Effect of lactobacillus on the gut microfiora and barrier function of the rats with abdominal infection

AIM: To investigate the effect of probiotics supplemented by gut on the tight junctions of epithelial cells, barrier function and the microflora of rats with abdominal infection. METHODS: After the model of cecal ligation and perforation established, SD rats were divided into two groups: parenteral nutrition (PN) group and PN+probiotics (probiotics) group, PN solution was supplemented by neck vein and probiotics was delivered via the jejunostomy tube for five days. Vena cava blood and the homogenated tissue of liver, lung and mesenteric lymph nodes were cultured to determine the bacterial translocation rate (BTR). The ultrastructure of epithelial tight junctions and microvilli of the gut were observed by electron microscopy; occluding expression was measured by indirect-immune fluorescence method; anaerobic bacterial growth by anaerobic culture and DNA fingerprint of bacterial colonies of the feces by PCR. RESULTS: The quantity of lactobacteria and bifydobacteria in probiotics group was higher than that of PN group. The profiles of DNA fingerprint expression in probiotics group were similar to that in the normal group, a new 16S rDNA sequence appeared in the profile in PN group. The occludin expression, the integrality of the gut epithelial tight junctionand microvilli in probiotics group were improved as compared with PN group. The BTR and endotoxin in blood were reduced more significantly in probiotics group as compared with PN group.CONCLUSION: The probiotics could improve the gut microflora disturbance, increase occludin expression, maintain the gut epithelial tight junction and decrease the bacterial translocations rate.     

Effect of lactobacillus on the gut microflora and barrier function of the rats with abdominal infection

AIM: To investigate the effect of probiotics supplemented by gut on the tight junctions of epithelial cells, barrier function and the microflora of rats with abdominal infection. METHODS: After the model of cecal ligation and perforation established, SD rats were divided into two groups: parenteral nutrition (PN) group and PIM+probiotics (probiotics) group, PN solution was supplemented by neck vein and probiotics was delivered via the jejunostomy tube for five days. Vena cava blood and the homogenated tissue of liver, lung and mesenteric lymph nodes were cultured to determine the bacterial translocation rate (BTR). The ultra-structure of epithelial tight junctions and microvilli of the gut were observed by electron microscopy; occluding expression was measured by indirect-immune fluorescence method; anaerobic bacterial growth by anaerobic culture and DNA fingerprint of bacterial colonies of the feces by PCR. RESULTS: The quantity of lactobacteria and bifydobacteria in probiotics group was higher than that of PN group. The profiles of DNA fingerprint expression in probiotics group were similar to that in the normal group, a new 16S rDNA sequence appeared in the profile in PN group. The occludin expression, the integrality of the gut epithelial tight junction and microvilli in probiotics group were improved as compared with PN group. The BTR and endotoxin in blood were reduced more significantly in probiotics group as compared with PN group. CONCLUSION: The probiotics could improve the gut microflora disturbance, increase occludin expression, maintain the gut epithelial tight junction and decrease the bacterial translocations rate.     

Intestinal permeability in patients after surgical trauma and effect of enteral nutrition versus parenteral nutrition

AIM: To study the intestinal permeability (IP) following stress of abdominal operation and the different effects on IP of enteral nutrition (EN) and parenteral nutrition (PN).METHODS: Forty patients undergoing abdominal surgery were randomized into EN group and PN group. Each group received nutritional support of the same nitrogen and calorie from postoperative day (POD) 3 to POD 11. On the day before operation (POD-1), POD 7 and POD 12, 10 g of lactulose and 5 g of mannitol were given orally, and urine was collected for 6 hours. Urine excretion ratios of lactulose and mannitol (L/M) were measured.RESULTS: L/M ratios of EN group on POD-1, POD 7 and POD 12 were 0.026±0.017, 0.059±0.026, 0.027±0.017,respectively, and those of PN group were 0.025±0.013,0.080±0.032, 0.047±0.021, respectively. Patients of both groups had elevated L/M ratios on POD 7 vs. POD-1. However the ratio returned toward control level in EN group by POD 12.In contrast, PN group still had elevated L/M ratios on POD 12.CONCLUSION: L/M ratio increases for a period of time after surgical trauma and the loss of gut mucosal integrity can be reversed by substitution of enteral nutrition.     

Brain-derived neurotrophic factor preserves intestinal mucosal barrier function and alters gut microbiota in mice

The intestinal mucosal barrier (IMB) enables the intestine to provide adequate containment of luminal microorganisms and molecules while preserving the ability to absorb nutrients. In this study, we explored the effect of brain-derived neurotrophic factor (BDNF) on IMB function and gut microbiota in mice. BDNF gene knock-out mice (the BDNF^+/? group) and wild-type mice (the BDNF^+/+ group) were selected. The gut microbiota of these mice was analyzed by denaturing gradient gel electrophoresis (DGGE) assay. The ultrastructure of the ileum and the colonic epithelium obtained from decapitated mice were observed by transmission electron microscopy. The protein expression of epithelial tight junction proteins, zonula occludens-1 (ZO-1) and occludin was detected by immunohistochemistry staining. The protein expression of claudin-1 and claudin-2 was determined by Western blotting. The DGGE band patterns of gut microbiota in the BDNF^+/? group were significantly different from that in the BDNF^+/+ group, which indicated that the BDNF expression alters the gut microbiota in mice. Compared with the BDNF^+/+ group, the BDNF^+/? group presented no significant difference in the ultrastructure of ileal epithelium; however, a significant difference was observed in the colonic epithelial barrier, manifested by decreased microvilli, widening intercellular space and bacterial invasion. Compared with the BDNF^+/+ group, the expression of ZO-1 and occludin in the BDNF^+/? group was significantly decreased. The expression of claudin-1 in the BDNF^+/? group was significantly reduced, while the expression of claudin-2 was elevated. These findings indicate that BDNF preserves IMB function and modulates gut microbiota in mice.     

Intestinal barrier damage caused by trauma and lipopolysaccharide

AIM: To investigate the intestinal barrier function damage induced by trauma and infection in rats. METHODS: Experimental models of surgical trauma and infection were established in rats. Adult Sprague-Dawley rats were divided into 4 groups: control group (n = 8), EN group (n = 10), PN group (n = 9) and Sep group (n = 8). The rats in PN and Sep groups were made into PN models that received isonitrogenous, isocaloric and isovolumic TPN solution during the 7-d period. Rats in EN and Sep groups received laparotomy and cervical catheterization on day 1 and received lipopolysaccharide injection intraperitoneally on d 7. On the 7(th) day all the animals were gavaged with lactulose and mannitol to test the intestinal permeability. Twenty-four hours later samples were collected and examined. RESULTS: The inflammatory responses became gradually aggravated from EN group to Sep group. The mucosal structure of small intestine was markedly impaired in PN and Sep groups. There was a low response in IgA level in Sep group when compared with that of EN group. Lipopolysaccharide injection also increased the nitric oxide levels in the plasma of the rats. The intestinal permeability and bacterial translocation increased significantly in Sep group compared with that of control group. CONCLUSION: One wk of parenteral nutrition causes an atrophy of the intestinal mucosa and results in a moderate inflammatory reaction in the rats. Endotoxemia aggravates the inflammatory responses that caused by laparotomy plus TPN, increases the production of nitric oxide in the body, and damages the intestinal barrier function.     

Functional and morphological changes of the gut barrier during the restitution process after hemorrhagic shock

AIM: To investigate the functional, morphological changes of the gut barrier during the restitution process after hemorrhagic shock, and the regional differences of the large intestine and small intestine in response to ischemia/reperfusion injury. METHODS: Forty-seven Sprague-Dawley rats with body weight of 250-300 g were divided into two groups: control group (sham shock n = 5) and experimental group (n = 42). Experimental group was further divided into six groups (n = 7 each) according to different time points after the hemorrhagic shock, including 0(th) h group, 1st h group, 3rd h group, 6th h group, 12th h group and 24th h group. All the rats were gavaged with 2 mL of suspension of lactulose (L) (100 mg/2 mL) and mannitol (M) (50 mg/each) at the beginning and then an experimental rat model of hemorrhagic shock was set up. The specimens from jejunum, ileum and colon tissues and the blood samples from the portal vein were taken at 0, 1, 3, 6, 12 and 24 h after shock resuscitation, respectively. The morphological changes of the intestinal mucosa, including the histology of intestinal mucosa, the thickness of mucosa, the height of villi, the index of mucosal damage and the numbers of goblet cells, were determined by light microscope and/or electron microscope. The concentrations of the bacterial endotoxin lipopolysaccharides (LPS) from the portal vein blood, which reflected the gut barrier function, were examined by using Limulus test. At the same time point, to evaluate intestinal permeability, all urine was collected and the concentrations of the metabolically inactive markers such as L and M in urine were measured by using GC-9A gas chromatographic instrument. RESULTS: After the hemorrhagic shock, the mucosal epithelial injury was obvious in small intestine even at the 0(th) h, and it became more serious at the 1st and the 3rd h. The tissue restitution was also found after 3 h, though the injury was still serious. Most of the injured mucosal restitution was established after 6 h and completed in 24 h. Two distinct models of cell death-apoptosis and necrosis-were involved in the destruction of rat intestinal epithelial cells. The number of goblet cells on intestinal mucosa was reduced significantly from 0 to 24 h (the number from 243+/-13 to 157+/-9 for ileum, 310+/-19 to 248+/-18 for colon; r = -0.910 and -0.437 respectively, all P<0.001), which was the same with the large intestine, but the grade of injury was lighter with the values of mucosal damage index in 3 h for jejunum, ileum, and colon being 2.8, 2.6, 1.2, respectively. The mucosal thickness and the height of villi in jejunum and ileum diminished in 1 h (the average height decreased from 309+/-24 to 204+/-23 microm and 271+/-31 to 231+/-28 microm, r = -0.758 and -0.659, all P<0.001; the thickness from 547+/-23 to 418+/-28 microm and 483+/-45 to 364+/-35 microm, r = -0.898 and -0.829, all P<0.001), but there was no statistical difference in the colon (F = 0.296, P = 0.934). Compared with control group, the urine L/M ratio and the blood LPS concentration in the experimental groups raised significantly, reaching the peak in 3-6 h (L/M: control vs 3 h vs 6 h was 0.029+/-0.09 vs 0.063+/-0.012 vs 0.078+/-0.021, r = -0.786, P<0.001; LPS: control vs 3 h vs 6 h was 0.09+/-0.021 vs 0.063+/-0.012 vs 0.25+/-0.023, r = -0.623, P<0.001), and it kept increasing in 24 h. CONCLUSION: The gut barrier of the rats was seriously damaged at the early phase of ischemic reperfusion injury after hemorrhagic shock, which included the injury and atrophy in intestinal mucosa and the increasing of intestinal permeability. Simultaneously, the intestinal mucosa also showed its great repairing potentiality, such as the improvement of the intestinal permeability and the recovery of the morphology at different phases after ischemic reperfusion injury. The restitution of gut barrier function was obviously slower than that of the morphology and there was no direct correlation between them. Compared with the small intestine, the large intestine had stronger potentiality against injury. The reduction of the amount of intestinal goblet cells by injury did not influence the ability of intestinal mucosal restitution at a certain extent and it appeared to be intimately involved in the restitution of the epithelium.     

肿瘤坏死因子α对暴发性肝衰竭小鼠肠上皮细胞紧密连接蛋白表达的影响

目的 研究TNFα对暴发性肝衰竭（FHF）小鼠大肠上皮细胞紧密连接蛋白occludin表达的影响。方法 采用D-氨基半乳糖（GaIN）和内毒素（LPS）联合腹腔注射（ip）制备FHF小鼠动物模型,并设立TNFα组（TNFα10μs／kg,ip）,TNFα抗体组（注射LPS和GaIN前30min尾静脉注射TNFd抗体100μg/只）。在不同的时间点（6h,9h）处死动物,应用免疫组织化学技术、Westernblot及实时定量PCR检测各组小鼠大肠上皮细胞紧密连接蛋白occludin表达的变化。结果 在生理盐水（NS）对照组,几乎整张切片上均可见到沿大肠黏膜上皮细胞膜顶端呈线性分布的occludin阳性染色,但FHF组及TNFα组小鼠的阳性染色较之明显减弱,TNFα抗体组occludin的阳性反应与Ns对照组比较无明显减弱。Westernblot结果与免疫组织化学结果相一致,FHF组及TNFα组小鼠9h时occludin蛋白含量明显下降,与NS对照组相比差异有统计学意义（0．36±0．05,0．48±0．02比0．71±0．09,P〈0．05）,TNFα抗体组与NS对照组相比差异无统计学意义（0．74±0．03比0．71±0．09,P〉0．05）。实时定量PCR结果显示,FHF组与TNFα组小鼠6h时occludinmRNA水平最低,与NS对照组相比差异有统计学意义（0．72±0．04,0．81±0．03比1．00±0．05,P〈0．05）,TNFα抗体组与Ns对照组相比差异无统计学意义（1．01±0．10比1．00±0．05,P〉0．05）。结论 在小鼠暴发性肝衰竭过程中,TNFα介导大肠上皮细胞间紧密连接蛋白occludin表达的下降。     

Specific probiotics alleviate allergic rhinitis during the birch pollen season

AIM： To investigate whether birch pollen allergy symptoms are linked with gut microbiota changes and whether probiotics have an effect on these.
METHODS： Forty seven children with confirmed birch pollen allergy were randomized to receive either a probiotic combination of Lactobacillus acidophilus （L. acidophilus） NCFM^TM （ATCC 700396） and Bifidobacterium lactis （B. lactis） BI-04 （ATCC SD5219） or placebo in a double-blind manner for 4 mo, starting prior to onset of the birch pollen season. Symptoms were recorded in a diary. Blood samples were taken for analysis of cytokines and eosinophils. Fecal samples were analysed for microbiota components, calprotectin and IgA. Nasal swabs were taken for analysis of eosinophils.
RESULTS： The pollen season induced a reduction in Bifldobacterium , Clostridium and Bacteroides which could not be prevented by the probiotic intervention. During the intervention, significantly higher numbers ofB. lactis 11.2 × 10^7 ± 4.2 ×10^7 vs 0.1 × 10^7 ± 0.1 × 10^7 bacteria/g feces （P 〈 0.0001） and L. acidophilus NCFMTM 3.5 × 10^6 ± 1.3 × 10^6 vs 0.2 × 10^6 ±0.1 × 10^6 bacteria/g feces （P 〈 0.0001） were observed in the probiotic group compared to the placebo group.During May, there was a tendency for fewer subjects, （76.2% vs 95.2%, P = 0.078） to report runny nose, while during June, fewer subjects, 11.1% vs 33.3%, reported nasal blocking in the probiotics group （P = 0.101）. Concomitantly, fewer subjects in the probiotic group had infiltration of eosinophils in the nasal mucosa compared to the placebo group, 57.1% vs 95% （P = 0.013）. Eye symptoms tended to be slightly more frequent in the probiotic group, 12.5 d [interquartile range （IQR） 6-18] vs 7.5 d （IQR 0-11.5） （P = 0.066） during May. Fecal IgA was increased in the placebo group during the pollen season; this increase was prevented by the probiotics （P = 0.028）.
CONCLUSION： Birch pollen allergy was shown to be associated with changes in fecal microbiota composition. The specific combination of probiotics used was shown to prevent the pollen-induced infiltration of eosinophils into the nasal mucosa, and indicated a trend for reduced nasal symptoms.     

The influence of Enteral Nutrition in postoperative patients with poor liver function

AIM: To investigate the safety, rationality and the practicality of enteral nutritional (EN) support in the postoperative patients with damaged liver function and the protective effect of EN on the gut barrier.METHODS: 135 patients with liver function of Child B or C grade were randomly allocated to enteral nutrition group (EN, 65 cases), total parenteral nutrition group (TPN, 40cases) and control group (CON, 30 cases). Nutritional parameters, hepatic and kidney function indexes were measured at the day before operation, 5th and 10th day after the operation respectively. Comparison was made to evaluate the efficacy of different nutritional support. Urinary concentrations of lactulose(L) and mannitol(M) were measured by pulsed electrochemical detection(HPLC-PED)and the L/M ratio calculated to evaluate their effectiveness on protection of gut barrier.RESULTS: No significant damages in hepatic and kidney function were observed in both EN and TPN groups between pre- and postoperatively. EN group was the earliest one reaching the positive nitrogen balance after operation and with the lowest loss of body weight and there was no change in L/M ratio after the operation (0.026±0.004) at the day 1before operation, 0.030±0.004 at the day 5 postoperative and 0.027±0.005 at the day 10 postoperative), but the change in TPN group was significant at the day 5postoperative (0.027±0.003 vs 0.038±0.009,P＜0.01).CONCLUSION: EN is a rational and effective method in patients with hepatic dysfunction after operation and has significant protection effect on the gut barrier.     

Gut dysbiosis and irritable bowel syndrome: The potential role of probiotics

Objective

To discuss the role of gut dysbiosis in the development of irritable bowel syndrome (IBS) and the impact of probiotics as a potential therapeutic measure.

水通道蛋白3与紧密连接的相关性

目的:探讨水通道蛋白3(aquaporin 3,AQP3)对肠黏膜上皮细胞间紧密连接(tight junction,TJ)的影响,并探讨其可能的作用机制.方法:应用Caco-2细胞系在体外构建肠黏膜上皮屏障,构建沉默AQP3的shRNA慢病毒载体,建立稳定转染细胞系.实验分为3组:空白对照组(BLANK)、阴性对照组(NC)、AQP3干扰组(AQP3 shRNA).Western blot验证TJ相关蛋白Occludin以及Claudin-1的表达情况;并且采用免疫细胞化学法观察TJ相关蛋白的分布和结构变化.结果:RT-PCR及Western blot结果显示在Caco-2细胞系中成功沉默AQP3的表达.干扰组与对照组相比下降约75%.Western blot结果显示AQP3干扰组TJ相关蛋白Occludin以及Claudin-1的表达明显降低.免疫细胞化学结果显示Caco-2细胞间Occludin以及Claudin-1主要表达在细胞膜和/或胞浆中,Occludin和Claudin-1细胞间棕褐色颗粒减少,结构变模糊.相邻Caco-2细胞间TJ结构遭到破坏.结论:靶向AQP3的shRNA技术可以引起TJ的结构变化和相关蛋白的表达分布的异常.     

Modulation of immunity and inflammatory gene expression in the gut,in inflammatory diseases of the gut and in the liver by probiotics

The potential for the positive manipulation of the gut microbiome through the introduction of beneficial microbes, as also known as probiotics, is currently an active area of investigation. The FAO/WHO define probiotics as live microorganisms that confer a health benefit to the host when administered in adequate amounts. However, dead bacteria and bacterial molecular components may also exhibit probiotic properties. The results of clinical studies have demonstrated the clinical potential of probiotics in many pathologies, such as allergic diseases, diarrhea, inflammatory bowel disease and viral infection. Several mechanisms have been proposed to explain the beneficial effects of probiotics, most of which involve gene expression regulation in specific tissues, particularly the intestine and liver. Therefore, the modulation of gene expression mediated by probiotics is an important issue that warrants further investigation. In the present paper, we performed a systematic review of the probiotic-mediated modulation of gene expression that is associated with the immune system and inflammation. Between January 1990 to February 2014, Pub Med was searched for articles that were published in English using the Me SH terms “probiotics " and " gene expression " combined with “intestines", "liver", "enterocytes", "antigen-presenting cells", "dendritic cells", "immune system", and "inflammation". Two hundred and five original articles matching these criteria were initially selected, although only those articles that included specific gene expression results(77) were later considered for this review and separated into three major topics: the regulation of immunity and inflammatory gene expression in the gut, in inflammatory diseases of the gut and in the liver. Particular strains of Bifidobacteria, Lactobacilli, Escherichia coli, Propionibacterium, Bacillus and Saccharomyces influence the gene expression of mucins, Toll-like receptors, caspases, nuclear factor-κB, and interleukins and lead mainly to an anti-inflammatory response in cultured enterocytes. In addition, the interaction of commensal bacteria and probiotics with the surface of antigenpresenting cells in vitro results in the downregulation of pro-inflammatory genes that are linked to inflammatory signaling pathways, whereas other anti-inflammatory genes are upregulated. The effects of probiotics have been extensively investigated in animal models ranging from fish to mice, rats and piglets. These bacteria induce a tolerogenic and hyporesponsive immune response in which many genes that are related to the immune system, in particular those genes expressing anti-inflammatory cytokines, are upregulated. By contrast, information related to gene expression in human intestinal cells mediated by the action of probiotics is scarce. There is a need for further clinical studies that evaluate the mechanism of action of probiotics both in healthy humans and in patients with chronic diseases. These types of clinical studies are necessary for addressing the influence of these microorganisms in gene expression for different pathways, particularly thosethat are associated with the immune response,and to better understand the role that probiotics might have in the prevention and treatment of disease.     

Bioflora Probiotic in Immunomodulation and Prophylaxis of Intestinal Bacterial Translocation in Rats

The immunomodulator effect of Bioflora probiotic on T (CD4+) and B (CD20) lymphocytes in gastrointestinal mucosa and intestinal bacterial translocation was studied using Wistar rats (n = 10 per group). Two experiments were used: (I) stress with immobilization and water immersion at 22 degrees C for 7 h plus the application of indomethacin (Indo) 10 mg/kg SC every 24 h for 3 days (comparator group), and (II) stress experiment I with the addition of 1 mL of Bioflora applied through a orogastric tube every 12 h for 3 days. At the 4th day, in asepsis, a dissection laparotomy of liver, spleen, mesenteric lymphatic nodes, and cecum was performed for microbiological culture, and stomach, ileum, and colon were also dissected for immunohistochemical and quantification of CD4+ and CD20. Findings in experiment I revealed cecum bacterial overdevelopment of 6 x 10(10) +/- 2.3 x 10(9) colony-forming units (CFU) (P < 0.01) and positive cultures in liver, spleen, and all mesenteric lymphatic nodes. On the other hand, in the group treated with Probiotic Bioflora, cecum without overdevelopment (3 x 10(6) +/- 1.3 x 10(5) CFU), negative cultures in liver and spleen, and in lymphatic nodes two positive and eight negative cultures for E. coli and P. vulgaris (P < 0.01) were observed. Immunohistochemistry revealed a relevant increase of T lymphocytes (CD4+) in ileum and colon. Conclusions Bioflora probiotic was shown to be an intestinal immunomodulator that induced increased T (CD4+) lymphocytes that also offer prophylaxis of intestinal bacterial translocation in a stressed rat model.     

三种营养支持治疗高龄心力衰竭并发肾功能不全患者疗效的比较

目的:观察和比较肠内肠外联合营养、单纯肠内营养、单纯肠外营养3种营养支持方法对高龄心力衰竭并发肾功能不全患者疗效的影响及安全性。方法:入选86例急性心力衰竭并发肾功能不全患者,随机分为3组:肠内肠外联合营养组(联合组,33例),肠内营养组(肠内组,27例),肠外营养组(肠外组,26例)。采用随机、开放、无安慰剂对照、无长期随访和预后终点的方法采集和统计信息,3组营养支持后7 d,进行临床疗效分析和安全性评价。结果:同组治疗前后比较,血清白蛋白(ALB)、前白蛋白(PAB)、转铁蛋白(TF)、N末端B型钠尿肽原(NT-proBNP)下降,均有统计学意义(P0.05),联合组与肠内组、肠外组治疗后比较,PAB、ALB、TF、NT-proBNP均有统计学意义(P0.05),肠内组与肠外组治疗后比较,PAB、ALB、TF、NT-proBNP均无统计学意义。3组治疗前及治疗后肌酐无显著差异。与治疗前比较,3组治疗后每搏输出量(SV)、左室射血分数(LVEF)、左室舒张末内径(LVEDD)、左室收缩末内径(LVESD)均显著改善(P0.05);联合组治疗后较肠内组、肠外组改善更显著(P0.05);肠内组与肠外组治疗后比较无统计学差异。3组临床症状改善率和不良事件发生率均无显著差异。结论:对于高龄心衰并发肾功能不全的患者,联合营养支持的近期疗效显著优于单独肠内营养支持和肠外营养支持。     

The Immunostimulatory Effect of Lactic Acid Bacteria in a Rat Model

Background: Probiotics are “live”, beneficial microbes that provide important health benefits in their hosts. There is significant interest in the modulation and regulation of the immune function by probiotics. Objective: To investigate the immunomodulatory effects of a probiotic mixture, including Lactobacillus and Bifidobacterium species, by detecting serum cytokine and immunoglobulin levels. Methods: The rats were randomly divided into 4 groups. The first group was “Control group” and other 3 groups were probiotic application groups who received different doses of probiotics. The probiotic mixture included 12 probiotic bacteria, mostly Lactobacillus and Bifidobacterium strains. Probiotic mixture was administered to rats for 12 consecutive days. TNF-α, TGF-β, IL-1-β, IL-6, and IL-10 levels as well as serum IgG and IgA concentrations were detected in the sera after 12 days. Results: Probiotics led to a decrease in the levels of TNF-α, IL-6 and TGF-β; however, they led to increase in the serum levels of IL-10, IgG and IgA. There were significant differences between control group and probiotic application groups (p<0.05). Conclusion: These data suggest that the commensal microbiota are important for stimulating both proinflammatory and regulatory responses in order to rapidly clear infections and minimize inflammation-associated tissue damage.     

Changes in tight junctional resistance of the cervical epithelium are associated with modulation of content and phosphorylation of occludin 65-kilodalton and 50-kilodalton forms

Treatment of human cervical epithelial CaSki cells with ATP or with the diacylglyceride sn-1,2-dioctanoyl diglyceride (diC8) induced a staurosporine-sensitive transient increase, followed by a late decrease, in tight-junctional resistance (R(TJ)). CaSki cells express two immunoreactive forms of occludin, 65 and 50 kDa. Treatments with ATP and diC8 decreased the density of the 65-kDa form and increased the density of the 50-kDa form. ATP also decreased threonine phosphorylation of the 65-kDa form and increased threonine phosphorylation of the 50-kDa form and tyrosine phosphorylation of the 65- and 50-kDa forms. Staurosporine decreased acutely threonine and tyrosine phosphorylation of the two isoforms and in cells pretreated with staurosporine ATP increased acutely the density of the 65-kDa form and threonine phosphorylation of the 65-kDa form. Treatment with N-acetyl-leucinyl-leucinyl-norleucinal increased the densities of the 65- and 50-kDa forms. Pretreatment with N-acetyl-leucinyl-leucinyl-norleucinal attenuated the late decreases in R(TJ) induced by ATP and diC8 and the decrease in the 65-kDa and increase in the 50-kDa forms induced by ATP. Correlation analyses showed that high levels of R(TJ) correlated with the 65-kDa form, whereas low levels of R(TJ) correlated negatively with the 65-kDa form and positively with the 50-kDa form. The results suggest that in CaSki cells 1) occludin determines gating of the tight junctions, 2) changes in occludin phosphorylation status and composition regulate the R(TJ), 3) protein kinase-C-mediated, threonine dephosphorylation of the 65-kDa occludin form increases the resistance of assembled tight junctions, 4) the early stage of tight junction disassembly involves calpain-mediated breakdown of occludin 65-kDa form to the 50-kDa form, and 5) increased levels of the 50-kDa form interfere with occludin gating of the tight junctions.     

Cellular immunolocalization of occludin during embryonic and postnatal development of the mouse testis and epididymis.

Cellular junctions in the testis and epididymis play crucial roles for the development and maturation of spermatozoa. In the testis, tight junctions between Sertoli cells form a functional blood testis barrier between 10 and 16 days of age, whereas the tight junctional blood epididymal barrier between adjacent epithelial cells is formed between days 18 and 21. In the present study, occludin, a constituent integral membrane protein of tight junctions, was localized by immunofluorescent confocal microscopy in embryonic (days 13.5-18.5), postnatal (days 5-23) and adult (day 70) mouse testes and epididymides to correlate its expression with the onset of tight junctions and eventual formation of these barriers. At embryonic days 13.5 and 16.5, low diffuse cytoplasmic levels of occludin were observed in cells of the testicular cords. By embryonic day 18.5, the level of occludin was still low but appeared as a filiform-like network streaming toward the center of the cord. At postnatal days 5 and 7 immunostaining became more intense and appeared to outline the periphery of Sertoli cells of seminiferous tubules. Postnatal day 14 marked the appearance of an intense, focal band-like localization of occludin at the base of the tubules, correlating with the appearance of a functional blood-testis barrier. By day 23 and in adults, expression of occludin was noted at the base of the tubule appearing as intense, wavy, discontinuous bands similar in appearance irrespective of the stage of the seminiferous epithelium cycle. In the developing epididymis, intense cytoplasmic immunostaining was present in epithelial cells of many epididymal tubules at embryonic day 13.5. By embryonic day 16.5, intense occludin immunostaining appeared along the lateral plasma membranes of epithelial cells, whereas at embryonic day 18.5, immunostaining was punctate and apically located, suggesting the presence of tight junctions by this age; similar immunostaining was noted at postnatal days 5 and 7. In the adult epididymis, distinct punctate apical staining was observed between adjacent principal cells of all epididymal regions except the proximal initial segment, where occludin was found only in association with narrow cells. These results indicate that in the epididymis, the appearance of occludin at apical sites between adjacent epithelial cells occurs during embryonic development suggesting that tight junctions form earlier than in the testis. While occludin was expressed in a similar pattern between Sertoli cells at all stages of the cycle in the adult testis, its expression in the adult epididymis was cell- and region-specific. Taken together these data suggest that different factors regulate occludin expression in the testis and epididymis.     

急性脑卒中患者补充肠外营养的短期预后分析

目的探讨急性脑卒中患者出现胃肠功能紊乱时,补充肠外营养(PN)对患者短期预后的影响。方法将吞咽困难而出现胃肠功能紊乱的48例急性脑卒中患者随机分为2组:肠内营养(EN)组21例.营养支持停留在胃肠可以耐受的程度,平均热量为(58.5±15.0)kJ/(kg·d),蛋白质(40.4±11.8)g/d;EN+PN组27例,平均热量为(101.9±22.2)kJ/(kg·d),蛋白质(62.8±9.7)g/d。观察2组营养支持前和营养支持2周后血红蛋白、生化全项、美国国立卫生研究院脑卒中量表(NIHSS)评分及并发症。结果与营养支持前比较,营养支持2周后,EN组患者白蛋白、前白蛋白、血红蛋白和总蛋白明显下降,有显著差异(P0.01);EN+PN组患者总蛋白、白蛋白、前白蛋白虽有下降,但无显著差异(P0.05)。与EN组比较,营养支持2周后,EN+PN组患者血红蛋白、红细胞压积、白蛋白、前白蛋白明显升高、NIHSS评分明显下降(P0.05);EN组泌尿系统感染发生率明显高于EN+PN组(P0.05)。结论对于吞咽困难的脑卒中患者,在给予EN制剂时,如果遇到患者胃肠功能紊乱,应及时PN补充,可以有效地防止低蛋白血症的发生,对神经功能恢复有良好的促进作用。     

益生元对急性胰腺炎大鼠肠屏障功能的影响

目的 研究肠道补充益生元对急性胰腺炎肠上皮细胞紧密连接和屏障功能的影响.方法 Wistar大鼠腹腔注射左精氨酸建立AP模型,按随机数字法分组法分成对照组、AP组、益生元组.益生元组于造模前7 d开始给予益生元4 g·kg-1·d-1.建模后12 h处死大鼠,心脏取血检测血浆二胺氧化酶(DAO)活性,观察空肠病理变化、采用免疫组化方法检测紧密结合蛋白Occludin的含量.结果 AP组的血浆DAO活性为(7.29±0.68)U/L,显著高于对照组的(2.01±0.34)U/L(P＜0.01)和益生元组的(3.44±0.59)U/L(P＜0.05).AP组肠上皮occludin蛋白表达的灰度值为95.1±9.2,明显高于对照组的44.7±8.2和益生元组的59.7±7.8(P＜0.01).益生元组occludin表达也显著高于对照组(P＜0.05).结论 AP大鼠补充益生菌可增加肠上皮occludin蛋白表达,维持肠上皮紧密连接,维持正常的肠道通透性,从而达到保护肠黏膜屏障的效果.