Influence of nicotine and cotinine on retinal phospholipase A2 and its significance to macular function

The macula is a constituent of the sensory retina that is necessary for sharp contrast and color vision. A significant relationship has been found between tobacco smoking and age-related macular degeneration. Opsin, rhodopsin and phospholipase A2 (PLA2) are located in excitable membranes of retina which are enriched with polyunsaturated fatty acids (PUFA). A question may arise as to whether nicotine and its major metabolite cotinine influence PLA2 so that arachidonic acid (AA) and proinflammatory prostaglandins (PG) are produced in the retina. Therefore, the effects of nicotine and cotinine on the retinal PLA2 were studied. PLA2 activity of rat retinal sonicates was assayed using 1-palmitoyl-2[1-14C]arachidonyl-Phosphatidylethanolamine (PE, 2.2 nmol) as a substrate in Tris buffer (10 mM, pH 7.4) at 37 degrees C with and without nicotine or cotinine in the assay medium. These studies gave the following results: (1) Rat retinal PLA2 activity was 4.2+/-0.8 pmol PE hydrolyzed/100 ng protein/hr. (2) Nicotine in low concentrations (1-150 nM) activated PLA2 (EC50 5 nM). (3) Cotinine also activated PLA2 (EC50 300 nM). (4) Only high concentrations of nicotine (> 1.0 microM) and cotinine (> 25 microM) exhibit inhibition of PLA2. (5) All three known PLA2 inhibitors, mepacrine, 4-bromophenacyl bromide and bromoacetylcholine bromide (IC50: 0.5mM, 88 microM, 30 mM, respectively) inhibited retinal PLA2 activity. These observations suggest that polyunsaturated fatty acids are cleaved, and arachidonic acid, the precursor for prostaglandins and related pro-inflammatory mediators, is liberated by nicotine and cotinine. Oxidative stress (reduced levels of antioxidants), vascular insufficiency, as well as activation of PLA2 by nicotine and cotinine may contribute for retinal degeneration in smokers during aging.     

Acute effects of high-dose intragastric nicotine on mucosal defense mechanisms: an analysis of nicotine, prostaglandin E2, phospholipase A2, and phospholipids

Peptic ulcer disease is overrepresented among smokers; they also heal slowly and relapse frequently. Data are accumulating that smoking is detrimental to gastroduodenal mucosal cytoprotection. This study was designed to assess acute effects of high-dose intragastric nicotine, as it has been shown that nicotine is accumulated in gastric juice when smoking. Seven healthy smokers were given nicotine base (6 mg) as tablets, which yielded very high intragastric concentrations and plasma levels comparable to those seen when smoking. In addition to nicotine analysis, concentration levels of prostaglandin E2 (PGE2), phospholipase A2 (PLA2), and phospholipid classes were measured before and after nicotine administration. Nicotine inhibited PGE2 levels by 27-81%, whereas PLA2 and total phospholipids were unaffected. Lysolecithin, a degradation product of the main constituent of gastric surfactant, ie, phosphatidylcholine, tended to increase, but this was not reflected in intragastric phosphatidylcholine levels. In conclusion, nicotine acutely inhibits PGE2 and may thus impair mucosal cytoprotection. The present findings do not imply a central role of surface-active phospholipids with respect to nicotine and gastric cytoprotection, but the chronic effects of nicotine remain to be investigated.     

Motilin in human milk: identification and stability during digestion

The presence of motilin in human milk and the influence of human milk on the degradation of [125I][Nle13] porcine motilin by gastric and duodenal fluids were investigated. Milk and plasma samples were collected from 14 mothers, and motilin was measured by radioimmunoassay. Plasma levels were 416 +/- 37 pg/mL. In 8 defatted samples the motilin level was 105 +/- 14 pg/mL, in the six others levels were above 1000 pg/mL but dilution curves were non-linear. After solid-phase extraction milk levels were 108 +/- 21 pg/mL in 13 samples, in 1 sample the dilution curve was still non-linear. The stability of motilin after ingestion was studied in vitro by incubating [121I][Nle13] porcine motilin with gastric and intestinal juices obtained from newborns (10 times diluted). Incubations were performed at 37 degrees C at pH 1.8, 3.2 and 5.8 for the gastric fluid and at pH 7.4 for the duodenal fluid. After different times of intervals (5, 10, 20 and 30 minutes) intact motilin was precipitated with trichloroacetic acid and the radioactivity of the supernatant was determined. Motilin was rapidly degraded by gastric juice. The breakdown was greatest at pH 3.2 (74% after 30 minutes) and lowest at pH 5.8 (29%), the pH after milk feeding in neonates. Degradation by intestinal juice at pH 7.4 was also very rapid (77% after 30 minutes). Human milk and BSA inhibited partially the gastric digestion at pH 3.2 (17 and 29%, respectively). Digestion by intestinal juice was not affected by human milk and BSA. These results suggest that digestion of motilin in the stomach may be sufficiently retarded by human milk in the newborn to exert a biological role.     

Isolation and characterization of a cytosolic phospholipase A2 from bovine adrenal medulla

We have recently demonstrated that bovine adrenal medulla contains a soluble phospholipase A2 (PLA2), which is localized in the cytosol. In the present study, this PLA2 was purified 1,097-fold using sequential concanavalin A, hydrophobic interaction, anion exchange, gel filtration, and an additional anion exchange chromatography. The enzyme is activated over the range of 20-1,000 microM Ca2+ and has a pH optimum near 8.0. On sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the protein has a molecular mass of 26 kDa and an isoelectric point of 4.6 as revealed by isoelectric focusing. The cytosolic PLA2 is not inhibited by NaCl, and the enzymatic activity is stimulated at low concentrations of Triton X-100 (0.01%) and deoxycholate (1 mM) but inhibited at higher concentrations (0.1% and 3 mM, respectively) of these detergents. Furthermore, heat treatment (57 degrees C, 5 min) reduced the enzymatic activity by 80%, whereas glycerol (30%) increased the activity. p-Bromophenacylbromide, a frequently used irreversible inhibitor of type II PLA2, has little effect until 100 microM, and 2-10 mM dithiothreitol totally inactivated the enzyme. The purified PLA2 displays a preference for phosphatidylcholine as a substrate but hydrolyzes phospholipid substrates with arachidonic acid or linoleic acid esterified at the sn-2 position to the same extent. It is concluded that the chromaffin cell cytosolic PLA2, which was isolated and characterized in this study, represents a type of PLA2 that has not been formerly reported in chromaffin cells. Additional research on the chromaffin cell cytosolic PLA2 will help to reveal whether the enzyme is important for exocytosis.     

Survival of Escherichia coli O157:H7 in synthetic gastric fluid after cold and acid habituation in apple juice or trypticase soy broth acidified with hydrochloric acid or organic acids

Extreme acid tolerance of Escherichia coli O157:H7 has raised doubts about the safety of acidic foods. This study examined whether prior storage in acidic and/or cold conditions enhanced survival of E. coli O157:H7 in synthetic gastric fluid (SGF). Three E. coli O157:H7 strains were stored in trypticase soy broth (TSB; acidified with HCl, malic acid, citric acid, or lactic acid) or pH 3.5 and 6.5 (nonacidic control) apple juice at 4 and 21 degrees C for < or = 7 days and then were incubated in pH 2.5 SGF at 37 degrees C for 4 h. Cells survived better in apple juice than in TSB containing organic acids, suggesting that juice constituents other than organic acids protect E. coli O157:H7. Refrigeration combined with low pH best protected cells in apple juice and acidified TSB, but, compared to the nonacidic control, only acidified TSB enhanced subsequent survival in pH 2.5 SGF. Equal survival in SGF occurred after storage in pH 3.5 or 6.5 apple juice at 4 degrees C, suggesting that low temperature alone in apple juice enhanced acid tolerance. Two strains stored at 4 degrees C in TSB containing malic or citric acid subsequently survived better in SGF than cells stored in nonacidified TSB but poorer than cells stored in the presence of HCl. These differences reflect the higher pKa of these organic acids. However, subsequent survival of these strains in SGF was poorer after refrigerated storage in apple juice than in TSB containing citric or malic acids. Cells stored in lactic acid were most likely to be completely eliminated upon transfer to SGF. Differences in survival in storage media or SGF related to strain, storage conditions, or acidifier were consistent and often statistically significant (P < 0.05). Although the survival of E. coli O157:H7 in refrigerated acidic beverages may not be affected by the type of acidifier used, the subsequent survival in SGF of this pathogen may be critically dependent on this factor.     

Once-daily gentamicin therapy for experimental Escherichia coli meningitis

BACKGROUND: Eradication of Helicobacter pylori by antibiotics in combination with gastric acid inhibition can result in overgrowth of non-H. pylori bacterial flora. This may confound the histological detection of H. pylori at eradication control if non-specific staining methods are used. OBJECTIVE AND METHODS: In 18 patients treated with amoxycillin (2 weeks) and omeprazole (6 weeks), endoscopically obtained gastric juice was cultured and two biopsies of corpus, antrum and duodenum were taken before and after eradication therapy (with gastric acid inhibition still going on) for culture and for histology to assess the intragastric bacterial flora. By histology, modified Giemsa (MG) and an H. pylori-specific immunohistochemical stain (IMM) were evaluated. RESULTS: Median pH of gastric juice was 1.5 (n = 18) before and 7 (n = 17) after eradication therapy, when patients were still on omeprazole. After therapy, culture showed a significant decrease (P < 0.05) in mean amount of H. pylori in corpus, antral and duodenal biopsies and a significant increase of non-H. pylori flora (P < 0.05) in gastric juice, corpus, antral and duodenal mucosa. With culture as a standard, 16 and 4 biopsy specimens were scored falsely positive for H. pylori by MG and IMM, respectively, and H. pylori was not detected in 23 and 13 biopsy specimens when culture was H. pylori-positive. CONCLUSION: Because of the possible presence of non-H. pylori flora after eradication therapy, the use of IMM is recommended in this situation for the histological detection of H. pylori, especially in those patients with ongoing gastric acid inhibitory therapy.     

Phospholipid interactions affect substrate hydrolysis by bovine brain phospholipase A1

The specificity of substrate hydrolysis by bovine brain phospholipase A1 (PLA1) was examined. In the presence of Mg2+, using pH values of 7 to 9, the purified enzyme deacylated 1-palmitoyl-2-[1-14C]arachidonoyl-phosphatidylethanolamine yielding 2-[1-14C]arachidonoyl-lysophosphatidylethanolamine at a rate of 70 mumol/min per mg. In the absence of Mg2+, however, the reaction rate slowed at pH values above 7.25. In contrast, brain PLA1 slowly (3.8 mumol/min per mg) hydrolyzed 1-palmitoyl-2-[1-14C]arachidonoyl-phosphatidylcholine (PAPC) unless phosphatidylserine (PS) was included. Maximal PAPC hydrolyzing activity required a PAPC/PS molar ratio of 2.5:1, Mg2+, and a pH value of 8.5-9.5. Replacing PS with phosphatidylethanolamine (PE) or phosphatidic acid (PA), but not phosphatidylinositol (PI), produced a similar effect. Moreover, hydrolysis of either arachidonoyl-substituted or dipalmitoyl-substituted PC at pH 7.5 was enhanced by increasing the mol fraction of PE. Brain PLA1 also hydrolyzed 1-stearoyl-[1-14C]arachidonoyl-PI with high velocity, but only if the substrate was dispersed in PE vesicles. In contrast, the velocity of PS, 1-palmitoyl-lyso-PC or diacylglycerol hydrolysis was low and unaffected by PE. In summary, PLA1 hydrolyzed PE with high velocity and specificity, whereas a high rate of PC or PI hydrolysis was observed only if PS, PE, or PA was present. In addition, PLA1 activity was greatly influenced by pH and Mg2+, implying that the substrate conformation is important to the catalytic efficiency of PLA1. Finally, the high rate of PE, PC or PI hydrolysis suggests PLA1 significantly contributes to the turnover of these phospholipids in the brain.     

Duodenogastric reflux potentiates the injurious effects of gastroesophageal reflux

Experimental studies have shown that the severity of esophageal mucosal injury in gastroesophageal reflux disease is related to the reflux of both gastric and duodenal juice. The purpose of this study was to determine whether duodenal juice potentiates esophageal injury in patients with reflux disease or, in fact, causes no harm allowing acid and pepsin to do the damage. A total of 148 consecutive patients who had no previous gastric or esophageal surgery underwent endoscopy and biopsy, manometry, and 24-hour esophageal pH and bilirubin monitoring. Esophageal injury was defined by the presence of erosive esophagitis, stricture, or biopsy-proved Barrett's esophagus. Exposure to duodenal juice, identified by the absorbance of bilirubin, was defined as an exposure time exceeding the ninety-fifth percentile measured in 35 volunteers. To separate the effects of gastric and duodenal juice, patients were stratified according to their acid exposure time. One hundred patients had documented acid reflux on pH monitoring, and in 63 of them it was combined with reflux of duodenal juice. Patients with combined reflux (50 of 63) were more likely to have injury than patients without combined reflux (22 of 37; P < 0.05). When the acid exposure time was greater than 10%, patients with injury (n = 40) had a greater exposure to duodenal juice (median exposure time 17.2% vs. 1.1%, P = 0.006) than patients without injury (n = 5), but there was no difference in their acid exposure (16.9% vs. 13.4%). Patients with dysplasia of Barrett's epithelium (n = 9) had a greater exposure to duodenal juice (median exposure time 30.2% vs. 7.2%, P = 0.04) compared to patients without complications (n = 25), whereas acid exposure was the same (16.4% vs. 15%). Duodenal juice adds a noxious component to the refluxed gastric juice and potentiates the injurious effects of gastric juice on the esophageal mucosa.     

Gastric acidity influences the blood response to a beta-carotene dose in humans

The effect of gastric acidity on the blood response to a single dose of 120 mg beta-carotene in humans was investigated in 12 normal subjects (5 women, 7 men) aged 23-68 y. Omeprazole was used for 7 d to obliterate gastric acid secretion and to raise gastric pH to > 4.5. In a crossover design, six subjects were randomly assigned to take beta-carotene with omeprazole either at the beginning (day 9) or at the end (day 26) of the study. The beta-carotene response in blood was not altered by the experimental order. Results from the high-gastric-pH phase (ie, with omeprazole) were analyzed together and compared with the results from the low-gastric-pH phase (ie, without omeprazole). The increases of serum concentrations of both trans beta-carotene and cis beta-carotene 6 and 24 h after the beta-carotene dose were significantly greater at a low gastric pH (pH = 1.3 +/- 0.1, ie, without omeprazole) than those at a high gastric pH (pH = 6.4 +/- 0.3, ie, with omeprazole), P < 0.02. Similarly, 24 h after beta-carotene administration, the area under the blood beta-carotene response curve (trans plus cis beta-carotene) was significantly greater at a low gastric pH (6825 +/- 760 nmol.h/L) than at a high gastric pH (3390 +/- 550 nmol.h/L), P < 0.002. In investigations of bacterial overgrowth, gelatin capsule disintegration and isomeric profiles associated with high and low pH, we could not identify factors to explain the differences observed in the blood response curves between low-gastric-pH and high-gastric-pH conditions. A suppressed blood response of beta-carotene at a high intraluminal pH may have been due to the slower movement of negatively charged micelles through the unstirred water layer and cell membrane.     

Accumulation of aliphatic amines in gastric juice of acute renal failure patients. Possible cause of hypergastrinemia associated with uremia

In this study we analyzed by gas chromatographic headspace analysis the composition and concentration of gastrin-stimulatory volatile aliphatic amines in the gastric juice of healthy subjects and acute renal failure patients. We demonstrated that although these aliphatic amines are present in the gastric juice of normal subjects in trace amounts, they accumulate in the gastric juice of uremic subjects. This 30-40-fold elevation in gastric juice amine concentration agreed favorably with the 40-50-fold augmentation in serum gastrin levels in acute renal failure, with a significant association (r = 0.87) existing between these two parameters. It was also determined that a 2-hr hemodialysis procedure resulted in a modest nonparallel decline in both gastric amine and serum gastrin levels. These results support the hypothesis that the accumulation of volatile aliphatic amines in the gastric juice of uremic individuals may induce an activation of the antral G cells, resulting in hypergastrinemia.     

Comparative Study of the Chemical Resistance of Titanium Nitride and Stainless Steel in Media of the Oral Cavity

We have shown that TiN powders practically do not react with biochemical media in the body. Using the polarization curve method, x-ray phase analysis, and Auger-electron spectroscopy of the electrode surface, we have studied the reaction of titanium nitride and 12Kh18N10T stainless steel with distilled water, a 3% solution of NaCl, HCl, and gastric juice. We have demonstrated the high corrosion resistance of both materials in H₂O (which is also typical for saliva); but in the presence of salt and when gastric juice is ejected from the esophagus into the oral cavity, the corrosion resistance of steel may decrease by several orders of magnitude compared with TiN. In the case of a titanium nitride electrode, we have also established the mechanism of the protective (inhibiting) action of organic reagents contained in the biochemical media, while stainless steel dissolves in gastric juice significantly more rapidly than in hydrochloric acid with the same pH 2.0.

     

Comparative Study of the Chemical Resistance of Titanium Nitride and Stainless Steel in Media of the Oral Cavity

We have shown that TiN powders practically do not react with biochemical media in the body. Using the polarization curve method, x-ray phase analysis, and Auger-electron spectroscopy of the electrode surface, we have studied the reaction of titanium nitride and 12Kh18N10T stainless steel with distilled water, a 3% solution of NaCl, HCl, and gastric juice. We have demonstrated the high corrosion resistance of both materials in H₂O (which is also typical for saliva); but in the presence of salt and when gastric juice is ejected from the esophagus into the oral cavity, the corrosion resistance of steel may decrease by several orders of magnitude compared with TiN. In the case of a titanium nitride electrode, we have also established the mechanism of the protective (inhibiting) action of organic reagents contained in the biochemical media, while stainless steel dissolves in gastric juice significantly more rapidly than in hydrochloric acid with the same pH 2.0.     

Simultaneous determinations of pancreatic phospholipase A2 and prophospholipase A2 in various pancreatic diseases

Pancreatic phospholipase A2 (PLA2) is secreted into the pancreatic juice by pancreatic acinar cells as a proenzyme (proPLA2), which is activated by trypsin. Radioimmunoassays with monoclonal antibodies to PLA2 and proPLA2 were used to examine the serum PLA2 and proPLA2 levels simultaneously in patients with various pancreatic diseases. In healthy subjects, proPLA2 proved to be the major form of the enzyme. The serum PLA2 level were found to be significantly increased in patients with acute pancreatitis, the active phase of chronic relapsing pancreatitis, and the early stage of pancreatic cancer. In the terminal stage of pancreatic cancer the serum PLA2 level became low. In patients with chronic pancreatitis, significant correlations were observed between the levels of factors evaluated by the secretin test and the serum total PLA2 and proPLA2 level, but not the PLA2 level. The serum PLA2 and proPLA2 concentrations, and the proportion of proPLA2 in the total, were within normal ranges in patients with liver cirrhosis, hepatocellular carcinoma, and chronic renal failure. These results suggest that simultaneous measurements of serum PLA2 and proPLA2 are clinically useful for diagnosis and monitoring of the active phase of pancreatitis.     

Transplantation of hepatocytes for prevention of intracranial hypertension in pigs with ischemic liver failure

BACKGROUND: How Helicobacter pylori infection affects gastric acid secretion is still unclear. METHODS: Gastric juice pH, ammonia concentration in gastric juice, serum gastrin level, and grade of gastritis in accordance with the Sydney System were determined for patients with gastric ulcer (GU) and duodenal ulcer (DU) before and after treatment with lansoprazole and amoxicillin, and results were compared with those of H. pylori-negative controls. RESULTS: Scores for H. pylori density, atrophy, metaplasia, and activity of gastritis in the corpus were higher in patients with GU, especially those with proximally located GU, than in those with DU. Gastric juice pH was significantly higher in GU patients than in DU patients and controls. After H. pylori eradication, gastric juice pH and serum gastrin levels in both GU and DU patients were significantly decreased to control levels. In patients without eradication, no significant changes in these factors were observed. CONCLUSIONS: These findings suggest that H. pylori infection and gastritis in the corpus suppress acid secretion and increase gastric juice pH, resulting in hypergastrinemia, and that eradication of H. pylori normalizes acid secretion and serum gastrin levels.     

Gastric juice neopterin in Helicobacter pylori infection

Neopterin, a pteridine compound produced by macrophages activated by interferon-gamma, is widely used to assess the activation of cellular immunity. An elevation in serum or urinary neopterin reflects immune activation in many different disorders, including viral infections, cancer, autoimmune diseases or acute myocardial infarction, but less attention has been paid to neopterin concentration in other biological fluids. The aim of the present study was to examine neopterin concentration in gastric juice. An association with the presence of Helicobacter pylori, a bacterium linked to the most common disorders of upper digestive tract, was also investigated. Gastric juice was obtained at endoscopy from 61 patients. Neopterin was determined by a radioimmunoassay and the presence of H. pylori was examined by urease test. The macroscopic finding of bile in gastric juice was associated with significantly higher neopterin levels compared to patients where no bile was noted (15.5 +/- 15.6 vs. 2.1 +/- 3.0 nmol/l, P < 0.001). However, similar concentrations were observed in the H. pylori positive and H. pylori negative patients (7.6 +/- 12.0 vs. 11.1 +/- 14.9 nmol/l). Even in the absence of macroscopic bile contamination, no significant difference could be found between the infected and uninfected patients (2.3 +/- 3.2 vs. 1.3 +/- 1.9 nmol/l), and the patients with duodenal ulcer and normal findings (3.8 +/- 4.6 vs 1.6 +/- 1.9 nmol/l). The contamination of gastric juice with bile represents the limitation for the use of neopterin as a marker of immune activation in the gastric mucosa. Rather than an index of immune activation, gastric juice neopterin concentration represents a marker of duodenogastric reflux.     

Hemoperitoneum due to splenic rupture in a CAPD patient with chronic myelogenous leukemia

BACKGROUND: Due to the expression of urease, Helicobacter pylori is able to establish itself in the human stomach under acidic conditions. A novel host defence mechanism was recently proposed, suggesting that the formation of salivary nitrite in symbiosis with facultative anaerobic bacteria in the oropharynx, is aimed at enhancing the antimicrobial activity of gastric juice. AIMS: To investigate whether the addition of nitrite in physiological concentrations influences the resistance of H pylori to acid. METHODS: H pylori cultured from fresh gastric Biopsy specimens was exposed for 30 minutes to normal saline and to HCl/KCl buffer (0.2M) at pH 2 with urea (5 mM) added. The influence of potassium nitrite (50-1000 mumol/l) on bacterial survival was determined. RESULTS: Addition of nitrite (1 mM) to acidic solutions (pH 2) resulted in complete kill of H pylori within 30 minutes exposure time whereas acid alone allowed the organism to survive (p < 0.001). The antimicrobial effect of nitrite at pH 2 against H pylori was dose dependent and complete kill of organisms occurred at concentrations > or = 500 mumol/l. CONCLUSION: Acidified nitrite has anti-bacterial activity against H pylori. This should prompt further research into the effect of salivary nitrite on the survival of H pylori in the human stomach.     

Elevation of the phospholipase A2 activity in peritoneal fluid cells from women with endometriosis

OBJECTIVE: To assess the prostaglandin (PG) production on peritoneal fluid (PF) cells, phospholipase A2 (PLA2) activity of those cells in women with endometriosis was measured and compared with that of women without endometriosis. DESIGN: Prospective clinical controlled study. PATIENTS: Women who underwent laparoscopy and were found either to have endometriosis (n = 15) or not (n = 9) were included in this study. Mononuclear cells obtained from the patients at laparoscopy were immediately separated by a Ficoll-Paque technique, lysed by nitrogen cavitation, and stored at -80 degrees C. INTERVENTIONS: Phospholipase A2 activity was measured by Dole assay using 1-palmitoyl-2-[1-14C] palmitoyl phosphatidyl choline and assessed on a protein basis and a cell number basis. RESULTS: There were at least four measurable kinds of PLA2 activity detected in the cells: two calcium-dependent pH optima 7.0 and 9.0 activities and two calcium-independent pH optima 7.5 and 8.5 activities. A calcium-dependent and pH optima 9.0 activity was the highest, and it was significantly higher in women with endometriosis when compared with those who did not have endometriosis. CONCLUSION: These results indicate that the increase in the PGs in PF with endometriosis may be produced by PF cells in which PLA2 activity is elevated.     

Gastroduodenal mucosal protection

The barrier that protects the undamaged gastroduodenal mucosa from autodigestion by gastric juice is a dynamic multicomponent system. The major elements of this barrier are the adherent mucus gel layer, which is percolated by the HCO3- secretion from the underlying epithelial cells; the epithelial layer itself, which provides a permeability barrier and can rapidly repair superficial damage by a process of cell migration referred to as reepithelization or restitution; and a specially adapted vasculature, which provides a supply of HCO3- for transcellular transport and/or diffusion into the mucus layer. Passive diffusion of intestinal HCO3- into the lumen is particularly important when there is superficial damage resulting in increased leakiness of the mucosal epithelium. The process of reepithelization occurs by the migration of performed cells from gastric pits or duodenal crypts. This process is quite distinct from the wound healing and associated inflammatory response that accompany more severe injury or chronic damage. The adherent mucus gel acts as a physical barrier against luminal pepsin and provides a stable unstirred layer that supports surface neutralization of acid by mucosal HCO3-. Surface neutralization by mucosal HCO3- provides a major mechanism of protection against acid in the proximal duodenum. In the stomach, where luminal acidity can fall to around pH 1, other mechanisms of protection must exist, since the surface pH gradient is reported to collapse when luminal H+ exceeds approximately 10 mM. This collapse of the surface pH gradients may reflect, at least in part, that such studies have been mostly performed on non-acid-secreting mucosa where the supply of HCO3- to the interstitium from the parietal cells will be reduced. However, because the gastric mucosa can withstand prolonged exposure to acid without apparent damage, this implies an intrinsic resistance of the epithelial apical surface. This is amply illustrated within the gastric glands that do not secrete mucus and HCO3- yet are exposed to undiluted pepsin and an isotonic solution of HCl. Bicarbonate and mucus secretions together with mucosal blood flow are under paracrine, endocrine, and neural control. The rate of reepithelialization will depend on local chemotactic factors, adhesion mechanisms, and the creation of an acid/pepsin/irritant-free environment under a protective gelatinous or mucoid cap. If optimal conditions are met, then the rate of reepithelialization appears to depend primarily on the intrinsic properties of the migrating cells themselves rather than control by exogenous mediators.(ABSTRACT TRUNCATED AT 400 WORDS)     

Properties of phospholipase A1/transacylase in the white muscle of bonito Euthynnus pelamis (Linnaeus)

The properties of phospholipase A1 (PLA1) obtained from the white muscle of bonito, Euthynnus pelamis (Linnaeus), were examined. The PLA1 activity had a pH optimum from 6.5 to 7.0 for phosphatidylcholine (PC), and calcium ion was not required. The optimum temperature was from 20 to 30 degrees C. When a fatty alcohol was used as an acceptor, a wax ester was produced by transferring a fatty acid at the sn-1 position of the donor's PC. The maximum production of lysophosphatidylcholine was shifted by 0.5 pH units to the acidic side and the pH optimum of wax ester synthesis was from 6.0 to 6.5. The synthesis was independent of calcium ion and Coenzyme A. The transacylation was also observed when 1-lyso-2-acyl-sn-glycero-3-phosphocholine was used as an acceptor. Fatty acid at the sn-1 position of the donor PC was transferred to the unoccupied hydroxy group of the acceptor at the sn-1 position. When 2,3-dipalmitoyl-sn-glycero-1-phosphocholine was used as the acyl donor, a similar amount of palmitic acid was transferred as in the case of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine. However, 1-acyl-2-lyso-sn -glycero-3-phosphocholine, a positional isomer, was a poor acceptor. These results indicate that the transacylation by the PLA1 from bonito muscle is not stereospecific, but is position-specific both for the acyl donor and acceptor.     

Arachidonic acid protects against hypoxic injury in rat proximal tubules

Free fatty acids (FFA) and lysophospholipids accumulate during hypoxia (H) in rat proximal tubular epithelial cells partly as a result of increased phospholipase A2 (PLA2) activity. The role of FFA in mediating hypoxic injury and modulating PLA2 activity is not clear. In the present study, the effect of several FFA including arachidonic acid (AA, 20:4) on hypoxia-induced injury and PLA2 activity was assessed in freshly isolated rat proximal tubules. Hypoxia (H) was induced in the presence of either an unsaturated free fatty acid (uFFA) [AA or linoleic acid (LA, 18:2)] or a saturated FFA (sFFA) [palmitic (PA, 16:0) or myristic acid (MA, 14:0)]. Cell membrane injury was assessed by measuring lactate dehydrogenase release (LDH). AA markedly reduced LDH release during hypoxia in a dose dependent manner, while sFFA had no protective effect. LA showed similar protection to that observed with AA. AA did not affect buffer calcium concentration, buffer pH, intracellular pH or intracellular calcium concentration. Neither inhibiting the cyclooxygenase pathway with indomethacin, nor the lipoxygenase pathway with nordihydroguaiaretic acid (NDGA) had any effect on the AA observed cytoprotection. In vitro PLA2 activity in the control tubular extracts was compared to that following addition of AA or PA. PLA2 activity decreased significantly with AA but not with PA in a dose dependent manner. These data suggest that: (1) AA protects against hypoxic injury in rat proximal tubules. (2) This cytoprotection is not specific for AA and other uFFA have a similar effect. (3) AA significantly inhibits PLA2 activity, (4) AA induced cytoprotection may be related to a negative feedback inhibition of PLA2 activity.