Transmural intestinal wall permeability in severe ischemia after enteral protease inhibition.

In intestinal ischemia, inflammatory mediators in the small intestine's lumen such as food byproducts, bacteria, and digestive enzymes leak into the peritoneal space, lymph, and circulation, but the mechanisms by which the intestinal wall permeability initially increases are not well defined. We hypothesize that wall protease activity (independent of luminal proteases) and apoptosis contribute to the increased transmural permeability of the intestine's wall in an acutely ischemic small intestine. To model intestinal ischemia, the proximal jejunum to the distal ileum in the rat was excised, the lumen was rapidly flushed with saline to remove luminal contents, sectioned into equal length segments, and filled with a tracer (fluorescein) in saline, glucose, or protease inhibitors. The transmural fluorescein transport was determined over 2 hours. Villi structure and epithelial junctional proteins were analyzed. After ischemia, there was increased transmural permeability, loss of villi structure, and destruction of epithelial proteins. Supplementation with luminal glucose preserved the epithelium and significantly attenuated permeability and villi damage. Matrix metalloproteinase (MMP) inhibitors (doxycycline, GM 6001), and serine protease inhibitor (tranexamic acid) in the lumen, significantly reduced the fluorescein transport compared to saline for 90 min of ischemia. Based on these results, we tested in an in-vivo model of hemorrhagic shock (90 min 30 mmHg, 3 hours observation) for intestinal lesion formation. Single enteral interventions (saline, glucose, tranexamic acid) did not prevent intestinal lesions, while the combination of enteral glucose and tranexamic acid prevented lesion formation after hemorrhagic shock. The results suggest that apoptotic and protease mediated breakdown cause increased permeability and damage to the intestinal wall. Metabolic support in the lumen of an ischemic intestine with glucose reduces the transport from the lumen across the wall and enteral proteolytic inhibition attenuates tissue breakdown. These combined interventions ameliorate lesion formation in the small intestine after hemorrhagic shock

Automated segmentation and quantification of airway mucus with endobronchial optical coherence tomography

We propose a novel suite of algorithms for automatically segmenting the airway lumen and mucus in endobronchial optical coherence tomography (OCT) data sets, as well as a novel approach for quantifying the contents of the mucus. Mucus and lumen were segmented using a robust, multi-stage algorithm that requires only minimal input regarding sheath geometry. The algorithm performance was highly accurate in a wide range of airway and noise conditions. Mucus was classified using mean backscattering intensity and grey level co-occurrence matrix (GLCM) statistics. We evaluated our techniques in vivo in asthmatic and non-asthmatic volunteers

Removal of luminal content protects the small intestine during hemorrhagic shock but is not sufficient to prevent lung injury.

The small intestine plays a key role in the pathogenesis of multiple organ failure following circulatory shock. Current results show that reduced perfusion of the small intestine compromises the mucosal epithelial barrier, and the intestinal contents (including pancreatic digestive enzymes and partially digested food) can enter the intestinal wall and transport through the circulation or mesenteric lymph to other organs such as the lung. The extent to which the luminal contents of the small intestine mediate tissue damage in the intestine and lung is poorly understood in shock. Therefore, rats were assigned to three groups: No-hemorrhagic shock (HS) control and HS with or without a flushed intestine. HS was induced by reducing the mean arterial pressure (30 mmHg; 90 min) followed by return of shed blood and observation (3 h). The small intestine and lung were analyzed for hemorrhage, neutrophil accumulation, and cellular membrane protein degradation. After HS, animals with luminal contents had increased neutrophil accumulation, bleeding, and destruction of E-cadherin in the intestine. Serine protease activity was elevated in mesenteric lymph fluid collected from a separate group of animals subjected to intestinal ischemia/reperfusion. Serine protease activity was elevated in the plasma after HS but was detected in lungs only in animals with nonflushed lumens. Despite removal of the luminal contents, lung injury occurred in both groups as determined by elevated neutrophil accumulation, permeability, and lung protein destruction. In conclusion, luminal contents significantly increase intestinal damage during experimental HS, suggesting transport of luminal contents across the intestinal wall should be minimized

Proteolytic processing and activation of Clostridium perfringens epsilon toxin by caprine small intestinal contents.

Epsilon toxin (ETX), a pore-forming toxin produced by type B and D strains of Clostridium perfringens, mediates severe enterotoxemia in livestock and possibly plays a role in human disease. During enterotoxemia, the nearly inactive ETX prototoxin is produced in the intestines but then must be activated by proteolytic processing. The current study sought to examine ETX prototoxin processing and activation ex vivo using the intestinal contents of a goat, a natural host species for ETX-mediated disease. First, this study showed that the prototoxin has a KEIS N-terminal sequence with a molecular mass of 33,054 Da. When the activation of ETX prototoxin ex vivo by goat small intestinal contents was assessed by SDS-PAGE, the prototoxin was processed in a stepwise fashion into an ~27-kDa band or higher-molecular-mass material that could be toxin oligomers. Purified ETX corresponding to the ~27-kDa band was cytotoxic. When it was biochemically characterized by mass spectrometry, the copresence of three ETX species, each with different C-terminal residues, was identified in the purified ~27-kDa ETX preparation. Cytotoxicity of each of the three ETX species was then demonstrated using recombinant DNA approaches. Serine protease inhibitors blocked the initial proteotoxin processing, while carboxypeptidase inhibitors blocked further processing events. Taken together, this study provides important new insights indicating that, in the intestinal lumen, serine protease (including trypsin and possibly chymotrypsin) initiates the processing of the prototoxin but other proteases, including carboxypeptidases, then process the prototoxin into multiple active and stable species. Importance: Processing and activation by intestinal proteases is a prerequisite for ETX-induced toxicity. Previous studies had characterized the activation of ETX using only arbitrarily chosen amounts of purified trypsin and/or chymotrypsin. Therefore, the current study examined ETX activation ex vivo by natural host intestinal contents. These analyses demonstrated that (i) ETX processing in host intestinal contents occurs in an ordered, stepwise fashion, (ii) processing of prototoxin by host intestinal contents results in higher-molecular-mass material and 3 distinct ~27-kDa ETX species, and (iii) serine proteases, such as trypsin, chymotrypsin, and other proteases, including carboxypeptidases, play a role in the activation of ETX by intestinal contents. These studies provide new insights into the activation and processing of ETX and demonstrate that this process is more complicated than previously appreciated

Kraft cooking of birch wood chips: Differences between the dissolved organic material in pore and bulk liquor

The delignification of birch chips during kraft pulping was investigated, targeting both the impregnation and cooking steps. Wood chips were impregnated using white liquor, white liquor + NaCl, water or NaCl aqueous solution. Then, the chips were cooked in batch autoclaves applying the same constant composition cooking conditions for all samples. Pulp and two fractions of black liquor (bulk liquor and centrifuged liquor representing the liquor inside the wood chips and fibers) were collected after different pulping times and analyzed for lignin and carbohydrate content. The dissolved wood components were precipitated from selected samples and characterized with respect to composition, molecular weight distribution and structural motifs. Cooking chemicals in the impregnation liquors led to faster delignification and xylan removal during cooking. Higher contents of lignin and xylan were measured in the lumen than in the bulk. The concentration profiles also showed accumulation of dissolved material in the lumen over time, suggesting significant mass transport limitation from lumen to bulk. Further analysis revealed higher fragmentation/degradation of dissolved material with increasing pulping time and in the bulk when compared to the lumen liquor, as demonstrated by the lower molecular weights and the changes in chemical shifts in the NMR spectra

Secretion from Human Apocrine Glands: An Electron Microscopic Study

Electron microscopic examination of apocrine glands revealed three types of secretion: merocrine, apocrine, and possibly holocrine. In the merocrine type of secretion numerous vesicles originating in the Golgi area discharged their granular contents into the lumen of the gland. In the apocrine type of secretion three stages were observed: (1) formation of an apical cap; (2) formation of a dividing membrane at the base of the apical cap; and (3) formation of tubules above the dividing membrane that extended parallel to the membrane and led to a separation of the apical cap from the underlying cell. In the holocrine type of secretion individual secretory cells or even strands of secretory cells were discharged into the lumen of the gland

Gut inflammation provides a respiratory electron acceptor for Salmonella.

Salmonella enterica serotype Typhimurium (S. Typhimurium) causes acute gut inflammation by using its virulence factors to invade the intestinal epithelium and survive in mucosal macrophages. The inflammatory response enhances the transmission success of S. Typhimurium by promoting its outgrowth in the gut lumen through unknown mechanisms. Here we show that reactive oxygen species generated during inflammation react with endogenous, luminal sulphur compounds (thiosulphate) to form a new respiratory electron acceptor, tetrathionate. The genes conferring the ability to use tetrathionate as an electron acceptor produce a growth advantage for S. Typhimurium over the competing microbiota in the lumen of the inflamed gut. We conclude that S. Typhimurium virulence factors induce host-driven production of a new electron acceptor that allows the pathogen to use respiration to compete with fermenting gut microbes. Thus the ability to trigger intestinal inflammation is crucial for the biology of this diarrhoeal pathogen

Catechesis in Europe during the 20th century

During the 20th century, the teaching of the Catechism, and all the other forms of religious education were undergoing very serious changes and transformations. As a result of these transformations, we get the gradual introduction of the Bible as one of the main sources of the pastoral activity of the Church, especially catechesis. The Bible emerged from a period of hibernation into which it had entered for several centuries. In this work, the catechetical endeavours of Germany, of France and of Malta are highlighted. In this respect, we find the Munich method and to a much larger extent the Kerygmatic method, in Germany. In France, the use of the Bible is more linked to individual persons who, each in his or her own way tried to give a more central place to the Bible, thus opening the way to a more biblical catechesis. In Malta, we get Fr Ġorġ PRECA, who through his writings of a catechetical and educational nature gave the Word of God to the Maltese people in the vernacular.peer-reviewe

Studies on the sperm reservoir of the pig oviduct

During sperm transport in the female pig a proportion of spermatozoa are arrested, often for 24 h or more, in a particular segment of the utero-tubal junction (UTJ) and the adjacent tubal isthmus, where a sperm reservoir (SR) is built up. The SR maintains, via its lining epithelium and the fluid it produces, sperm viability pre-ovulation. It also controls the release of potentially fertilising spermatozoa so that only a small sub-population reaches the site of fertilisation, thus diminishing the risk of polyspermy. In vitro research has focused on sperm binding as the main mechanism of sperm storage, sperm release and modulation of capacitation, but little attention has hitherto been paid to the isthmic oviductal fluid (IOF), its composition and the control of capacitation in vivo. This thesis aimed to study the intra-luminal milieu of the SR in sexually cycling gilts and sows, its contents of glycosaminoglycans (GAGs) — particularly of the non-sulphated hyaluronan (HA) — and the presence and expression of the specific HA receptor CD44 and of HA synthases. Both non-inseminated (controls) and inseminated animals were studied during specific moments of oestrus, in relation to spontaneous ovulation. Ultimately, the study aimed to determine the capacitation status of SR-stored spermatozoa and the effect of IOF and HA on capacitation of the SR-stored spermatozoa. The results showed that SR-stored spermatozoa are entrapped in a mucus-like IOF pre-ovulation. This IOF contains fluctuating levels of sulphated GAGs and HA. Hyaluronan is synthesised in the lining epithelium by HA synthase 3 (has3). Both the ligand and the specific HA-receptor CD44 were particularly present in the deep furrows of the SR, where most spermatozoa are trapped. Massive sperm capacitation does not occur in vivo in the porcine SR under spontaneous standing oestrus, particularly during pre- and peri-ovulation, but SR spermatozoa capacitate if exposed to the effector bicarbonate. Exposure of SR spermatozoa to IOF (or its component HA) in vitro was seen to reverse the bicarbonate influence during pre- and peri-ovulation but to potentiate capacitation post-ovulation, suggesting an active role for the intra-tubal fluid and/or HA in modulating sperm capacitation in pigs. The findings support the concept that the oviductal SR keeps the potentially fertile spermatozoa viable and uncapacitated during their pre-ovulatory arrest. The findings may help improve sperm preparation protocols for porcine in vitro fertilisation (IVF) and preservation of boar semen