细胞凋亡在急性坏死型胰腺炎早期肠黏膜上皮细胞死亡中的作用

目的观察急性坏死型胰腺炎(ANP)大鼠早期肠黏膜上皮细胞凋亡的发生及其演变规律,探讨其在肠黏膜屏障功能障碍中的作用.方法Spraque-Dawley大鼠48只,采用胆胰管内逆行注入5%牛磺胆酸钠溶液诱导大鼠ANP模型.假手术组(shamoperation,SO)大鼠仅作剖腹术.术后3、6、12和24h分批处死大鼠,取末端回肠组织,分别应用DNA琼脂糖凝胶电泳,异硫氰基荧光素(FITC)结合的Annexin-V和碘化丙啶(PI)标记细胞作流式细胞仪(FCM)检测和免疫组化(TUNEL法)等技术,研究ANP大鼠肠黏膜上皮细胞凋亡.结果DNA琼脂糖凝胶电泳结果显示,SO组仅于12h出现"梯形(ladder)”条带;ANP大鼠,各时点均可见"梯形”条带.FCM检测结果表明,术后3、6、12和24hSO组肠黏膜上皮脱落细胞凋亡比例分别为(53.7±3.7)%、(27.6±6.0)%、(39.0±4.8)%和(29.0±11.3)%;ANP组分别为(50.3±11.3)%、(79.7±9.2)%、(47.8±17.3)%和(49.6±9.5)%,术后6h达峰值,且较SO组明显增高(P＜0.01),凋亡与坏死细胞(A/N)比值为27.7±17.9.TUNEL法显示,凋亡细胞呈细胞核固缩、断片状,术后3、6、12和24hSO组凋亡指数为6.2±2.4、7.5±1.7、10.5±0.9和5.3±0.8;ANP组为17.5±3.5、20.4±2.9、14.8±1.7和14.2±2.1,各时点均较SO组明显增高(P＜0.01),且于术后6h达峰值.结论ANP大鼠早期肠黏膜上皮细胞凋亡为细胞死亡的主要模式;肠黏膜上皮细胞凋亡在ANP大鼠早期肠黏膜屏障功能障碍发生中可能起重要作用.     

Regulation of iron homeostasis through the erythroferrone-hepcidin axis in sickle cell disease

Sickle cell disease (SCD) has a distinct pattern of transfusional iron overload (IO) when compared to transfusion-dependent β-thalassaemia major (TDT). We conducted a single institution prospective study to evaluate plasma biomarkers of iron regulation and inflammation in patients with SCD with IO (SCD IO cases, n = 22) and without IO (SCD non-IO cases, n = 11), and non-SCD controls (n = 13). Hepcidin was found to be inappropriately low, as evidenced by a significantly higher median hepcidin/ferritin ratio in non-SCD controls compared to SCD IO cases (0·3 vs. 0·02, P < 0·0001) and SCD non-IO cases (0·3 vs. 0·02, P < 0·0001), suggesting that certain inhibitory mechanism (s) work to suppress hepcidin in SCD. As opposed to the SCD non-IO state, where hepcidin shows a strong significant positive correlation with ferritin (Spearman ρ = 0·7, P = 0·02), this correlation was lost when IO occurs (Spearman ρ = −0·2, P = 0·4). Although a direct non-linear correlation between erythroferrone (ERFE) and hepcidin did not reach statistical significance both in the IO (Spearman ρ = −0·4, P = 0·08) and non-IO state (Spearman ρ = −0·6, P = 0·07), patients with highest ERFE had low hepcidin levels, suggesting that ERFE contributes to hepcidin regulation in some patients. Our results suggest a multifactorial mechanism of hepcidin regulation in SCD.     

Glutamine depletion induces murine neonatal melena with increased apoptosis of the intestinal epithelium

AIM:To investigate the possible biological outcome and effect of glutamine depletion in neonatal mice and rodent intestinal epithelial cells.METHODS:We developed three kinds of artificial milk with different amounts of glutamine;Complete amino acid milk (CAM),which is based on maternal mouse milk,glutamine-depleted milk (GDM),and glutaminerich milk (GRM).GRM contains three-fold more glutamine than CAM.Eighty-seven newborn mice were divided into three groups and were fed with either of CAM,GDM,or GRM via a r...     

Increased small intestinal apoptosis in coeliac disease.

BACKGROUND: Coeliac disease (CD) mucosa is flattened despite epithelial hyperproliferation. AIMS: To establish mechanisms of cell loss in CD. PATIENTS: 14 controls, 17 active CD patients, and 16 maintained with gluten free diet. METHODS: Programmed cell death was examined in small intestinal biopsy specimens by staining fragmented DNA using terminal uridine deoxynucleotidyl nick end labelling (TUNEL), in comparison with haematoxylin and eosin stained adjacent sections. Double staining with anti-CD45 antibodies determined the origin of apoptotic cells. Apoptosis was graded from 1-3 (< 5, 5-20, > 20% respectively). Proliferating cells, immunostained by Ki-67 (MIB-1) antibody, were counted. RESULTS: Apoptotic cells were seen rarely by haematoxylin and eosin but more readily by TUNEL. In controls, 1.4 +/- 0.2% of epithelial cells were apoptotic (mean grade 1.1), mainly located in the upper villus. In active CD, frequent apoptotic cells were distributed throughout the crypt-villus unit (mean grade 2.4), decreasing after treatment to 1.1 (p < 0.001) even when still histologically abnormal. CD45 antibodies rarely stained apoptotic cells in active CD. The number of TUNEL positive cells correlated with proliferating cell number (p < 0.001). CONCLUSION: Enterocyte apoptosis is greatly increased in untreated CD, correlates with proliferation, and falls to normal with a gluten free diet, before histological improvement. Increased apoptosis may be responsible for villous atrophy in CD.     

Regulation of homeostasis and inflammation in the intestine

The gastrointestinal tract is the largest immune interface with the environment. Exposure to large numbers of dietary and microbial antigens requires complex and highly regulated immune responses by different mucosal cell types, which result in the induction and maintenance of intestinal homeostasis. Defects in this equilibrium can disrupt the homeostatic mechanisms and lead to chronic intestinal inflammation. We review the cell populations and mechanisms involved in the control of intestinal homeostasis and inflammation, focusing on inflammatory bowel diseases. We describe some aspects of gut immunity that could alter the delicate balance between inflammatory and tolerogenic responses and result in chronic gastrointestinal tract inflammation in patients.     

The microvilli of the small intestinal surface epithelium in coeliac disease and in idiopathic steatorrhoea

This paper records the electron microscopic appearances in patients with coeliac disease and idiopathic steatorrhoea, and draws attention to the changes in the microvilli.     

Presence of bacteria and innate immunity of intestinal epithelium in childhood celiac disease

OBJECTIVES: Exposure to gliadin and related prolamins and appropriate HLA-DQ haplotype are necessary but not sufficient for contracting celiac disease (CD). Aberrant innate immune reactions could be contributing risk factors. Therefore, jejunal biopsies were screened for bacteria and the innate immune status of the epithelium investigated. METHODS: Children with untreated, treated, challenged CD, and controls were analyzed. Bacteria were identified by scanning electron microscopy. Glycocalyx composition and mucin and antimicrobial peptide production were studied by quantitative RT-PCR, antibody and lectin immunohistochemistry. RESULTS: Rod-shaped bacteria were frequently associated with the mucosa of CD patients, with both active and inactive disease, but not with controls. The lectin Ulex europaeus agglutinin I (UEAI) stained goblet cells in the mucosa of all CD patients but not of controls. The lectin peanut agglutinin (PNA) stained glycocalyx of controls but not of CD patients. mRNA levels of mucin-2 (MUC2), alpha-defensins HD-5 and HD-6, and lysozyme were significantly increased in active CD and returned to normal in treated CD. Their expression levels correlated to the interferon-gamma mRNA levels in intraepithelial lymphocytes. MUC2, HD-5, and lysozyme proteins were seen in absorptive epithelial cells. beta-defensins hBD-1 and hBD-2, carcinoembryonic antigen (CEA), CEA cell adhesion molecule-1a (CEACAM1a), and MUC3 were not affected. CONCLUSIONS: Unique carbohydrate structures of the glycocalyx/mucous layer are likely discriminating features of CD patients. These glycosylation differences could facilitate bacterial adhesion. Ectopic production of MUC2, HD-5, and lysozyme in active CD is compatible with goblet and Paneth cell metaplasia induced by high interferon-gamma production by intraepithelial lymphocytes.     

Regulation of endothelial cell survival and apoptosis during angiogenesis

The process of angiogenesis plays an important role in many physiological and pathological conditions. Inhibition of endothelial cell (EC) apoptosis providing EC survival is thought to be an essential mechanism during angiogenesis. Many of the angiogenic growth factors inhibit EC apoptosis. In addition, the adhesion of ECs to the extracellular matrix or intercellular adhesion promotes EC survival. In contrast, increasing evidence suggests that the induction of EC apoptosis may counteract angiogenesis. In this review, we focus on the regulation of EC survival and apoptosis during angiogenesis and especially on the effects and intracellular signaling promoted by angiogenic growth factors, endogenous angiogenic inhibitors (such as angiostatin, endostatin, and thrombospondin-1), and the adhesion to the extracellular matrix. Furthermore, we discuss the effects of cross talk between adhesion molecules and growth factors. Understanding the molecular mechanisms involved in the regulation of EC survival and apoptosis may provide new targets for the development of new therapies to enhance angiogenesis in the case of tissue-ischemia (eg, the neovascularization of myocardium) or to inhibit angiogenesis in the case of neovascularization-dependent disease (eg, tumor, diabetic retinopathy).     

Regulation of neutrophil homeostasis

PURPOSE OF REVIEW: Neutrophils are an essential component of the innate immune response and a major contributor to inflammation. Consequently, neutrophil number in the blood is tightly regulated. Herein, we review recent studies that have greatly advanced our understanding of the mechanisms controlling neutrophil homeostasis. RECENT FINDINGS: Accumulating evidence shows that stromal derived factor-1 (CXCL12) through interaction with its major receptor CXCR4 provides a key retention signal for neutrophils in the bone marrow. Granulocyte colony-stimulating factor induces neutrophil release from the bone marrow, in major part, by disrupting stromal derived factor-1/CXCR4 signaling. Granulocyte colony-stimulating factor expression is regulated by a novel feedback loop that senses neutrophil emigration into tissues. Specifically, engulfment of apoptotic neutrophils by tissue phagocytes initiates a cytokine cascade that includes interleukin-23, interleukin-17, and ultimately granulocyte colony-stimulating factor. SUMMARY: Granulocyte colony-stimulating factor plays a central role in the dynamic regulation of neutrophil production and release from the bone marrow in response to environmental stresses. Recent studies have begun to elucidate both the pathways linking neutrophil clearance to granulocyte colony-stimulating factor expression and the mechanisms by which the factor induces neutrophil release from the bone marrow. These studies may lead to novel strategies to modulate neutrophil responses in host defense and inflammation.     

Chemokines and chemokine receptors in mucosal homeostasis at the intestinal epithelial barrier in inflammatory bowel disease

Chemokines, a large family of small chemoattractive cytokines, and their receptors play an integral role in the regulation of the immune response and homeostasis. The ability of chemokines to attract specific populations of immune cells sets them apart from other chemoattractants. Chemokines produced within the gastrointestinal mucosa are critical players in directing the balance between physiological and pathophysiological inflammation in health, inflammatory bowel disease (IBD), and the progression to colon cancer. In addition to the well-characterized role of chemokines in directed trafficking of immune cells to the gut mucosa, the expression of chemokine receptors on the cells of the epithelium makes them active participants in the chemokine signaling network. Recent findings demonstrate an important role for chemokines and chemokine receptors in epithelial barrier repair and maintenance as well as an intricate involvement in limiting metastasis of colonic carcinoma. Increased recognition of the association between barrier defects and inflammation and the subsequent progression to cancer in IBD thus implicates chemokines as key regulators of mucosal homeostasis and disease pathogenesis.     

Role of the thyroid gland in blood calcium homeostasis during intestinal calcium absorption in man

An oral load of calcium with a radiocalcium trace dose was administered to totally thyroidectomized and control subjects in order to evaluate the role of the thyroid gland in the control of postprandial hypercalcemia. The following results were obtained: (1) During calcium absorption thyroidectomized subjects showed blood calcium increments significantly higher than controls. (2) Plasma radioactivity appearance curves in both thyroidectomized and control subjects were similar. (3) Any difference in the intestinal absorption of calcium was not found between the experimental groups. From the results, a difference in intestinal calcium absorption and/or in the size and turnover rate of the dilutional compartment of calcium can be ruled out as possible causes of higher blood calcium increments in thyroidectomized subjects. Therefore, impaired homeostatic control caused by the lack of calcitonin appeared to be the most satisfactory explanation for these findings. However, considering that there is only a slight difeerence in calcium increments and that calcium elevations were constantly observed also in controls, the effect of the hormone on blood calcium homeostasis appears to be of limited importance. The bone calcium-sparing effect of the hormone during post-prandial hypercalcemia in the adult can probably be considered physiologically more significant than protection against hypercalcemia.     

Probiotics maintain the intestinal microbiome homeostasis of the sailors during a long sea voyage

ABSTRACT

The challenging conditions encountered during long sea voyages increase the risk of health-threatening physiological and psychological stress for sailors compared with land-based workers. However, how the intestinal microbiota responds to a long sea voyage and whether there is a feasible approach for protecting gut health during sea voyage are still unexplored. Here, we designed a 30-d longitudinal study including a placebo group (n = 42) and a probiotic group (n = 40) and used shotgun metagenomic sequencing to explore the impacts of sea voyage on the intestinal microbiome of sailors. By comparing the intestinal microbiome of subjects in the placebo group at baseline (d 0) and at the end of the sea voyage (d 30), we observed an alteration in the intestinal microbiome during the long sea voyage based on the microbial structure; the results revealed an increase in the species Streptococcus gordonii and Klebsiella pneumoniae as well as a decrease in some functional features. However, the change in the microbial structure of sailors in the probiotic group between d 0 and d 30 was limited, which indicated a maintenance effect of probiotics on intestinal microbiome homeostasis. At the metagenomic strain level, a generally positive correlation was observed between probiotics and the strains belonging to Bifidobacterium longum and Bifidobacterium animalis, whereas a common negative correlation was observed between probiotics and Clostridium leptum; this result revealed the potential mechanism of maintaining intestinal microbiome homeostasis by probiotics. The present study provided a feasible approach for protecting gut health during a long sea voyage.     

Localization of endotoxin in the rat intestinal epithelium

High levels of circulating lipopolysaccharide (LPS) cause intestinal inflammation and increased permeability to bacteria and toxins. To better understand the effects of LPS on the gut, confocal microscopy and immunofluorescence staining were used to examine the distribution of LPS in the rat intestine after intravenous or enteral administration. LPS was localized in macrophages in the lamina propria from 1 h to >28 days after intravenous injection. LPS was also detected in the epithelial cells from 8 h to 7 days after injection. In contrast, LPS administered enterally was found in the gut lumen in close proximity to the mucosa but was not detected in enterocytes at any time. The concentration of LPS in enterocytes near the villus tip provides a mechanism for the clearance of endotoxin, by the turnover and shedding of LPS-containing enterocytes into the gut lumen, that has not been previously described.     

Phasic study of intestinal homeostasis disruption in experimental intestinal obstruction

AIM: To investigate the phasic alteration of intestinal homeostasis in an experimental model of intestinal obstruction.METHODS: A rabbit model of intestinal obstruction was established by transforming parts of an infusion set into an in vivo pulled-type locking clamp and creating a uniform controllable loop obstruction in the mesenteric non-avascular zone 8 cm from the distal end of the ileum. The phasic alteration of intestinal homeostasis was studied after intestinal obstruction. The changes in goblet cells, intraepithelial lymphocytes, lamina propria lymphocytes, and intestinal epithelium were quantified from periodic acid-Schiff-stained sections. Ornithine decarboxylase (ODC) activity and serum citrulline levels were measured by high-performance liquid chromatography. Claudin 1 mRNA expression was examined by real-time polymerase chain reaction analysis. Intestinal microorganisms, wet/dry weight ratios, pH values, and endotoxin levels were determined at multiple points after intestinal obstruction. Furthermore, the number and ratio of CD3⁺, CD4⁺ and CD8⁺ T cells were determined by flow cytometry, and secretory IgA levels were measured with an enzyme-linked immunosorbent assay.RESULTS: A suitable controllable rabbit model of intestinal obstruction was established. Intestinal obstruction induced goblet cell damage and reduced cell number. Further indicators of epithelial cell damage were observed as reduced serum citrulline levels and claudin 1 gene expression, and a transient increase in ODC activity. In addition, the wet/dry weight ratio and pH of the intestinal lumen were also dramatically altered. The ratio of Bacillus bifidus and enterobacteria was reversed following intestinal obstruction. The number and area of Peyer’s patches first increased then sharply decreased after the intestinal obstruction, along with an alteration in the ratio of CD4/CD8⁺ T cells, driven by an increase in CD3⁺ and CD8⁺ T cells and a decrease in CD4⁺ T cells. The number of lamina propria lymphocytes also gradually decreased with prolonged obstruction.CONCLUSION: Intestinal obstruction can induce disruption of intestinal homeostasis.