Long-term intake of resistant starch improves colonic mucosal integrity and reduces gut apoptosis and blood immune cells

OBJECTIVE: The potential effect of a long-term intake of resistant starch on colonic fermentation and on gut morphologic and immunologic indices of interest in bowel conditions in humans was studied in pigs. METHODS: Sixteen growing pigs were meal fed for 14 wk on a diet containing a large amount of raw potato starch (RPS; resistant starch) or corn starch (CS; digestible starch). Effects were assessed in the colon from the physicochemical properties of digesta and in the intestinal morphology, including lymphocytic infiltration, apoptosis, and proliferation activities. Hematologic and blood leukocyte cell subsets analysis were performed. RESULTS: After 97 d, the digestive content from RPS pigs was heavier than for CS pigs, producing a hypertrophy of tunica muscularis (P < 0.05). The proportion of butyrate was two-fold higher in proximal colon digesta in RPS pigs (P < 0.05). RPS-fed pigs had reduced apoptosis in the crypts, lamina propria and lymphoid nodules in the colon, and ileal Peyer's patches (P < 0.05). Fermentation of RPS reduced indices associated with damage to epithelial cells, such as crypt cell proliferation and magnesium excretion, whereas mucin sulfuration was increased, which promotes epithelial protection. The numbers of intraepithelial T cells and of blood leukocytes, neutrophils, and lymphocytes, mainly T-helper lymphocytes, were reduced in RPS pigs (P < 0.05). CONCLUSION: Long-term intake of RPS induces pronounced changes in the colonic environment, reduces damage to colonocytes, and improves mucosal integrity, reducing colonic and systemic immune reactivity, for which health benefits in inflammatory conditions are likely to be associated.     

Lactobacillus acidophilus 74-2 and Butyrate Induce Cyclooxygenase (COX)-1 Expression in Gastric Cancer Cells

Cyclo-oxygenase (COX) profile predicts prognosis of gastric cancer; COX-2 positive tumors are more often aggressive, and COX-2 suppression is protective against gastric cancer. In contrast, COX-1 suppression is harmful to the intestinal mucosa. The COX-1, COX-2, and COX-1ir expression profiles were measured with real-time PCR in primary (AGS) and metastatic (NCI-N87) gastric adenocarcinoma cell lines treated with butyrate, hyperosmolar medium, and, in the case of NCI-N87, cell-free supernatants of probiotic bacteria Lactobacillus acidophilus 74-2 and Bifidobacterium lactis 420. The cell lines showed differences in the profile when treated with either hyperosmolar medium or butyrate. In NCI-N87 COX-2 expression was higher but only COX-1 expression was significantly upregulated by butyrate. Similarly to butyrate, the cell-free supernatant of L. acidophilus 74-2 upregulated COX-1, while COX-2 expression remained unchanged. COX-1ir, including COX-3, was upregulated by probiotics and osmotic stress. In conclusion, consumption of L. acidophilus 74-2 could be beneficial for the expression of cytoprotective COX-1.     

Butyrate Inhibits NF-κB Activation in Lamina Propria Macrophages of Patients with Ulcerative Colitis

Background: In ulcerative colitis (UC) the activation (i.e. nuclear translocation) of nuclear factor kappa B (NF- &#115 B) is an important step in the regulation of cytokines secreted by lamina propria macrophages. Clinical trials suggest anti-inflammatory effects of locally administered butyrate in UC. The potential effects of butyrate on NF- &#115 B activation in lamina propria macrophages of UC patients were investigated. Methods: Eleven patients with distal UC were treated for up to 8 weeks with butyrate at 100 mM ( n = 6) or placebo ( n = 5) enemas. At entry and after 4 and 8 weeks, clinical status was noted and intestinal inflammation was graded endoscopically and histologically. Double-staining with antibodies against NF &#115 B (p65) and CD68 was employed to detect NF- &#115 B and macrophages, respectively. Results: In untreated patients, nuclear translocation of NF- &#115 B was detectable in virtually all macrophages. Butyrate treatment for 4 and 8 weeks resulted in a significant reduction in the number of macrophages being positive for nuclear translocated NF- &#115 B. In addition, butyrate significantly reduced both the number of neutrophils in crypt and surface epithelia and of the lamina propria lymphocytes/plasma cells. These findings correlated with a significant decrease in the Disease Activity Index (DAI). Conclusions: The decrease in DAI and mucosal inflammation in butyrate-treated patients is associated with a reduction of NF- &#115 B translocation in lamina propria macrophages. Since the inflammatory process in UC is mainly sustained by macrophage-derived cytokines, the known anti-inflammatory effects of butyrate may in part be mediated by an inhibition of NF- &#115 B activation in these macrophages.     

Probiotics and prebiotics and health in ageing populations

In healthy adults microbial communities that colonise different regions of the human colon contribute nutrients and energy to the host via the fermentation of non-digestible dietary components in the large intestine. A delicate balance of microbial species is required to maintain healthy metabolism and immune function. Disturbance in this microbial balance can have negative consequences for health resulting in elevated inflammation and infection, that are contributory factors in diabetes and cancer. There is a growing awareness that the microbial balance in the colon may become increasingly perturbed with aging and therefore hasten the onset of certain diseases. Societal and dietary factors influence microbial community composition both in the short and long term in the elderly (>65 years old) whilst immunosenescence may also be linked to a perturbed distal gut microbiota and frailty in the elderly. Significant progress has been made in defining some of the dominant members of the microbial community in the healthy large intestine and in identifying their roles in metabolism. There is therefore an urgent need for better awareness of the impact of diet, prebiotic and probiotic strategies in driving human colonic microbial composition in order to understand the possibilities for maintaining healthy gut function and well-being in an increasingly elderly population. Here we review gut microbial changes associated with aging and how diet, prebiotics and probiotics may modulate the gut microbiota to maintain health in the elderly.     

Cultures of Human Colonic Epithelial Cells Isolated from Endoscopical Biopsies from Patients with Inflammatory Bowel Disease. Effect of IFNγ, TNFα and IL-1β on Viability,Butyrate Oxidation and IL-8 Secretion

Cytokjne-mediated impairment of viability and metabolic function of epithelial cells has been suggested as a possible early pathogenic event in the development of inflammatory bowel disease (IBD). It is currently unknown whether pro-inflammatory cytokines have a direct effect on human nontransformed colonic epithelial cells. We investigated the effects of TNFα, IFNγ and IL-1β on viability, short chain fatty acid (butyrate) oxidation and IL-8 secretion in human colonic epithelial cell cultures in vitro obtained from macroscopically normal mucosa from IBD patients and controls. Colonic crypts were isolated from endoscopical biopsies by ultra-short (10 min) EDTA/EGTA treatment, and exposed to TNFα, IFNγ and IL-1β for 24 hours. The combination of TNFoc+IFNγ induced a significant decrease in cell viability as judged by methyltetrazoleum (MTT) metabolism which decreased to median 68 % of unexposed cultures (P<0.01). This effect was more pronounced than that observed after addition of TNFα (median 88 %) (P<0.05), but not IFNγ alone (median 78 %), whereas IL-1(3 had no significant effect. Cells from IBD patients were significantly less sensitive to TNFα + IFNγ exposure (median 74 %) compared to cells from controls (median 58 %) (P < 0.05). Butyrate oxidation, as measured by entrapment of CO₂, was not inhibited in cells exposed to TNFα + IFNγ, neither from controls (median 112%) nor from IBD patients (median 108%), suggesting a relative increase of this specific metabolic function in living cells in response to immunoinflammatory stress. IL-8 levels in cell supernatants were increased by TNFα + IFNγ, supporting the role of the epithelium in signalling between luminal factors and mucosal immune cells. In conclusion, we report that TNFα and IFNγ damage and influence human colonic epithelial cell function in vitro and that such mechanisms, if operative in vivo, also may be involved in the pathogenesis of IBD     

丁酸在溃疡性结肠炎中的作用概述

溃疡性结肠炎是以慢性复发性肠道炎症损伤为特征的一种疾病,虽然发病原因与病理机制还没有完全清楚,但许多证据表明其与肠黏膜对肠道微生物的免疫混乱有关,肠道微生物被认为是溃疡性结肠炎发生发展的主要因素之一。丁酸作为肠道膳食纤维由微生物发酵的主要终末产物之一,是肠上皮细胞重要能量来源,它维持肠道内稳态,在诸多方面如调节宿主的免疫应答与氧化应激反应、增强结肠黏膜的防御屏障与改善通透性等影响着溃疡性结肠炎的发生发展过程。     

丁酸钠对结肠癌细胞SW480凋亡及Survivin基因和VEGF表达水平的影响

目的:观察丁酸钠对结肠癌细胞株SW480的生长抑制、凋亡情况以及对生成素(Survivin)表达水平的影响,探讨其预防结肠癌的作用机制。方法:人传代结肠癌细胞株SW480经不同浓度丁酸钠处理后,在不同时段收集细胞,分别用四唑蓝法检测肿瘤细胞生长增殖率,流式细胞仪检测细胞周期和凋亡变化,免疫细胞化学技术观察对Survivin基因和血管内皮生长因子(VEGF)表达水平。结果:随着培养液中丁酸浓度的不断升高和培养时间的延长,SW480细胞的生长逐渐受到抑制。G1期细胞明显增多,S期细胞明显减少,细胞凋亡率逐渐增高。Sur-vivin基因和VEGF随着丁酸浓度的不断升高而表达下降。结论:丁酸钠能抑制SW480细胞株增生,诱导凋亡,并抑制Survivin基因和VEGF表达。     

Plasma pharmacokinetics of butyrate after intravenous administration of sodium butyrate or oral administration of tributyrin or sodium butyrate to mice and rats

Purpose: To define the plasma concentrations of butyrate achieved and the profile of plasma butyrate concentrations versus time in mice and rats treated with tributyrin or sodium butyrate. Methods: Female CD₂F₁ mice were treated with tributyrin by oral gavage or with sodium butyrate by i.v. bolus or oral gavage. Oral tributyrin doses delivered to mice were 3.1, 5.2, 7.8, and 10.3 g/kg. Intravenous sodium butyrate doses were 0.31, 0.62, 0.94, and 1.25 g/kg. Oral sodium butyrate was given to mice at 5 g/kg. Subsequently, similar studies were performed in female Sprague-Dawley rats. Rats were given tributyrin by oral gavage at doses of 3.6, 5.2, or 10.3 g/kg or sodium butyrate i.v. at a dose of 500 mg/kg. Plasma butyrate concentrations were determined by gas chromatography. Results: In mice, oral dosing with tributyrin resulted in detectable plasma butyrate concentrations as early as at 5 min after treatment and produced peak plasma butyrate concentrations at between 15 and 60 min after dosing. Peak plasma butyrate concentrations increased proportionally with increasing tributyrin dose, but as the oral tributyrin dose increased there was a greater than proportional increase in the area under the curve of plasma butyrate concentrations versus time (AUC). At a tributyrin dose of 10.3 g/kg, plasma butyrate concentrations peaked at approximately 1.75 mM and remained ≥1 mM for between 10 and 60 min after dosing. However, approximately 10% of mice treated with this dose died acutely. At a tributyrin dose of 7.8 g/kg, plasma butyrate concentrations reached approximately 1 mM by 15 min after dosing and remained between 0.8 and 1 mM until 60 min after dosing. No mouse treated with this dose died acutely. Mice given tributyrin doses of 5.2 and 3.1 g/kg achieved peak plasma butyrate concentrations of approximately 0.9 and 0.5 mM, respectively, by 45 min after dosing. Plasma butyrate concentrations in these mice remained above 0.1 mM until 120 and 90 min after dosing, respectively. The four i.v. doses of sodium butyrate resulted in plasma concentration-time profiles that also indicated nonlinear pharmacokinetics and were well described by a one-compartment model with saturable elimination. Values recorded for the Michaelis-Menten constant (K _m) and the maximal velocity of the process (V_max) ranged between 1.02 and 5.65 mM and 0.60 and 1.82 mmol/min, respectively. Values noted for the volume of the central compartment (V_c) varied between 0.48 and 0.72 l/kg. At 1.25 g/kg, i.v. sodium butyrate produced peak plasma butyrate concentrations of 10.5–17.7 mM, and plasma butyrate concentrations remained above 1 mM for 20–30 min. Sodium butyrate delivered orally to mice at 5 g/kg produced peak plasma butyrate concentrations of approximately 9 mM at 15 min after dosing and plasma butyrate concentrations exceeding 1 mM for 90 min after dosing. In rats the 10.3-g/kg oral dose of tributyrin produced peak plasma butyrate concentrations of approximately 3 mM by 75 min after dosing and butyrate concentrations excedding 1 mM from 30 to 90 min after dosing. The plasma butyrate concentrations produced in rats by 5.2- and 3.6-g/kg doses were appropriately lower than those produced by the 10.3-g/kg dose, and there was no evidence of nonlinearity. The 500-mg/kg i.v. dose of sodium butyrate produced peak plasma butyrate concentrations in rats of approximately 11 mM, and the decline in plasma butyrate concentrations with time after dosing was consistent with saturable clearance. Conclusion: These studies document the ability to use oral administration of tributyrin to achieve pharmacologically relevant concentrations of butyrate in rodent plasma. They also document the nonlinear nature of butyrate clearance. These data are being used in the design of clinical trials of oral tributyrin in patients with malignancies and hemoglobinopathies. Received: 27 July 1998 / Accepted: 3 November 1998     

Current trends in the development and application of molecular technologies for cancer epigenetics

Current progress in epigenetic research supports the view that diet and dietary components are important in cancer etiology by enhancing or inhibiting carcinogenesis.Since diet and dietary factors may significantly contribute to the causation and progression of many cancers,it is important to find the molecular mechanisms of action of such dietary factors for cancer prevention and treatment.Recently,the role of epigenetic mechanisms in the cancer development and progression has attracted more attention as additional evidence along with traditional DNA sequence based mechanisms such as mutations and structural re-arrangements.Such an increasing interest in cancer epigenetics has also accelerated the development and application of molecular assays and tools for DNA methylation detection and histone modification enrichment analysis.In this paper,key assays and methods for epigenetic research are reviewed and discussed in terms of their utility and usability.In addition,more advanced methods for genome-wide analysis are introduced as part of upcoming research trends and directions.