Colonic Mucosal Blood Flow Research Articles

Participation of the intestinal microbiota in the mechanism of beneficial effect of treatment with synbiotic Syngut on experimental colitis under stress conditions.

Gut-brain axis plays a central role in the regulation of stress related diseases such as irritable bowel syndrome (IBS) or inflammatory bowel disease (IBD). It is increasingly recognized that stress modulates gut microbiota community structure and activity and represents an important causal factor in dysbiosis. This study was designed to determine the effect of daily treatment with synbiotic (Syngut) containing inulin, Lactobacillus acidophilus, Bifidobacterium lactis W51, Lactobacillus plantarum W21 and Lactococcus lactis applied i.g. at a dose of 50 mg/kg i.g. on the colonic damage and colonic mucosal blood flow in rats with experimentally induced TNBS-colitis that were additionally exposed or not to acute stress (episodes of cold restraint stress every other day before colitis induction). Control rats received daily treatment with vehicle (saline, i.g.) or mesalazine (50 mg/kg-d i.g.), the standard drug recommended in therapy of IBD. At the termination of TNBS colitis, the histologic evaluation of colonic mucosa, mucosal malonyldialdehyde (MDA) level and plasma concentrations of proinflammatory cytokines (TNF-α, IL-1β) and adipokine adiponectin were assessed. the samples of colonic mucosa not involving colonic lesions and surrounding the flared mucosa were excised for the determination of mRNA expression for proinflammatory biomarkers TNF-α, IL-1β, IL-10 and COX-2 as well as antioxidazing factors SOD-1 and SOD-2. Finally, the gut microbial profiles were analyzed by 16S rRNA sequencing at phylum, family and genus level. Episodes of cold stress significantly aggravated the course of TNBS colitis, and significantly increased the release of proinflammatory cytokines as well as the significant increase in the MDA concentration has been observed as compared with non-stressed TNBS rats. These changes were followed by the significant fall in the CBF and plasma adiponectin levels and by the overexpression of mRNA of proinflammatory biomarkers. Synbiotic treatment with Syngut significantly reduced the area of colonic lesions observed macroscopically and microscopically in rats with TNBS colitis with or without exposure to cold stress, significantly increased the CBF, normalized plasma adiponectin levels and significantly attenuated the release and colonic expression of proinflammatory cytokines and biomarkers. the analysis of the gut microbiota showed a significant reduction of microbial diversity (Shannon index) in rats with TNBS colitis with or without exposure to stress. The therapy with Syngut failed to significantly affect the alpha diversity. At the phylum level, the significant rise in Proteobacteria has been observed in stressed rats with TNBS colitis and this effects was attenuated by treatment with Syngut. At family level, TNBS colitis alone or in combination with stress led to a significant decrease of SCFA producing bacterial taxa such as Ruminococaceae and Lachnospiraceae and Syngut counteracted this effect. We conclude that: 1) cold stress exacerbates the gastrointestinal inflammation in experimental colitis; 2) the synbiotic therapy with Syngut ameliorates the gut inflammation in rats with TNBS colitis combined with cold stress; 3) the beneficial effect of Syngut is accompanied by increase of anti-inflammatory taxa such as Ruminococaceae and Lachnospiraceae, and 4) the modulation of gut microbiota with Syngut alleviates stress-related intestinal inflammation suggesting a potential usefulness of synbiotic therapy in intestinal disorders accompanied by stress in patients with IBD.

ENOS involved in colitis-induced mucosal blood flow increase

The role of NO in inflammatory bowel disease is controversial. Studies indicate that endothelial nitric oxide synthase (eNOS) might be involved in protecting the mucosa against colonic inflammation. The aim of this study was to investigate the involvement of nitric oxide (NO) in regulating colonic mucosal blood flow in two different colitis models in rats. In anesthetized control and colitic rats, the distal colon was exteriorized and the mucosa visualized. Blood flow (laser-Doppler flowmetry) and arterial blood pressure were continuously monitored throughout the experiments, and vascular resistance was calculated. Trinitrobenzene sulfonic acid (TNBS) or dextran sulfate sodium (DSS) was used to induce colitis. All groups were given the NOS inhibitor N(omega)-nitro-l-arginine (l-NNA) or the inducible NOS (iNOS) inhibitor l-N(6)-(1-iminoethyl)-lysine (l-NIL). iNOS, eNOS, and neuronal NOS (nNOS) mRNA in colonic samples were investigated with real-time RT-PCR. Before NOS inhibition, colonic mucosal blood flow, expressed as perfusion units, was higher in both colitis models compared with the controls. The blood flow was reduced in the TNBS- and DSS-treated rats during l-NNA administration but was not altered in the control group. Vascular resistance increased more in the TNBS- and DSS-treated rats than in the control rats, indicating a higher level of vasodilating NO in the colitis models. l-NIL did not alter blood pressure or blood flow in any of the groups. iNOS and eNOS mRNA increased in both colitis models, whereas nNOS remained at the control level. TNBS- and DSS-induced colitis results in increased colonic mucosal blood flow, most probably due to increased eNOS activity.

Role Of Nitric Oxide Synthase Inhibitor In Experimental Colitis Induced By 2,4,6‐Trinitrobenzene Sulphonic Acid In Rats

1. The present study was designed to investigate the role of nitric oxide (NO) in modulating 2,4,6-trinitrobenzene sulphonic acid (TNBS)-induced colitis in rats. 2. Damage scores and NO synthase (NOS) activity were measured. 3. The damage scores and NOS activity reached a peak on the 4th day after administration of TNBS solution (day 0), thereafter gradually decreasing, and were significantly higher than in the group treated with saline throughout the experimental period. 4. Subsequently, we divided the stage of colitis into two groups, one from day 0 to day 3 after induction of colitis, and the other from day 4 onwards. We evaluated the effects of the NOS inhibitor N(G)-monomethyl-L-arginine (L-NMMA), on TNBS-hapten-induced colitis and colonic mucosal blood flow. Two different methods of L-NMMA administration, from day 0 to day 3, and from day 4 onwards, were undertaken. 5. The damage score in the early L-NMMA treatment group was significantly higher than in the group without L-NMMA on day 14. In contrast, the damage score in the late L-NMMA treatment group was not significantly different from the group without L-NMMA. Colonic mucosal blood flow in the early L-NMMA treatment group was not significantly different from that in the late L-NMMA treatment group. 6. These data suggest that NO is important for inhibiting inflammation during the early stages.

Effects of n–3 Polyunsaturated Fatty Acids and Vitamin E on Colonic Mucosal Leukotriene Generation, Lipid Peroxidation, and Microcirculation in Rats with Experimental Colitis

Aims: We investigated the effect of n–3 polyunsaturated fatty acids (PUFAs) on mucosal levels of leukotrienes (LTs) and lipid peroxide (LPO), and on mucosal microcirculation, in rats with experimental colitis induced by dextran sulfate sodium (DSS). Methods: We fed Wistar rats a perilla oil-enriched diet containing α-linolenic acid (63.2% of total fatty acids) with various doses of vitamin E for 4 weeks, with 4% DSS added to the drinking water during the last week. Control rats were fed a diet produced from soybean oil containing α-linolenic acid (5.1% of total fatty acids). Colonic mucosal blood flow was measured with a laser Doppler flowmeter. Results: The mucosal level of arachidonic acid was significantly lower and that of eicosapentaenoic acid was significantly higher in the experimental group. The mucosal level of LPO in the experimental group fed a trace or ordinary dose of vitamin E was significantly higher than that of the controls. The production of LTB₄ and LTC₄ from the colonic mucosa in the experimental group was significantly lower than that in controls. However, only the experimental group fed a vitamin E dose 4-fold higher than that given to the controls showed a significant increase in mucosal blood flow. Conclusion: These results suggest that n–3 PUFAs increase mucosal blood flow by inhibiting LT production when there is sufficient vitamin E to inhibit lipid peroxidation in rats with experimental colitis.

Epidermal growth factor increases basal mucosal blood flow in the rat colon, a prostaglandin dependent effect.

Non-mitogenic biological activity such as modulation of mucosal blood flow is suspected to convey the protective effects of epidermal growth factor (EGF) in vivo. The aims of our present study were to determine the effects of EGF on colonic mucosal blood flow and injury induced hyperaemia in rats. Rats were pretreated with i.p. injections of vehicle, EGF, or indomethacin and EGF prior to mucosal injury. Basal mucosal blood flow and injury induced hyperaemia at the border of the damaged mucosa was determined by using reflectance spectrophotometry. EGF significantly increased basal mucosal blood flow but did not further enhance injury induced hyperaemia. The EGF induced increase in basal mucosal blood flow was completely abolished by indomethacin pretreatment. EGF induces an increase of basal mucosal blood flow through induction of prostaglandin synthesis. We hypothesize that the increase in mucosal blood flow contributes to the ability of EGF to protect the colonic mucosa against injury.

Mucosal blood flow and generation of superoxide in rat experimental colitis induced by succinic acid.

As we consider succinic acid to be an exacerbating factor in ulcerative colitis, we investigated its influence on rat colonic mucosa in terms of mucosal blood flow and superoxide generation. We measured mucosal blood flow by the hydrogen gas clearance method and superoxide generation by the chemiluminescence method, and observed histopathological findings to determine the effects of succinic acid. After the instillation of succinic acid of any concentration tested to the colon, mucosal blood flow decreased. Histopathologically, the higher the concentration of succinic acid, the greater was the erosion formation in the colonic mucosa, while significant polymorpho-nuclear cell infiltration superoxide generation from colon tissue were observed with 0.01% succinic acid compared with higher or lower concentrations. Succinic acid, at fecal concentrations found in active stage ulcerative colitis, appears to be implicated in mucosal injury, mediated by a decrease in colonic mucosal blood flow and infiltration of superoxide-generating polymorpho-nuclear cells into the mucosa.

Portal hypertension triggers local activation of inducible nitric oxide synthase gene in colonic mucosa

Recently a new clinical entity “portal hypertensive colopathy” has been reported. It involves vascular abnormalities and bleeding. Because nitric oxide may mediate these changes, we studied whether portal hypertension affects nitric oxide synthase in portal hypertensive colonic mucosa. In portal hypertensive and sham-operated rats the following studies were done: (1) colonic mucosal blood flow, (2) quantitative histologic examination, (3) reverse transcription—polymerase chain reaction for nitric oxide synthase mRNA, (4) nitric oxide synthase activity assay, and (5) immunostaining for nitric oxide synthase. In portal hypertensive rats, colonic mucosal blood flow and the number of submucosal veins were significantly increased in comparison to sham-operated rats. The mRNA expression and enzyme activity for inducible nitric oxide synthase (but not constitutive nitric oxide synthase) were significantly increased in portal hypertensive rats. Fluorescence signal intensity for inducible nitric oxide synthase in endothelia of mucosal and submucosal veins was significantly higher in portal hypertensive rats than in sham-operated rats. Portal hypertension activates inducible nitric oxide synthase gene and protein in colonic mucosal vessels. The excess of nitric oxide generated by overexpressed inducible nitric oxide synthase may play an important role in the development of vascular and hemodynamic abnormalities characterizing portal hypertensive colopathy.

Hemodynamics in the colonic mucosa of rats with dextran sulfate-induced colitis in the early phase.

We investigated hemodynamics in the colonic mucosa of rats with experimental colitis induced by the administration of dextran sulfate sodium (DSS). As parameters of hemodynamics, we determined the indices of mucosal hemoglobin concentration (IHb) and mucosal oxygen saturation (ISO2), measured by reflectance spectrophotometry, and an index of colonic mucosal blood flow (Flow), measured by laser-Doppler flowmetry. In the ascending colon, each parameter was measured by a combination of these methods after 1, 3, 5, 7, and 10 days of DSS administration. Histopathological examination was also performed. IHb in the DSS group increased with time; on the 7th day, the value was 126.9 +/- 8.32, while that in the control group was 85.0 +/- 4.14, IHb in the DSS group being significantly increased (P < 0.02). ISO2 in the DSS group was lower than that in the control group, and on the 7th day, was significantly lower in the DSS group (25.7 +/- 1.34) than in the control group (33.4 +/- 1.77) (P < 0.01). No changes in Flow were observed in either the DSS or the control group during the administration period, and no significant difference in Flow was found between the two groups. On histopathological examination, we observed a time-dependent increase in the infiltration of inflammatory cells in the ascending colon of rats treated with DSS, but changes such as erosion and ulceration were not found in the superficial layer of the mucosa. No histopathological changes were found in the control animals. In the early phase of the experimental colitis, hemodynamic alterations in the colonic mucosa were already present at the time the slight histopathological changes developed. These observations seemed to indicate the involvement of hemodynamic alterations in the subsequent tissue injury.

Colonic Mucosal Blood Flow in Ischemic Colitis

Abstract:Altered colonic mucosal blood flow (CMBF) is thought to be involved in the onset mechanism of ischemic colitis (IC). This study was designed to clarify the features of CMBF in six patients with IC (average age, 61 yr) and 10 control subjects (44.6yr). In the IC patients, CMBF was measured at both the affected site (ulcer margin and surrounding mucosa) and at a normal appearing site, during the active, healing, or recovery phase, by the laser doppler method. In the control group, CMBF was measured in various segments of the colon. The CMBF around the ulcer site during the active phase was significantly lower than that at the homologous segment in normal controls, while CMBF at normal sites in IC patients did not differ markedly from that of normal controls. CMBF in reddened portions of the ulcer margin in IC patients was significantly higher than that of the surrounding mucosa. A significant increase in CMBF was also observed during PGEi administration as compared to that before administration. This evidence clearly indicates that decreased CMBF is involved in the pathogenic mechanism of IC. The CMBF at sites of active mucosal restoration was somewhat increased. Continuous intravenous administration of PGEi was found to increase CMBF in IC patients.

Mural blood flow distribution in the large colon of horses during low-flow ischemia and reperfusion

SUMMARY Six horses were subjected to 3 hours of low-flow ischemia and 3 hours of reperfusion of the large colon. After induction of anesthesia, the large colon was exteriorized through a ventral midline celiotomy. Colonic blood flow was measured continuously, using Doppler ultrasonic flow probes placed on the colonic arteries supplying the dorsal and ventral colons and was allowed to stabilize for 15 to 30 minutes after instrumentation. Low-flow ischemia was induced by reducing colonic arterial blood flow to 20% of baseline (bl) flow. Colonic mucosal, seromuscular, and full-thickness blood flow were determined on a tissue-weight basis by injecting colored microspheres proximally into the colonic artery supplying the ventral colon. Reference blood samples were obtained at a known flow rate from the colonic artery and vein at a site more distal to the site of injection. Left ventral colon biopsy specimens were harvested at bl, 3 hours of ischemia, and 15 minutes of reperfusion. Blood and tissue samples were digested and filtered to collect the microspheres, and dimethylformamide was added to release the colored dyes. Dye concentration in blood and tissue samples was measured by use of spectrophotometry, and tissue-blood flow was calculated. Data were analyzed, using two-way anova for repeated measures; statistical significance was set at P &lt; 0.05. Doppler blood flow decreased to approximately 20% of BL, whereas microsphere blood flow ranged between 13.7 and 15.5% of bl at 3 hours of ischemia. Doppler-determined blood flow increased immediately on restoration of blood flow, reached 183% of bl at 15 minutes of reperfusion, and remained at or above bl throughout 3 hours of reperfusion. This reactive hyperemia was also detected, using the colored microspheres; blood flow increased to 242 and 327% of bl at 15 minutes of reperfusion in the mucosal and seromuscular layers, respectively. Mucosal blood flow was not different from seromuscular blood flow at any time, indicating relatively equal distribution of blood flow between these 2 layers. As determined from the venous reference samples, there was no evidence of arteriovenous anastomoses.

Mucosal vascular stasis precedes loss of viability of endothelial cells in rat acetic acid colitis

The hypothesis that a significant reduction in colonic mucosal perfusion, and hence ischemic injury, precedes the development of mucosal ulceration and inflammation is tested in this report. The microcirculatory changes in the rat colonic mucosa within 1 hr of topical exposure to 10% acetic acid were assessed. Colonic mucosal blood flow signals measured by laser Doppler flowmetry were significantly reduced to 61 +/- 8, 52 +/- 10, and 37 +/- 13% (mean +/- SEM) of baseline values at 1 min, 4 min, and 10 min after the colonic mucosa was exposed to 10% acetic acid, respectively, but not in controls exposed to saline. After the start of application of 10% acetic acid (for 4 min), in vivo microscopy studies demonstrated that colonic mucosal ischemia (stasis of the red blood cells in the mucosal capillaries) occurred at 9 +/- 5 min (mean +/- SEM). Evidence of endothelial cell death (failure to exclude a fluorescent dye, propidium iodide, by endothelial cells) developed at 25 +/- 10 min (mean +/- SEM). These findings indicate that within minutes after contact of the colonic mucosa with 10% acetic acid, colonic mucosal ischemia develops, followed shortly by death of endothelial cells. The data do not establish a cause-and-effect relationship between the reductions in mucosal blood flow and loss of endothelial cell viability in response to acetic acid. Nevertheless, because these events occur at such an early time point, they may play a pathogenetic role in the development of the subsequent inflammatory and ulcerative changes in this animal model of colitis. Further studies to define the potential causal relationships between these parameters are warranted.

Endoscopic Measurements of Canine Colonic, Mucosal Blood Flow Using Hydrogen Gas Clearance

We have examined the feasibility of hydrogen (H2) clearance for endoscopic measurements of colonic mucosal blood flow in anesthetized dogs. In 6 animals, measurements of H2 clearance did not differ significantly in different regions of the sigmoid colon and they were highly reproducible (p less than 0.001) on different days. In a total of 12 dogs, measurements of H2 clearance correlated closely with those obtained using radioactive microspheres under resting conditions and, in 4 dogs, during infusion of vasopressin (slope = 0.94, p less than 0.001). In 8 dogs, ligation of the major arteries supplying the sigmoid colon resulted in an acute 60% decrease in sigmoid mucosal blood flow (p less than 0.001); however, in 5 animals that survived the procedure, mucosal blood flow returned nearly to control levels as early as 3 days after operation. Endoscopic H2 clearance thus appears to be feasible for measuring mucosal blood flow in the colon. Serial measurements of H2 clearance may prove useful in characterizing the role of mucosal blood flow in the pathogenesis of various forms of human colonic disease.

Colon blood flow.

1. The intramural blood vessels of the human colon have been described. 2. Five methods that have been used to study colonic mucosal blood flow in patients with colostomies or with normal colons were briefly outlined along with some of the limitations of each technic. 3. It has been shown that the colonic mucosal blood flow in the human responds to heating or cooling an extremity, eating a meal, pharmacologic agents, and emotionally stressful situations. 4. Adequate evaluation of colonic mucosal blood flow requires the development of quantitative technics of measurement. It may be that methods of monitoring radioactive indicators with critically shielded detectors may meet some of these needs16 and offer a way of studying the intact colon of an unanesthetized human subject.