, but it had an early role in repressing development towards the hepatic lineage. When hindgut stage cells were further cultured in 3D, they formed self-renewing organoid structures containing all major intestinal cell types even without exogenous R-spondin1 (RSPO1), a crucial factor for the culture of epithelial organoids derived from adult intestine. This may be explained by the presence of a mesenchymal compartment in the hPSC-derived organoids. Addition of WNT3A increased the expression of the Paneth cell marker Lysozyme in hPSC-derived organoid cultures, whereas FGF4 inhibited both the formation and maturation of intestinal-like organoids. Similar hindgut and organoid cultures were established from human induced pluripotent stem cells, implying that this approach can be used to create patient-specific intestinal tissue models for disease modeling in vitro.

  4. Intestinal Capillariasis

    DTIC Science & Technology

    1987-12-01

    bhIll inenais, the tiny nematode causing Intestinal capillariasis In humans, Is a Iunique parasite. It is one of the newest parasites that has been...Capillariaphilippinensis, the tiny nematode causing intestinal capillariasis in humans, is a unique parasite. It is one of the newest parasites that has been shown to...stichocytes surrounding the oesophagus. The posterior half of the nematode is wider than the anterior half and contains the digestive tract and the

  5. Mast cell expression of the serotonin1A receptor in guinea pig and human intestine.

    PubMed

    Wang, Guo-Du; Wang, Xi-Yu; Zou, Fei; Qu, Meihua; Liu, Sumei; Fei, Guijun; Xia, Yun; Needleman, Bradley J; Mikami, Dean J; Wood, Jackie D

    2013-05-15

    Serotonin [5-hydroxytryptamine (5-HT)] is released from enterochromaffin cells in the mucosa of the small intestine. We tested a hypothesis that elevation of 5-HT in the environment of enteric mast cells might degranulate the mast cells and release mediators that become paracrine signals to the enteric nervous system, spinal afferents, and secretory glands. Western blotting, immunofluorescence, ELISA, and pharmacological analysis were used to study expression of 5-HT receptors by mast cells in the small intestine and action of 5-HT to degranulate the mast cells and release histamine in guinea pig small intestine and segments of human jejunum discarded during Roux-en-Y gastric bypass surgeries. Mast cells in human and guinea pig preparations expressed the 5-HT1A receptor. ELISA detected spontaneous release of histamine in guinea pig and human preparations. The selective 5-HT1A receptor agonist 8-hydroxy-PIPAT evoked release of histamine. A selective 5-HT1A receptor antagonist, WAY-100135, suppressed stimulation of histamine release by 5-HT or 8-hydroxy-PIPAT. Mast cell-stabilizing drugs, doxantrazole and cromolyn sodium, suppressed the release of histamine evoked by 5-HT or 8-hydroxy-PIPAT in guinea pig and human preparations. Our results support the hypothesis that serotonergic degranulation of enteric mast cells and release of preformed mediators, including histamine, are mediated by the 5-HT1A serotonergic receptor. Association of 5-HT with the pathophysiology of functional gastrointestinal disorders (e.g., irritable bowel syndrome) underlies a question of whether selective 5-HT1A receptor antagonists might have therapeutic application in disorders of this nature.

  6. Human intestinal parasites in non-biting synanthropic flies in Ogun State, Nigeria.

    PubMed

    Adenusi, Adedotun Adesegun; Adewoga, Thomas O Sunday

    2013-01-01

    Filth-feeding and breeding, non-biting synanthropic flies have been incriminated in the dissemination of human enteropathogens in the environment. This study determined the species of non-biting synanthropic flies associated with four filthy sites in Ilishan, Ogun State, southwest Nigeria, and assessed their potentials for mechanical transmission of human intestinal parasites. 7190 flies identified as Musca domestica (33.94%), Chrysomya megacephala (26.01%), Musca sorbens (23.23%), Lucilia cuprina (8.76%), Calliphora vicina (4.59%), Sarcophaga sp. (2.78%) and Fannia scalaris (0.70%) were examined for human intestinal parasites by the formol-ether concentration and modified Ziehl-Neelsen techniques. Eggs of the following parasites: Ascaris lumbricoides (34.08%), Trichuris trichiura (25.87%), hookworms (20.45%), Taenia sp. (2.36%), Hymenolepis nana (1.11%), Enterobius vermicularis (0.56%), Strongyloides stercoralis (larvae; 3.89%) and cysts of Entamoeba histolytica/dispar (27.26%), Entamoeba coli (22.67%), Giardia lamblia (3.34%) and Cryptosporidium sp. (1.81%) were isolated from the body surfaces and or gut contents of 75.24% of 719 pooled fly batches. The helminths A. lumbricoides and T. trichiura and the protozoans, E. histolytica/dispar and E. coli were the dominant parasites detected, both on body surfaces and in the gut contents of flies. C. megacephala was the highest carrier of parasites (diversity and number). More parasites were isolated from the gut than from body surfaces (P < 0.05). Flies from soiled ground often carried more parasites than those from abattoir, garbage or open-air market. Synanthropic fly species identified in this study can be of potential epidemiological importance as mechanical transmitters of human intestinal parasites acquired naturally from filth and carried on their body surfaces and or in the gut, because of their vagility and feeding mechanisms.

  7. Human Milk Oligosaccharides in Premature Infants: Absorption, Excretion and Influence on the Intestinal Microbiota

    PubMed Central

    Underwood, Mark A.; Gaerlan, Stephanie; De Leoz, M. Lorna A.; Dimapasoc, Lauren; Kalanetra, Karen M.; Lemay, Danielle G.; German, J. Bruce; Mills, David A.; Lebrilla, Carlito B.

    2015-01-01

    Background Human milk oligosaccharides (HMOs) shape the intestinal microbiota in term infants. In premature infants, alterations in the intestinal microbiota (dysbiosis) are associated with risk of necrotizing enterocolitis and sepsis and the influence of HMOs on the microbiota is unclear. Methods Milk, urine, and stool specimens from 14 mother-premature infant dyads were investigated by mass spectrometry for HMO composition. The stools were analyzed by next-generation sequencing (NGS) to complement a previous analysis. Results Percentages of fucosylated and sialylated HMOs were highly variable between individuals but similar in urine, feces and milk within dyads. Differences in urine and fecal HMO composition suggest variability in absorption. Secretor status of the mother correlated with the urine and fecal content of specific HMO structures. Trends toward higher levels of Proteobacteria and lower levels of Firmicutes, were noted in premature infants of non-secretor mothers. Specific HMO structures in the milk, urine and feces were associated with alterations in fecal Proteobacteria and Firmicutes. Conclusion HMOs may influence the intestinal microbiota in premature infants. Specific HMOs, for example those associated with secretor mothers, may have a protective effect by decreasing pathogens associated with sepsis and necrotizing enterocolitis while other HMOs may increase dysbiosis in this population. PMID:26322410

  8. Infection with fully mature Corynosoma cf. validum causes ulcers in the human small intestine.

    PubMed

    Takahashi, Keitaro; Ito, Takahiro; Sato, Tomonobu; Goto, Mitsuru; Kawamoto, Toru; Fujinaga, Akihiro; Yanagawa, Nobuyuki; Saito, Yoshinori; Nakao, Minoru; Hasegawa, Hideo; Fujiya, Mikihiro

    2016-06-01

    Corynosoma is a parasite that can normally be found in the intestinal tract of fish-eating mammals, particularly in seals and birds. The present case proposed that Corynosoma could attain full maturity in the human intestine. A 70-year-old female complained of abdominal pain. A computed tomography (CT) scan revealed a swelling of the intraperitoneal lymph nodes with no responsible lesion. Video capsule endoscopy and double-balloon endoscopy detected several ulcerations and one parasite in the ileum, which was tightly attached at the bottom of the ulcerations. The parasite was cylindrical and measured approximately 10 mm (long) x 3 mm (wide). Pathologically, the worm had a four-layered body wall and contained embryonated eggs. The sequences of the parasite-derived nuclear ribosomal DNA fragment and mitochondrial DNA fragment of cox1 were almost identical to those of Corynosoma validum. The patient's abdominal pain immediately improved after the administration of pyrantel pamoate (1,500 mg). Corynosoma was possibly the responsible disease in a patient who complained of abdominal pain and in whom no responsible lesion was detected by CT, gastroduodenoscopy or colonoscopy. Examinations of the small intestines should be aggressively performed in such cases.

  9. N-nitrosation of medicinal drugs catalysed by bacteria from human saliva and gastro-intestinal tract, including Helicobacter pylori.

    PubMed

    Ziebarth, D; Spiegelhalder, B; Bartsch, H

    1997-02-01

    Micro-organisms commonly present in human saliva and three DSM strains (Helicobacter pylori, Campylobacter jejuni and Neisseria cinerea), which can be isolated from the human gastro-intestinal tract, were assayed in vitro for their capacity to catalyse N-nitrosation of a series of medicinal drugs and other compounds. Following incubation at pH 7.2 in the presence of nitrate (or nitrite) for up to 24 (48) h, the yield of N-nitroso compounds (NOC) was quantified by HPLC equipped with a post-column derivatization device, allowing the sensitive detection of acid-labile and acid-stable NOC. Eleven out of the 23 test compounds underwent bacteria-catalysed nitrosation by salivary bacteria, the yield of the respective nitrosation products varying 800-fold. 4-(Methylamino)antipyrine exhibited the highest rate of nitrosation, followed by dichlofenac > metamizole > piperazine > five other drugs, whilst L-proline and L-thioproline had the lowest nitrosation rate. Ten drugs including aminophenazone, cimetidine and nicotine, did not inhibit bacterial growth, allowing transitory nitrite to be formed, but no N-nitroso derivatives were detected. Three drugs inhibited the proliferation of bacteria and neither nitrite nor any NOC were formed. Using metamizole as an easily nitrosatable precursor, two strains, Campylobacter jejuni and Helicobacter pylori, were shown to catalyse nitrosation in the presence of nitrite at pH 7.2. As compared to Neisseria cinerea used as a nitrosation-proficient control strain, H. pylori was 30-100 times less effective, whilst C. jejuni had intermediary activity. The results of our sensitive nitrosation assay further confirm that bacteria isolated from human sources, possessing nitrate reductase and/or nitrosating enzymes such as cytochrome cd1-nitrite reductase (Calmels et al., Carcinogenesis, 17, 533-536, 1996), can contribute to intragastric nitrosamine formation in the anacidic stomach when nitrosatable precursors from exogenous and endogenous sources

  10. The Impact of Handling and Storage of Human Fecal Material on Bacterial Activity.

    PubMed

    Karatza, Eleni; Vertzoni, Maria; Muenster, Uwe; Reppas, Christos

    2016-11-01

    Fecal material prepared from human stools is frequently used for the assessment of bacterial degradation of active pharmaceutical ingredients as relevant data are useful for evaluating the potential for colonic drug delivery. The impact of handling and storage of human fecal material on bacterial activity was assessed by evaluating the degradation characteristics of metronidazole and olsalazine. Multiple freeze (-70°C)-thaw cycles should be avoided. Incubation of frozen material for about 2 h in the anaerobic workstation ensures regeneration of the highest possible bacterial activity. Material could be stored at -70°C for at least 12 months.

  11. Functional modulation of human intestinal epithelial cell responses by Bifidobacterium infantis and Lactobacillus salivarius

    PubMed Central

    O'Hara, Ann M; O'Regan, Padraig; Fanning, Áine; O'Mahony, Caitlin; MacSharry, John; Lyons, Anne; Bienenstock, John; O'Mahony, Liam; Shanahan, Fergus

    2006-01-01

    Intestinal epithelial cells (IECs) and dendritic cells (DCs) play a pivotal role in antigen sampling and the maintenance of gut homeostasis. However, the interaction of commensal bacteria with the intestinal surface remains incompletely understood. Here we investigated immune cell responses to commensal and pathogenic bacteria. HT-29 human IECs were incubated with Bifidobacterium infantis 35624, Lactobacillus salivarius UCC118 or Salmonella typhimurium UK1 for varying times, or were pretreated with a probiotic for 2 hr prior to stimulation with S. typhimurium or flagellin. Gene arrays were used to examine inflammatory gene expression. Nuclear factor (NF)-κB activation, interleukin (IL)-8 secretion, pathogen adherence to IECs, and mucin-3 (MUC3) and E-cadherin gene expression were assayed by TransAM assay, enzyme-linked immunosorbent assay (ELISA), fluorescence, and real-time reverse transcriptase–polymerase chain reaction (RT-PCR), respectively. IL-10 and tumour necrosis factor (TNF)-α secretion by bacteria-treated peripheral blood-derived DCs were measured using ELISA. S. typhimurium increased expression of 36 of the 847 immune-related genes assayed, including NF-κB and IL-8. The commensal bacteria did not alter expression levels of any of the 847 genes. However, B. infantis and L. salivarius attenuated both IL-8 secretion at baseline and S. typhimurium-induced pro-inflammatory responses. B. infantis also limited flagellin-induced IL-8 protein secretion. The commensal bacteria did not increase MUC3 or E-cadherin expression, or interfere with pathogen binding to HT-29 cells, but they did stimulate IL-10 and TNF-α secretion by DCs. The data demonstrate that, although the intestinal epithelium is immunologically quiescent when it encounters B. infantis or L. salivarius, these commensal bacteria exert immunomodulatory effects on intestinal immune cells that mediate host responses to flagellin and enteric pathogens. PMID:16771855

  12. Human milk oligosaccharides: evolution, structures and bioselectivity as substrates for intestinal bacteria.

    PubMed

    German, J Bruce; Freeman, Samara L; Lebrilla, Carlito B; Mills, David A

    2008-01-01

    Human milk contains a high concentration of diverse soluble oligosaccharides, carbohydrate polymers formed from a small number of monosaccharides. Novel methods combining liquid chromatography with high resolution mass spectrometry have identified approximately 200 unique oligosaccharides structures varying from 3 to 22 sugars. The increasing complexity of oligosaccharides follows the general pattern of mammalian evolution though the concentration and diversity of these structures in homo sapiens are strikingly. There is also diversity among human mothers in oligosaccharides. Milks from randomly selected mothers contain as few as 23 and as many as 130 different oligosaccharides. The functional implications of this diversity are not known. Despite the role of milk to serve as a sole nutrient source for mammalian infants, the oligosaccharides in milk are not digestible by human infants. This apparent paradox raises questions about the functions of these oligosaccharides and how their diverse molecular structures affect their functions. The nutritional function most attributed to milk oligosaccharides is to serve as prebiotics - a form of indigestible carbohydrate that is selectively fermented by desirable gut microflora. This function was tested by purifying human milk oligosaccharides and providing these as the sole carbon source to various intestinal bacteria. Indeed, the selectively of providing the complex mixture of oligosaccharides pooled from human milk samples is remarkable. Among a variety of Bifidobacteria tested only Bifidobacteria longum biovar infantis was able to grow extensively on human milk oligosaccharides as sole carbon source. The genomic sequence of this strain revealed approximately 700 genes that are unique to infantis, including a variety of co-regulated glycosidases, relative to other Bifidobacteria, implying a co-evolution of human milk oligosaccharides and the genetic capability of select intestinal bacteria to utilize them. The goal of

  13. Metabolism of Kaempferia parviflora polymethoxyflavones by human intestinal bacterium Bautia sp. MRG-PMF1.

    PubMed

    Kim, Mihyang; Kim, Nayoung; Han, Jaehong

    2014-12-24

    Poylmethoxyflavones (PMFs) are major bioactive flavonoids, which exhibit various biological activities, such as anticancer effects. The biotransformation of PMFs and characterization of a PMF-metabolizing human intestinal bacterium were studied herein for the first time. Hydrolysis of aryl methyl ether functional groups by human fecal samples was observed from the bioconversion of various PMFs. Activity-guided screening for PMF-metabolizing intestinal bacteria under anaerobic conditions resulted in the isolation of a strict anaerobic bacterium, which was identified as Blautia sp. MRG-PMF1. The isolated MRG-PMF1 was able to metabolize various PMFs to the corresponding demethylated flavones. The microbial conversion of bioactive 5,7-dimethoxyflavone (5,7-DMF) and 5,7,4'-trimethoxyflavone (5,7,4'-TMF) was studied in detail. 5,7-DMF and 5,7,4'-TMF were completely metabolized to 5,7-dihydroxyflavone (chrysin) and 5,7,4'-trihydroxyflavone (apigenin), respectively. From a kinetics study, the methoxy group on the flavone C-7 position was found to be preferentially hydrolyzed. 5-Methoxychrysin, the intermediate of 5,7-DMF metabolism by Blautia sp. MRG-PMF1, was isolated and characterized by nuclear magnetic resonance spectroscopy. Apigenin was produced from the sequential demethylation of 5,7,4'-TMF, via 5,4'-dimethoxy-7-hydroxyflavone and 7,4'-dihydroxy-5-methoxyflavone (thevetiaflavone). Not only demethylation activity but also deglycosylation activity was exhibited by Blautia sp. MRG-PMF1, and various flavonoids, including isoflavones, flavones, and flavanones, were found to be metabolized to the corresponding aglycones. The unprecedented PMF demethylation activity of Blautia sp. MRG-PMF1 will expand our understanding of flavonoid metabolism in the human intestine and lead to novel bioactive compounds.

  14. Combined Effects of Lipophilic Phycotoxins (Okadaic Acid, Azapsiracid-1 and Yessotoxin) on Human Intestinal Cells Models

    PubMed Central

    Ferron, Pierre-Jean; Dumazeau, Kevin; Beaulieu, Jean-François; Le Hégarat, Ludovic; Fessard, Valérie

    2016-01-01

    Phycotoxins are monitored in seafood because they can cause food poisonings in humans. Phycotoxins do not only occur singly but also as mixtures in shellfish. The aim of this study was to evaluate the in vitro toxic interactions of binary combinations of three lipophilic phycotoxins commonly found in Europe (okadaic acid (OA), yessotoxin (YTX) and azaspiracid-1 (AZA-1)) using the neutral red uptake assay on two human intestinal cell models, Caco-2 and the human intestinal epithelial crypt-like cells (HIEC). Based on the cytotoxicity of individual toxins, we studied the interactions between toxins in binary mixtures using the combination index-isobologram equation, a method widely used in pharmacology to study drug interactions. This method quantitatively classifies interactions between toxins in mixtures as synergistic, additive or antagonistic. AZA-1/OA, and YTX/OA mixtures showed increasing antagonism with increasing toxin concentrations. In contrast, the AZA-1/YTX mixture showed increasing synergism with increasing concentrations, especially for mixtures with high YTX concentrations. These results highlight the hazard potency of AZA-1/YTX mixtures with regard to seafood intoxication. PMID:26907345

  15. Transepithelial transport of ambroxol hydrochloride across human intestinal Caco-2 cell monolayers.

    PubMed

    Stetinová, Vera; Smetanová, Libuse; Kholová, Dagmar; Svoboda, Zbynek; Kvetina, Jaroslav

    2009-09-01

    This study aimed i) to characterize the transepithelial transport of the mucolytic agent ambroxol hydrochloride across the intestinal barrier, ii) to classify the ambroxol according to Biopharmaceutics Classification System (BCS) and iii) to predict ambroxol absorption in humans. Transport of ambroxol (100, 300 and 1000 micromol/l) was studied in a human colon carcinoma cell line Caco-2 in apical to basolateral and basolateral to apical direction, under iso-pH 7.4 and pH-gradient (6 vs. 7.4) conditions. The relative contribution of the paracellular route was estimated using Ca2+-free transport medium. Ambroxol samples from receiver compartments were analysed by HPLC with UV detection (242 nm). Results showed that ambroxol transport is linear with time, pH-dependent and direction-independent, displays non-saturable (first-order) kinetics. Thus, the transport seems to be transcellular mediated by passive diffusion. Estimated high solubility and high permeability (P(app) = 45 x 10(-6) cm/s) of ambroxol rank it among well absorbed compounds and class I of BCS. It can be expected that the oral dose fraction of ambroxol absorbed in human intestine is high.

  16. Initiation of an inflammatory response in resident intestinal lamina propria cells -use of a human organ culture model.

    PubMed

    Schröder-Braunstein, Jutta; Gras, Judith; Brors, Benedikt; Schwarz, Sonja; Szikszai, Timea; Lasitschka, Felix; Wabnitz, Guido; Heidtmann, Antje; Lee, Young-Seon; Schiessling, Serin; Leowardi, Christine; Al-Saeedi, Mohammed; Ulrich, Alexis; Engelke, Antonia; Winter, Johannes; Samstag, Yvonne; Giese, Thomas; Meuer, Stefan

    2014-01-01

    Resident human lamina propria immune cells serve as powerful effectors in host defense. Molecular events associated with the initiation of an intestinal inflammatory response in these cells are largely unknown. Here, we aimed to characterize phenotypic and functional changes induced in these cells at the onset of intestinal inflammation using a human intestinal organ culture model. In this model, healthy human colonic mucosa was depleted of epithelial cells by EDTA treatment. Following loss of the epithelial layer, expression of the inflammatory mediators IL1B, IL6, IL8, IL23A, TNFA, CXCL2, and the surface receptors CD14, TLR2, CD86, CD54 was rapidly induced in resident lamina propria cells in situ as determined by qRT-PCR and immunohistology. Gene microarray analysis of lamina propria cells obtained by laser-capture microdissection provided an overview of global changes in gene expression occurring during the initiation of an intestinal inflammatory response in these cells. Bioinformatic analysis gave insight into signalling pathways mediating this inflammatory response. Furthermore, comparison with published microarray datasets of inflamed mucosa in vivo (ulcerative colitis) revealed a significant overlap of differentially regulated genes underlining the in vivo relevance of the organ culture model. Furthermore, genes never been previously associated with intestinal inflammation were identified using this model. The organ culture model characterized may be useful to study molecular mechanisms underlying the initiation of an intestinal inflammatory response in normal mucosa as well as potential alterations of this response in inflammatory bowel disease.

  17. Transesterification of a series of 12 parabens by liver and small-intestinal microsomes of rats and humans.

    PubMed

    Fujino, Chieri; Watanabe, Yoko; Uramaru, Naoto; Kitamura, Shigeyuki

    2014-02-01

    Hydrolytic transformation of parabens (4-hydroxybenzoic acid esters; used as antibacterial agents) to 4-hydroxybenzoic acid and alcohols by tissue microsomes is well-known both in vitro and in vivo. Here, we investigated transesterification reactions of parabens catalyzed by rat and human microsomes, using a series of 12 parabens with C1-C12 alcohol side chains. Transesterification of parabens by rat liver and small-intestinal microsomes occurred in the presence of alcohols in the microsomal incubation mixture. Among the 12 parabens, propylparaben was most effectively transesterified by rat liver microsomes with methanol or ethanol, followed by butylparaben. Relatively low activity was observed with longer-side-chain parabens. In contrast, small-intestinal microsomes exhibited higher activity towards moderately long side-chain parabens, and showed the highest activity toward octylparaben. When parabens were incubated with liver or small-intestinal microsomes in the presence of C1-C12 alcohols, ethanol and decanol were most effectively transferred to parabens by rat liver microsomes and small-intestinal microsomes, respectively. Human liver and small-intestinal microsomes also exhibited significant transesterification activities with different substrate specificities, like rat microsomes. Carboxylesterase isoforms, CES1b and CES1c, and CES2, exhibited significant transesterification activity toward parabens, and showed similar substrate specificity to human liver and small-intestinal microsomes, respectively.

  18. Intestinal parasite co-infection among pulmonary tuberculosis cases without human immunodeficiency virus infection in a rural county in China.

    PubMed

    Li, Xin-Xu; Chen, Jia-Xu; Wang, Li-Xia; Tian, Li-Guang; Zhang, Yu-Ping; Dong, Shuang-Pin; Hu, Xue-Guang; Liu, Jian; Wang, Feng-Feng; Wang, Yue; Yin, Xiao-Mei; He, Li-Jun; Yan, Qiu-Ye; Zhang, Hong-Wei; Xu, Bian-Li; Zhou, Xiao-Nong

    2014-01-01

    Epidemiologic studies of co-infection with tuberculosis (TB) and intestinal parasites in humans have not been extensively investigated in China. A cross-section study was conducted in a rural county of Henan Province, China. Pulmonary TB (PTB) case-patients receiving treatment for infection with Mycobacterium tuberculosis and healthy controls matched for geographic area, age, and sex were surveyed by using questionnaires. Fecal and blood specimens were collected for detection of intestinal parasites, routine blood examination, and infection with human immunodeficiency virus. The chi-square test was used for univariate analysis and multivariate logistic regression models were used to adjust for potential confounding factors. A total of 369 persons with PTB and 366 healthy controls were included; all participants were negative for human immunodeficiency virus. The overall prevalence of intestinal parasites in persons with PTB was 14.9%, including intestinal protozoa (7.9%) and helminthes (7.6%). The infection spectrum of intestinal parasites was Entamoeba spp. (1.4%), Blastocystis hominis (6.2%), Trichomonas hominis (0.3%), Clonorchis sinensis (0.3%), Ascaris lumbricoides (0.5%), Trichuris trichiura (2.2%), and hookworm (4.6%). The prevalence of intestinal parasites showed no significant difference between persons with PTB and healthy controls after adjusting for potential confounding factors. There was no factor that affected infection rates for intestinal parasites between the two groups. Infection with intestinal parasites of persons with PTB was associated with female sex (adjusted odds ratio [AOR] = 2.05, 95% confidence interval [CI] = 1.01-4.17), body mass index ≤ 19 (AOR = 3.02, 95% CI = 1.47-6.20), and anemia (AOR = 2.43, 95% CI = 1.17-5.03). Infection of healthy controls was only associated with an annual labor time in farmlands > 2 months (AOR = 4.50, 95% CI = 2.03-10.00). In addition, there was no significant trend between rates of infection with

  19. In vitro and in vivo small intestinal metabolism of CYP3A and UGT substrates in preclinical animals species and humans: species differences.

    PubMed

    Komura, Hiroshi; Iwaki, Masahiro

    2011-11-01

    Intestinal first-pass metabolism has a great impact on the bioavailability of cytochrome P450 3A4 (CYP3A) and/or uridine 5'-diphosphate (UDP)-glucoronosyltranferase (UGT) substrates in humans. In vitro and in vivo intestinal metabolism studies are essential for clarifying pharmacokinetics in animal species and for predicting the effects of human intestinal metabolism. We review species differences in intestinal metabolism both in vitro and in vivo. Based on mRNA expression levels, the major intestinal CYP3A isoform is CYP3A4 for humans, CYP3A4 (3A8) for monkeys, CYP3A9 for rats, cyp3a13 for mice, and CYP3A12 for dogs. Additionally, the intestinal-specific UGT would be UGT1A10 for humans, UGT1A8 for monkeys, and UGT1A7 for rats. In vitro and in vivo intestinal metabolism of CYP3A substrates were larger in monkeys than in humans, although a correlation in intestinal availability between monkeys and humans has been reported. Little information is available regarding species differences in in vitro and in vivo UGT activities; however, UGT-mediated in vivo intestinal metabolism has been demonstrated for raloxifene in humans and for baicalein in rats. Further assessment of intestinal metabolism, particularly for UGT substrates, is required to clarify the entire picture of species differences.

  20. May We Strengthen the Human Natural Defenses with Bacterial Lysates?

    PubMed Central

    2010-01-01

    During the last twenty years bacterial lysates have gained a new interest and their use has obtained a progressively larger consensus in the medical practice. They are commonly used as immunomodulators, in order to up-regulate immune responses against infectious damages. As a matter of fact, the role of these lysate seems relevant in upper and lower respiratory tract infections prevention, frequently observed both in paediatric and elder ages, and which represent a relevant problem also in terms of socio-economical implications. The effects of bacterial lysates as immunostimulatory agents have become the central point of many studies. The aim of those in vivo and in vitro studies was to understand and evaluate the capacity of this kind of treatments to create a better answer of the immune system against microbial infections, eventually leading to a reduction in their number. All the in vivo and in vitro findings analyzed support the evidence that bacterial lysates are powerful inducers of a specific immune response against bacterial infections. Both in paediatric and adult clinical trials, a positive trend has been found in terms of overall reduction of infection rates and duration, beneficial effect on symptoms, reduction in antibiotics use and possibility to improve the patient's quality of life in several diseases. Further well-designed trials in terms of blinding and randomization procedures and including a higher number of patients, selected according to the disease and its severity, are needed. PMID:23282746

  1. Failure of d-psicose absorbed in the small intestine to metabolize into energy and its low large intestinal fermentability in humans.

    PubMed

    Iida, Tetsuo; Hayashi, Noriko; Yamada, Takako; Yoshikawa, Yuko; Miyazato, Shoko; Kishimoto, Yuka; Okuma, Kazuhiro; Tokuda, Masaaki; Izumori, Ken

    2010-02-01

    Experiments with rats have produced data on the metabolism and energy value of d-psicose; however, no such data have been obtained in humans. The authors assessed the availability of d-psicose absorbed in the small intestine by measuring carbohydrate energy expenditure (CEE) by indirect calorimetry. They measured the urinary excretion rate by quantifying d-psicose in urine for 48 hours. To examine d-psicose fermentation in the large intestine, the authors measured breath hydrogen gas and fermentability using 35 strains of intestinal bacteria. Six healthy subjects participated in the CEE test, and 14 participated in breath hydrogen gas and urine tests. d-Psicose fermentation subsequent to an 8-week adaptation period was also assessed by measuring hydrogen gas in 8 subjects. d-Psicose absorbed in the small intestine was not metabolized into energy, unlike glucose, because CEE did not increase within 3 hours of d-psicose ingestion (0.35 g/kg body weight [BW]). The accumulated d-psicose urinary excretion rates were around 70% for 0.34, 0.17, and 0.08 g/kg BW of ingested d-psicose. Low d-psicose fermentability was observed in intestinal bacteria and breath hydrogen gas tests, in which fructooligosaccharide (0.34, 0.17, and 0.08 g/kg BW) was used as a positive control because its available energy is known to be 8.4 kJ/g. Based on the results of the plot of breath hydrogen concentration vs calories ingested, the energy value of d-psicose was expected to be less than 1.6 kJ/g. Incremental d-psicose fermentability subsequent to an adaptation period was not observed.

  2. The identification of a bacterial strain BGI-1 isolated from the intestinal flora of Blattella germanica, and its anti-entomopathogenic fungi activity.

    PubMed

    Huang, Y H; Wang, X J; Zhang, F; Huo, X B; Fu, R S; Liu, J J; Sun, W B; Kang, D M; Jing, X

    2013-02-01

    A bacterial strain BGI-1 was isolated from the gut of German cockroaches (Blattella germanica L.) and was identified as Bacillus subtilis based on 16S rDNA sequence and morphological, physiological, and biochemical characters. The strain BGI-1 inhibited the growth of Beauveria bassiana; the diameter of the inhibition zone exceeded 30 mm. Vesicles were observed in B. bassiana hyphae on the edge of the inhibition zone. Fermentation of BGI-1 reduced the conidial germination rate by 12%. Further studies demonstrated that B. bassiana infections in German cockroaches orally treated with the extracts of BGI-1 fermentation were significantly weakened. Cumulative mortality rate was 49.5% in the treatment group at the 20 d, while that of the control group was 62.3%. The study intends to understand the relationship between the intestinal flora and the cockroach. Those microbes with anti-entomopathogenic fungi activity might contribute to resisting the infection of pathogenic fungi.

  3. Biotransformation and metabolic profile of catalpol with human intestinal microflora by ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry.

    PubMed

    Tao, Jin-hua; Zhao, Min; Wang, Dong-geng; Yang, Chi; Du, Le-Yue; Qiu, Wen-qian; Jiang, Shu

    2016-01-15

    Traditional Chinese medicine (TCM) has been used in clinical practice for thousands of years. Catalpol, an iridoid glucoside, abundantly found in the root of the common used herb medicine Rehmannia glutinosa Libosch, has been reported to show various biological effects and pharmacological activities. After oral administration, the active ingredient might have interactions with the intestinal bacteria, which could help unravel how the medicine was processed in vivo. In this work, different pure bacteria from healthy human feces were isolated and used to bioconvert catalpol. Ultra performance liquid chromatography/quadrupole-time-of-flight mass spectrometry (UPLC-Q-TOF/MS) technique combined with Metabolynx(™) software was applied to analyze catalpol metabolites. Compared with blank samples, parent compound (M0) and four metabolites (M1-M4) were detected and tentatively identified based on the characteristics of their protonated ions. The metabolites were likely to be: catalpol aglycone (M1), acetylated catalpol (M2), dimethylated and hydroxylated catalpol aglycone (M3), nitrogen-containing catalpol aglycone (M4). M1 and M4 were generated in the majority of the samples like Bacteroides sp. 45. M3 was obtained in several bacterial samples like Enterococcus sp. 8-2 and M2 was detected only in the sample of Enterococcus sp. 43-1. To our knowledge, the metabolic routes and metabolites of catalpol produced by human intestinal bacteria were all firstly reported.

  4. Characterization of Glycosaminoglycan (GAG) Sulfatases from the Human Gut Symbiont Bacteroides thetaiotaomicron Reveals the First GAG-specific Bacterial Endosulfatase*

    PubMed Central

    Ulmer, Jonathan E.; Vilén, Eric Morssing; Namburi, Ramesh Babu; Benjdia, Alhosna; Beneteau, Julie; Malleron, Annie; Bonnaffé, David; Driguez, Pierre-Alexandre; Descroix, Karine; Lassalle, Gilbert; Le Narvor, Christine; Sandström, Corine; Spillmann, Dorothe; Berteau, Olivier

    2014-01-01

    Despite the importance of the microbiota in human physiology, the molecular bases that govern the interactions between these commensal bacteria and their host remain poorly understood. We recently reported that sulfatases play a key role in the adaptation of a major human commensal bacterium, Bacteroides thetaiotaomicron, to its host (Benjdia, A., Martens, E. C., Gordon, J. I., and Berteau, O. (2011) J. Biol. Chem. 286, 25973–25982). We hypothesized that sulfatases are instrumental for this bacterium, and related Bacteroides species, to metabolize highly sulfated glycans (i.e. mucins and glycosaminoglycans (GAGs)) and to colonize the intestinal mucosal layer. Based on our previous study, we investigated 10 sulfatase genes induced in the presence of host glycans. Biochemical characterization of these potential sulfatases allowed the identification of GAG-specific sulfatases selective for the type of saccharide residue and the attachment position of the sulfate group. Although some GAG-specific bacterial sulfatase activities have been described in the literature, we report here for the first time the identity and the biochemical characterization of four GAG-specific sulfatases. Furthermore, contrary to the current paradigm, we discovered that B. thetaiotaomicron possesses an authentic GAG endosulfatase that is active at the polymer level. This type of sulfatase is the first one to be identified in a bacterium. Our study thus demonstrates that bacteria have evolved more sophisticated and diverse GAG sulfatases than anticipated and establishes how B. thetaiotaomicron, and other major human commensal bacteria, can metabolize and potentially tailor complex host glycans. PMID:25002587

  5. Study Bacteria-Host Interactions Using Intestinal Organoids.

    PubMed

    Zhang, Yong-Guo; Sun, Jun

    2016-08-19

    The intestinal epithelial cells function to gain nutrients, retain water and electrolytes, and form an efficient barrier against foreign microbes and antigens. Researchers employed cell culture lines derived from human or animal cancer cells as experimental models in vitro for understanding of intestinal infections. However, most in vitro models used to investigate interactions between bacteria and intestinal epithelial cells fail to recreate the differentiated tissue components and structure observed in the normal intestine. The in vitro analysis of host-bacteria interactions in the intestine has been hampered by a lack of suitable intestinal epithelium culture systems. Here, we present a new experimental model using an organoid culture system to study bacterial infection.

  6. Bacterial agents as a cause of infertility in humans.

    PubMed

    Ruggeri, Melania; Cannas, Sara; Cubeddu, Marina; Molicotti, Paola; Piras, Gennarina Laura; Dessole, Salvatore; Zanetti, Stefania

    2016-07-01

    Infertility is a problem affecting almost 15% of couples. There are many causes for this condition, among which urogenital bacterial infections seem to play an important role. Many studies have explained the mechanisms by which bacteria cause infertility both in men and women. Therefore we undertook this study to evaluate the presence of genito-urinary infections in infertile couples who sought counselling to investigate their condition. Microbiological analysis was performed on semen and vaginal/cervical samples of both partners of each couple. The percentage of individuals affected by a urogenital bacterial infection was between 14 and 20%. More significantly, most of the species isolated both in men and women have been described in the literature as potential causes of infertility.

  7. Effects of transgalactosylated disaccharides on the human intestinal microflora and their metabolism.

    PubMed

    Ito, M; Kimura, M; Deguchi, Y; Miyamori-Watabe, A; Yajima, T; Kan, T

    1993-06-01

    The effects of transgalactosylated disaccharide (TD) intake on human fecal microflora and their metabolism were investigated in 12 Japanese males. TD is a mixture of sugars, galactosyl galactose, and galactosyl glucose, synthesized from lactose through the transgalactosylation reaction of Streptococcus thermophilus beta-galactosidase. Volunteers took 15 g of the test sugar daily for 6 days. The TD ingestion increased the number of bifidobacteria and lactobacilli, but decreased the number of Bacteroidaceae and Candida spp. in the feces. The ratio of bifidobacteria to total bacteria increased from 0.28 to 0.51. TD decreased the fecal concentrations of propionic acid, isobutyric acid, isovaleric acid, and valeric acid. This sugar also lowered the fecal pH, and the concentrations of fecal ammonia, p-cresol, and indole. Moreover, a positive correlation was found between the concentration of ammonia, and that of branched-chain fatty acids (isobutyric acid and isovaleric acid), p-cresol, and indole. All of these compounds are produced from amino acids through deamination by the intestinal bacteria. The depression of amino acid fermentation by intestinal bacteria may be involved in the reduction of fecal ammonia. These results suggest that a part of the transgalactosylated disaccharides passes into the colon, inducing changes in the colonic microflora composition, hastening carbohydrate fermentation, and depressing amino acid fermentation in the human gut.

  8. Transport and function of syntaxin 3 in human epithelial intestinal cells.

    PubMed

    Breuza, L; Fransen, J; Le Bivic, A

    2000-10-01

    To follow the transport of human syntaxin (Syn) 3 to the apical surface of intestinal cells, we produced and expressed in Caco-2 cells a chimera made of the entire Syn3 coding sequence and the extracellular domain of the human transferrin receptor (TfR). This chimera (Syn3TfR) was localized to the apical membrane and was transported along the direct apical pathway, suggesting that this is also the case for endogenous Syn3. To test the potential role of Syn3 in apical transport, we overexpressed it in Caco-2 cells and measured the efficiency of apical and basolateral delivery of several endogenous markers. We observed a strong inhibition of apical delivery of sucrase-isomaltase (SI), an apical transmembrane protein, and of alpha-glucosidase, an apically secreted protein. No effect was observed on the basolateral delivery of Ag525, a basolateral antigen, strongly suggesting that Syn3 is necessary for efficient delivery of proteins to the apical surface of intestinal cells.

  9. Cockroaches as carriers of human intestinal parasites in two localities in Ethiopia.

    PubMed

    Kinfu, Addisu; Erko, Berhanu

    2008-11-01

    A study was undertaken to assess the role of cockroaches as potential carriers of human intestinal parasites in Addis Ababa and Ziway, Ethiopia. A total of 6480 cockroaches were trapped from the two localities from October 2006 to March 2007. All the cockroaches trapped in Addis Ababa (n=2240) and almost 50% (2100/4240) of those trapped in Ziway were identified as Blattella germanica. The rest of the cockroaches trapped in Ziway were identified as Periplaneta brunnea (24.52%), Pycnoscelus surinamensis (16.03%) and Supella longipalpa (9.90%). Microscopic examination of the external body washes of pooled cockroaches and individual gut contents revealed that cockroaches are carriers of Entamoeba coli and Entamoeba histolytica/dispar cysts as well as Enterobius vermicularis, Trichuris trichiura, Taenia spp. and Ascaris lumbricoides ova. Besides their role as a nuisance, the present study further confirms that cockroaches serve as carriers of human intestinal parasites. The possible association of cockroaches with allergic conditions such as asthma is also discussed. Hence, appropriate control measures should be taken particularly to make hotels and residential areas free of cockroaches as they represent a health risk.

  10. Histone Deacetylase Inhibitors Promote Mitochondrial Reactive Oxygen Species Production and Bacterial Clearance by Human Macrophages.

    PubMed

    Ariffin, Juliana K; das Gupta, Kaustav; Kapetanovic, Ronan; Iyer, Abishek; Reid, Robert C; Fairlie, David P; Sweet, Matthew J

    2015-12-28

    Broad-spectrum histone deacetylase inhibitors (HDACi) are used clinically as anticancer agents, and more isoform-selective HDACi have been sought to modulate other conditions, including chronic inflammatory diseases. Mouse studies suggest that HDACi downregulate immune responses and may compromise host defense. However, their effects on human macrophage antimicrobial responses are largely unknown. Here, we show that overnight pretreatment of human macrophages with HDACi prior to challenge with Salmonella enterica serovar Typhimurium or Escherichia coli results in significantly reduced intramacrophage bacterial loads, which likely reflect the fact that this treatment regime impairs phagocytosis. In contrast, cotreatment of human macrophages with HDACi at the time of bacterial challenge did not impair phagocytosis; instead, HDACi cotreatment actually promoted clearance of intracellular S. Typhimurium and E. coli. Mechanistically, treatment of human macrophages with HDACi at the time of bacterial infection enhanced mitochondrial reactive oxygen species generation by these cells. The capacity of HDACi to promote the clearance of intracellular bacteria from human macrophages was abrogated when cells were pretreated with MitoTracker Red CMXRos, which perturbs mitochondrial function. The HDAC6-selective inhibitor tubastatin A promoted bacterial clearance from human macrophages, whereas the class I HDAC inhibitor MS-275, which inhibits HDAC1 to -3, had no effect on intracellular bacterial loads. These data are consistent with HDAC6 and/or related HDACs constraining mitochondrial reactive oxygen species production from human macrophages during bacterial challenge. Our findings suggest that, whereas long-term HDACi treatment regimes may potentially compromise host defense, selective HDAC inhibitors may have applications in treating acute bacterial infections.

  11. Histone Deacetylase Inhibitors Promote Mitochondrial Reactive Oxygen Species Production and Bacterial Clearance by Human Macrophages

    PubMed Central

    Ariffin, Juliana K.; das Gupta, Kaustav; Kapetanovic, Ronan; Iyer, Abishek; Reid, Robert C.; Fairlie, David P.

    2015-01-01

    Broad-spectrum histone deacetylase inhibitors (HDACi) are used clinically as anticancer agents, and more isoform-selective HDACi have been sought to modulate other conditions, including chronic inflammatory diseases. Mouse studies suggest that HDACi downregulate immune responses and may compromise host defense. However, their effects on human macrophage antimicrobial responses are largely unknown. Here, we show that overnight pretreatment of human macrophages with HDACi prior to challenge with Salmonella enterica serovar Typhimurium or Escherichia coli results in significantly reduced intramacrophage bacterial loads, which likely reflect the fact that this treatment regime impairs phagocytosis. In contrast, cotreatment of human macrophages with HDACi at the time of bacterial challenge did not impair phagocytosis; instead, HDACi cotreatment actually promoted clearance of intracellular S. Typhimurium and E. coli. Mechanistically, treatment of human macrophages with HDACi at the time of bacterial infection enhanced mitochondrial reactive oxygen species generation by these cells. The capacity of HDACi to promote the clearance of intracellular bacteria from human macrophages was abrogated when cells were pretreated with MitoTracker Red CMXRos, which perturbs mitochondrial function. The HDAC6-selective inhibitor tubastatin A promoted bacterial clearance from human macrophages, whereas the class I HDAC inhibitor MS-275, which inhibits HDAC1 to -3, had no effect on intracellular bacterial loads. These data are consistent with HDAC6 and/or related HDACs constraining mitochondrial reactive oxygen species production from human macrophages during bacterial challenge. Our findings suggest that, whereas long-term HDACi treatment regimes may potentially compromise host defense, selective HDAC inhibitors may have applications in treating acute bacterial infections. PMID:26711769

  12. Bacterial Topography of the Healthy Human Lower Respiratory Tract

    PubMed Central

    Erb-Downward, John R.; Freeman, Christine M.; McCloskey, Lisa; Falkowski, Nicole R.

    2017-01-01

    ABSTRACT Although culture-independent techniques have refuted lung sterility in health, controversy about contamination during bronchoscope passage through the upper respiratory tract (URT) has impeded research progress. We sought to establish whether bronchoscopic sampling accurately reflects the lung microbiome in health and to distinguish between two proposed routes of authentic microbial immigration, (i) dispersion along contiguous respiratory mucosa and (ii) subclinical microaspiration. During bronchoscopy of eight adult volunteers without lung disease, we performed seven protected specimen brushings (PSB) and bilateral bronchoalveolar lavages (BALs) per subject. We amplified, sequenced, and analyzed the bacterial 16S rRNA gene V4 regions by using the Illumina MiSeq platform. Rigorous attention was paid to eliminate potential sources of error or contamination, including a randomized processing order and the inclusion and analysis of exhaustive procedural and sequencing control specimens. Indices of mouth-lung immigration (mouth-lung community similarity, bacterial burden, and community richness) were all significantly greater in airway and alveolar specimens than in bronchoscope contamination control specimens, indicating minimal evidence of pharyngeal contamination. Ecological indices of mouth-lung immigration peaked at or near the carina, as predicted for a primary immigration route of microaspiration. Bacterial burden, diversity, and mouth-lung similarity were greater in BAL than PSB samples, reflecting differences in the sampled surface areas. (This study has been registered at ClinicalTrials.gov under registration no. NCT02392182.) PMID:28196961

  13. The Intestinal Transport of Bovine Milk Exosomes Is Mediated by Endocytosis in Human Colon Carcinoma Caco-2 Cells and Rat Small Intestinal IEC-6 Cells123

    PubMed Central

    Wolf, Tovah; Baier, Scott R; Zempleni, Janos

    2015-01-01

    Background: MicroRNAs play essential roles in gene regulation. A substantial fraction of microRNAs in tissues and body fluids is encapsulated in exosomes, thereby conferring protection against degradation and a pathway for intestinal transport. MicroRNAs in cow milk are bioavailable in humans. Objective: This research assessed the transport mechanism of bovine milk exosomes, and therefore microRNAs, in human and rodent intestinal cells. Methods: The intestinal transport of bovine milk exosomes and microRNAs was assessed using fluorophore-labeled bovine milk exosomes in human colon carcinoma Caco-2 cells and rat small intestinal IEC-6 cells. Transport kinetics and mechanisms were characterized using dose-response studies, inhibitors of vesicle transport, carbohydrate competitors, proteolysis of surface proteins on cells and exosomes, and transepithelial transport in transwell plates. Results: Exosome transport exhibited saturation kinetics at 37°C [Michaelis constant (Km) = 55.5 ± 48.6 μg exosomal protein/200 μL of media; maximal transport rate = 0.083 ± 0.057 ng of exosomal protein · 81,750 cells−1 · h−1] and decreased by 64% when transport was measured at 4°C, consistent with carrier-mediated transport in Caco-2 cells. Exosome uptake decreased by 61–85% under the following conditions compared with controls in Caco-2 cells: removal of exosome and cell surface proteins by proteinase K, inhibition of endocytosis and vesicle trafficking by synthetic inhibitors, and inhibition of glycoprotein binding by carbohydrate competitors. When milk exosomes, at a concentration of 5 times the Km, were added to the upper chamber in transwell plates, Caco-2 cells accumulated miR-29b and miR-200c in the lower chamber, and reverse transport was minor. Transport characteristics were similar in IEC-6 cells and Caco-2 cells, except that substrate affinity and transporter capacity were lower and higher, respectively. Conclusion: The uptake of bovine milk exosomes is

  14. Incorporation of a non-human glycan mediates human susceptibility to a bacterial toxin

    SciTech Connect

    Byres, Emma; Paton, Adrienne W.; Paton, James C.; Löfling, Jonas C.; Smith, David F.; Wilce, Matthew C.J.; Talbot, Ursula M.; Chong, Damien C.; Yu, Hai; Huang, Shengshu; Chen, Xi; Varki, Nissi M.; Varki, Ajit; Rossjohn, Jamie; Beddoe, Travis

    2009-01-30

    AB{sub 5} toxins comprise an A subunit that corrupts essential eukaryotic cell functions, and pentameric B subunits that direct target-cell uptake after binding surface glycans. Subtilase cytotoxin (SubAB) is an AB{sub 5} toxin secreted by Shiga toxigenic Escherichia coli (STEC), which causes serious gastrointestinal disease in humans. SubAB causes haemolytic uraemic syndrome-like pathology in mice through SubA-mediated cleavage of BiP/GRP78, an essential endoplasmic reticulum chaperone. Here we show that SubB has a strong preference for glycans terminating in the sialic acid N-glycolylneuraminic acid (Neu5Gc), a monosaccharide not synthesized in humans. Structures of SubB-Neu5Gc complexes revealed the basis for this specificity, and mutagenesis of key SubB residues abrogated in vitro glycan recognition, cell binding and cytotoxicity. SubAB specificity for Neu5Gc was confirmed using mouse tissues with a human-like deficiency of Neu5Gc and human cell lines fed with Neu5Gc. Despite lack of Neu5Gc biosynthesis in humans, assimilation of dietary Neu5Gc creates high-affinity receptors on human gut epithelia and kidney vasculature. This, and the lack of Neu5Gc-containing body fluid competitors in humans, confers susceptibility to the gastrointestinal and systemic toxicities of SubAB. Ironically, foods rich in Neu5Gc are the most common source of STEC contamination. Thus a bacterial toxin's receptor is generated by metabolic incorporation of an exogenous factor derived from food.

  15. Role of intestinal epithelial cells in the host secretory response to infection by invasive bacteria. Bacterial entry induces epithelial prostaglandin h synthase-2 expression and prostaglandin E2 and F2alpha production.

    PubMed Central

    Eckmann, L; Stenson, W F; Savidge, T C; Lowe, D C; Barrett, K E; Fierer, J; Smith, J R; Kagnoff, M F

    1997-01-01

    Increased intestinal fluid secretion is a protective host response after enteric infection with invasive bacteria that is initiated within hours after infection, and is mediated by prostaglandin H synthase (PGHS) products in animal models of infection. Intestinal epithelial cells are the first host cells to become infected with invasive bacteria, which enter and pass through these cells to initiate mucosal, and ultimately systemic, infection. The present studies characterized the role of intestinal epithelial cells in the host secretory response after infection with invasive bacteria. Infection of cultured human intestinal epithelial cell lines with invasive bacteria, but not noninvasive bacteria, is shown to induce the expression of one of the rate-limiting enzymes for prostaglandin formation, PGHS-2, and the production of PGE2 and PGF2alpha. Furthermore, increased PGHS-2 expression was observed in intestinal epithelial cells in vivo after infection with invasive bacteria, using a human intestinal xenograft model in SCID mice. In support of the physiologic importance of epithelial PGHS-2 expression, supernatants from bacteria-infected intestinal epithelial cells were shown to increase chloride secretion in an in vitro model using polarized epithelial cells, and this activity was accounted for by PGE2. These studies define a novel autocrine/paracrine function of mediators produced by intestinal epithelial cells in the rapid induction of increased fluid secretion in response to intestinal infection with invasive bacteria. PMID:9218506

  16. Doxycycline protects human intestinal cells from hypoxia/reoxygenation injury: Implications from an in-vitro hypoxia model.

    PubMed

    Hummitzsch, Lars; Zitta, Karina; Berndt, Rouven; Kott, Matthias; Schildhauer, Christin; Parczany, Kerstin; Steinfath, Markus; Albrecht, Martin

    2017-04-15

    Intestinal ischemia/reperfusion (I/R) injury is a grave clinical emergency and associated with high morbidity and mortality rates. Based on the complex underlying mechanisms, a multimodal pharmacological approach seems necessary to prevent intestinal I/R injury. The antibiotic drug doxycycline, which exhibits a wide range of pleiotropic therapeutic properties, might be a promising candidate for also reducing I/R injury in the intestine. To investigate possible protective effects of doxycycline on intestinal I/R injury, human intestinal CaCo-2 cells were exposed to doxycycline at clinically relevant concentrations. In order to mimic I/R injury, CaCo-2 were thereafter subjected to hypoxia/reoxygenation by using our recently described two-enzyme in-vitro hypoxia model. Investigations of cell morphology, cell damage, apoptosis and hydrogen peroxide formation were performed 24h after the hypoxic insult. Hypoxia/reoxygenation injury resulted in morphological signs of cell damage, elevated LDH concentrations in the respective culture media (P<0.001) and increased protein expression of proapoptotic caspase-3 (P<0.05) in the intestinal cultures. These events were associated with increased levels hydrogen peroxide (P<0.001). Preincubation of CaCo-2 cells with different concentrations of doxycycline (5µM, 10µM, 50µM) reduced the hypoxia induced signs of cell damage and LDH release (P<0.001 for all concentrations). The reduction of cellular damage was associated with a reduced expression of caspase-3 (5µM, P<0.01; 10µM, P<0.01; 50µM, P<0.05), while hydrogen peroxide levels remained unchanged. In summary, doxycycline protects human intestinal cells from hypoxia/reoxygenation injury in-vitro. Further animal and clinical studies are required to prove the protective potential of doxycycline on intestinal I/R injury under in-vivo conditions.

  17. Potential Use of Bacterial Community Succession in Decaying Human Bone for Estimating Postmortem Interval.

    PubMed

    Damann, Franklin E; Williams, Daniel E; Layton, Alice C

    2015-07-01

    Bacteria are taphonomic agents of human decomposition, potentially useful for estimating postmortem interval (PMI) in late-stage decomposition. Bone samples from 12 individuals and three soil samples were analyzed to assess the effects of decomposition and advancing time on bacterial communities. Results indicated that partially skeletonized remains maintained a presence of bacteria associated with the human gut, whereas bacterial composition of dry skeletal remains maintained a community profile similar to soil communities. Variation in the UniFrac distances was significantly greater between groups than within groups (p < 0.001) for the unweighted metric and not the weighted metric. The members of the bacterial communities were more similar within than between decomposition stages. The oligotrophic environment of bone relative to soft tissue and the physical protection of organic substrates may preclude bacterial blooms during the first years of skeletonization. Therefore, community membership (unweighted) may be better for estimating PMI from skeletonized remains than community structure (weighted).

  18. Dysfunctions at human intestinal barrier by water-borne protozoan parasites: lessons from cultured human fully differentiated colon cancer cell lines.

    PubMed

    Liévin-Le Moal, Vanessa

    2013-06-01

    Some water-borne protozoan parasites induce diseases through their membrane-associated functional structures and virulence factors that hijack the host cellular molecules and signalling pathways leading to structural and functional lesions in the intestinal barrier. In this Microreview we analyse the insights on the mechanisms of pathogenesis of Entamoeba intestinalis, Giardia and Cryptosporidium observed in the human colon carcinoma fully differentiated colon cancer cell lines, cell subpopulations and clones expressing the structural and functional characteristics of highly specialized fully differentiated epithelial cells lining the intestinal epithelium and mimicking structurally and functionally an intestinal barrier.

  19. Free fucose is a danger signal to human intestinal epithelial cells.

    PubMed

    Chow, Wai Ling; Lee, Yuan Kun

    2008-03-01

    Fucose is present in foods, and it is a major component of human mucin glycoproteins and glycolipids. l-Fucose can also be found at the terminal position of many cell-surface oligosaccharide ligands that mediate cell-recognition and adhesion-signalling pathways. Mucin fucose can be released through the hydrolytic activity of pathogens and indigenous bacteria, leading to the release of free fucose into the intestinal lumen. The immunomodulating effects of free fucose on intestinal epithelial cells (enterocyte-like Caco-2) were investigated. It was found that the presence of l-fucose up regulated genes and secretion of their encoded proteins that are involved in both the innate and adaptive immune responses, possibly via the toll-like receptor-2 signalling pathway. These include TNFSF5, TNFSF7, TNF-alpha, IL12, IL17 and IL18. Besides modulating immune reactions in differentiated Caco-2 cells, fucose induced a set of cytokine genes that are involved in the development and proliferation of immune cells. These include the bone morphogenetic proteins (BMP) BMP2, BMP4, IL5, thrombopoietin and erythropoietin. In addition, the up regulated gene expression of fibroblast growth factor-2 may help to promote epithelial cell restitution in conjunction with the enhanced expression of transforming growth factor-beta mRNA. Since the exogenous fucose was not metabolised by the differentiated Caco-2 cells as a carbon source, the reactions elicited were suggested to be a result of the direct interaction of fucose and differentiated Caco-2 cells. The presence of free fucose may signal the invasion of mucin-hydrolysing microbial cells and breakage of the mucosal barrier. The intestinal epithelial cells respond by up regulation and secretion of cytokines, pre-empting the actual invasion of pathogens.

  20. Silencing of ecdysone receptor, insect intestinal mucin and sericotropin genes by bacterially produced double-stranded RNA affects larval growth and development in Plutella xylostella and Helicoverpa armigera.

    PubMed

    Israni, B; Rajam, M V

    2017-04-01

    RNA interference mediated gene silencing, which is triggered by double-stranded RNA (dsRNA), has become a important tool for functional genomics studies in various systems, including insects. Bacterially produced dsRNA employs the use of a bacterial strain lacking in RNaseIII activity and harbouring a vector with dual T7 promoter sites, which allow the production of intact dsRNA molecules. Here, we report an assessment of the functional relevance of the ecdysone receptor, insect intestinal mucin and sericotropin genes through silencing by dsRNA in two lepidopteran insect pests, Helicoverpa armigera and Plutella xylostella, both of which cause serious crop losses. Oral feeding of dsRNA led to significant reduction in transcripts of the target insect genes, which caused significant larval mortality with various moulting anomalies and an overall developmental delay. We also found a significant decrease in reproductive potential in female moths, with a drop in egg laying and compromised egg hatching from treated larvae as compared to controls. dsRNA was stable in the insect gut and was efficiently processed into small interfering RNAs (siRNAs), thus accounting for the phenotypes observed in the present work. The study revealed the importance of these genes in core insect processes, which are essential for insect development and survival.

  1. PGE2 is a direct and robust mediator of anion/fluid secretion by human intestinal epithelial cells

    PubMed Central

    Fujii, Satoru; Suzuki, Kohei; Kawamoto, Ami; Ishibashi, Fumiaki; Nakata, Toru; Murano, Tatsuro; Ito, Go; Shimizu, Hiromichi; Mizutani, Tomohiro; Oshima, Shigeru; Tsuchiya, Kiichiro; Nakamura, Tetsuya; Araki, Akihiro; Ohtsuka, Kazuo; Okamoto, Ryuichi; Watanabe, Mamoru

    2016-01-01

    Intestinal epithelial cells (IECs) play an indispensable role in maintaining body fluid balance partly through their ability to regulate anion/fluid secretion. Yet in various inflammatory gastrointestinal diseases, over-secretion of anions results in symptoms such as severe diarrhoea. Endogenous mediators, such as vasoactive intestinal peptide or prostaglandin E2 (PGE2), regulate intestinal anion/fluid secretion, but their direct effect on purified human IECs has never been described in detail. Based on a previously described intestinal organoid swelling model, we established a 3D-scanner-assisted quantification method to evaluate the anion/fluid secretory response of cultured human IECs. Among various endogenous secretagogues, we found that PGE2 had the lowest EC50 value with regard to the induction of swelling of the jejunal and colonic organoids. This PGE2-mediated swelling response was dependent on environmental Cl− concentrations as well as on several channels and transporters as shown by a series of chemical inhibitor studies. The concomitant presence of various inflammatory cytokines with PGE2 failed to modulate the PGE2-mediated organoid swelling response. Therefore, the present study features PGE2 as a direct and robust mediator of anion/fluid secretion by IECs in the human intestine. PMID:27827428

  2. Degradation of endogenous bacterial cell wall polymers by the muralytic enzyme mutanolysin prevents hepatobiliary injury in genetically susceptible rats with experimental intestinal bacterial overgrowth.

    PubMed Central

    Lichtman, S N; Okoruwa, E E; Keku, J; Schwab, J H; Sartor, R B

    1992-01-01

    Jejunal self-filling blind loops with subsequent small bowel bacterial overgrowth (SBBO) induce hepatobiliary injury in genetically susceptible Lewis rats. Lesions consist of portal tract inflammation, bile duct proliferation, and destruction. To determine the pathogenesis of SBBO-induced hepatobiliary injury, we treated Lewis rats with SBBO by using several agents with different mechanisms of activity. Buffer treatment, ursodeoxycholic acid, prednisone, methotrexate, and cyclosporin A failed to prevent SBBO-induced injury as demonstrated by increased plasma aspartate aminotransferase (AST) and elevated histology scores. However, hepatic injury was prevented by mutanolysin, a muralytic enzyme whose only known activity is to split the beta 1-4 N-acetylmuramyl-N-acetylglucosamine linkage of peptidoglycan-polysaccharide (PG-PS), a bacterial cell wall polymer with potent inflammatory and immunoregulatory properties. Mutanolysin therapy started on the day blind loops were surgically created and continued for 8 wk significantly diminished AST (101 +/- 37 U/liter) and liver histology scores (2.2 +/- 2.7) compared to buffer-treated rats (228 +/- 146 U/liter, P < 0.05, 8.2 +/- 1.9, P < 0.001 respectively). Mutanolysin treatment started during the early phase of hepatic injury, 16-21 d after surgery, decreased AST in 7 of 11 rats from 142 +/- 80 to 103 +/- 24 U/liter contrasted to increased AST in 9 of 11 buffer-treated rats from 108 +/- 52 to 247 +/- 142 U/liter, P < 0.05. Mutanolysin did not change total bacterial numbers within the loop, eliminate Bacteroides sp., have in vitro antibiotic effects, or diminish mucosal PG-PS transport. However, mutanolysin treatment prevented elevation of plasma anti-PG antibodies and tumor necrosis factor-alpha (TNF alpha) levels which occurred in buffer treated rats with SBBO and decreased TNF alpha production in isolated Kupffer cells stimulated in vitro with PG-PS. Based on the preventive and therapeutic activity of this highly specific

  3. Intestinal microbiology in early life: specific prebiotics can have similar functionalities as human-milk oligosaccharides.

    PubMed

    Oozeer, Raish; van Limpt, Kees; Ludwig, Thomas; Ben Amor, Kaouther; Martin, Rocio; Wind, Richèle D; Boehm, Günther; Knol, Jan

    2013-08-01

    Human milk is generally accepted as the best nutrition for newborns and has been shown to support the optimal growth and development of infants. On the basis of scientific insights from human-milk research, a specific mixture of nondigestible oligosaccharides has been developed, with the aim to improve the intestinal microbiota in early life. The mixture has been extensively studied and has been shown to be safe and to have potential health benefits that are similar to those of human milk. The specific mixture of short-chain galacto-oligosaccharides and long-chain fructo-oligosaccharides has been found to affect the development of early microbiota and to increase the Bifidobacterium amounts as observed in human-milk-fed infants. The resulting gut ecophysiology is characterized by high concentrations of lactate, a slightly acidic pH, and specific short-chain fatty acid profiles, which are high in acetate and low in butyrate and propionate. Here, we have summarized the main findings of dietary interventions with these specific oligosaccharides on the gut microbiota in early life. The gut ecophysiology in early life may have consequences for the metabolic, immunologic, and even neurologic development of the child because reports increasingly substantiate the important function of gut microbes in human health. This review highlights major findings in the field of early gut colonization and the potential impact of early nutrition in healthy growth and development.

  4. SREBP-2 negatively regulates FXR-dependent transcription of FGF19 in human intestinal cells.

    PubMed

    Miyata, Masaaki; Hata, Tatsuya; Yamazoe, Yasushi; Yoshinari, Kouichi

    2014-01-10

    Sterol regulatory element-binding protein-2 (SREBP-2) is a basic helix-loop-helix-leucine zipper transcription factor that positively regulates transcription of target genes involved in cholesterol metabolism. In the present study, we have investigated a possible involvement of SREBP-2 in human intestinal expression of fibroblast growth factor (FGF)19, which is an endocrine hormone involved in the regulation of lipid and glucose metabolism. Overexpression of constitutively active SREBP-2 decreased FGF19 mRNA levels in human colon-derived LS174T cells. In reporter assays, active SREBP-2 overexpression suppressed GW4064/FXR-mediated increase in reporter activities in regions containing the IR-1 motif (+848 to +5200) in the FGF19 gene. The suppressive effect disappeared in reporter activities in the region containing the IR-1 motif when the mutation was introduced into the IR-1 motif. In electrophoretic mobility shift assays, binding of the FXR/retinoid X receptor α heterodimer to the IR-1 motif was attenuated by adding active SREBP-2, but SREBP-2 binding to the IR-1 motif was not observed. In chromatin immunoprecipitation assays, specific binding of FXR to the IR-1-containing region of the FGF19 gene (+3214 to +3404) was increased in LS174T cells by treatment with cholesterol and 25-hydroxycholesterol. Specific binding of SREBP-2 to FXR was observed in glutathione-S-transferase (GST) pull-down assays. These results suggest that SREBP-2 negatively regulates the FXR-mediated transcriptional activation of the FGF19 gene in human intestinal cells.

  5. Supersaturation and Precipitation of Posaconazole Upon Entry in the Upper Small Intestine in Humans.

    PubMed

    Hens, Bart; Brouwers, Joachim; Corsetti, Maura; Augustijns, Patrick

    2016-09-01

    The purpose of this study was to explore gastrointestinal dissolution, supersaturation and precipitation of the weakly basic drug posaconazole in humans, and to assess the impact of formulation pH and type on these processes. In a cross-over study, two posaconazole suspensions (40 mg dispersed in 240 mL water at pH 1.6 and pH 7.1, respectively) were intragastrically administered; subsequently, gastric and duodenal fluids were aspirated. In parallel, blood samples were collected. Additionally, posaconazole was intragastrically administered as a solution (20 mg in 240 mL water, pH 1.6). When posaconazole was administered as an acidified suspension, supersaturated duodenal concentrations of posaconazole were observed for approximately 45 min. However, extensive intestinal precipitation was observed. Administration of the neutral suspension resulted in subsaturated concentrations with a mean duodenal AUC0-120 min and Cmax being approximately twofold lower than for the acidified suspension. The mean plasma AUC0-8 h of posaconazole was also twofold higher following administration of the acidified suspension. Similar to the acidified suspension, significant intestinal precipitation (up to 92%) was observed following intragastric administration of the posaconazole solution. This study demonstrated for the first time the gastrointestinal behavior of a weakly basic drug administered in different conditions, and its impact on systemic exposure.

  6. Aboral changes in D-glucose transport by human intestinal brush-border membrane vesicles.

    PubMed Central

    Bluett, M K; Abumrad, N N; Arab, N; Ghishan, F K

    1986-01-01

    D-Glucose transport was investigated in isolated brush-border membrane vesicles from human small intestine. Characteristics of D-glucose transport from the jejunum were compared with that in the mid and terminal ileum. Jejunal and mid-ileal D-glucose transport was Na+-dependent and electrogenic. The transient overshoot of jejunal D-glucose transport was significantly greater than corresponding values in mid-ileum. The terminal ileum did not exhibit Na+-dependent D-glucose transport, but did exhibit Na+-dependent taurocholate transport. Na+-glucose co-transport activity as measured by tracer-exchange experiments was greatest in the jejunum, and diminished aborally. We conclude that D-glucose transport in man is Na+-dependent and electrogenic in the proximal intestine and directly related to the activity of D-glucose-Na+ transporters present in the brush-border membranes. D-Glucose transport in the terminal ileum resembles colonic transport of D-glucose. PMID:3800877

  7. Stereomicroscopic 3D-pattern profiling of murine and human intestinal inflammation reveals unique structural phenotypes

    PubMed Central

    Rodriguez-Palacios, Alex; Kodani, Tomohiro; Kaydo, Lindsey; Pietropaoli, Davide; Corridoni, Daniele; Howell, Scott; Katz, Jeffry; Xin, Wei; Pizarro, Theresa T.; Cominelli, Fabio

    2015-01-01

    Histology is fundamental to assess two-dimensional intestinal inflammation; however, inflammatory bowel diseases (IBDs) are often indistinguishable microscopically on the basis of mucosal biopsies. Here, we use stereomicroscopy (SM) to rapidly profile the entire intestinal topography and assess inflammation. We examine the mucosal surface of >700 mice (encompassing >16 strains and various IBD-models), create a profiling catalogue of 3D-stereomicroscopic abnormalities and demonstrate that mice with comparable histological scores display unique sub-clusters of 3D-structure-patterns of IBD pathology, which we call 3D-stereoenterotypes, and which are otherwise indiscernible histologically. We show that two ileal IBD-stereoenterotypes (‘cobblestones' versus ‘villous mini-aggregation') cluster separately within two distinct mouse lines of spontaneous ileitis, suggesting that host genetics drive unique and divergent inflammatory 3D-structural patterns in the gut. In humans, stereomicroscopy reveals ‘liquefaction' lesions and hierarchical fistulous complexes, enriched with clostridia/segmented filamentous bacteria, running under healthy mucosa in Crohn's disease. We suggest that stereomicroscopic (3D-SMAPgut) profiling can be easily implemented and enable the comprehensive study of inflammatory 3D structures, genetics and flora in IBD. PMID:26154811

  8. Human intestinal Vdelta1+ lymphocytes recognize tumor cells of epithelial origin

    PubMed Central

    1996-01-01

    gammadelta T cells can be grouped into discrete subsets based upon their expression of T cell receptor (TCR) variable (V) region families, their tissue distribution, and their specificity. Vdelta2+ T cells constitute the majority of gammadelta T cells in peripheral blood whereas Vdelta1+T cells reside preferentially in skin epithelium and in the intestine. gammadelta T cells are envisioned as first line host defense mechanisms capable of providing a source of immune effector T cells and immunomodulating cytokines such as interleukin (IL) 4 or interferon (IFN) gamma. We describe here the fine specificity of three distinct gammadelta+ tumor-infiltrating lymphocytes (TIL) obtained from patients with primary or metastatic colorectal cancer, that could be readily expanded in vitro in the presence of IL-1beta and IL-7. Irrespective of donor, these individual gammadelta T cells exhibited a similar pattern of reactivity defined by recognition of autologous and allogeneic colorectal cancer cells, renal cell cancer, pancreatic cancer, and a freshly isolated explant from human intestine as measured by cytolytic T cell responses and by IFN-gamma release. In contrast, tumors of alternate histologies were not lysed, including lung cancer, squamous cell cancer, as well as the natural/lymphocyte-activated killer cell-sensitive hematopoietic cell lines T2, C1R, or Daudi. The cell line K562 was only poorly lysed when compared with colorectal cancer targets. Target cell reactivity mediated by Vdelta1+ T cells was partially blocked with Abs directed against the TCR, the beta2 or beta7 integrin chains, or fibronectin receptor. Marker analysis using flow cytometry revealed that all three gammadelta T cell lines exhibit a similar phenotype. Analysis of the gammadelta TCR junctional suggested exclusive usage of the Vdelta1/Ddelta3/Jdelta1 TCR segments with extensive (< or = 29 bp) N/P region diversity. T cell recognition of target cells did not appear to be a major histocompatibility

  9. Transcobalamin derived from bovine milk stimulates apical uptake of vitamin B12 into human intestinal epithelial cells.

    PubMed

    Hine, Brad; Boggs, Irina; Green, Ralph; Miller, Joshua W; Hovey, Russell C; Humphrey, Rex; Wheeler, Thomas T

    2014-11-01

    Intestinal uptake of vitamin B12 (hereafter B12) is impaired in a significant proportion of the human population. This impairment is due to inherited or acquired defects in the expression or function of proteins involved in the binding of diet-derived B12 and its uptake into intestinal cells. Bovine milk is an abundant source of bioavailable B12 wherein it is complexed with transcobalamin. In humans, transcobalamin functions primarily as a circulatory protein, which binds B12 following its absorption and delivers it to peripheral tissues via its cognate receptor, CD320. In the current study, the transcobalamin-B12 complex was purified from cows' milk and its ability to stimulate uptake of B12 into cultured bovine, mouse and human cell lines was assessed. Bovine milk-derived transcobalamin-B12 complex was absorbed by all cell types tested, suggesting that the uptake mechanism is conserved across species. Furthermore, the complex stimulated the uptake of B12 via the apical surface of differentiated Caco-2 human intestinal epithelial cells. These findings suggest the presence of an alternative transcobalamin-mediated uptake pathway for B12 in the human intestine other than that mediated by the gastric glycoprotein, intrinsic factor. Our findings highlight the potential for transcobalamin-B12 complex derived from bovine milk to be used as a natural bioavailable alternative to orally administered free B12 to overcome B12 malabsorption.

  10. Bacterial Human Virulence Genes across Diverse Habitats As Assessed by In silico Analysis of Environmental Metagenomes

    PubMed Central

    Søborg, Ditte A.; Hendriksen, Niels B.; Kilian, Mogens; Christensen, Jan H.; Kroer, Niels

    2016-01-01

    The occurrence and distribution of clinically relevant bacterial virulence genes across natural (non-human) environments is not well understood. We aimed to investigate the occurrence of homologs to bacterial human virulence genes in a variety of ecological niches to better understand the role of natural environments in the evolution of bacterial virulence. Twenty four bacterial virulence genes were analyzed in 46 diverse environmental metagenomic datasets, representing various soils, seawater, freshwater, marine sediments, hot springs, the deep-sea, hypersaline mats, microbialites, gutless worms and glacial ice. Homologs to 16 bacterial human virulence genes, involved in urinary tract infections, gastrointestinal diseases, skin diseases, and wound and systemic infections, showed global ubiquity. A principal component analysis did not demonstrate clear trends across the metagenomes with respect to occurrence and frequency of observed gene homologs. Full-length (>95%) homologs of several virulence genes were identified, and translated sequences of the environmental and clinical genes were up to 50–100% identical. Furthermore, phylogenetic analyses indicated deep branching positions of some of the environmental gene homologs, suggesting that they represent ancient lineages in the phylogeny of the clinical genes. Fifteen virulence gene homologs were detected in metatranscriptomes, providing evidence of environmental expression. The ubiquitous presence and transcription of the virulence gene homologs in non-human environments point to an important ecological role of the genes for the activity and survival of environmental bacteria. Furthermore, the high degree of sequence conservation between several of the environmental and clinical genes suggests common ancestral origins. PMID:27857707

  11. Cooperation between MEF2 and PPARγ in human intestinal β,β-carotene 15,15'-monooxygenase gene expression

    PubMed Central

    Gong, Xiaoming; Tsai, Shu-Whei; Yan, Bingfang; Rubin, Lewis P

    2006-01-01

    Background Vitamin A and its derivatives, the retinoids, are essential for normal embryonic development and maintenance of cell differentiation. β, β-carotene 15,15'-monooxygenase 1 (BCMO1) catalyzes the central cleavage of β-carotene to all-trans retinal and is the key enzyme in the intestinal metabolism of carotenes to vitamin A. However, human and various rodent species show markedly different efficiencies in intestinal BCMO1-mediated carotene to retinoid conversion. The aim of this study is to identify potentially human-specific regulatory control mechanisms of BCMO1 gene expression. Results We identified and functionally characterized the human BCMO1 promoter sequence and determined the transcriptional regulation of the BCMO1 gene in a BCMO1 expressing human intestinal cell line, TC-7. Several functional transcription factor-binding sites were identified in the human promoter that are absent in the mouse BCMO1 promoter. We demonstrate that the proximal promoter sequence, nt -190 to +35, confers basal transcriptional activity of the human BCMO1 gene. Site-directed mutagenesis of the myocyte enhancer factor 2 (MEF2) and peroxisome proliferator-activated receptor (PPAR) binding elements resulted in decreased basal promoter activity. Mutation of both promoter elements abrogated the expression of intestinal cell BCMO1. Electrophoretic mobility shift and supershift assays and transcription factor co-expression in TC-7 cells showed MEF2C and PPARγ bind to their respective DNA elements and synergistically transactivate BCMO1 expression. Conclusion We demonstrate that human intestinal cell BCMO1 expression is dependent on the functional cooperation between PPARγ and MEF2 isoforms. The findings suggest that the interaction between MEF2 and PPAR factors may provide a molecular basis for interspecies differences in the transcriptional regulation of the BCMO1 gene. PMID:16504037

  12. Up-regulating the human intestinal microbiome using whole plant foods, polyphenols, and/or fiber.

    PubMed

    Tuohy, Kieran M; Conterno, Lorenza; Gasperotti, Mattia; Viola, Roberto

    2012-09-12

    Whole plant foods, including fruit, vegetables, and whole grain cereals, protect against chronic human diseases such as heart disease and cancer, with fiber and polyphenols thought to contribute significantly. These bioactive food components interact with the gut microbiota, with gut bacteria modifying polyphenol bioavailability and activity, and with fiber, constituting the main energy source for colonic fermentation. This paper discusses the consequences of increasing the consumption of whole plant foods on the gut microbiota and subsequent implications for human health. In humans, whole grain cereals can modify fecal bacterial profiles, increasing relative numbers of bifidobacteria and lactobacilli. Polyphenol-rich chocolate and certain fruits have also been shown to increase fecal bifidobacteria. The recent FLAVURS study provides novel information on the impact of high fruit and vegetable diets on the gut microbiota. Increasing whole plant food consumption appears to up-regulate beneficial commensal bacteria and may contribute toward the health effects of these foods.

  13. The Yin and Yang of Bacterial Resilience in the Human Gut Microbiota

    PubMed Central

    Gibson, Molly K.; Pesesky, Mitchell W.; Dantas, Gautam

    2014-01-01

    The human gut is home to trillions of microbes that form a symbiotic relationship with the human host. During health, the intestinal microbiota provides many benefits to the host and is generally resistant to colonization by new species; however, disruption of this complex community can lead to pathogen invasion, inflammation, and disease. Restoration and maintenance of a healthy gut microbiota composition requires effective therapies to reduce and prevent colonization of harmful bacteria (pathogens) while simultaneously promoting growth of beneficial bacteria (probiotics). Here we review the mechanisms by which the host modulates the gut community composition during health and disease, and discuss prospects for antibiotic and probiotic therapy for restoration of a healthy intestinal community following disruption. PMID:24911583

  14. The yin and yang of bacterial resilience in the human gut microbiota.

    PubMed

    Gibson, Molly K; Pesesky, Mitchell W; Dantas, Gautam

    2014-11-25

    The human gut is home to trillions of microbes that form a symbiotic relationship with the human host. During health, the intestinal microbiota provides many benefits to the host and is generally resistant to colonization by new species; however, disruption of this complex community can lead to pathogen invasion, inflammation, and disease. Restoration and maintenance of a healthy gut microbiota composition requires effective therapies to reduce and prevent colonization of harmful bacteria (pathogens) while simultaneously promoting growth of beneficial bacteria (probiotics). Here we review the mechanisms by which the host modulates the gut community composition during health and disease, and we discuss prospects for antibiotic and probiotic therapy for restoration of a healthy intestinal community following disruption.

  15. Postprandial concentrations of free and conjugated bile acids down the length of the normal human small intestine

    PubMed Central

    Northfield, T. C.; McColl, I.

    1973-01-01

    Small intestinal samples were obtained by intubation from multiple sites along the small intestine in 11 subjects with no known gastrointestinal disease eating a normal diet and at laparotomy in a further three subjects. Free (unconjugated) bile acids were consistently demonstrated in ileal samples, and occasionally in lower jejunal samples, by thin-layer chromatography, supplemented in some cases by gas/liquid chromatography and by infrared spectroscopy. The free bile acid concentration, measured enzymically following thin-layer chromatography, reached a maximum (1 mM) in the lower ileum, where it represented half the total bile acid concentration. Following ampicillin, the concentration of free bile acids decreased markedly, suggesting that they resulted from bacterial deconjugation; at the same time the total bile acid concentration increased, suggesting impaired absorption due to the reduced concentration of the more rapidly absorbed free bile acids. Our results indicate that the presence of free bile acids in lower jejunal and ileal samples is a normal finding, and cannot be taken as evidence of abnormal bacterial overgrowth. They also suggest that bacterial deconjugation at these sites may be a factor contributing to the remarkable efficiency of bile salt reabsorption. ImagesFig 2 PMID:4729918

  16. An improved prediction of the human in vivo intestinal permeability and BCS class of drugs using the in vitro permeability ratio obtained for rat intestine using an Ussing chamber system.

    PubMed

    Li, Hong; Jin, Hyo-Eon; Shim, Won-Sik; Shim, Chang-Koo

    2013-10-01

    The Biopharmaceutics Classification System (BCS) was developed to facilitate estimation of the in vivo pharmacokinetic performance of drugs from human intestinal permeability and solubility. However, the measurement of human in vivo intestinal permeability, unlike that of solubility, is problematic and inefficient. Thus, rat in vitro intestinal permeability results obtained via the Ussing chamber technique are often used instead. However, these data could be unreliable due to difficulty in maintaining the viability of the dissected intestinal membrane in the Ussing chamber. Therefore, a more efficient method to obtain a reliable in vitro permeability is mandatory. Here, we propose a new approach by introducing a novel factor called the permeability ratio (PR). Basically, PR is a rat in vitro intestinal permeability obtained from the Ussing chamber, which is then corrected by the permeability of lucifer yellow, a paracellular permeability marker. To prove the validity of the method, 12 model drugs representing different BCS classes were tested, and the correlation with human in vivo intestinal permeability was high. More importantly, the new method perfectly classified all 12 model drugs. The results indicate that PR is a reliable factor with high correlation to human in vivo intestinal permeability, which can further be used to accurately predict the BCS classification.

  17. Exploiting Bacterial Operons To Illuminate Human Iron-Sulfur Proteins.

    PubMed

    Andreini, Claudia; Banci, Lucia; Rosato, Antonio

    2016-04-01

    Organisms from all kingdoms of life use iron-sulfur proteins (FeS-Ps) in a multitude of functional processes. We applied a bioinformatics approach to investigate the human portfolio of FeS-Ps. Sixty-one percent of human FeS-Ps bind Fe4S4 clusters, whereas 39% bind Fe2S2 clusters. However, this relative ratio varies significantly depending on the specific cellular compartment. We compared the portfolio of human FeS-Ps to 12 other eukaryotes and to about 700 prokaryotes. The comparative analysis of the organization of the prokaryotic homologues of human FeS-Ps within operons allowed us to reconstruct the human functional networks involving the conserved FeS-Ps common to prokaryotes and eukaryotes. These functional networks have been maintained during evolution and thus presumably represent fundamental cellular processes. The respiratory chain and the ISC machinery for FeS-P biogenesis are the two conserved processes that involve the majority of human FeS-Ps. Purine metabolism is another process including several FeS-Ps, in which BOLA proteins possibly have a regulatory role. The analysis of the co-occurrence of human FeS-Ps with other proteins highlighted numerous links between the iron-sulfur cluster machinery and the response mechanisms to cell damage, from repair to apoptosis. This relationship probably relates to the production of reactive oxygen species within the biogenesis and degradation of FeS-Ps.

  18. Evaluation of a Multiplex Real-Time PCR Assay for Detecting Major Bacterial Enteric Pathogens in Fecal Specimens: Intestinal Inflammation and Bacterial Load Are Correlated in Campylobacter Infections

    PubMed Central

    Wohlwend, Nadia; Tiermann, Sacha; Risch, Lorenz; Risch, Martin

    2016-01-01

    A total of 1,056 native or Cary-Blair-preserved stool specimens were simultaneously tested by conventional stool culturing and by enteric bacterial panel (EBP) multiplex real-time PCR for Campylobacter jejuni, Campylobacter coli, Salmonella spp., and shigellosis disease-causing agents (Shigella spp. and enteroinvasive Escherichia coli [EIEC]). Overall, 143 (13.5%) specimens tested positive by PCR for the targets named above; 3 coinfections and 109 (10.4%) Campylobacter spp., 17 (1.6%) Salmonella spp., and 20 (1.9%) Shigella spp./EIEC infections were detected. The respective positive stool culture rates were 75 (7.1%), 14 (1.3%), and 7 (0.7%). The median threshold cycle (CT) values of culture-positive specimens were significantly lower than those of culture-negative ones (CT values, 24.3 versus 28.7; P < 0.001), indicating that the relative bacterial load per fecal specimen was significantly associated with the culture results. In Campylobacter infections, the respective median fecal calprotectin concentrations in PCR-negative/culture-negative (n = 40), PCR-positive/culture-negative (n = 14), and PCR-positive/culture-positive (n = 15) specimens were 134 mg/kg (interquartile range [IQR], 30 to 1,374 mg/kg), 1,913 mg/kg (IQR, 165 to 3,813 mg/kg), and 5,327 mg/kg (IQR, 1,836 to 18,213 mg/kg). Significant differences were observed among the three groups (P < 0.001), and a significant linear trend was identified (P < 0.001). Furthermore, the fecal calprotectin concentrations and CT values were found to be correlated (r = −0.658). Our results demonstrate that molecular screening of Campylobacter spp., Salmonella spp., and Shigella spp./EIEC using the BD Max EBP assay will result in timely diagnosis and improved sensitivity. The determination of inflammatory markers, such as calprotectin, in fecal specimens may aid in the interpretation of PCR results, particularly for enteric pathogens associated with mucosal damage and colonic inflammation. PMID:27307458

  19. Evaluation of a Multiplex Real-Time PCR Assay for Detecting Major Bacterial Enteric Pathogens in Fecal Specimens: Intestinal Inflammation and Bacterial Load Are Correlated in Campylobacter Infections.

    PubMed

    Wohlwend, Nadia; Tiermann, Sacha; Risch, Lorenz; Risch, Martin; Bodmer, Thomas

    2016-09-01

    A total of 1,056 native or Cary-Blair-preserved stool specimens were simultaneously tested by conventional stool culturing and by enteric bacterial panel (EBP) multiplex real-time PCR for Campylobacter jejuni, Campylobacter coli, Salmonella spp., and shigellosis disease-causing agents (Shigella spp. and enteroinvasive Escherichia coli [EIEC]). Overall, 143 (13.5%) specimens tested positive by PCR for the targets named above; 3 coinfections and 109 (10.4%) Campylobacter spp., 17 (1.6%) Salmonella spp., and 20 (1.9%) Shigella spp./EIEC infections were detected. The respective positive stool culture rates were 75 (7.1%), 14 (1.3%), and 7 (0.7%). The median threshold cycle (CT) values of culture-positive specimens were significantly lower than those of culture-negative ones (CT values, 24.3 versus 28.7; P < 0.001), indicating that the relative bacterial load per fecal specimen was significantly associated with the culture results. In Campylobacter infections, the respective median fecal calprotectin concentrations in PCR-negative/culture-negative (n = 40), PCR-positive/culture-negative (n = 14), and PCR-positive/culture-positive (n = 15) specimens were 134 mg/kg (interquartile range [IQR], 30 to 1,374 mg/kg), 1,913 mg/kg (IQR, 165 to 3,813 mg/kg), and 5,327 mg/kg (IQR, 1,836 to 18,213 mg/kg). Significant differences were observed among the three groups (P < 0.001), and a significant linear trend was identified (P < 0.001). Furthermore, the fecal calprotectin concentrations and CT values were found to be correlated (r = -0.658). Our results demonstrate that molecular screening of Campylobacter spp., Salmonella spp., and Shigella spp./EIEC using the BD Max EBP assay will result in timely diagnosis and improved sensitivity. The determination of inflammatory markers, such as calprotectin, in fecal specimens may aid in the interpretation of PCR results, particularly for enteric pathogens associated with mucosal damage and colonic inflammation.

  20. Delphinidin Reduces Glucose Uptake in Mice Jejunal Tissue and Human Intestinal Cells Lines through FFA1/GPR40.

    PubMed

    Hidalgo, Jorge; Teuber, Stefanie; Morera, Francisco J; Ojeda, Camila; Flores, Carlos A; Hidalgo, María A; Núñez, Lucía; Villalobos, Carlos; Burgos, Rafael A

    2017-04-05

    Anthocyanins are pigments with antihyperglycemic properties, and they are potential candidates for developing functional foods for the therapy or prevention of Diabetes mellitus type 2 (DM2). The mechanism of these beneficial effects of anthocyanins are, however, hard to explain, given their very low bioavailability due to poor intestinal absorption. We propose that free fatty acid receptor 1 (FFA1, also named GPR40), is involved in an inhibitory effect of the anthocyanidin delphinidin over intestinal glucose absorption. We show the direct effects of delphinidin on the intestine using jejunum samples from RF/J mice, and the human intestinal cell lines HT-29, Caco-2, and NCM460. By the use of specific pharmacological antagonists, we determined that delphinidin inhibits glucose absorption in both mouse jejunum and a human enterocytic cell line in a FFA1-dependent manner. Delphinidin also affects the function of sodium-glucose cotransporter 1 (SGLT1). Intracellular signaling after FFA1 activation involved cAMP increase and cytosolic Ca(2+) oscillations originated from intracellular Ca(2+) stores and were followed by store-operated Ca(2+) entry. Taken together, our results suggest a new GPR-40 mediated local mechanism of action for delphinidin over intestinal cells that may in part explain its antidiabetic effect. These findings are promising for the search for new prevention and pharmacological treatment strategies for DM2 management.

  1. Bifidobacteria Prevent Tunicamycin-Induced Endoplasmic Reticulum Stress and Subsequent Barrier Disruption in Human Intestinal Epithelial Caco-2 Monolayers

    PubMed Central

    Akiyama, Takuya; Oishi, Kenji

    2016-01-01

    Endoplasmic reticulum (ER) stress is caused by accumulation of unfolded and misfolded proteins in the ER, thereby compromising its vital cellular functions in protein production and secretion. Genome wide association studies in humans as well as experimental animal models linked ER stress in intestinal epithelial cells (IECs) with intestinal disorders including inflammatory bowel diseases. However, the mechanisms linking the outcomes of ER stress in IECs to intestinal disease have not been clarified. In this study, we investigated the impact of ER stress on intestinal epithelial barrier function using human colon carcinoma-derived Caco-2 monolayers. Tunicamycin-induced ER stress decreased the trans-epithelial electrical resistance of Caco-2 monolayers, concomitant with loss of cellular plasma membrane integrity. Epithelial barrier disruption in Caco-2 cells after ER stress was not caused by caspase- or RIPK1-dependent cell death but was accompanied by lysosomal rupture and up-regulation of the ER stress markers Grp78, sXBP1 and Chop. Interestingly, several bifidobacteria species inhibited tunicamycin-induced ER stress and thereby diminished barrier disruption in Caco-2 monolayers. Together, these results showed that ER stress compromises the epithelial barrier function of Caco-2 monolayers and demonstrate beneficial impacts of bifidobacteria on ER stress in IECs. Our results identify epithelial barrier loss as a potential link between ER stress and intestinal disease development, and suggest that bifidobacteria could exert beneficial effects on this phenomenon. PMID:27611782

  2. Effect of variations in the amounts of P-glycoprotein (ABCB1), BCRP (ABCG2) and CYP3A4 along the human small intestine on PBPK models for predicting intestinal first pass.

    PubMed

    Bruyère, Arnaud; Declèves, Xavier; Bouzom, Francois; Ball, Kathryn; Marques, Catie; Treton, Xavier; Pocard, Marc; Valleur, Patrice; Bouhnik, Yoram; Panis, Yves; Scherrmann, Jean-Michel; Mouly, Stephane

    2010-10-04

    It is difficult to predict the first-pass effect in the human intestine due to a lack of scaling factors for correlating in vitro and in vivo data. We have quantified cytochrome P450/3A4 (CYP3A4) and two ABC transporters, P-glycoprotein (P-gp, ABCB1) and the breast cancer resistant protein BCRP (ABCG2), throughout the human small intestine to determine the scaling factors for predicting clearance from intestinal microsomes and develop a physiologically based pharmacokinetic (PBPK) model. CYP3A4, P-gp and BCRP proteins were quantified by Western blotting and/or enzyme activities in small intestine samples from 19 donors, and mathematical trends of these expressions with intestinal localization were established. Microsome fractions were prepared and used to calculate the amount of microsomal protein per gram of intestine (MPPGI). Our results showed a trend in CYP3A4 expression decrease from the upper to the lower small intestine while P-gp expression is increasing. In contrast, BCRP expression did not vary significantly with position, but varied greatly between individuals. The MPPGI (mg microsomal protein per centimeter intestine) remained constant along the length of the small intestine, at about 1.55 mg/cm. Moreover, intrinsic clearance measured with specific CYP3A4 substrates (midazolam and an in-house Servier drug) and intestinal microsomes was well correlated with the amount of CYP3A4 (R(2) > 0.91, p < 0.01). In vivo data were more accurately predicted using PBPK models of blood concentrations of these two substrates based on the segmental distributions of these enzymes and MPPGI determined in this study. Thus, these mathematical trends can be used to predict drug absorption at different intestinal sites and their metabolism can be predicted with the MPPGI.

  3. Vasoactive intestinal peptide (VIP) induces malignant transformation of the human prostate epithelial cell line RWPE-1.

    PubMed

    Fernández-Martínez, Ana B; Bajo, Ana M; Isabel Arenas, M; Sánchez-Chapado, Manuel; Prieto, Juan C; Carmena, María J

    2010-12-18

    The carcinogenic potential of vasoactive intestinal peptide (VIP) was analyzed in non-tumor human prostate epithelial cells (RWPE-1) and in vivo xenografts. VIP induced morphological changes and a migratory phenotype consistent with stimulation of expression/activity of metalloproteinases MMP-2 and MMP-9, decreased E-cadherin-mediated cell-cell adhesion, and increased cell motility. VIP increased cyclin D1 expression and cell proliferation that was blocked after VPAC(1)-receptor siRNA transfection. Similar effects were seen in RWPE-1 tumors developed by subcutaneous injection of VIP-treated cells in athymic nude mice. VIP acts as a cytokine in RWPE-1 cell transformation conceivably through epithelial-mesenchymal transition (EMT), reinforcing VIP role in prostate tumorigenesis.

  4. Automatic segmentation and classification of human intestinal parasites from microscopy images.

    PubMed

    Suzuki, Celso T N; Gomes, Jancarlo F; Falcão, Alexandre X; Papa, João P; Hoshino-Shimizu, Sumie

    2013-03-01

    Human intestinal parasites constitute a problem in most tropical countries, causing death or physical and mental disorders. Their diagnosis usually relies on the visual analysis of microscopy images, with error rates that may range from moderate to high. The problem has been addressed via computational image analysis, but only for a few species and images free of fecal impurities. In routine, fecal impurities are a real challenge for automatic image analysis. We have circumvented this problem by a method that can segment and classify, from bright field microscopy images with fecal impurities, the 15 most common species of protozoan cysts, helminth eggs, and larvae in Brazil. Our approach exploits ellipse matching and image foresting transform for image segmentation, multiple object descriptors and their optimum combination by genetic programming for object representation, and the optimum-path forest classifier for object recognition. The results indicate that our method is a promising approach toward the fully automation of the enteroparasitosis diagnosis.

  5. Human chorionic gonadotropin promotes expression of protein absorption factors in the intestine of goldfish (Carassius auratus).

    PubMed

    Zhou, Y; Hao, G; Zhong, H; Wu, Q; Lu, S Q; Zhao, Q; Liu, Z

    2015-07-27

    Protein use is crucial for the ovulation and spawning of fish. Currently, limited information is available regarding the expression of protein absorption factors during the breeding seasons of teleosts and thus how various proteins involved in this process is not well-understood. The expression of CDX2, CREB, gluatamate dehydrogenase, LAT2, aminopeptidase N, PepT1, and SP1 were significantly elevated from the non-breeding season to the breeding season in female goldfish, and all proteins except PepT1 and SP1 were elevated in male goldfish. Injection of human chorionic gonadotropin upregulated the expression of all proteins except for aminopeptidase N in female goldfish and SP1 in male goldfish, suggesting a luteinizing hormone-inductive effect on protein absorption factors. Protein use in the intestine is increased during the breeding seasons as a result of increased luteinizing hormone.

  6. Substrate specificity and some properties of phenol sulfotransferase from human intestinal Caco-2 cells

    SciTech Connect

    Baranczyk-Kuzma, A.; Garren, J.A.; Hidalgo, I.J.; Borchardt, R.T. )

    1991-01-01

    The phase 2 metabolic reactions, sulfation and glucuronidation, were studied in a human colon carcinoma cell line (Caco-2), which has been developed as a model of intestinal enterocytes. Phenol sulfotransferase was isolated from Caco-2 cells cultured for 7, 14 and 21 days. The enzyme catalyzed the sulfation of both p-nitrophenol and catecholamines as well as most catecholamine metabolites. The affinity (K{sub m}) of PST for dopamine was much higher than for p-nitrophenol, and the specific activity of PST with both substrates increased with the age of the cells. The thermal stability of Caco-2 PST increased with cell age and was not dependent on the acceptor substrate used. The thermolabile PST from 7-day old cells was more sensitive to NEM than was the thermostable enzyme from 21-day old cells. No UDP-glucuronyltransferase activity was detected in 7-, 14- and 21-day old Caco-2 cells with any of the methods used.

  7. Activation of Intestinal Human Pregnane X Receptor Protects against Azoxymethane/Dextran Sulfate Sodium–Induced Colon Cancer

    PubMed Central

    Cheng, Jie; Fang, Zhong-Ze; Nagaoka, Kenjiro; Okamoto, Minoru; Qu, Aijuan; Tanaka, Naoki; Kimura, Shioko

    2014-01-01

    The role of intestinal human pregnane X receptor (PXR) in colon cancer was determined through investigation of the chemopreventive role of rifaximin, a specific agonist of intestinal human PXR, toward azoxymethane (AOM)/dextran sulfate sodium (DSS)–induced colon cancer. Rifaximin treatment significantly decreased the number of colon tumors induced by AOM/DSS treatment in PXR-humanized mice, but not wild-type or Pxr-null mice. Additionally, rifaximin treatment markedly increased the survival rate of PXR-humanized mice, but not wild-type or Pxr-null mice. These data indicated a human PXR–dependent therapeutic chemoprevention of rifaximin toward AOM/DSS-induced colon cancer. Nuclear factor κ-light-chain-enhancer of activated B cells–mediated inflammatory signaling was upregulated in AOM/DSS-treated mice, and inhibited by rifaximin in PXR-humanized mice. Cell proliferation and apoptosis were also modulated by rifaximin treatment in the AOM/DSS model. In vitro cell-based assays further revealed that rifaximin regulated cell apoptosis and cell cycle in a human PXR-dependent manner. These results suggested that specific activation of intestinal human PXR exhibited a chemopreventive role toward AOM/DSS-induced colon cancer by mediating anti-inflammation, antiproliferation, and proapoptotic events. PMID:25277138

  8. Innervation of enteric mast cells by primary spinal afferents in guinea pig and human small intestine.

    PubMed

    Wang, Guo-Du; Wang, Xi-Yu; Liu, Sumei; Qu, Meihua; Xia, Yun; Needleman, Bradley J; Mikami, Dean J; Wood, Jackie D

    2014-10-01

    Mast cells express the substance P (SP) neurokinin 1 receptor and the calcitonin gene-related peptide (CGRP) receptor in guinea pig and human small intestine. Enzyme-linked immunoassay showed that activation of intramural afferents by antidromic electrical stimulation or by capsaicin released SP and CGRP from human and guinea pig intestinal segments. Electrical stimulation of the afferents evoked slow excitatory postsynaptic potentials (EPSPs) in the enteric nervous system. The slow EPSPs were mediated by tachykinin neurokinin 1 and CGRP receptors. Capsaicin evoked slow EPSP-like responses that were suppressed by antagonists for protease-activated receptor 2. Afferent stimulation evoked slow EPSP-like excitation that was suppressed by mast cell-stabilizing drugs. Histamine and mast cell protease II were released by 1) exposure to SP or CGRP, 2) capsaicin, 3) compound 48/80, 4) elevation of mast cell Ca²⁺ by ionophore A23187, and 5) antidromic electrical stimulation of afferents. The mast cell stabilizers cromolyn and doxantrazole suppressed release of protease II and histamine when evoked by SP, CGRP, capsaicin, A23187, electrical stimulation of afferents, or compound 48/80. Neural blockade by tetrodotoxin prevented mast cell protease II release in response to antidromic electrical stimulation of mesenteric afferents. The results support a hypothesis that afferent innervation of enteric mast cells releases histamine and mast cell protease II, both of which are known to act in a diffuse paracrine manner to influence the behavior of enteric nervous system neurons and to elevate the sensitivity of spinal afferent terminals.

  9. Innervation of enteric mast cells by primary spinal afferents in guinea pig and human small intestine

    PubMed Central

    Wang, Guo-Du; Wang, Xi-Yu; Liu, Sumei; Qu, Meihua; Xia, Yun; Needleman, Bradley J.; Mikami, Dean J.

    2014-01-01

    Mast cells express the substance P (SP) neurokinin 1 receptor and the calcitonin gene-related peptide (CGRP) receptor in guinea pig and human small intestine. Enzyme-linked immunoassay showed that activation of intramural afferents by antidromic electrical stimulation or by capsaicin released SP and CGRP from human and guinea pig intestinal segments. Electrical stimulation of the afferents evoked slow excitatory postsynaptic potentials (EPSPs) in the enteric nervous system. The slow EPSPs were mediated by tachykinin neurokinin 1 and CGRP receptors. Capsaicin evoked slow EPSP-like responses that were suppressed by antagonists for protease-activated receptor 2. Afferent stimulation evoked slow EPSP-like excitation that was suppressed by mast cell-stabilizing drugs. Histamine and mast cell protease II were released by 1) exposure to SP or CGRP, 2) capsaicin, 3) compound 48/80, 4) elevation of mast cell Ca2+ by ionophore A23187, and 5) antidromic electrical stimulation of afferents. The mast cell stabilizers cromolyn and doxantrazole suppressed release of protease II and histamine when evoked by SP, CGRP, capsaicin, A23187, electrical stimulation of afferents, or compound 48/80. Neural blockade by tetrodotoxin prevented mast cell protease II release in response to antidromic electrical stimulation of mesenteric afferents. The results support a hypothesis that afferent innervation of enteric mast cells releases histamine and mast cell protease II, both of which are known to act in a diffuse paracrine manner to influence the behavior of enteric nervous system neurons and to elevate the sensitivity of spinal afferent terminals. PMID:25147231

  10. Profiling human gut bacterial metabolism and its kinetics using [U-13C]glucose and NMR.

    PubMed

    de Graaf, Albert A; Maathuis, Annet; de Waard, Pieter; Deutz, Nicolaas E P; Dijkema, Cor; de Vos, Willem M; Venema, Koen

    2010-01-01

    This study introduces a stable-isotope metabolic approach employing [U-(13)C]glucose that, as a novelty, allows selective profiling of the human intestinal microbial metabolic products of carbohydrate food components, as well as the measurement of the kinetics of their formation pathways, in a single experiment. A well-established, validated in vitro model of human intestinal fermentation was inoculated with standardized gastrointestinal microbiota from volunteers. After culture stabilization, [U-(13)C]glucose was added as an isotopically labeled metabolic precursor. System lumen and dialysate samples were taken at regular intervals. Metabolite concentrations and isotopic labeling were determined by NMR, GC, and enzymatic methods. The main microbial metabolites were lactate, acetate, butyrate, formate, ethanol, and glycerol. They together accounted for a (13)C recovery rate as high as 91.2%. Using an NMR chemical shift prediction approach, several minor products that showed (13)C incorporation were identified as organic acids, amino acids, and various alcohols. Using computer modeling of the (12)C contents and (13)C labeling kinetics, the metabolic fluxes in the gut microbial pathways for synthesis of lactate, formate, acetate, and butyrate were determined separately for glucose and unlabeled background substrates. This novel approach enables the study of the modulation of human intestinal function by single nutrients, providing a new rational basis for achieving control of the short-chain fatty acids profile by manipulating substrate and microbiota composition in a purposeful manner.

  11. Listeria monocytogenes Inhibits Serotonin Transporter in Human Intestinal Caco-2 Cells.

    PubMed

    Latorre, E; Pradilla, A; Chueca, B; Pagán, R; Layunta, E; Alcalde, A I; Mesonero, J E

    2016-10-01

    Listeria monocytogenes is a Gram-positive bacterium that can cause a serious infection. Intestinal microorganisms have been demonstrated to contribute to intestinal physiology not only through immunological responses but also by modulating the intestinal serotonergic system. Serotonin (5-HT) is a neuromodulator that is synthesized in the intestinal epithelium and regulates the whole intestinal physiology. The serotonin transporter (SERT), located in enterocytes, controls intestinal 5-HT availability and therefore serotonin's effects. Infections caused by L. monocytogenes are well described as being due to the invasion of intestinal epithelial cells; however, the effect of L. monocytogenes on the intestinal epithelium remains unknown. The main aim of this work, therefore, was to study the effect of L. monocytogenes on SERT. Caco2/TC7 cell line was used as an enterocyte-like in vitro model, and SERT functional and molecular expression assays were performed. Our results demonstrate that living L. monocytogenes inhibits serotonin uptake by reducing SERT expression at the brush border membrane. However, neither inactivated L. monocytogenes nor soluble metabolites were able to affect SERT. The results also demonstrate that L. monocytogenes yields TLR2 and TLR10 transcriptional changes in intestinal epithelial cells and suggest that TLR10 is potentially involved in the inhibitory effect observed on SERT. Therefore, L. monocytogenes, through TLR10-mediated SERT inhibition, may induce increased intestinal serotonin availability and potentially contributing to intestinal physiological changes and the initiation of the inflammatory response.

  12. Culturable Aerobic and Facultative Anaerobic Intestinal Bacterial Flora of Black Cobra (Naja naja karachiensis) in Southern Pakistan

    PubMed Central

    Iqbal, Junaid; Sagheer, Mehwish; Tabassum, Nazneen; Siddiqui, Ruqaiyyah; Khan, Naveed Ahmed

    2014-01-01

    Using morphological analysis and biochemical testing, here for the first time, we determined the culturable gut bacterial flora (aerobes and facultative anaerobes) in the venomous Black Cobra (Naja naja karachiensis) from South Asia. The findings revealed that these snakes inhabit potentially pathogenic bacteria including Serratia marcescens, Pseudomonas aeruginosa, Shewanella putrefaciens, Aeromonas hydrophila, Salmonella sp., Moraxella sp., Bacillus sp., Ochrobactrum anthropi, and Providencia rettgeri. These findings are of concern, as injury from snake bite can result in wound infections and tissue necrosis leading to sepsis/necrotizing fasciitis and/or expose consumers of snake meat/medicine in the community to infections. PMID:25002979

  13. Ultrasound Elasticity Imaging for Detecting Intestinal Fibrosis and Inflammation in Rats and Humans With Crohn’s Disease

    PubMed Central

    Stidham, Ryan W.; Xu, Jingping; Johnson, Laura A.; Kim, Kang; Moons, David S.; Mckenna, Barbara J.; Rubin, Jonathan M.; Higgins, Peter D. R.

    2016-01-01

    BACKGROUND Intestinal fibrosis causes many complications of Crohn’s disease (CD). Available biomarkers and imaging modalities lack sufficient accuracy to distinguish intestinal inflammation from fibrosis. Transcutaneous ultrasound elasticity imaging (UEI) is a promising, noninvasive approach for measuring tissue mechanical properties. We hypothesized that UEI could differentiate inflammatory from fibrotic bowel wall changes in both animal models of colitis and humans with CD. METHODS Female Lewis rats underwent weekly trinitrobenzene sulfonic acid enemas yielding models of acute inflammatory colitis (n = 5) and chronic intestinal fibrosis (n = 6). UEI scanning used a novel speckle-tracking algorithm to estimate tissue strain. Resected bowel segments were evaluated for evidence of inflammation and fibrosis. Seven consecutive patients with stenotic CD were studied with UEI and their resected stenotic and normal bowel segments were evaluated by ex vivo elastometry and histopathology. RESULTS Transcutaneous UEI normalized strain was able to differentiate acutely inflamed (−2.07) versus chronic fibrotic (−1.10) colon in rat models of inflammatory bowel disease (IBD; P = .037). Transcutaneous UEI normalized strain also differentiated stenotic (−0.87) versus adjacent normal small bowel (−1.99) in human CD (P = .0008), and this measurement also correlated well with ex vivo elastometry (r = −0.81). CONCLUSIONS UEI can differentiate inflammatory from fibrotic intestine in rat models of IBD and can differentiate between fibrotic and unaffected intestine in a pilot study in humans with CD. UEI represents a novel technology with potential to become a new objective measure of progression of intestinal fibrosis. Prospective clinical studies in CD are needed. PMID:21784048

  14. Presence of drug resistance in intestinal lactobacilli of dairy and human origin in Turkey.

    PubMed

    Cataloluk, Osman; Gogebakan, Bulent

    2004-07-01

    The prevalence of different resistance genes was investigated in lactobacilli of human and dairy origin by PCR. The presence of erm, van, tet, and cat-TC genes were determined in 16 raw milk, 15 cream, 10 yogurt, 50 hand-made cheese, and 20 industrially produced white-cheese samples of dairy origin and 16 mouth, 32 fecal, and 36 vaginal samples from different subjects of human origin. Lactobacilli of dairy and human origin were found to carry only erm(B) and tet(M) genes. The majority of the isolates, Lactobacillus crispatus (61), Lactobacillus gasseri (49), Lactobacillus plantarum (80) studied were found to harbor either erm(B) or tet(M) gene or both. No resistant lactobacilli was found in raw-milk and cream samples. All the human fecal samples and the majority of vaginal (29 of 36) and mouth (10 of 14) samples were found to carry the resistance genes. While a third of the hand-made cheeses carried resistant lactobacilli only one industrially produced cheese was found to carry resistant lactobacilli. Furthermore, the genes were found in the non-starter species, Lactobacillus acidophilus and Lb. plantarum, indicating that industrially produced cheeses in this respect could be considered more favorable. These results indicate that drug resistance seems to be very common in Turkey. Even though the number of dairy samples harboring the resistance genes (17 of 111) is smaller in regards to human samples, 10% of them were still found to carry the resistance genes as well. The presence of the resistance genes in majority of the samples of human origin and in minority of the samples of dairy origin indicates that drug resistance may be acquired in the intestinal tract during passage and spread to dairy products by the hands of workers during production.

  15. Improved biocompatibility of small intestinal submucosa (SIS) following conditioning by human endothelial cells.

    PubMed

    Woods, A M; Rodenberg, E J; Hiles, M C; Pavalko, F M

    2004-02-01

    Small intestinal submucosa (SIS) is a naturally occurring tissue matrix composed of extracellular matrix proteins and various growth factors. SIS is derived from the porcine jejunum and functions as a remodeling scaffold for tissue repair. While SIS has proven to be a useful biomaterial for implants in vivo, problems associated with endothelialization and thrombogenicity of SIS implants may limit its vascular utility. The goal of this study was to determine if the biological properties of SIS could be improved by growing human umbilical vein endothelial cells (HUVEC) on SIS and allowing these cells to deposit human basement membrane proteins on the porcine substrate to create what we have called "conditioned" SIS (c-SIS). Using an approach in which HUVEC were grown for 2 weeks on SIS and then removed via a technique that leaves behind an intact basement membrane, we hypothesized that the surface properties of SIS might be improved. We found that when re-seeded on c-SIS, HUVEC exhibited enhanced organization of cell junctions and had increased metabolic activity compared to cells on native SIS (n-SIS). Furthermore, HUVEC grown on c-SIS released lower amounts of the pro-inflammatory prostaglandin PGI2 into the media compared to cells grown on n-SIS. Additionally, we found that adhesion of resting or activated human platelets to c-SIS was significantly decreased compared to n-SIS suggesting that, in addition to improved cell growth characteristics, conditioning SIS with human basement membrane proteins might decrease its thrombogenic potential. In summary, conditioning of porcine SIS by human endothelial cells improves key biological properties of the material that may improve its usefulness as remodeling scaffold for tissue repair. Identification of critical modifications of SIS by human endothelial cells should help guide future efforts to develop more biocompatible vascular grafts.

  16. Differential modulation of human intestinal bifidobacterium populations after consumption of a wild blueberry (Vaccinium angustifolium) drink.

    PubMed

    Guglielmetti, Simone; Fracassetti, Daniela; Taverniti, Valentina; Del Bo', Cristian; Vendrame, Stefano; Klimis-Zacas, Dorothy; Arioli, Stefania; Riso, Patrizia; Porrini, Marisa

    2013-08-28

    Bifidobacteria are gaining increasing interest as health-promoting bacteria. Nonetheless, the genus comprises several species, which can exert different effects on human host. Previous studies showed that wild blueberry drink consumption could selectively increase intestinal bifidobacteria, suggesting an important role for the polyphenols and fiber present in wild blueberries. This study evaluated the modulation of the most common and abundant bifidobacterial taxonomic groups inhabiting the human gut in the same fecal samples. The analyses carried out showed that B. adolescentis, B. breve, B. catenulatum/pseudocatelulatum, and B. longum subsp. longum were always present in the group of subjects enrolled, whereas B. bifidum and B. longum subsp. infantis were not. Furthermore, it was found that the most predominant bifidobacterial species were B. longum subsp. longum and B. adolescentis. The results obtained revealed a high interindividual variability; however, a significant increase of B. longum subsp. infantis cell concentration was observed in the feces of volunteers after the wild blueberry drink treatment. This bifidobacterial group was shown to possess immunomodulatory abilities and to relieve symptoms and promote the regression of several gastrointestinal disorders. Thus, an increased cell concentration of B. longum subsp. infantis in the human gut could be considered of potential health benefit. In conclusion, wild blueberry consumption resulted in a specific bifidogenic effect that could positively affect certain populations of bifidobacteria with demonstrated health-promoting properties.

  17. Alteration of a human intestinal microbiota under extreme life environment in the Antarctica.

    PubMed

    Jin, Jong-Sik; Touyama, Mutsumi; Yamada, Shin; Yamazaki, Takashi; Benno, Yoshimi

    2014-01-01

    The human intestinal microbiota (HIM) settles from birth and continues to change phenotype by some factors (e.g. host's diet) throughout life. However, the effect of extreme life environment on human HIM composition is not well known. To understand HIM fluctuation under extreme life environment in humans, fecal samples were collected from six Japanese men on a long Antarctic expedition. They explored Antarctica for 3 months and collected their fecal samples at once-monthly intervals. Using terminal restriction fragment length polymorphism (T-RFLP) and real time polymerase chain reaction (PCR) analysis, the composition of HIM in six subjects was investigated. Three subjects presented restoration of HIM after the expedition compared versus before and during the expedition. Two thirds samples collected during the expedition belonged to the same cluster in dendrogram. However, all through the expedition, T-RFLP patterns showed interindividual variability. Especially, Bifidobacterium spp. showed a tendency to decrease during and restore after the expedition. A reduction of Bifidobacterium spp. was observed in five subjects the first 1 month of the expedition. Bacteroides thetaiotaomicron, which is thought to proliferate during emotional stress, significantly decreased in one subject, indicating that other factors in addition to emotional stress may affect the composition of HIM in this study. These findings could be helpful to understand the effect of extreme life environment on HIM.

  18. Carboxylated nanodiamonds are neither cytotoxic nor genotoxic on liver, kidney, intestine and lung human cell lines.

    PubMed

    Paget, V; Sergent, J A; Grall, R; Altmeyer-Morel, S; Girard, H A; Petit, T; Gesset, C; Mermoux, M; Bergonzo, P; Arnault, J C; Chevillard, S

    2014-08-01

    Although nanodiamonds (NDs) appear as one of the most promising nanocarbon materials available so far for biomedical applications, their risk for human health remains unknown. Our work was aimed at defining the cytotoxicity and genotoxicity of two sets of commercial carboxylated NDs with diameters below 20 and 100 nm, on six human cell lines chosen as representative of potential target organs: HepG2 and Hep3B (liver), Caki-1 and Hek-293 (kidney), HT29 (intestine) and A549 (lung). Cytotoxicity of NDs was assessed by measuring cell impedance (xCELLigence® system) and cell survival/death by flow cytometry while genotoxicity was assessed by γ-H2Ax foci detection, which is considered the most sensitive technique for studying DNA double-strand breaks. To validate and check the sensitivity of the techniques, aminated polystyrene nanobeads were used as positive control in all assays. Cell incorporation of NDs was also studied by flow cytometry and luminescent N-V center photoluminescence (confirmed by Raman microscopy), to ensure that nanoparticles entered the cells. Overall, we show that NDs effectively entered the cells but NDs do not induce any significant cytotoxic or genotoxic effects on the six cell lines up to an exposure dose of 250 µg/mL. Taken together these results strongly support the huge potential of NDs for human nanomedicine but also their potential as negative control in nanotoxicology studies.

  19. Total Body Irradiation in the "Hematopoietic" Dose Range Induces Substantial Intestinal Injury in Non-Human Primates.

    PubMed

    Wang, Junru; Shao, Lijian; Hendrickson, Howard P; Liu, Liya; Chang, Jianhui; Luo, Yi; Seng, John; Pouliot, Mylene; Authier, Simon; Zhou, Daohong; Allaben, William; Hauer-Jensen, Martin

    2015-11-01

    The non-human primate has been a useful model for studies of human acute radiation syndrome (ARS). However, to date structural changes in various parts of the intestine after total body irradiation (TBI) have not been systematically studied in this model. Here we report on our current study of TBI-induced intestinal structural injury in the non-human primate after doses typically associated with hematopoietic ARS. Twenty-four non-human primates were divided into three groups: sham-irradiated control group; and total body cobalt-60 (60Co) 6.7 Gy gamma-irradiated group; and total body 60Co 7.4 Gy gamma-irradiated group. After animals were euthanized at day 4, 7 and 12 postirradiation, sections of small intestine (duodenum, proximal jejunum, distal jejunum and ileum) were collected and fixed in 10% formalin. The intestinal mucosal surface length, villus height and crypt depths were assessed by computer-assisted image analysis. Plasma citrulline levels were determined using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Total bone marrow cells were counted and hematopoietic stem/progenitor cells in bone marrow were analyzed by flow cytometer. Histopathologically, all segments exhibited conspicuous disappearance of plicae circulares and prominent atrophy of crypts and villi. Intestinal mucosal surface length was significantly decreased in all intestinal segments on day 4, 7 and 12 after irradiation (P < 0.02-P < 0.001). Villus height was significantly reduced in all segments on day 4 and 7 (P = 0.02-0.005), whereas it had recovered by day 12 (P > 0.05). Crypt depth was also significantly reduced in all segments on day 4, 7 and 12 after irradiation (P < 0.04-P < 0.001). Plasma citrulline levels were dramatically reduced after irradiation, consistent with intestinal mucosal injury. Both 6.7 and 7.4 Gy TBI reduced total number of bone marrow cells. And further analysis showed that the number and function of CD45(+)CD34(+) hematopoietic stem/progenitors in bone

  20. Metformin Transport by a Newly Cloned Proton-Stimulated Organic Cation Transporter (Plasma Membrane Monoamine Transporter) Expressed in Human Intestine

    PubMed Central

    Zhou, Mingyan; Xia, Li; Wang, Joanne

    2009-01-01

    Metformin is a widely used oral antihyperglycemic drug for the treatment of type II diabetes mellitus. The intestinal absorption of metformin is dose-dependent and involves an active, saturable uptake process. Metformin has been shown to be transported by the human organic cation transporters 1 and 2 (hOCT1–2). We recently cloned and characterized a novel proton-activated organic cation transporter, plasma membrane monoamine transporter (PMAT). We previously showed that PMAT transports many classic organic cations (e.g., monoamine neurotransmitters, 1-methyl-4-phenylpyridinium) in a pH-dependent manner and its mRNA is expressed in multiple human tissues. The goal of this study is to investigate whether metformin is a substrate of PMAT and whether PMAT plays a role in the intestinal uptake of metformin. Using Madin-Darby canine kidney cells stably expressing human PMAT, we showed that metformin is avidly transported by PMAT, with an apparent affinity (Km = 1.32 mM) comparable to those reported for hOCT1–2. Interestingly, the concentration-velocity profile of PMAT-mediated metformin uptake is sigmoidal, with a Hill coefficient of 2.64. PMAT-mediated metformin transport is greatly stimulated by acidic pH, with the uptake rate being ~4-fold higher at pH 6.6 than at pH 7.4. Using a polyclonal antibody against PMAT, we showed that the PMAT protein (58 kDa) was expressed in human small intestine and concentrated on the tips of the mucosal epithelial layer. Taken together, our results suggest that PMAT transports metformin, is expressed in human intestine, and may play a role in the intestinal absorption of metformin and possibly other cationic drugs. PMID:17600084

  1. Bacterial fermentation affects net mineral flux in the large intestine of pigs fed diets with viscous and fermentable nonstarch polysaccharides.

    PubMed

    Metzler-Zebeli, B U; Hooda, S; Mosenthin, R; Gänzle, M G; Zijlstra, R T

    2010-10-01

    The impact of colonic fermentation on postileal absorption of Ca, Mg, P, Cu, Fe, Mn, and Zn was investigated in 8 ileally cannulated grower pigs (initial BW = 29.1 ± 1.6 kg) according to a double 4 × 4 Latin square. A semi-purified diet was supplemented with 5.20% low viscous, low fermentable cellulose (CEL), 6.25% high viscous, low fermentable carboxymethylcellulose (CMC), 8.95% low viscous, high fermentable oat beta-glucan (LG), or 9.25% high viscous, high fermentable oat beta-glucan (HG), resulting in 5% actual added nonstarch polysaccharides (NSP) in the diets. Because of the intrinsic mineral content in LG and HG, pigs receiving the LG and HG diets had a greater (P < 0.05) daily intake of Mg, P, Cu, Fe, Mn, and Zn, and also Ca for the HG diet compared with the CEL and CMC diets. Different amounts of minerals reached the large intestine for the 4 diets as indicated by the 60 to 86% less (P < 0.05) ileal flow of Ca, Mg, P, and Fe for CMC compared with CEL and HG. Apparent mineral retention was generally less (P < 0.05) for CEL compared with CMC. Regression analyses indicated that postileal flux of Ca, Cu, and Zn were related (R(2) = 0.24 to 0.99; P < 0.05) to short-chain fatty acid (SCFA) concentrations in feces. Postileal Ca absorption was negatively related (R(2) = 0.24; P < 0.05) to fecal total SCFA concentrations when SCFA concentrations were greater than 95.6 mmol/kg of DM. Furthermore, postileal Zn (R(2) = 0.99; P < 0.001) and Cu secretion (R(2) = 0.94; P < 0.001) decreased with increasing total SCFA concentrations in feces. Additionally, postileal secretion of Fe increased (R(2) = 0.20; P < 0.05) with increasing 16S rRNA gene copies of Enterobacteriaceae in feces, whereas the secretion of Cu decreased (R(2) = 0.25; P < 0.01) with increasing gene copies of Enterobacteriaceae. Overall, the apparent retention of Ca, Mg, and P was 27 to 85% less (P < 0.05) for CEL and HG than for CMC, whereas the apparent retention of Fe, Mn, and Zn was less (P < 0.05) for

  2. Human monocyte-derived dendritic cells from leukoreduction system chambers after plateletpheresis are functional in an in vitro co-culture assay with intestinal epithelial cells.

    PubMed

    Tiscornia, Inés; Sánchez-Martins, Viviana; Hernández, Ana; Bollati-Fogolín, Mariela

    2012-10-31

    The dendritic cells (DC) found in the intestine are involved both in the maintenance of tolerance towards commensal microbiota, and in the generation of protective immune responses against pathogens, thus contributing to gut immune homeostasis. There is an increasing interest in the use of lactic acid bacteria (LAB) as probiotics; among their beneficial effects we highlight the modulation of the immune system which is one of their fundamental properties. As these effects are strain-dependent, it is important to have in vitro systems that include DC and intestinal epithelial cells (IEC), which are crucial for intestinal homeostasis, to identify candidates by means of bacterial screening. Obtaining enough human cells, necessary to simultaneously test several bacteria, is a major challenge for researchers. In this study we analyzed the usefulness of the cellular fraction retained in leukoreduction system chambers following plateletpheresis (PP) as a source of DC. We compared the capacity of peripheral blood mononuclear cells (PBMC) from buffy coats (BC) or PP to generate DC using a short differentiation protocol. The functionality of the DC obtained was analyzed in co-cultures together with intestinal epithelial HT-29 cells, stimulating with LPS alone or with two LAB commonly used in the food industry, Streptococcus thermophilus and Lactobacillus delbrueckii. DC surface markers CD86, HLA-DR and cytokine production were measured. The behavior of DC derived from PP was similar to the behavior observed for DC derived from BC. When we tested the response of DC to bacteria, we found significant differences in cytokine secretion, especially for IL-10, suggesting that the system has the ability to discriminate LAB with different immunomodulatory properties. We also found that DC derived from both sources displayed a similar ability to phagocyte bacteria. In conclusion, we hereby propose a modification of the two-day protocol for obtaining human DC previously described, using

  3. Effects of dietary antibiotic growth promoter and Saccharomyces cerevisiae fermentation product on production, intestinal bacterial community, and nonspecific immunity of hybrid tilapia (Oreochromis niloticus female x Oreochromis aureus male).

    PubMed

    He, S; Zhou, Z; Meng, K; Zhao, H; Yao, B; Ringø, E; Yoon, I

    2011-01-01

    To investigate the effects of a dietary antibiotic growth promoter (florfenicol) and a Saccharomyces cerevisiae fermentation product (DVAQUA) on growth, G:F, daily feed intake, intestinal bacterial community, and nonspecific immunity of hybrid tilapia (Oreochromis niloticus ♀ × Oreochromis aureus ♂), a 16-wk feeding trial was conducted in a recirculating aquaculture system. Four feeding regimens were evaluated: control, dietary florenicol (0.02 g/kg; 16 wk), dietary DVAQUA (0.5 g/kg; 16 wk), and sequential use of florenicol (0.02 g/kg; 8 wk), and DVAQUA (0.5 g/kg; 8 wk). Each regimen had 4 replicate tanks (0.5 × 0.5 × 0.5 m) and each tank contained 12 fish (initial BW: 46.88 ± 0.38 g). Dietary florfenicol improved growth (P = 0.089), G:F (P = 0.036), and serum complement component concentrations (P < 0.001) of hybrid tilapia. However, the compound decreased the estimated intestinal bacterial count estimated by rpoB quantitative PCR (P < 0.001) and bacterial diversity (visual band numbers, Shannon diversity index, and Shannon equitability index based on 16S rDNA V3 denaturing gradient gel electrophoresis fingerprints) compared with the control. Although sequential use of florfenicol and DVAQUA improved growth and G:F numerically to a similar extent as dietary florfenicol, and increased intestinal bacterial count to normal quantities, the sequential use of florenicol and DVAQUA decreased intestinal bacterial diversity (visual band numbers, Shannon diversity index, and Shannon equitability index) as well as serum complement component concentrations (P < 0.001) compared with their respective use and the control. These findings might be negatively related to disease control and host defense, and the sequential use of florenicol and DVAQUA should be practiced with caution. Feeding DAVQUA to the fish improved nonspecific immunity and increased intestinal bacterial count and bacterial diversity, but further research, including challenge studies, should be conducted

  4. Epidemiology of infections with intestinal parasites and human immunodeficiency virus (HIV) among sugar-estate residents in Ethiopia.

    PubMed

    Fontanet, A L; Sahlu, T; Rinke de Wit, T; Messele, T; Masho, W; Woldemichael, T; Yeneneh, H; Coutinho, R A

    2000-04-01

    Intestinal parasitic infections could play an important role in the progression of infection with human immunodeficiency virus (HIV), by further disturbing the immune system whilst it is already engaged in the fight against HIV. HIV and intestinal parasitic infections were investigated in 1239, randomly selected individuals, aged 15-54 years, living on a sugar estate in central Ethiopia. Intestinal parasites were identified in faecal samples (one/subject) using direct, concentration, and (for Strongyloides stercoralis larvae) Baermann methods. HIV serological status was determined using ELISA, with ELISA-positive samples confirmed as positive by western blotting. Most (70.1%) of the subjects were infected with at least one intestinal parasite and 3.1% were seropositive (but asymptomatic) for HIV. The intestinal parasites identified in the study population were amoebic parasites (Entamoeba histolytica/Enta. dispar) (24.6%), hookworms (23.8%), Ascaris lumbricoides (22.2%), Trichuris trichiura (19.5%), S. stercoralis (13.0%), Taenia saginata (4.5%), Giardia lamblia (3.0%), and Enterobius vermicularis (1.3%). Overall, the HIV-positives were no more or less likely to carry intestinal parasites than the HIV-negatives (76.2% v. 69.9%; P > 0.05). However, when each parasite was considered separately, amoebic parasites were found to be more common in the HIV-positives than the HIV-negatives (43.7% v. 24.0%; P < 0.05). This difference remained significant in a multivariate analysis, after controlling for the socio-demographic characteristics of the study participants. In conclusion, there was moderate interaction between intestinal parasites and HIV at the asymptomatic stage of HIV infection. The observed association between amoebic and HIV infections requires confirmation in a prospective study, allowing for the analysis of biological mechanisms involved in the association.

  5. Physical stress and bacterial colonization

    PubMed Central

    Otto, Michael

    2014-01-01

    Bacterial surface colonizers are subject to a variety of physical stresses. During the colonization of human epithelia such as on the skin or the intestinal mucosa, bacteria mainly have to withstand the mechanical stress of being removed by fluid flow, scraping, or epithelial turnover. To that end, they express a series of molecules to establish firm attachment to the epithelial surface, such as fibrillar protrusions (pili) and surface-anchored proteins that bind to human matrix proteins. In addition, some bacteria – in particular gut and urinary tract pathogens – use internalization by epithelial cells and other methods such as directed inhibition of epithelial turnover to ascertain continued association with the epithelial layer. Furthermore, many bacteria produce multi-layered agglomerations called biofilms with a sticky extracellular matrix, providing additional protection from removal. This review will give an overview over the mechanisms human bacterial colonizers have to withstand physical stresses with a focus on bacterial adhesion. PMID:25212723

  6. Novel GM1 ganglioside-like peptide mimics prevent the association of cholera toxin to human intestinal epithelial cells in vitro.

    PubMed

    Yu, Robert K; Usuki, Seigo; Itokazu, Yutaka; Wu, Han-Chung

    2016-01-01

    Cholera is an acute diarrheal disease caused by infection in the gastrointestinal tract by the gram-negative bacterium, Vibrio cholerae, and is a serious public health threat worldwide. There has not been any effective treatment for this infectious disease. Cholera toxin (CT), which is secreted by V. cholerae, can enter host cells by binding to GM1, a monosialoganglioside widely distributed on the plasma membrane surface of various animal epithelial cells. The present study was undertaken to generate peptides that are conformationally similar to the carbohydrate epitope of GM1 for use in the treatment of cholera and related bacterial infection. For this purpose, we used cholera toxin B (CTB) subunit to select CTB-binding peptides that structurally mimic GM1 from a dodecamer phage-display library. Six GM1-replica peptides were selected by biopanning based on CTB recognition. Five of the six peptides showed inhibitory activity for GM1 binding to CTB. To test the potential of employing the peptide mimics for intervening with the bacterial infection, those peptides were examined for their binding capacity, functional inhibitory activity and in vitro effects using a human intestinal epithelial cell line, Caco-2 cells. One of the peptides, P3 (IPQVWRDWFKLP), was most effective in inhibiting cellular uptake of CTB and suppressing CT-stimulated cyclic adenosine monophosphate production in the cells. Our results thus provide convincing evidence that GM1-replica peptides could serve as novel agents to block CTB binding on epithelial cells and prevent the ensuing physiological effects of CT.

  7. PCR DGGE and RT-PCR DGGE show diversity and short-term temporal stability in the Clostridium coccoides-Eubacterium rectale group in the human intestinal microbiota.

    PubMed

    Maukonen, Johanna; Mättö, Jaana; Satokari, Reetta; Söderlund, Hans; Mattila-Sandholm, Tiina; Saarela, Maria

    2006-12-01

    As the Clostridium coccoides-Eubacterium rectale (Erec; clostridial phylogenetic cluster XIVa) group is one of the major groups of the human intestinal microbiota, DNA- and RNA-based population analysis techniques (denaturing gradient gel electrophoresis; DGGE) were developed and applied to assess the diversity and temporal stability (6 months-2 years) of this faecal clostridial microbiota in 12 healthy adults. The stability of the Erec group was compared with the stability of the predominant bacterial microbiota, which was also assessed with PCR-DGGE. In addition, the Erec group was quantified with a hybridization-based method. According to our results, the Erec group was diverse in each subject, but interindividual uniqueness was not as clear as that of the predominant bacteria. The Erec group was found to be temporally as stable as the predominant bacteria. Over 200 clones obtained from two samples proved the developed method to be specific. However, the amount of bacteria belonging to the Erec group was not related to the diversity of that same bacterial group. In conclusion, the newly developed DGGE method proved to be a valuable and specific tool for the direct assessment of the stability of the Erec group, demonstrating diversity in addition to short-term stability in most of the subjects studied.

  8. Effects of dietary inclusion of silymarin on performance, intestinal morphology and ileal bacterial count in aflatoxin-challenged broiler chicks.

    PubMed

    Jahanian, E; Mahdavi, A H; Asgary, S; Jahanian, R

    2017-01-04

    This study was conducted to investigate the effect of dietary supplementation of silymarin on performance, jejunal morphology and ileal bacterial population in broiler chicks intoxicated with a mix of aflatoxins. A total of three hundred thirty six 7-day-old Ross broiler chicks were randomly distributed between seven experimental groups with four replicates of 12 birds each. Experimental treatments consisted of a control group (unchallenged), and a 2 × 3 factorial arrangement, including two aflatoxin levels (0.5 and 2 ppm) and three levels of silymarin (0, 500 and 1000 ppm). Birds were challenged with a mix of aflatoxins from 7 to 28 days of age. Results showed that increasing aflatoxin level resulted in decreased average daily feed intake (ADFI) and weight gain (ADWG), consequently impaired feed conversion ratio (FCR) throughout the trial period. Dietary supplementation of silymarin resulted in the marked increases in ADFI and ADWG, and improved FCR values in aflatoxin-challenged chicks. Ileal bacterial populations at days 28 and 42 of age were increased by incremental levels of aflatoxins. On the other hand, dietary silymarin supplementation suppressed ileal populations of Escherichia coli, Salmonella, Klebsiella and total negative bacteria in aflatoxicated birds. Increase in dietary aflatoxin level resulted in the decreased villi height, villi height-to-crypt depth ratio (VH:CD), villi surface area and apparent villi absorptive area, while it increased crypt depth, goblet cell count and lymphoid follicular diameter. Feeding silymarin at the level of 1000 ppm increased villi height and VH:CD in aflatoxicated birds. Present results indicate that dietary inclusion of silymarin could improve performance by suppressing ileal bacteria and enhancing absorptive surface area in aflatoxin-challenged broiler chicks.

  9. Evaluation of physicochemical properties and intestinal permeability of six dietary polyphenols in human intestinal colon adenocarcinoma Caco-2 cells.

    PubMed

    Rastogi, Himanshu; Jana, Snehasis

    2016-02-01

    Phenolic compounds are common ingredients in many dietary supplements and functional foods. However, data concerning physicochemical properties and permeability of polyphenols on the intestinal epithelial cells are scarce. The aims of this study were to determine the experimental partition coefficient (Log P), and parallel artificial membrane permeability assay (PAMPA), to characterize the bi-directional transport of six phenolic compounds viz. caffeic acid, chrysin, gallic acid, quercetin, resveratrol and rutin in Caco-2 cells. The experimental Log P values of six polyphenols were correlated (R (2) = 0.92) well with the calculated Log P values. The apparent permeability (P app) range of all polyphenols in PAMPA for the apical (AP) to basolateral (BL) was 1.18 ± 0.05 × 10(-6) to 5.90 ± 0.16 × 10(-6) cm/s. The apparent Caco-2 permeability (P app) range for the AP-BL was 0.96 ± 0.03 × 10(-6) to 3.80 ± 0.45 × 10(-6) cm/s. The efflux ratio of P app (BL → AP) to P app (AP → BL) for all phenolics was <2, suggesting greater permeability in the absorptive direction. Six compounds exhibited strong correlations between Log P and PAMPA/Caco-2 cell monolayer permeation data. Dietary six polyphenols were poorly absorbed through PAMPA and Caco-2 cells, and their transepithelial transports were mainly by passive diffusion.

  10. Studies on the bioavailability of zinc in humans: intestinal interaction of tin and zinc.

    PubMed

    Solomons, N W; Marchini, J S; Duarte-Favaro, R M; Vannuchi, H; Dutra de Oliveira, J E

    1983-04-01

    Mineral/mineral interactions at the intestinal level are important in animal nutrition and toxicology, but only limited understanding of their extent or importance in humans has been developed. An inhibitory interaction of dietary tin on zinc retention has been recently described from human metabolic studies. We have explored the tin/zinc interaction using the change-in-plasma-zinc-concentration method with a standard dosage of 12.5 mg of zinc as zinc sulfate in 100 ml of Coca-Cola. Sn/Zn ratios of 2:1, 4:1, and 8:1, constituted by addition of 25, 50, and 100 mg of tin as stannous chloride, had no significant overall effect on zinc uptake. The 100-mg dose of tin produced noxious gastrointestinal symptoms. Addition of iron as ferrous sulfate to form ratios of Sn/Fe/Zn of 1:1:1 and 2:2:1 with the standard zinc solution and the appropriate doses of tin produced a reduction of zinc absorption not dissimilar from that seen previously with zinc and iron alone, and addition of picolinic acid did not influence the uptake of zinc from the solution with the 2:2:1 Sn/Fe/Zn ratio.

  11. Bidirectional intragraft alloreactivity drives the repopulation of human intestinal allografts and correlates with clinical outcome.

    PubMed

    Zuber, Julien; Shonts, Brittany; Lau, Sai-Ping; Obradovic, Aleksandar; Fu, Jianing; Yang, Suxiao; Lambert, Marion; Coley, Shana; Weiner, Joshua; Thome, Joseph; DeWolf, Susan; Farber, Donna L; Shen, Yufeng; Caillat-Zucman, Sophie; Bhagat, Govind; Griesemer, Adam; Martinez, Mercedes; Kato, Tomoaki; Sykes, Megan

    2016-10-01

    A paradigm in transplantation states that graft-infiltrating T cells are largely non-alloreactive "bystander" cells. However, the origin and specificity of allograft T cells over time has not been investigated in detail in animals or humans. Here, we use polychromatic flow cytometry and high throughput TCR sequencing of serial biopsies to show that gut-resident T cell turnover kinetics in human intestinal allografts are correlated with the balance between intra-graft host-vs-graft (HvG) and graft-vs-host (GvH) reactivities and with clinical outcomes. In the absence of rejection, donor T cells were enriched for GvH-reactive clones that persisted long-term in the graft. Early expansion of GvH clones in the graft correlated with rapid replacement of donor APCs by the recipient. Rejection was associated with transient infiltration by blood-like recipient CD28+ NKG2D(Hi) CD8+ alpha beta T cells, marked predominance of HvG clones, and accelerated T cell turnover in the graft. Ultimately, these recipient T cells acquired a steady state tissue-resident phenotype, but regained CD28 expression during rejections. Increased ratios of GvH to HvG clones were seen in non-rejectors, potentially mitigating the constant threat of rejection posed by HvG clones persisting within the tissue-resident graft T cell population.

  12. Bidirectional intragraft alloreactivity drives the repopulation of human intestinal allografts and correlates with clinical outcome

    PubMed Central

    Zuber, Julien; Obradovic, Aleksandar; Fu, Jianing; Yang, Suxiao; Lambert, Marion; Coley, Shana; Weiner, Joshua; Thome, Joseph; DeWolf, Susan; Farber, Donna L.; Shen, Yufeng; Caillat-Zucman, Sophie; Bhagat, Govind; Griesemer, Adam; Martinez, Mercedes; Kato, Tomoaki; Sykes, Megan

    2016-01-01

    A paradigm in transplantation states that graft-infiltrating T cells are largely non-alloreactive “bystander” cells. However, the origin and specificity of allograft T cells over time has not been investigated in detail in animals or humans. Here, we use polychromatic flow cytometry and high throughput TCR sequencing of serial biopsies to show that gut-resident T cell turnover kinetics in human intestinal allografts are correlated with the balance between intra-graft host-vs-graft (HvG) and graft-vs-host (GvH) reactivities and with clinical outcomes. In the absence of rejection, donor T cells were enriched for GvH-reactive clones that persisted long-term in the graft. Early expansion of GvH clones in the graft correlated with rapid replacement of donor APCs by the recipient. Rejection was associated with transient infiltration by blood-like recipient CD28+ NKG2DHi CD8+ alpha beta T cells, marked predominance of HvG clones, and accelerated T cell turnover in the graft. Ultimately, these recipient T cells acquired a steady state tissue-resident phenotype, but regained CD28 expression during rejections. Increased ratios of GvH to HvG clones were seen in non-rejectors, potentially mitigating the constant threat of rejection posed by HvG clones persisting within the tissue-resident graft T cell population. PMID:28239678

  13. Molecular Paleoparasitological Hybridization Approach as Effective Tool for Diagnosing Human Intestinal Parasites from Scarce Archaeological Remains

    PubMed Central

    Jaeger, Lauren Hubert; Iñiguez, Alena Mayo

    2014-01-01

    Paleoparasitology is the science that uses parasitological techniques for diagnosing parasitic diseases in the past. Advances in molecular biology brought new insights into this field allowing the study of archaeological material. However, due to technical limitations a proper diagnosis and confirmation of the presence of parasites is not always possible, especially in scarce and degraded archaeological remains. In this study, we developed a Molecular Paleoparasitological Hybridization (MPH) approach using ancient DNA (aDNA) hybridization to confirm and complement paleoparasitological diagnosis. Eight molecular targets from four helminth parasites were included: Ascaris sp., Trichuris trichiura, Enterobius vermicularis, and Strongyloides stercoralis. The MPH analysis using 18th century human remains from Praça XV cemetery (CPXV), Rio de Janeiro, Brazil, revealed for the first time the presence E. vermicularis aDNA (50%) in archaeological sites of Brazil. Besides, the results confirmed T. trichiura and Ascaris sp. infections. The prevalence of infection by Ascaris sp. and E. vermicularis increased considerably when MPH was applied. However, a lower aDNA detection of T. trichiura (40%) was observed when compared to the diagnosis by paleoparasitological analysis (70%). Therefore, based on these data, we suggest a combination of Paleoparasitological and MPH approaches to verify the real panorama of intestinal parasite infection in human archeological samples. PMID:25162694

  14. Soluble Human Intestinal Lactoferrin Receptor: Ca(2+)-Dependent Binding to Sepharose-Based Matrices.

    PubMed

    Oshima, Yuta; Seki, Kohei; Shibuya, Masataka; Naka, Yuki; Yokoyama, Tatsuya; Sato, Atsushi

    2016-01-01

    A soluble form of human intestinal lactoferrin receptor (shLFR) is identical to human intelectin-1 (hITLN-1), a galactofuranose-binding protein that acts as a host defense against invading pathogenic microorganisms. We found that recombinant shLFR, expressed in mammalian cells (CHO DG44, COS-1, and RK13), binds tightly to Sepharose 4 Fast Flow (FF)-based matrices in a Ca(2+)-dependent manner. This binding of shLFR to Sepharose 4 FF-based matrices was inhibited by excess D-galactose, but not by D-glucose, suggesting that shLFR recognizes repeating units of α-1,6-linked D-galactose in Sepharose 4 FF. Furthermore, shLFR could bind to both Sepharose 4B- and Sepharose 6B-based matrices that were not crosslinked in a similar manner as to Sepharose 4 FF-based matrices. Therefore, shLFR (hITLN-1) binds to Sepharose-based matrices in a Ca(2+)-dependent manner. This binding property is most likely related to the ability, as host defense lectins, to recognize sepharose (agarobiose)-like structures present on the surface of invading pathogenic microorganisms.

  15. Effect of the artificial sweetener, sucralose, on small intestinal glucose absorption in healthy human subjects.

    PubMed

    Ma, Jing; Chang, Jessica; Checklin, Helen L; Young, Richard L; Jones, Karen L; Horowitz, Michael; Rayner, Christopher K

    2010-09-01

    It has been reported that the artificial sweetener, sucralose, stimulates glucose absorption in rodents by enhancing apical availability of the transporter GLUT2. We evaluated whether exposure of the proximal small intestine to sucralose affects glucose absorption and/or the glycaemic response to an intraduodenal (ID) glucose infusion in healthy human subjects. Ten healthy subjects were studied on two separate occasions in a single-blind, randomised order. Each subject received an ID infusion of sucralose (4 mM in 0.9% saline) or control (0.9% saline) at 4 ml/min for 150 min (T = - 30 to 120 min). After 30 min (T = 0), glucose (25 %) and its non-metabolised analogue, 3-O-methylglucose (3-OMG; 2.5 %), were co-infused intraduodenally (T = 0-120 min; 4.2 kJ/min (1 kcal/min)). Blood was sampled at frequent intervals. Blood glucose, plasma glucagon-like peptide-1 (GLP-1) and serum 3-OMG concentrations increased during ID glucose/3-OMG infusion (P < 0.005 for each). However, there were no differences in blood glucose, plasma GLP-1 or serum 3-OMG concentrations between sucralose and control infusions. In conclusion, sucralose does not appear to modify the rate of glucose absorption or the glycaemic or incretin response to ID glucose infusion when given acutely in healthy human subjects.

  16. Challenges of culturing human norovirus in three-dimensional organoid intestinal cell culture models.

    PubMed

    Papafragkou, Efstathia; Hewitt, Joanne; Park, Geun Woo; Greening, Gail; Vinjé, Jan

    2014-01-01

    Human noroviruses are the most common cause of acute gastroenteritis worldwide. Recently, cell culture systems have been described using either human embryonic intestinal epithelial cells (Int-407) or human epithelial colorectal adenocarcinoma cells (Caco-2) growing on collagen-I porous micro carrier beads in a rotating bioreactor under conditions of physiological fluid shear. Here, we describe the efforts from two independent laboratories to implement this three dimensional (3D) cell culture system for the replication of norovirus. Int-407 and Caco-2 were grown in a rotating bioreactor for up to 28 days. Prior to infection, cells were screened for the presence of microvilli by electron microscopy and stained for junction proteins (zonula occludens-1, claudin-1, and β-catenin). Differentiated 3D cells were transferred to 24-well plates and infected with bacteria-free filtrates of various norovirus genotypes (GI.1, GI.3, GI.8, GII.2, GII.4, GII.7, and GII.8). At 12 h, 24 h, and 48 h post inoculation, viral RNA from both cells and supernatants were collected and analyzed for norovirus RNA by real-time reverse transcription PCR. Despite observations of high expression of junction proteins and microvilli development in stained thin sections, our data suggest no significant increase in viral titer based on norovirus RNA copy number during the first 48 h after inoculation for the different samples and virus culture conditions tested. Our combined efforts demonstrate that 3D cell culture models using Int-407 or Caco-2 cells do not support norovirus replication and highlight the complexity and difficulty of developing a reproducible in vitro cell culture system for human norovirus.

  17. Systemic and mucosal immune responses following oral adenoviral delivery of influenza vaccine to the human intestine by radio controlled capsule

    PubMed Central

    Kim, Leesun; Martinez, C. Josefina; Hodgson, Katie A.; Trager, George R.; Brandl, Jennifer R.; Sandefer, Erik P.; Doll, Walter J.; Liebowitz, Dave; Tucker, Sean N.

    2016-01-01

    There are several benefits of oral immunization including the ability to elicit mucosal immune responses that may protect against pathogens that invade through a mucosal surface. Our understanding of human immune biology is hampered by the difficulty in isolating mucosal cells from humans, and the fact that animal models may or may not completely mirror human intestinal immunobiology. In this human pharmacodynamic study, a novel adenovirus vector-based platform expressing influenza hemagglutinin was explored. We used radio-controlled capsules to deliver the vaccine to either the jejunum or the ileum. The resulting immune responses induced by immunization at each of the intestinal sites were investigated. Both intestinal sites were capable of inducing mucosal and systemic immune responses to influenza hemagglutinin, but ileum delivery induced higher numbers of antibody secreting cells of IgG and IgA isotypes, increased mucosal homing B cells, and higher number of vaccine responders. Overall, these data provided substantial insights into human mucosal inductive sites, and aided in the design and selection of indications that could be used with this oral vaccine platform. PMID:27881837

  18. Identification of household bacterial community and analysis of species shared with human microbiome.

    PubMed

    Jeon, Yoon-Seong; Chun, Jongsik; Kim, Bong-Soo

    2013-11-01

    Microbial populations in indoor environments, where we live and eat, are important for public health. Various bacterial species reside in the kitchen, and refrigerators, the major means of food storage within kitchens, can be a direct source of food borne illness. Therefore, the monitoring of microbiota in the refrigerator is important for food safety. We investigated and compared bacterial communities that reside in the vegetable compartment of the refrigerator and on the seat of the toilet, which is recognized as highly colonized by microorganisms, in ten houses using high-throughput sequencing. Proteobacteria, Firmicutes, Actinobacteria, and Bacteroidetes were predominant in refrigerator and toilet samples. However, Proteobacteria was more abundant in the refrigerator, and Firmicutes was more abundant in the toilet. These household bacterial communities were compared with those of human skin and gut to identify potential sources of household bacteria. Bacterial communities from refrigerators and toilets shared more species in common with human skin than gut. Opportunistic pathogens, including Propionibacterium acnes, Bacteroides vulgatus, and Staphylococcus epidermidis, were identified as species shared with human skin and gut microbiota. This approach can provide a general background of the household microbiota and a potential method of source-tracking for public health purposes.

  19. Characterization of the diversity and temporal stability of bacterial communities in human milk.

    PubMed

    Hunt, Katherine M; Foster, James A; Forney, Larry J; Schütte, Ursel M E; Beck, Daniel L; Abdo, Zaid; Fox, Lawrence K; Williams, Janet E; McGuire, Michelle K; McGuire, Mark A

    2011-01-01

    Recent investigations have demonstrated that human milk contains a variety of bacterial genera; however, as of yet very little work has been done to characterize the full diversity of these milk bacterial communities and their relative stability over time. To more thoroughly investigate the human milk microbiome, we utilized microbial identification techniques based on pyrosequencing of the 16S ribosomal RNA gene. Specifically, we characterized the bacterial communities present in milk samples collected from 16 women at three time-points over four weeks. Results indicated that milk bacterial communities were generally complex; several genera represented greater than 5% of the relative community abundance, and the community was often, yet not always, stable over time within an individual. These results support the conclusion that human milk, which is recommended as the optimal nutrition source for almost all healthy infants, contains a collection of bacteria more diverse than previously reported. This finding begs the question as to what role this community plays in colonization of the infant gastrointestinal tract and maintaining mammary health.

  20. Microbiota/Host Crosstalk Biomarkers: Regulatory Response of Human Intestinal Dendritic Cells Exposed to Lactobacillus Extracellular Encrypted Peptide

    PubMed Central

    Al-Hassi, Hafid O.; Mann, Elizabeth R.; Urdaci, María C.; Knight, Stella C.; Margolles, Abelardo

    2012-01-01

    The human gastrointestinal tract is exposed to a huge variety of microorganisms, either commensal or pathogenic; at this site, a balance between immunity and immune tolerance is required. Intestinal dendritic cells (DCs) control the mechanisms of immune response/tolerance in the gut. In this paper we have identified a peptide (STp) secreted by Lactobacillus plantarum, characterized by the abundance of serine and threonine residues within its sequence. STp is encoded in one of the main extracellular proteins produced by such species, which includes some probiotic strains, and lacks cleavage sites for the major intestinal proteases. When studied in vitro, STp expanded the ongoing production of regulatory IL-10 in human intestinal DCs from healthy controls. STp-primed DC induced an immunoregulatory cytokine profile and skin-homing profile on stimulated T-cells. Our data suggest that some of the molecular dialogue between intestinal bacteria and DCs may be mediated by immunomodulatory peptides, encoded in larger extracellular proteins, secreted by commensal bacteria. These peptides may be used for the development of nutraceutical products for patients with IBD. In addition, this kind of peptides seem to be absent in the gut of inflammatory bowel disease patients, suggesting a potential role as biomarker of gut homeostasis. PMID:22606249

  1. 4-Nonylphenol reduces cell viability and induces apoptosis and ER-stress in a human epithelial intestinal cell line.

    PubMed

    Lepretti, M; Paolella, G; Giordano, D; Marabotti, A; Gay, F; Capaldo, A; Esposito, C; Caputo, I

    2015-10-01

    4-Nonylphenol is a widely diffused and stable environmental contaminant, originating from the degradation of alkyl phenol ethoxylates, common surfactants employed in several industrial applications. Due to its hydrophobic nature, 4-nonylphenol can easily accumulate in living organisms, including humans, where it displays a wide range of toxic effects. Since the gastrointestinal tract represents the main route by which 4-nonylphenol enters the body, the intestine may be one of the first organs to be damaged by chronic exposure to this pollutant through the diet. In the present study, we investigated the effects of 4-nonylphenol on a human intestinal epithelial cell line (Caco-2 cells). We demonstrated that 4-nonylphenol was cytotoxic to cells, as revealed by a decrease of the cell number and the decrement of mitochondrial functionality after 24 h of treatment. 4-Nonylphenol also reduced the number of cells entering into S-phase and interfered with epidermal growth factor signalling, with consequent negative effects on cell survival. In addition, 4-nonylphenol induced apoptosis, involving the activation of caspase-3, and triggered an endoplasmic reticulum-stress response, as revealed by over-expression of GRP78 (78 kDa glucose-regulated protein) and activation of XBP1 (X-box binding protein-1). Together, these findings support the hypothesis that prolonged exposure to 4-nonylphenol through the diet may lead to local damage at the level of intestinal mucosa, with potentially negative consequences for intestinal homeostasis and functionality.

  2. Microbiota/host crosstalk biomarkers: regulatory response of human intestinal dendritic cells exposed to Lactobacillus extracellular encrypted peptide.

    PubMed

    Bernardo, David; Sánchez, Borja; Al-Hassi, Hafid O; Mann, Elizabeth R; Urdaci, María C; Knight, Stella C; Margolles, Abelardo

    2012-01-01

    The human gastrointestinal tract is exposed to a huge variety of microorganisms, either commensal or pathogenic; at this site, a balance between immunity and immune tolerance is required. Intestinal dendritic cells (DCs) control the mechanisms of immune response/tolerance in the gut. In this paper we have identified a peptide (STp) secreted by Lactobacillus plantarum, characterized by the abundance of serine and threonine residues within its sequence. STp is encoded in one of the main extracellular proteins produced by such species, which includes some probiotic strains, and lacks cleavage sites for the major intestinal proteases. When studied in vitro, STp expanded the ongoing production of regulatory IL-10 in human intestinal DCs from healthy controls. STp-primed DC induced an immunoregulatory cytokine profile and skin-homing profile on stimulated T-cells. Our data suggest that some of the molecular dialogue between intestinal bacteria and DCs may be mediated by immunomodulatory peptides, encoded in larger extracellular proteins, secreted by commensal bacteria. These peptides may be used for the development of nutraceutical products for patients with IBD. In addition, this kind of peptides seem to be absent in the gut of inflammatory bowel disease patients, suggesting a potential role as biomarker of gut homeostasis.

  3. Glutamine pretreatment reduces IL-8 production in human intestinal epithelial cells by limiting IkappaBalpha ubiquitination.

    PubMed

    Hubert-Buron, Aurélie; Leblond, Jonathan; Jacquot, Arnaud; Ducrotté, Philippe; Déchelotte, Pierre; Coëffier, Moïse

    2006-06-01

    Glutamine, the most abundant amino acid in the human body, plays several important roles in the intestine. Recent studies showed that glutamine regulates protein metabolism and intestinal inflammation among other mechanisms by reducing proinflammatory cytokine release. Because regulation of the inflammatory response was shown to be linked to proteolysis regulation, we hypothesized that glutamine pretreatment could act on IL-8 production in human intestinal epithelial cells through the regulation of inhibitor kappaB (IkappaB) ubiquitination. The HCT-8 cells were pretreated for 24 h with 0.6, 2, or 10 mmol/L glutamine. IL-8 concentration and IkappaB (free and ubiquitinated) expressions were assessed by ELISA and immunoblotting, respectively. A pretreatment with 10 mmol/L glutamine decreased IL-8 production under both basal and proinflammatory conditions (both P < 0.05). In the presence of a proteasome inhibitor (MG132), the ubiquitin-IkappaBalpha complex expression was not significantly modified by glutamine under basal conditions but decreased significantly under proinflammatory conditions (P < 0.05). After the addition of 10 mmol/L of glutamine, the free IkappaBalpha expression increased under basal and stimulated conditions (both P < 0.05). A glutamine pretreatment of 10 mmol/L did not affect ubiquitin expression or proteasome activity. This study indicates that glutamine pretreatment may reduce the intestinal inflammatory response by limiting the proteolysis of IkappaBalpha.

  4. Assessment of the mode of action underlying development of rodent small intestinal tumors following oral exposure to hexavalent chromium and relevance to humans

    PubMed Central

    Proctor, Deborah M.; Suh, Mina; Haws, Laurie C.; Kirman, Christopher R.; Harris, Mark A.

    2013-01-01

    Chronic exposure to high concentrations of hexavalent chromium (Cr(VI)) in drinking water causes intestinal adenomas and carcinomas in mice, but not in rats. Cr(VI) causes damage to intestinal villi and crypt hyperplasia in mice after only one week of exposure. After two years of exposure, intestinal damage and crypt hyperplasia are evident in mice (but not rats), as are intestinal tumors. Although Cr(VI) has genotoxic properties, these findings suggest that intestinal tumors in mice arise as a result of chronic mucosal injury. To better understand the mode of action (MOA) of Cr(VI) in the intestine, a 90-day drinking water study was conducted to collect histological, biochemical, toxicogenomic and pharmacokinetic data in intestinal tissues. Using MOA analyses and human relevance frameworks proposed by national and international regulatory agencies, the weight of evidence supports a cytotoxic MOA with the following key events: (a) absorption of Cr(VI) from the intestinal lumen, (b) toxicity to intestinal villi, (c) crypt regenerative hyperplasia and (d) clonal expansion of mutations within the crypt stem cells, resulting in late onset tumorigenesis. This article summarizes the data supporting each key event in the MOA, as well as data that argue against a mutagenic MOA for Cr(VI)-induced intestinal tumors. PMID:23445218

  5. Streptococcus massiliensis in the human mouth: a phylogenetic approach for the inference of bacterial habitats.

    PubMed

    Póntigo, F; Silva, C; Moraga, M; Flores, S V

    2015-12-29

    Streptococcus is a diverse bacterial lineage. Species of this genus occupy a myriad of environments inside humans and other animals. Despite the elucidation of several of these habitats, many remain to be identified. Here, we explore a methodological approach to reveal unknown bacterial environments. Specifically, we inferred the phylogeny of the Mitis group by analyzing the sequences of eight genes. In addition, information regarding habitat use of species belonging to this group was obtained from the scientific literature. The oral cavity emerged as a potential, previously unknown, environment of Streptococcus massiliensis. This phylogeny-based prediction was confirmed by species-specific polymerase chain reaction (PCR) amplification. We propose employing a similar approach, i.e., use of bibliographic data and molecular phylogenetics as predictive methods, and species-specific PCR as confirmation, in order to reveal other unknown habitats in further bacterial taxa.

  6. Comparative analysis of the intestinal bacterial and RNA viral communities from sentinel birds placed on selected broiler chicken farms

    Technology Transfer Automated Retrieval System (TEKTRAN)

    There is a great deal of interest in characterizing the complex microbial communities in the poultry gut, and in understanding the effects of these dynamic communities on poultry performance, disease status, animal welfare, and microbes with human health significance. Investigations characterizing ...

  7. Comparative analysis of the intestinal bacterial and RNA viral communities from sentinel birds placed on selected broiler chicken farms

    Technology Transfer Automated Retrieval System (TEKTRAN)

    There is a great deal of interest in characterizing the complex microbial communities in the poultry gut, and in understanding the effects of these dynamic communities on poultry performance, disease status, animal welfare, and microbes with human health significance. Investigations characterizing t...

  8. Omp85 genosensor for detection of human brain bacterial meningitis.

    PubMed

    Dash, Sandip Kumar; Sharma, Minakshi; Khare, Shashi; Kumar, Ashok

    2013-06-01

    The 5'-thiolated DNA probe based on specific virulence gene, Omp85, was immobilized onto a screen-printed gold electrode followed by hybridization with 6-100 ng/6 μl (5.9 × 10(5)-9.3 × 10(6 )c.f.u.) of Neisseria meningitidis single stranded genomic DNA (ssG-DNA) for 10 min at 25 °C from the cerebrospinal fluid (CSF) of a meningitis patient. The Omp85 genosensor can detect as little as 6 ng ssG-DNA in 6 μl CSF of a human brain meningitis patient in 30 min including a response time of 1 min by cyclic voltammetry, differential pulse voltammetry (DPV) and electrochemical impedance. The sensitivity of the genosensor electrode was 2.6(μA/cm(2))/ng using DPV with regression coefficient (R(2)) 0.954. The genosensor was characterized using Fourier transform infrared spectroscopy and atomic force microscopy. Omp85 genosensor was stable for 12 months at 4 °C with 12 % loss in DPV current.

  9. Vasoactive intestinal polypeptide immunoreactivity in the human cerebellum: qualitative and quantitative analyses

    PubMed Central

    Benagiano, Vincenzo; Flace, Paolo; Lorusso, Loredana; Rizzi, Anna; Bosco, Lorenzo; Cagiano, Raffaele; Ambrosi, Glauco

    2009-01-01

    Although autoradiographic, reverse transcription-polymerase chain reaction and immunohistochemical studies have demonstrated receptors for vasoactive intestinal polypeptide (VIP) in the cerebellum of various species, immunohistochemistry has never shown immunoreactivity for VIP within cerebellar neuronal bodies and processes. The present study aimed to ascertain whether VIP immunoreactivity really does exist in the human cerebellum by making a systematic analysis of samples removed post-mortem from all of the cerebellar lobes. The study was carried out using light microscopy immunohistochemical techniques based on a set of four different antibodies (three polyclonal and one monoclonal) against VIP, carefully selected on the basis of control tests performed on human colon. All of the antibodies used showed VIP-immunoreactive neuronal bodies and processes distributed in the cerebellar cortex and subjacent white matter of all of the cerebellum lobes, having similar qualitative patterns of distribution. Immunoreactive neurons included subpopulations of the main neuron types of the cortex. Statistical analysis of the quantitative data on the VIP immunoreactivity revealed by the different antibodies in the different cerebellar lobes did not demonstrate any significant differences. In conclusion, using four different anti-VIP antibodies, the first evidence of VIP immunoreactivity is herein supplied in the human post-mortem cerebellum, with similar qualitative/quantitative patterns of distribution among the different cerebellum lobes. Owing to the function performed by VIP as a neurotransmitter/neuromodulator, it is a candidate for a role in intrinsic and extrinsic (projective) circuits of the cerebellum, in agreement with previous demonstrations of receptors for VIP in the cerebellar cortex and nuclei. As VIP signalling pathways are implicated in the regulation of cognitive and psychic functions, cerebral blood flow and metabolism, processes of histomorphogenesis

  10. Structural and Biochemical Characterization of a Novel Aminopeptidase from Human Intestine*

    PubMed Central

    Tykvart, Jan; Bařinka, Cyril; Svoboda, Michal; Navrátil, Václav; Souček, Radko; Hubálek, Martin; Hradilek, Martin; Šácha, Pavel; Lubkowski, Jacek; Konvalinka, Jan

    2015-01-01

    N-acetylated α-linked acidic dipeptidase-like protein (NAALADase L), encoded by the NAALADL1 gene, is a close homolog of glutamate carboxypeptidase II, a metallopeptidase that has been intensively studied as a target for imaging and therapy of solid malignancies and neuropathologies. However, neither the physiological functions nor structural features of NAALADase L are known at present. Here, we report a thorough characterization of the protein product of the human NAALADL1 gene, including heterologous overexpression and purification, structural and biochemical characterization, and analysis of its expression profile. By solving the NAALADase L x-ray structure, we provide the first experimental evidence that it is a zinc-dependent metallopeptidase with a catalytic mechanism similar to that of glutamate carboxypeptidase II yet distinct substrate specificity. A proteome-based assay revealed that the NAALADL1 gene product possesses previously unrecognized aminopeptidase activity but no carboxy- or endopeptidase activity. These findings were corroborated by site-directed mutagenesis and identification of bestatin as a potent inhibitor of the enzyme. Analysis of NAALADL1 gene expression at both the mRNA and protein levels revealed the small intestine as the major site of protein expression and points toward extensive alternative splicing of the NAALADL1 gene transcript. Taken together, our data imply that the NAALADL1 gene product's primary physiological function is associated with the final stages of protein/peptide digestion and absorption in the human digestive system. Based on these results, we suggest a new name for this enzyme: human ileal aminopeptidase (HILAP). PMID:25752612

  11. Influence of rifampicin on the expression and function of human intestinal cytochrome P450 enzymes

    PubMed Central

    Glaeser, H; Drescher, S; Eichelbaum, M; Fromm, M F

    2005-01-01

    List of nonstandard abbreviations mlpc multilumen perfusion catheter TMPD: transmucosal potential difference a.u. arbitrary unit Aims To investigate the potential induction by rifampicin of intestinal CYP2C8, CYP2C9, CYP2D6 and CYP3A4 using preparations of human enterocytes. Methods Using a multilumen perfusion catheter shed human enterocytes were collected from 6 healthy subjects before and after 10 days of 600 mg day−1 oral rifampicin administration. The protein expression of CYP2C8, CYP