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PubMed2003

Optimization of in-gel protein digestion system in combination with thin-gel separation and negative staining in 96-well plate format.

Improvement of in-gel digestion efficiency is highly desirable for one- or two-dimensional gel electrophoretic separation and mass spectrometric (MS) analysis in proteomics, because the resultant increases in sequence coverage and MS signal intensity lead to higher confidence in protein identification. Here an optimized in-gel digestion system, in combination with thin-gel separation and negative staining in a high-throughput format using 96-well plates, is described. The combination of negative staining and protein separation on a 0.9 mm thick gel showed a clear improvement in in-gel digestion efficiency in comparison with the more typical protocols such as the combination of silver staining and a 1.0 mm gel. In addition, the use of 96-well plates to increase throughput did not decrease the efficiency of this strategy when the stirring of the gel pieces in processes such as destaining, washing, gel-shrinking and peptide extraction was performed by sonication instead of shaking the plates. This procedure was optimized and applied to identify proteins of the postsynaptic density fraction; 105 proteins were identified after SDS-PAGE separation.

Animals↗
PubMed2001

Phenotypic flexibility in digestive system structure and function in migratory birds and its ecological significance.

Birds during migration must satisfy the high energy and nutrient demands associated with repeated, intensive flight while often experiencing unpredictable variation in food supply and food quality. Solutions to such different challenges may often be physiologically incompatible. For example, increased food intake and gut size are primarily responsible for satisfying the high energy and nutrient demands associated with migration in birds. However, short-term fasting or food restriction during flight may cause partial atrophy of the gut that may limit utilization of ingested food energy and nutrients. We review the evidence available on the effects of long- and short-term changes in food quality and quantity on digestive performance in migratory birds, and the importance of digestive constraints in limiting the tempo of migration in birds. Another important physiological consequence of feeding in birds is the effect of diet on body composition dynamics during migration. Recent evidence suggests that birds utilize and replenish both protein and fat reserves during migration, and diet quality influences the rate of replenishment of both these reserves. We conclude that diet and phenotypic flexibility in both body composition and the digestive system of migratory birds are important in allowing birds to successfully overcome the often-conflicting physiological challenges of migration.

Animals↗
PubMed1996

Immunocytochemical detection of islet hormones in the digestive system of Protopterus annectens.

The presence, distribution, and interrelationships of the four typical pancreatic islet hormones were investigated in the digestive system of Protopterus annectens by single and double immunohistochemical methods. Insulin-, glucagon-, and somatostatin-immunoreactive (IR) elements were detected in both the pancreas and the gut. Pancreatic polypeptide (PP)-IR endocrine cells were always present in the gut, but were only present in the pancreas of a few specimens. Some of the latter cells also seemed to react with glucagon antiserum. In the pancreas the immunopositive cells were organized into islets of different sizes, and their organizations were studied by the double immunohistochemical techniques. In the few large islets insulin-IR cells were present in the central zone, glucagon- and PP-IR cells at the periphery, and somatostatin-IR cells intermingled with both the peripheral and the central endocrine cells. In the smaller islets, the number and the staining intensity of glucagon- and PP-IR endocrine cells varied markedly. In the gut, insulin-, somatostatin-, and PP-IR cells were of the open type; glucagon-containing cells were very few and had no luminal contact. They were differently distributed along the intestinal epithelium. Somatostatin-IR nerve fibers and somatostatin-IR neuron cell bodies were also observed in the intestinal wall. The organization of pancreatic endocrine cells in P. annectens is similar to that observed in the majority of teleosts even if a different topographical association can be found. Furthermore, islets of different sizes seem to display a different metabolic turnover, and the detection of pancreatic PP-immunoreactivity varied according to the specimens utilized. In the intestinal portion insulin-IR cells, in addition to PP-, glucagon- and somatostatin-IR cells are present: this suggests that intestinal insulin-like immunoreactivity may be more widespread than previously supposed.

Animals↗
PubMedDate not supplied

[Digestive system status after prolonged space flights].

In the crewmembers of four Salyut-6 long-term flights, enzyme excretory function of the gastro-intestinal tract was investigated. These studies included: gastric proenzyme, pepsinogen, and pancreatic enzymes, amylase and lipase, in blood and urine, trypsin in blood, intestinal enzymes, invertase and glycyl-L-leucine dipeptidase in feces, and lipids in feces. The results obtained demonstrated a correlation between changes in enzyme excretion and space flight duration and profile. After the 140- and 175-day flight the most marked changes in the digestive organs were seen; they manifested as a simultaneous increase in secretory function of the stomach and the pancreas. However, after the 185-day flight, in which advanced countermeasures were used, the above changes were less distinct.

Digestive System↗
PubMed2004

Interaction of two Bacillus thuringiensis delta-endotoxins with the digestive system of Lygus hesperus.

The active-toxin form of CrylAc (65 kDa) or Cry2Ab was fed to a non-susceptible insect, Lygus hesperus, in an artificial diet. Biochemical and immunocytochemical methods were used to determine the distribution of ingested toxin. The toxins did not elicit a feeding deterrent response. CrylAc and Cry2Ab were ingested; small amounts were absorbed into the hemolymph as holoproteins, but most was excreted. SDS-PAGE analysis of CrylAc and Cry2Ab incubations with salivary gland homogenate showed a small decrease in the molecular weight of the active toxins. Proteolytic processing of the toxins also occurred in vivo, within the digestive system of L. hesperus. Excreted CrylAc and Cry2Ab retained activity toward lepidopteran larvae. Immunocytochemical in vivo localization studies showed negligible association of CrylAc with L. hesperus tissues. In contrast, strong extracellular association of Cry2Ab was observed with L. hesperus midgut brush border microvilli and basement membrane, as well as with cellular outlines within the hemolymph and fat body.

Animals↗
PubMed1993

Specializations of the human upper respiratory and upper digestive systems as seen through comparative and developmental anatomy.

The human upper respiratory, or aerodigestive, tract serves as the crossroads of our breathing, swallowing and vocalizing pathways. Accordingly, developmental or evolutionary change in any of these functions will, of necessity, affect the others. Our studies have shown that the position in the neck of the mammalian larynx is a major factor in determining function in this region. Most mammals, such as our closest relatives the nonhuman primates, exhibit a larynx positioned high in the neck. This permits an intranarial larynx to be present and creates largely separate respiratory and digestive routes. While infant humans retain this basic mammalian pattern, developmental descent of the larynx considerably alters this configuration. Adult humans have, accordingly, lost separation of the respiratory and digestive routes, but have gained an increased supralaryngeal region of the pharynx which allows for the production of the varied sounds of human speech. How this region has changed during human evolution has been difficult to assess due to the absence of preserved soft-tissue structures. Our studies have shown that the relationship between basicranial shape and laryngeal position in living mammals can be a valuable guide to reconstruct the region in ancestral humans. Based on these findings we have examined the basicrania of fossil ancestors--from over two million years ago to near recent times--and have reconstructed the position of the larynx and pharyngeal region in these early forms. This has allowed us insight into how our ancestors may have breathed and swallowed, and when the anatomy necessary for human speech evolved.

Anatomy, Comparative↗
PubMed2004

Immunolocalization of protein 4.1B in the rat digestive system.

Protein 4.1 family proteins are thought to interact with membrane proteins and also membrane skeletons. In this study, immunohistochemical studies by light and electron microscopy were performed with a specific antibody against protein 4.1B. Specific protein 4.1B immunolabeling was observed in simple columnar epithelium in the adult rat large intestine, small intestine and stomach. Protein 4.1B immunolabeling was localized along the membranes facing the adjacent cells (lateral portion) and also facing the extracellular matrix (basal portion). Moreover, a spatial protein 4.1B expression gradient was observed along the crypt-villus axis of the rat small and large intestinal epithelium: strong protein 4.1B expression was present within the villus, with the crypt showing barely any detectable protein 4.1B. The expression of protein 4.1B was not detected in the stratified squamous epithelium in the forestomach or the esophagus. By immunoelectron microscopy, the immunolabeling of the cells was observed to be restricted to the cytoplasmic side just beneath the plasma membrane, including the membranes adjacent to the next cells, except for the tight junctions. We conclude that the protein 4.1B expression pattern is related to the maturation of simple columnar epithelium in the rat digestive system, probably by the effect of adhesion.

Animals↗
PubMed1980

Penetrability of proteins through the digestive system of Sarcophaga falculata blowfly.

On the background of the previous findings concerning the passage of a low molecular basic protein from cobra snake venom through the gut of a blowfly (Primor, N., Teitelbaum, Z. and Zlotkin, E. (1980) Biochim, Biophys, Acta 627, 71--81) several additional proteins were tested in the present study. With the aid of toxicity tests, radioiodinated proteins, gel filtration chromatography, and the usage of Sarcophaga falculata flies as test animals, the following information was obtained: 1. Out of the five species (representing four orders) of insects tested, only flies demonstrated the lethality by oral application of cobra snake venom. 2. The absence of oral toxicity of flies to phospholipase A2 (highly lethan by injection of cobra venom with molecular weight of about 13,000) was due to its impermeability through the digestive system. 3. About 2.2% of the orally applied low molecular weight basic neurotoxin (Mr 7000) from cobra venom crossed the gut and was found in the insects' hemolymph. 4. Bovine serum albumin (Mr 68,000) and an immunoglobulin (Mr 180,000), under the present experimental conditions, were found to be gut impermeable.

Animals↗
PubMed1999

Immunohistochemical detection of alpha1E voltage-gated Ca(2+) channel isoforms in cerebellum, INS-1 cells, and neuroendocrine cells of the digestive system.

Polyclonal antibodies were raised against a common and a specific epitope present only in longer alpha1E isoforms of voltage-gated Ca(2+) channels, yielding an "anti-E-com" and an "anti-E-spec" serum, respectively. The specificity of both sera was established by immunocytochemistry and immunoblotting using stably transfected HEK-293 cells or membrane proteins derived from them. Cells from the insulinoma cell line INS-1, tissue sections from cerebellum, and representative regions of gastrointestinal tract were stained immunocytochemically. INS-1 cells expressed an alpha1E splice variant with a longer carboxy terminus, the so-called alpha1Ee isoform. Similarily, in rat cerebellum, which was used as a reference system, the anti-E-spec serum stained somata and dendrites of Purkinje cells. Only faint staining was seen throughout the cerebellar granule cell layer. After prolonged incubation times, neurons of the molecular layer were stained by anti-E-com, suggesting that a shorter alpha1E isoform is expressed at a lower protein density. In human gastrointestinal tract, endocrine cells of the antral mucosa (stomach), small and large intestine, and islets of Langerhans were stained by the anti-E-spec serum. In addition, staining by the anti-E-spec serum was observed in Paneth cells and in the smooth muscle cell layer of the lamina muscularis mucosae. We conclude that the longer alpha1Ee isoform is expressed in neuroendocrine cells of the digestive system and that, in pancreas, alpha1Ee expression is restricted to the neuroendocrine part, the islets of Langerhans. alpha1E therefore appears to be a common voltage-gated Ca(2+) channel linked to neuroendocrine and related systems of the body.

Animals↗
PubMed1991

Risk factors for second cancers of the upper respiratory and digestive systems: a case-control study.

A case-control study of determinants of multiple cancers of the upper respiratory and digestive system (URDS) was conducted using a patient cohort-nested design. We analyzed demographic and risk factor information and clinical variables related to the index cancer for 85 cases of multiple cancers and 170 controls matched on sub-site of the index cancer of the case and date of hospital admission. Follow-up information for the control group was used to infer the person-years-at-risk for the cohort of 1977 patients. URDS cancer patients experienced a 10.7 times (95% confidence interval: 8.5-13.2) higher risk of additional related cancers than the general population. Although controls had cancers of the same sites as those of cases and thus, strongly tobacco and alcohol-related, there were marked residual effects for these two risks factors. In addition, characteristics related to the extension and clinical course of the index cancer were strongly associated with the patient's risk of developing additional cancers.

Adult↗
PubMed1999

[Comparison of effects of Herba Pogostmonis from gaoyao and wuchuan on digestive system].

The effects of the three extracts (decoction, oil-free decoction and volatile oil) of Herba Pogostmonis from Gaoyao and Wuchuan on digestive system were compared. The results demonstrated that the decotion and oil-free decoction from Gaoyao and Wuchuan could depressed gastric evacuation in mice. All the three extracts from Gaoyao and Wuchuan could inhibit the ACh-as well as BaCl2-induced spasmodic and automatic contraction of isolated rabbit intestine. The decoction and oil-free decoction from either Gaoyao or Wuchuan inhibited the normal intestinal propulsion. They also increased gastric secretion of acid and activity of pepsin and amylase. These suggest that both of Herba pogostmonis relief gastrointestinal spasm and improve the digestion. The total effect of Herba pogostmonis from Gaoyao was more potent that from Wuchuan.

Animals↗
PubMed1992

Calcitonin gene-related peptide-containing neurons supplying the rat digestive system: differential distribution and expression pattern.

In the enteric nervous system, calcitonin gene-related peptide (CGRP) immunoreactivity is localized to a substantial number of capsaicin-sensitive afferent fibers and to intrinsic neurons and processes. CGRP immunoreactivity detected by immunohistochemistry represents the expression of two distinct genes, the calcitonin/alpha-CGRP and the beta-CGRP genes, which have different tissue distributions. In the present study, we used (1) in situ hybridization histochemistry and ribonucleic acid (RNA) blot hybridization with RNA probes complementary to the divergent sequences of alpha- and beta-CGRP messenger RNAs (mRNAs) to differentiate which CGRP gene was expressed in enteric and afferent neurons; and (2) axonal transport approaches in combination with CGRP immunohistochemistry to define the location of CGRP-containing afferent neurons supplying the digestive system. In situ hybridization histochemistry with [35S]-labeled RNA probes indicated that in the gastrointestinal tract beta-CGRP mRNA, but not alpha-CGRP mRNA, was expressed in enteric neurons confined to the myenteric and submucous plexuses of the small and large intestine. In dorsal root and vagal sensory ganglia, mRNAs for alpha-CGRP and beta-CGRP were both present in a vast population of neurons, with an overlapping pattern, even though the alpha-CGRP signal appeared more intense. RNA blot hybridization analysis showed a single band of hybridization at 1.2 Kb with the beta-CGRP RNA probe in RNA extracts from muscle layer-myenteric plexus and submucosal layer preparations of the ileum, and from dorsal root ganglia; it also showed a single band at 1.3 Kb with the alpha-CGRP RNA probe in extracts from dorsal root ganglia, but not from the intestine. These findings further support the differential expression of alpha- and beta-CGRP mRNAs. Retrograde transport of fast blue or fluorogold coupled with CGRP immunohistochemistry demonstrated that the vast majority of CGRP-containing afferent neurons supplying the stomach, proximal duodenum, and pancreas were located in dorsal root ganglia at the middle and lower thoracic and at the upper lumbar levels, and represented a major component of the afferent innervation of these viscera (up to 89%). Approximately 50% of CGRP-immunoreactive afferent neurons also expressed tachykinin (TK) immunoreactivity, as shown by triple labeling. Only a minor component of the afferent innervation of the stomach, duodenum, and pancreas derived from vagal CGRP-containing neurons (less than 8%). A large portion of these neurons (an average of 62%) also contained TK immunoreactivity.(ABSTRACT TRUNCATED AT 400 WORDS)

Afferent Pathways↗
PubMed2004

Performance of a UASB-digester system treating domestic wastewater.

The anaerobic treatment of raw domestic wastewater by a novel technology consisting of an Up-flow Anaerobic Sludge Bed (UASB) reactor combined with a completely mixed digester for the stabilisation of the UASB sludge was assessed. A pilot-scale plant of the so-called UASB-Digester system was located at the municipal wastewater treatment facility of Santiago de Compostela (Northwest of Spain). The main aim of the Digester was to enhance the biodegradation of influent solids retained in the UASB reactor at low temperatures, then increasing its specific methanogenic activity. The sludge drawn from the middle zone of the UASB entered the upper zone of the Digester and then circulated from the bottom of the Digester to the UASB bottom. Circulating in an automated semi-continuous way, the flow of this sludge stream was selected in order to set a previously defined hydraulic retention time (HRT) (16-27 d) in the digester. The Digester temperature was set at an optimum value ranging from 25 to 35 degrees C. The steady state efficiency of the UASB system, at 6-8 h of HRT, 15-16 degrees C of temperature and 330-360 mg l(-1) of influent total chemical oxygen demand (TCOD) was 79% of total suspended solids (TSS) removal, 52% of TCOD removal and 60% of biological oxygen demand (BOD5) removal. The hydrolysis of retained solids reached 85%, while excess sludge generation was only 7% of influent TCOD. A stable anaerobic (pre)treatment of diluted domestic wastewater was reached as the sludge concentration in the reactor remained mainly constant and the specific methanogenic activity showed a slight increase.

Bacteria, Anaerobic↗
PubMed1985

Second cancer following cancer of the digestive system in Denmark, 1943-80.

Among 4,184 patients with cancer of the esophagus, 55 second primary cancers were observed, whereas 64 were expected [relative risk (RR) = 0.86]. The absence of an excess risk of alcohol- and tobacco-related cancers was not anticipated. A significant 19% deficit of second cancers was found among 30,843 patients with stomach cancer. Cancer of the rectum, kidney, and lung all occurred significantly below expectation. An excess risk of ovarian cancer (RR = 1.9) was seen in women. Reasons for these findings are not entirely clear. Cancer of the small intestine is rare, and despite a relatively short survival expectation, a moderate excess of second cancers was seen among 868 patients (36 vs. 26.8). Only cancers of the liver and gallbladder were significantly elevated, and the possibility of misclassified metastases is discussed. Colon cancer is one of the most common cancers in Denmark, and 29,490 patients with this disease were at slightly lower risk for development of second cancer (RR = 0.96; 95% confidence interval = 0.9-1.0) than the general Danish population, excluding secondary colon cancers. Esophageal, stomach, and liver cancers occurred less frequently than expected. That cancers of the uterine corpus and ovary were significantly increased supports the notion that common risk factors, such as diet and endogenous hormones, influence the development of these cancers. A significant 23% deficit of second cancers was also found among 26,597 patients with cancer of the rectum, excluding secondary rectal cancer. Significant deficits were seen for cancers of the stomach (RR = 0.5), lung (RR = 0.8), and brain (RR = 0.5), and for multiple myeloma (RR = 0.4). The likelihood of underreporting of second cancers, especially of the digestive system, is discussed. However, cancer of sites previously reported to be associated with rectal cancer, e.g., the colon, breast, and uterus, did not occur below expectation. Cancers of the liver and biliary tract occurred in 4,453 patients; their average survival was only 1 year. Except for a slight excess of cancer of the ovary (5 vs. 1.6), the risk of second cancer development for all sites was consistent with unity (RR = 0.90). The risk of second cancers among 7,752 persons with cancer of the pancreas was not greater than expected (88 vs. 85.2). Males were at significant risk of kidney cancer (RR = 3.2), whereas females showed elevated rates of cancers of the uterine corpus (RR = 3.2) and ovary (RR = 3.1). No site occurred significantly below expectation.

Colonic Neoplasms↗