Efficacy profiles for different concentrations of Lactobacillus acidophilus in experimental colitis

AIM:To determine the efficacy profiles of different concentrations of Lactobacillus acidophilus(L.acidophilus)for treating colitis using an experimental murine model.METHODS:Colitis was established in 64 BALB/c mice by adding 5%dextran sodium sulfate(DSS)to the drinking water and allowing ad libitum access for 7 d.The mice were then randomly divided into the following control and experimental model groups(n=8 each;day 0):untreated model control;negative-treatment model control(administered gavage of 1 mL/10 g normal saline);experimental-treatment models C4-C8(administered gavage of 104,105,106,107,or 108CFU/10 g L.acidophilus,respectively);positive-treatment model control(administration of the anti-inflammatory agent prednisone acetate at 45 g/10 g).Eight mice given regular water(no DSS)and no subsequent treatments served as the normal control group.Body weight,fecal traits,and presence of fecal occult blood were assessed daily.All animals were sacrificed on post-treatment day7 to measure colonic length,perform histological scoring,and quantify the major bacteria in the proximal and distal colon.Intergroup differences were determined by one-way ANOVA and post-hoc Student-Newman-Keuls comparison.RESULTS:All treatments(L.acidophilus and prednisone acetate)protected against colitis-induced weight loss(P<0.05 vs model and normal control groups).The extent of colitis-induced colonic shortening was significantly reduced by all treatments(prednisone acetate>C4>C5>C7>C8>C6;P<0.05 vs untreated model group),and the C6 group showed colonic length similar to that of the normal control group(P>0.05).The C6 group also had the lowest disease activity index scores among the model groups.The bacterial profiles in the proximal colon were similar between all of the experimental-treatment model groups(all P>0.05).In contrast,the bacterial profile in the distal colon of the C6 group showed the distinctive features(P<0.05 vs all other experimental-treatment model groups)of Lactobacillus sp.and Bifidobacterium sp.being the most abundant bacteria and Staphylococcus aureus being the least abundant bacteria.CONCLUSION:The most therapeutically efficacious concentration of L.acidophilus(106CFU/10 g)may exert its effects by modulating the bacterial profile in the distal colon.

Lactobacillus species shift in distal esophagus of high-fat-diet-fed rats

AIM:To analyze the microbiota shift in the distal esophagus of Sprague-Dawley rats fed a high-fat diet.METHODS:Twenty Sprague-Dawley rats were divided into high-fat diet and normal control groups of 10 rats each.The composition of microbiota in the mucosa from the distal esophagus was analyzed based on selective culture.A variety of Lactobacillus species were identified by molecular biological techniques.Bacterial DNA from Lactobacillus colonies was extracted,and 16S rDNA was amplified by PCR using bacterial universal primers.The amplified 16S rDNA products were separated by denaturing gradient gel electrophoresis(DGGE).Every single band was purified from the gel and sent to be sequenced.RESULTS:Based on mucosal bacterial culturing in the distal esophagus,Staphylococcus aureus was absent,and total anaerobes and Lactobacillus species were decreased significantly in the high-fat diet group compared with the normal control group(P < 0.01).Detailed DGGE analysis on the composition of Lactobacillus species in the distal esophagus revealed that Lactobacillus crispatus,Lactobacillus gasseri(L.gasseri) and Lactobacillus reuteri(L.reuteri) comprised the Lactobacillus species in the high-fat diet group,while the composition of Lactobacillus species in the normal control group consisted of L.gasseri,Lactobacillus jensenii and L.reuteri.CONCLUSION:High-fat diet led to a mucosal microflora shift in the distal esophagus in rats,especially the composition of Lactobacillus species.

Lactobacillus crispatus M206119 exacerbates murine DSS-colitis by interfering with inflammatory responses

AIM:To investigate the role of Lactobacillus crispatus(L.crispatus) strain China Center for Type Culture Collection(CCTCC) M206119 in intestinal inflammation.METHODS:Forty 8-wk-old Balb/c mice(20 ± 2 g) were divided into four groups of 10 mice each.Three groups that had received dextran sulfate sodium(DSS) were administered normal saline,sulfasalazine or CCTCC M206119 strain,and the fourth group received none of these.We assessed the severity of colitis using a disease activity index,measured the colon length and weight,collected stools and mesenteric lymph nodes for bacterial microflora analysis.One centimeter of the proximal colon,middle colon and distal colon were collected and fixed in 10% buffered formalin,dehydrated in ethanol,and embedded in paraffin.Interleukin(IL)1β,IL-6 and tumor necrosis factor(TNF)-α expression was detected using reverse transcription polymerase chain reaction.Protective factors zonula occludens(ZO)-1 and β-defensin 2 were detected by immunoblotting.The features of CCTCC M206119 strain were identified based on morphology,biochemical profile,and 16S RNA sequencing.RESULTS:DSS-colitis animals treated with CCTCC M206119 had markedly more severe disease,with greater weight loss,diarrhea,fecal bleeding,and shortened colon length.In addition,the CCTCC-M206119treated group had comparatively higher histological scores and more neutrophil infiltration than the controls.Expression of protective factors ZO-1 and β-defensin 2 was downregulated due to destruction of the mucosal barrier after CCTCC M206119 strain treatment.An in vitro assay demonstrated that CCTCC M206119 strain increased the nuclear translocation of nuclear factor-κB in epithelial cells.Intestinal proinflammatory or anti-inflammatory cytokine responses were evaluated.Proinflammatory colonic cytokine(IL-1β,IL-6 and TNF-α) levels were clearly increased in CCTCCM206119-treated animals,whereas anti-inflammatory colonic cytokine(IL-10) level was lowered compared with saline or 5-aminosalicylic-acid-treated DSS-colitis mice.Next,CCTCC M206119 strain was characterized as L.crispatus by microscopic morphology,biochemical tests and 16S rRNA gene level.

Effects of different diets on intestinal microbiota and nonalcoholic fatty liver disease development

AIM To study the effects of different diets on intestinal microbiota and nonalcoholic fatty liver disease(NAFLD) development at the same caloric intake.METHODS Thirty male Sprague-Dawley rats were randomized into five groups(six rats each). The control diet(CON) group and free high-fat diet(FFAT) group were allowed ad libitum access to a normal chow diet and a highfat diet, respectively. The restrictive high-fat diet(RFAT) group, restrictive high-sugar diet(RSUG) group, and high-protein diet(PRO) group were fed a highfat diet, a high-sugar diet, and a high-protein diet, respectively, in an isocaloric way. All rats were killed at 12 wk. Body weight, visceral fat index(visceral fat/body weight), liver index(liver/body weight), insulin resistance, portal lipopolysaccharide(LPS), serum alanine aminotransferase(ALT), serum aspartate aminotransferase(AST), and liver triglycerides were measured. The intestinal microbiota in the different groups of rats was sequenced using high-throughput sequencing technology.RESULTS The FFAT group had higher body weight, visceral fat index, liver index, peripheral insulin resistance, portal LPS, serum ALT, serum AST, and liver triglycerides compared with all other groups(P < 0.05). Taking the same calories, the RFAT and RSUG groups demonstrated increased body weight, visceral fat index, peripheral insulin resistance and liver triglycerides compared with the PRO group(P < 0.05). The RFAT group also showed increased portal LPS compared with the PRO group(P < 0.05). Unweighted Uni Frac principal coordinates analysis of the sequencing data revealed that the intestinal microbiota structures of the CON, FFAT, RSUG and PRO groups were roughly separated away from each other. Taxon-based analysis showed that, compared with the CON group, the FFAT group had an increased abundance of Firmicutes, Roseburia and Oscillospira bacteria, a higher ratio of Firmicutes to Bacteroidetes, and a decreased abundance of Bacteroidetes, Bacteroides and Parabacteroides bacteria(P < 0.05). The RFAT group showed an increased abundance of Firmicutes and decreased abundance of Parabacteroides bacteria(P < 0.05). The RSUG group showed an increased abundance of Bacteroidetes and Sutterella bacteria, higher ratio of Bacteroidetes to Firmicutes, and a decreased abundance of Firmicutes(P < 0.05). The PRO group showed an increased abundance of Bacteroidetes, Prevotella, Oscillospira and Sutterella bacteria, and a decreased abundance of Firmicutes(P < 0.05). Compared with the FFAT group, the RFAT group had an increased abundance of Bacteroidetes, higher ratio of Bacteroidetes to Firmicutes, and decreased abundance of Firmicutes and Oscillospira bacteria(P < 0.05).CONCLUSION Compared with the high-protein diet, the NAFLDinducing effects of high-fat and high-sugar diets are independent from calories, and may be associated with changed intestinal microbiota.

Ulcerative colitis as a polymicrobial infection characterized by sustained broken mucus barrier

To reduce medication for patients with ulcerative colitis(UC),we need to establish the etiology of UC.The intestinal microbiota of patients with inflammatory bowel disease(IBD)has been shown to differ from that of healthy controls and abundant data indicate that it changes in both composition and localization.Small intestinal bacterial overgrowth is significantly higher in IBD patients compared with controls.Probiotics have been investigated for their capacity to reduce the severity of UC.The luminal surfaces of the gastrointestinal tract are covered by a mucus layer.This normally acts as a barrier that does not allow bacteria to reach the epithelial cells and thus limits the direct contact between the host and the bacteria.The mucus layer in the colon comprises an inner layer that is firmly adherent to the intestinal mucosa,and an outer layer that can be washed off with minimal rinsing.Some bacteria can dissolve the protective inner mucus layer.Defects in renewal and formation of the inner mucus layer allow bacteria to reach the epithelium and have implications for the causes of colitis.In this review,important elements of UC pathology are thought to be the intestinal bacteria,gut mucus,and the mucosa-associated immune system.

Therapeutic effects of four strains of probiotics on experimental colitis in mice

AIM:To investigate the therapeutic effects of four strains of probiotics(E.feacalis,L.acidophilus,C.butyricum and B.adolescentis) on dextran sulphate sodium(DSS)-induced experimental colitis in Balb/c mice.METHODS:Eighty Balb/c mice were randomly divided into 8 groups.Weight-loss,fecal character,fecal occult blood and hematochezia were recorded daily.Disease activity index(DAI) scores were also evaluated everyday.Length of colon was measured and histological scores were evaluated on the 13th day.Myeloperoxidase(MPO) activity was detected.Interleukin-1(IL-1) and IL-4 expression was detected by ELISA and RT-PCR.RESULTS:The four strains of probiotics relieved the inflammatory condition of DSS-induced experimental colitis in mice.Weight loss was slowed down in all probiotics-treated mice.Even weight gain was observed by the end of probiotics treatment.The DAI and histological scores of probiotics-treated mice were lower than those of mice in the control group(1.9 ± 0.2 vs 8.6 ± 0.4,P < 0.05 for E.faecalis).The length of colon of probiotics-treated mice was longer than that of mice in the control group(10.3 ± 0.34 vs 8.65 ± 0.77,P < 0.05 for E.faecalis).The four strains of probiotics decreased the MP activity and the IL-1 expression,but increased the IL-4 expression.E.faecalis had a better effect on DSS-induced experimental colitis in mice than the other three strains.CONCLUSION:The four strains of probiotics have beneficial effects on experimental colitis in mice.E.faecalis has a better effect on DSS-induced experimental colitis in mice than the other three strains.Supplement of probiotics provides a new therapy for UC.