OBJECTIVE: To investigate the changes in bacterial flora in fecal samples, at the tumor loci and in adjacent mucosa in patients with colorectal cancer (CRC). METHODS: We collected fecal samples from 13 patients with CRC and 20 healthy individuals and tumor and adjacent mucosa samples from 6 CRC patients. The differences in bacterial composition between the fecal and mucosa samples were analyzed with 16S rDNA sequencing and bioinformatics methods. We also detected the total number of bacteria in the feces using flow cytometry, isolated and identified the microorganisms in the fecal and mucosa samples using common bacterial culture media. We further tested the effects of 7 isolated bacterial strains on apoptosis of 3 CRC cell lines using lactate dehydrogenase detection kit. RESULTS: The bacterial α-diversity in the feces of healthy individuals and in adjacent mucosa of CRC patients was significantly higher than that in the feces and tumor mucosa in CRC patients (P < 0.05). Lactobacillaceae is a specific bacteria in the feces, while Escherichia, Enterococcus, and Fusobacterium are specific bacteria in tumor mucosa of CRC patients as compared with healthy individuals. Cell experiment with3 CRC cell lines showed that Bacteroides fragilis isolated from the tumor mucosa of CRC patients produced significant inhibitory effects on cell proliferation (P < 0.0001), while the isolated strain Fusobacterium nucleatum obviously promoted the proliferation of the cell lines (P < 0.001). CONCLUSION The bacterial flora in the feces, tumor mucosa and adjacent mucosa of CRC patients is significantly different from that in the feces of healthy individuals, and the fecal flora of CRC patients can not represent the specific flora of the tumor mucosa. Inhibition of F. nucleatum colonization in the tumor mucosa and promoting B. fragilis colonization may prove beneficial for CRC treatment.
Bacteria ; Colorectal Neoplasms/pathology* ; Feces/microbiology* ; Gastrointestinal Microbiome ; Humans ; Intestinal Mucosa

Houttuynia cordata polysaccharide (HCP) is extracted from Houttuynia cordata, a key traditional Chinese medicine. The study was to investigate the effects of HCP on intestinal barrier and microbiota in H1N1 virus infected mice. Mice were infected with H1N1 virus and orally administrated HCP at a dosage of 40 mg(kg(d. H1N1 infection caused pulmonary and intestinal injury and gut microbiota imbalance. HCP significantly suppressed the expression of hypoxia inducible factor-1α and decreased mucosubstances in goblet cells, but restored the level of zonula occludens-1 in intestine. HCP also reversed the composition change of intestinal microbiota caused by H1N1 infection, with significantly reduced relative abundances of Vibrio and Bacillus, the pathogenic bacterial genera. Furthermore, HCP rebalanced the gut microbiota and restored the intestinal homeostasis to some degree. The inhibition of inflammation was associated with the reduced level of Toll-like receptors and interleukin-1β in intestine, as well as the increased production of interleukin-10. Oral administration of HCP alleviated lung injury and intestinal dysfunction caused by H1N1 infection. HCP may gain systemic treatment by local acting on intestine and microbiota. This study proved the high-value application of HCP.
Animals ; Cytokines ; metabolism ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; therapeutic use ; Gastrointestinal Microbiome ; drug effects ; Houttuynia ; chemistry ; Hypoxia-Inducible Factor 1, alpha Subunit ; metabolism ; Inflammation ; drug therapy ; pathology ; Influenza A Virus, H1N1 Subtype ; pathogenicity ; Intestinal Mucosa ; drug effects ; metabolism ; microbiology ; pathology ; Lung ; drug effects ; metabolism ; pathology ; Male ; Mice, Inbred BALB C ; Orthomyxoviridae Infections ; drug therapy ; pathology ; physiopathology ; Plant Extracts ; chemistry ; Polysaccharides ; chemistry ; pharmacology ; therapeutic use ; Toll-Like Receptors ; metabolism ; Zonula Occludens-1 Protein ; metabolism

OBJECTIVETo investigate the change in the expression of tight junction protein ZO-1 in intestinal epithelial cells (Caco-2 cells) and the protective effect of eicosapentaenoic acid (EPA) after adherent-invasive Escherichia coli (E.coli) LF82 infection.

METHODSThe Caco-2 cell line was used to establish an in vitro model of tight junction of intestinal epithelial cells. Caco-2 cells were divided into EPA treatment groups (0, 25, 50, 100, and 200 μmol/L EPA) and EPA (0, 25, 50, 100, and 200 μmol/L EPA)+E.coli LF82 treatment (0, 6, and 12 hours) groups. A microscope was used to observe the morphological characteristics of the cells. MTT assay was used to determine the cell growth curve. The activity of alkaline phosphatase (ALP) at both sides of the cell membrane was compared to evaluate the Caco-2 cell model. MTT assay and flow cytometry were used to investigate the effects of different concentrations of EPA on the survival rate and apoptosis rate of Caco-2 cells. RT-qPCR was used to measure the mRNA expression of ZO-1 in Caco-2 cells after EPA and/or E.coli LF82 treatment. ELISA was used to measure the change in the level of tumor necrosis factor-α (TNF-α) in culture supernatant.

RESULTSAfter EPA treatment (25 and 50 μmol/L), the proliferation of Caco-2 cells was induced in a dose-dependent manner. The survival rates of the cells were significantly higher than those in the control group (P<0.05). The EPA treatment (100 and 200 μmol/L) groups had a significant inhibitory effect on the proliferation of Caco-2 cells in a dose-dependent manner. The survival rates of the cells were significantly lower than those in the control group (P<0.05). The EPA treatment (100 and 200 μmol/L) groups had a significant increase in cell apoptosis rate compared with the control group (P<0.05). The 6- and 12-hour E.coli LF82 treatment groups had decreasing mRNA expression of ZO-1 in Caco-2 cells over the time of treatment and had significantly lower mRNA expression of ZO-1 than the untreated group (P<0.05). The Caco-2 cells treated with E.coli LF82 and 25 or 50 μmol/L EPA for 6 or 12 hours showed an increase in the mRNA expression of ZO-1 with the increasing concentration of EPA, as well as significantly higher mRNA expression of ZO-1 than the Caco-2 cells treated with E.coli LF82 alone (P<0.05). The Caco-2 cells treated with E.coli LF82 alone for 6 or 12 hours had increasing secretion of TNF-α over the time of treatment and had significantly higher secretion than the untreated Caco-2 cells (P<0.05). The Caco-2 cells treated with E.coli LF82 and 25 or 50 μmol/L EPA for 6 or 12 hours showed a reduction in the secretion of TNF-α with the increasing concentration of EPA and had significantly lower secretion than the Caco-2 cells treated with E.coli LF82 alone (P<0.05).

CONCLUSIONSEPA can effectively prevent the destruction of tight junction of intestinal epithelial cells induced by E.coli LF82 infection and inhibit the secretion of inflammatory factors. Therefore, it has a certain protective effect on intestinal mucosal barrier.

Apoptosis ; drug effects ; Caco-2 Cells ; Eicosapentaenoic Acid ; pharmacology ; Escherichia coli ; pathogenicity ; Humans ; Intestinal Mucosa ; metabolism ; microbiology ; RNA, Messenger ; analysis ; Tight Junctions ; drug effects ; Tumor Necrosis Factor-alpha ; secretion ; Zonula Occludens-1 Protein ; genetics

This paper is to explore changes of intestinal mucosal barrier, intestinal flora, and bacterial translocation in rats with severe acute pancreatitis (SAP). Twenty four male SD rats were randomly divided into the control group (n = 10) and the experimental group (n = 14). The model of severe acute pancreatitis of rats was induced by the method of injecting adversely 5% sodium taurocholate into the common biliary-pancreatic duct. All of the rats were killed after 24 hours and the level of the serum amylase and the plasma endotoxin was determined after that. The pathological changes of pancreas and small intestine were observed through hematoxylin-eosin staining (HE staining) and the abdominal viscera bacterial translocation rates were tested. With the method of real-time polymerase chain reaction (RT-PCR) the quantity of the intestinal flora was analyzed. In the control group, the level of Escherichia coli, Lactobacillus and Bifidobacterium were 2.08 ± 1.29, 11.04 ± 7.55 and 12.21 ± 4.95, respectively. On the contrast, the level of Escherichia coli in the cecum contents was much higher (9.72 ± 3.58, P < 0.01), while the Lactobacillus number was decreased significantly (0.67 ± 0.34, P < 0.01), and the Bifidobacterium number was also decreased (4.59 ± 3.42, P < 0.05) in the experimental group, so the ratio of Bifidobacterium/Escherichia coli was reversed. Besides, in the experimental group, the plasma endotoxin positive rates and the bacterial translocation rates were much higher (P < 0.01 or P < 0.05) and the pathology scores of pancreas and small intestines were also significantly higher (P < 0.01) than those in the control group. These results indicated that in severe acute pancreatitis rats, the intestinal mucosal barrier was severely damaged and the dysbacteriosis occurs in the intestinal canal. And these might relate to the occurrence and development of multiple organ infection.
Animals ; Bacterial Translocation ; Endotoxins ; Intestinal Mucosa ; pathology ; Intestines ; microbiology ; Male ; Pancreas ; pathology ; Pancreatitis ; microbiology ; pathology ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo observe the ultrastructural change of the route of gut bacterial translocation in a rat with spinal cord injury (SCI).

METHODSForty Wistar rats were divided into the following groups: control group and 3 SCI groups (10 in each group). The rats in the SCI groups were established SCI model at 24 h, 48 h, and 72 h after SCI. Small intestine mucous membrane tissue was identified and assayed by transmission electron microscope, scanning electron microscope and immunofluorescence microscopy.

RESULTSSmall intestine mucous membrane tissue in control group was not damaged significantly, but those in SCI groups were damaged significantly. Proliferation bacteria in gut lumen attached on microvilli. The extracellular bacteria torn the intestinal barrier and perforated into the small intestinal mucosal epithelial cell. The bacteria and a lot of particles of the seriously damaged region penetrated into the lymphatic system and the blood system directly. Some bacteria were internalized into the goblet cell through the apical granule. Some bacteria and particles perforated into the submucosa of the M cell running the long axis of M cells through the tight junctions. In the microcirculation of mucosa, the bacteria that had already broken through the microvilli into blood circulation swim accompanying with erythrocytes.

CONCLUSIONThe routes of bacterial translocation interact and format a vicious circle. At early step, the transcellular pathway of bacterial translocation is major. Following with the destroyed small intestine mucous, the routes of bacterial translocation through the lymphatic system and the blood system become direct pathways. The goblet cell-dendritic cell and M cell pathway also play an important role in the bacterial translocation.

Animals ; Bacteria ; Bacterial Translocation ; Epithelial Cells ; microbiology ; Goblet Cells ; microbiology ; Intestinal Mucosa ; microbiology ; pathology ; ultrastructure ; Intestine, Small ; microbiology ; pathology ; ultrastructure ; Microvilli ; microbiology ; Rats ; Rats, Wistar ; Spinal Cord Injuries ; microbiology

Radiation therapy is an important treatment modality for abdominal or pelvic cancer, but there is a common and serious complication such as radiation-induced enteritis. Probiotics is reported to have positive effects against radiation-induced enteropathy. In this study, morphological changes of bowel mucosa were analyzed in rats to presume the effect of probiotics on radiation-induced enteritis and its correlation with radiation dose. A total of 48 adult male Sprague-Dawley rats were randomly assigned to two groups and received a solution containing 1.0x108 colony-forming units of Lactiobacillus acidophilus or water once daily for 10 days. Each of two groups was divided into three subgroups and abdomino-pelvic area of each subgroup was irradiated with 10, 15, and 20 Gy, respectively on the seventh day of feeding the solutions. All rats were sacrificed 3 days after irradiation and the mucosal thickness and villus height of jejunum, ileum and colon were measured. The morphological parameters of the small intestine represented significant differences between two solution groups irradiated 10 or 15 Gy, except for villus height of jejunum in 15 Gy-subgroup (P=0.065). There was no significant morphometric difference between two groups irradiated with 20 Gy of radiation. Probiotics appear to be effective for the morphological shortening of small intestinal mucosa damaged by radiation less than or equal to 15 Gy.
Animals ; Colon/pathology ; Disease Models, Animal ; Enteritis/pathology/prevention & control ; Ileum/pathology ; Intestinal Mucosa/microbiology/*radiation effects ; Intestine, Small ; Jejunum/pathology ; Lactobacillus acidophilus/*metabolism ; Male ; Probiotics/administration & dosage/*pharmacology ; Radiation Injuries, Experimental/*prevention & control/therapy ; Radiation Protection/*methods ; Random Allocation ; Rats ; Rats, Sprague-Dawley

Toll-like receptors (TLRs) family may play important roles in inflammatory bowel disease. This study examined the expression of TLR2, TLR4 and TLR9 in the colonic tissues of patients with ulcerative colitis (UC) and explored their roles in the pathogenesis of UC. Colonic biopsies were taken from the colon of 30 patients with mild or moderate UC (at active phase) and 10 healthy controls during colonoscopy. TLR2, TLR4 and TLR9 protein expression levels were immunohistochemically detected. The mRNA expression levels of TLR2, TLR4 and TLR9 were assessed by reverse transcription polymerase chain reaction (RT-PCR). The disease activity index (DAI), colonoscopic and histologic grades and fecal microbial flora were determined. Histological examination showed that the intestinal mucous membrane of UC patients underwent acute inflammation changes. Immunohistochemistry exhibited that the expression levels of TLR2, TLR4 and TLR9 in colon epithelia and inflammatory cells were higher in UC patients than in control group (P<0.01). The mRNA expression levels of TLR2, TLR4 and TLR9 were increased in UC patients but were not detected in the normal controls. Expression levels of TLR2, TLR4 and TLR9 were positively correlated, and bore close correlation with DAI, colonoscopic and histologic grades and fecal microbial flora. An important mechanism of UC might be that abnormal activation of mucosal immunity by intestinal dysbacteriosis caused dysregulation of TLRS that mediates innate immunity.
Colitis, Ulcerative ; genetics ; metabolism ; pathology ; Colon ; metabolism ; microbiology ; Colonoscopy ; Feces ; microbiology ; Female ; Gene Expression ; Humans ; Immunohistochemistry ; Intestinal Mucosa ; metabolism ; microbiology ; Male ; Reverse Transcriptase Polymerase Chain Reaction ; Severity of Illness Index ; Toll-Like Receptor 2 ; biosynthesis ; genetics ; Toll-Like Receptor 4 ; biosynthesis ; genetics ; Toll-Like Receptor 9 ; biosynthesis ; genetics

No abstract available.
Bacteria/*isolation & purification ; Female ; Gastric Mucosa/*microbiology ; Gastritis/*microbiology ; Humans ; Intestinal Diseases/*microbiology ; Male ; *Microbiota

Toll-like receptors (TLRs) are key components of the innate immune system which trigger antimicrobial host defense responses. This study aimed to investigate the impact of probiotics (Lactobacillus, Bifidobacterium) on the expression of TLR4 and tumor necrosis factor-alpha (TNF-α) in the colon mucosa of rat experimental ulcerative colitis model induced by trinitrobenzene sulfonic acid (TNBS)/ethanol and immune complexes. The gross and histological changes of the colonic mucosa were observed and assessed by the means-standard deviation and independent samples t-test. The protein expression levels of TLR4 and TNF-α were detected by using immunohistochemistry and Western blotting, respectively. It was revealed that there was visible infiltration of inflammatory cells, formation of crypt abscess, and the reduction of goblet cells in the colon tissue of experimental models. As compared with the control group, the levels of TLR4 and TNF-α protein were significantly increased in the model group (P<0.01 for both). No significant difference was found in the expression of TLR4 and TNF-α between the two-week probiotics treatment group and the model group (P>0.05), whereas significant reductions were shown in rats which were treated with probiotics for four weeks as compared with the model group (P<0.01). There was no significant difference between two probiotics-treated groups. Our results implied that probiotics were likely to play a key role in protecting ulcerative colitis by reducing the inflammatory factor TNF-α expression through inhibiting the TLR4 expression in the colon tissue of experimental models.
Animals ; Bifidobacterium ; physiology ; Blotting, Western ; Colitis, Ulcerative ; chemically induced ; metabolism ; Colon ; drug effects ; metabolism ; microbiology ; Immunohistochemistry ; Intestinal Mucosa ; drug effects ; metabolism ; microbiology ; Lactobacillus ; physiology ; Male ; Probiotics ; pharmacology ; Rabbits ; Rats ; Rats, Sprague-Dawley ; Time Factors ; Toll-Like Receptor 4 ; biosynthesis ; Trinitrobenzenesulfonic Acid ; Tumor Necrosis Factor-alpha ; metabolism

The human intestinal microbiota is a community of 10(13)-10(14) microorganisms that harbor in the intestine and normally participate in a symbiotic relationship with human. Technical and conceptual advances have enabled rapid progress in characterizing the taxonomic composition, metabolic capacity and immunomodulatory activity of the human intestinal microbiota. Their collective genome, defined as microbiome, is estimated to contain > or =150 times as many genes as 2.85 billion base pair human genome. The intestinal microbiota and its microbiome form a diverse and complex ecological community that profoundly impact intestinal homeostasis and disease states. It is becoming increasingly evident that the large and complex bacterial population of the large intestine plays an important role in colorectal carcinogenesis. Numerous studies show that gut immunity and inflammation have impact on the development of colorectal cancer. Additionally, bacteria have been linked to colorectal cancer by the production of toxic and genotoxic bacterial metabolite. In this review, we discuss the multifactorial role of intestinal microbiota in colorectal cancer and role for probiotics in the prevention of colorectal cancer.
Animals ; Bacteroides/metabolism ; Colorectal Neoplasms/immunology/*microbiology ; Fatty Acids, Nonesterified/metabolism ; Humans ; Hydrogen Sulfide/metabolism ; Intestinal Mucosa/immunology/microbiology ; Metagenome ; *Probiotics ; Reactive Oxygen Species/metabolism ; Toxins, Biological/metabolism