Cellular mechanism of intestinal permeability alterations produced by chelation depletion.

The absorption of phenolsulfonphthalein (phenol red) was used as a measure in vivo of intestinal permeability in anesthetized rats. A chelating agent, sodium ethylenediaminetetraacetate (NaEDTA), placed in the lumen evoked a fivefold increase in membrane permeability; at the same time the mucosal content of magnesium and calcium decreased significantly. Making either magnesium or calcium available to the luminal surface of the membrane in isotonic solution restored normal permeability and brought the cation contents above the original levels. Electron micrographs of tissues treated in vivo with NaEDTA revealed (a) rounded swellings on the microvilli in the area of the junctional complexes between adjacent epithelial cells, (b) widening of intercellular channels particularly in the region of the intermediate junctions (zonulae adhaerentes), and (c) loss of architectural detail in the region of the desmosomes (maculae adhaerentes) with separation of their dense borders. All of these alterations in fine structure could be reversed by in vivo cation replacements which reinstated normal permeability. The implications of these findings on mechanisms of fluid transport across epithelial membranes are discussed, and a working hypothesis for the role of divalent cations in membrane permeability regulation is presented.

Ultrastructural modifications of intestinal and colonic mucosa induced by free or bound bile acids.

There is substantial evidence that bile acids may enhance the colon tumorigenesis induced by chemical carcinogens and that agents stimulating increased bile acid excretion may show similar promoting or enhancing activity. To test the premise that these agents might modify topographical ultrastructure of the small intestine and colon in the absence of carcinogens, rats were fed for 6 weeks on chemically defined diets containing 2% levels of three commercial bile acid sequestrants or 15% levels of wheat brain, cellulose, pectin, or alfalfa. Major qualitative and quantitative deviations from normal morphology were observed with each of the three sequestrants. Similar but less dramatic modifications occurred with diets containing alfalfa or pectin, both of which either "bind" bile acids in vitro or result in increased bile acid excretion. Bran and cellulose which neither "bind" bile acids nor increase their fecal excretion, were without significant effects on intestinal or colonic morphology. The morphological deviations observed with bile acid sequestrants were shown to be a direct response to free or bound bile acids by comparing the morphological modifications resulting from daily intracolonic infusions of free bile acids, sequestrant-bound bile acids, or the sequestrant alone.

Etiology of the Austin Flint murmur.

OBJECTIVES: The aim of the study was to determine the mechanism of the Austin Flint murmur. BACKGROUND: More than 100 years after the initial description of the Austin Flint murmur, the etiology of the murmur remains unclear. METHODS: M-mode and two-dimensional echocardiography, conventional and color flow Doppler study, and cine nuclear magnetic resonance (cine NMR) imaging were performed in 24 patients with clinically moderate or severe aortic regurgitation. Mitral valve area was determined by planimetry and pressure half-time measurement. Overlap of the aortic regurgitation and mitral inflow jets was graded 0 (no overlap) to 4 (marked overlap) by Doppler study and cine NMR imaging. The volume of signal loss resulting from turbulent blood flow secondary to the aortic regurgitation jet was determined on cine NMR images, and the extent of contact with the left ventricular endocardium was graded 0 (no contact) to 4 (extensive contact). RESULTS: The presence of an Austin Flint murmur did not correlate with mitral valve area (2.7 +/- 0.8 cm2 with the murmur vs. 2.5 +/- 0.7 cm2 without), overlap of the aortic regurgitation and mitral flow jets (3 +/- 1 vs. 2.3 +/- 1.2), diastolic mitral regurgitation (50% vs. 71%) or fluttering of the anterior mitral valve leaflet (70% vs. 50%). The presence of an Austin Flint murmur correlated best with the volume of signal loss associated with the aortic regurgitation jet on cine NMR imaging (65 +/- 16 ml with the murmur. vs. 38 +/- 11 ml without, p less than 0.001) and the extent of contact of this signal loss with the left ventricular endocardium (2.9 +/- 0.5 vs. 1.5 +/- 0.4, p less than 0.0001). CONCLUSIONS: The Austin Flint murmur is caused by the aortic regurgitation jet abutting the left ventricular endocardium, resulting in the generation of a low-pitched diastolic rumbling.

Captopril protects against myocardial injury induced by magnesium deficiency.

We have previously reported that antioxidant drug intervention protects against magnesium deficiency-induced myocardial lesions. In the present study, Golden Syrian male hamsters were fed either a magnesium-deficient diet or a magnesium-supplemented diet. Animals from each group received sulfhydryl-containing angiotensin converting enzyme inhibitors: captopril, epi-captopril (a stereoisomer of captopril), and zofenopril* (arginine blend of zofenopril containing a free SH group); another group of animals received the non-sulfhydryl-containing angiotensin converting enzyme inhibitor enalaprilat. The animals were killed after 14 days, and their hearts were isolated for morphological and morphometric analyses. Hematoxylin and eosin-stained sections were examined by a computer image analysis system for a morphometric determination of the severity of myocardial injury. Captopril reduced both the density of lesions, from 0.32 to 0.08 lesions/(mm2) (p less than 0.01), and the area fraction of lesions, from 7.42 x 10(-4) to 2.03 x 10(-4) lesion area/(mm2) (p less than 0.01), as well as the degree of inflammatory infiltration around the blood vessels. Epi-captopril and zofenopril* were virtually equipotent to captopril, but enalaprilat afforded only slight (nonsignificant) protection. These results indicate that a significant component of the protective effect of captopril in this model was attributable to its sulfhydryl moiety, rather than solely due to the inhibition of the angiotensin converting enzyme. These data further support our previous findings of possible free radical participation in cardiomyopathy due to magnesium deficiency.

Propranolol preserves ultrastructure in adult cardiocytes exposed to anoxia/reoxygenation: a morphometric analysis.

The protective effect of d,l-propranolol was studied using freshly isolated canine ventricular cardiocytes (1.5 x 10(6)/mL) exposed to 30 min anoxia (95% N2/5% CO2) and 0, 3, 20, and 45 min of reoxygenation (95% O2/5% CO2). In addition to preventing lipid peroxide formation, propranolol maintained cellular viability, and minimized ultrastructural alterations. In the absence of propranolol, the outer mitochondria become swollen and rounded up within the first few minutes of reoxygenation. The perinuclear mitochondrial area increased only slightly. We observed that the cellular injury process proceeded differentially from the exterior to the interior, with a mitochondrial area increase and outer membrane rupture. Sarcolemmal damage was also observed with prevalent blebbing and membrane loss. The Z-lines became wider and more diffuse with reoxygenation. Injury to the nuclear double membrane was observed. Incubation with propranolol showed significant protection during postanoxia reoxygenation. In contrast, the more water soluble beta-blocker atenolol only exhibited slight protection. In addition, d-propranolol (the non beta-blocking isomer) and the antioxidant enzymes, SOD and catalase, showed significant protection. These data support previous findings concerning the antioxidant properties of propranolol which appear to be independent of beta-receptor blockade.

Stimulation of intestinal cytokinetics and mucin turnover in rats fed wheat bran or cellulose.

Rats were fed defined diets containing no fiber, 10% wheat bran or 10% cellulose, and intestinal morphology and cytokinetics were assessed by light microscopy and autoradiography, respectively. In bran-fed animals, there were no differences in morphological appearance of the jejunum, in the number of cells/villus column or in numbers of goblet cells compared to controls. Autoradiographic analysis, at one and 24 h after [3H]thymidine, however, suggested an increased turnover and villus transit of intestinal cells. There was also a 2.5 fold increase in incorporation of labeled sulfate, and a 2-fold increase in [3H]glucose incorporation into total intestinal glycoproteins and mucins. Similar, albeit less dramatic results were obtained in rats fed diets containing cellulose. These studies provide evidence that diets containing certain fiber derivatives can alter aspects of intestinal cell turnover, and support the earlier morphological observations suggesting increased goblet cell secretory activity in response to feeding these fiber derivatives.

Comparative effects of chitosan and cholestyramine on lymphatic absorption of lipids in the rat.

The effects of chitosan and cholestyramine on intestinal absorption of oleic acid and cholesterol were assessed in adult male rats with cannulae in the thoracic duct lymphatic channel. In acute studies, 50 mg of either cholestyramine or chitosan were included in the test emulsion for intragastric administration. Over a 24-h lymph collection period, cholestyramine caused a 47% depression of cholesterol absorption and a 32% interference with oleic acid absorption. Chitosan had a similar effect on the absorption of both lipids under these conditions (51 and 41% depression, respectively). Studies were also conducted in animals fed for 4 wk on defined diets containing 1 and 5% levels of the test materials. Absorption of lipids from the standard aqueous emulsion was tested in lymph duct cannulated rats which had been fasted overnight. In these animals, prefeeding either cholestyramine or chitosan at the 1% level caused an 18 to 28% reduction in absorption of both lipids with greater variability in the chitosan-fed group. When either test material was fed at the 5% level, absorption of cholesterol was reduced by 63 to 69% and absorption of oleic acid by 58 to 62%. Although these agents may act by different mechanisms, the data suggest that chitosan is as effective as cholestyramine in its acute effects on lipid absorption and that, with chronic feeding, both materials cause equivalent adaptive changes in intestinal transport of administered fatty acid and cholesterol.

Dietary fiber and intestinal adaptation: effects on intestinal and pancreatic digestive enzyme activities.

Male Wistar rats were fed for four weeks on defined diets containing no fiber additions, 10% levels of insoluble fiber derivatives (cellulose or alfalfa), or 5% levels of viscous fiber derivatives (pectin, guar gum, or metamucil). After an overnight fast, the pancreas was assayed for protein, amylase, lipase, trypsin, and chymotrypsin. Homogenates of small intestinal mucosa were analyzed for protein, alkaline phosphatase, invertase and thymidine kinase. There were, with few exceptions, no dietary effects on the exocrine pancreatic enzymes. The specific activities of the villus marker enzymes (invertase and alkaline phosphatase) tended to be higher in the proximal (but not middle or distal) intestines of the fiber-fed groups, while total activities were the same in all groups. In contrast, the activity of the crypt marker, thymidine kinase, was highest in the distal intestinal segments, and even higher in animals given the alfalfa, guar gum or metamucil-supplemented diets.

Dietary fiber and intestinal adaptation: effects on lipid absorption and lymphatic transport in the rat.

Adult male rats fed defined diets containing various fiber supplements or cholestyramine for 4 wk were surgically provided with lymphatic drainage catheters and starved overnight. After duodenal administration of a standard lipid test emulsion, absorption rates and lipoprotein distributions of cholesterol and oleic acid were determined. Prefeeding diets containing cellulose or alfalfa had no significant effect on oleic acid absorption. Diets containing pectin, guar gum, metamucil, mixed fibers (Fibyrax), or cholestyramine caused decreased lymphatic recovery in the initial period; except for the metamucil diet, no decrease was caused in the 24-h recovery, suggesting delayed but not impaired absorption. Fatty acid distribution among lipoproteins and chylomicron size were not altered by diet. All supplements caused a significant reduction in cholesterol absorption during the initial period, and cholesterol absorption remained depressed in animals prefed pectin, guar gum, mixed fibers, metamucil, and cholestyramine.

Interactions of alfalfa plant and sprout saponins with cholesterol in vitro and in cholesterol-fed rats.

The in vitro interactions of saponins from alfalfa plant and alfalfa sprouts with cholesterol and the effects of alfalfa plant and sprout and saponin-free alfalfa plant on diet-induced liver cholesterol accumulation, bile acid excretion, and jejunal and colonic morphology were examined. Cholesterol-saponin interactions have been suggested as mechanisms for the observed hypocholesterolemic effects of alfalfa as well as the changes in intestinal morphology. Alfalfa plant saponins bound significant quantities of cholesterol both from ethanol solution and from micellar suspension. Alfalfa sprout saponins interacted with cholesterol to a lesser but significant extent. Sprout saponins also inhibited growth of Trichoderma viride significantly, another measure of saponin-cholesterol interaction. Bile acid adsorption was greatest for alfalfa plant and was not reduced by removal of saponins from the plant material. The ability of alfalfa to reduce liver cholesterol accumulation in cholesterol-fed rats was enhanced by removal of saponins and alfalfa sprouts did not prevent accumulation. Removal of saponins from alfalfa reduced the changes in intestinal morphology previously reported, but interaction with membrane cholesterol did not appear to be the cause of this effect of saponins. Saponin-cholesterol interaction is an important part of the hypocholesterolemic action of alfalfa but interaction of bile acids with other components of alfalfa may be of equal importance.

Effect of chronic intake of dietary fibers on the ultrastructural topography of rat jejunum and colon: a scanning electron microscopy study.

This report is an attempt to quantitate the observable topographical characteristics of small and large intestine after a specific dietary regimen under well-defined states of lipid absorption and metabolism. Alfalfa, white wheat bran, cellulose, and pectin were fed for 6 wk at a level of 15 g/100 g diet to four groups of rats (12 rats per dietary group). A 5th control group was maintained on Purina Rat Chow. Three animals from each group were blind-coded for morphological assessment. After anesthesia, the jejunum and mid-colon were removed and processed for scanning electron microscopy. Beginning with the mildest mucosal surface changes, the observed order in terms of increasing severity is bran less than cellulose less than pectin less than alfalfa. Our observations suggest that altered rates of cell loss in intestinal tract cytokinetics may be occurring with particular feeding patterns and should be considered as a possible mechanism in the nutritional consequences of dietary fiber intake.

In vitro clearance of chylomicron triglycerides containing (omega-3) eicosapentaenoate.

Rat mesenteric lymph chylomicrons, containing triglycerides enriched with either [14C]oleic acid or [14C]eicosapentaenoic acid, were prepared by ultracentrifugation of lymph samples collected for 6 h after a single duodenal infusion of an emulsion containing 0.3 mmol of either fatty acid. After determination of protein and of total fatty acid content and composition, enriched chylomicrons were suspended in Krebs-bicarbonate buffer. Non-working hearts were perfused in a recirculating system for 45 min using the enriched chylomicron preparations. At 15 min intervals during perfusion, the media were assayed for total radioactivity, 14CO2 and 14C-labeled fatty acids associated with triglycerides, unesterified fatty acids, phospholipids, mono- and diglycerides. After perfusion, the hearts were extracted and assayed for total lipid radioactivity and isotope distribution among heart lipid fractions. With this membrane-supported lipoprotein lipase system, clearances of chylomicron triglycerides containing either fatty acid were identical, as were the myocardial uptakes of the fatty acids and oxidations to 14CO2. Furthermore, except for a significantly greater incorporation of eicosapentaenoate into myocardial phospholipids, tissue isotope distributions of the two labeled fatty acids were also the same. These studies suggest that at least the initial phases of peripheral clearance of chylomicrons enriched in omega-3 fatty acids is as efficient as with those containing oleate.

Effects of cardiac rehabilitation and exercise training programs in women with depression.

Depression is prevalent in women with coronary artery disease, and increases morbidity and mortality following major coronary events. We demonstrated that women with depression had markedly abnormal overall cardiovascular risk profiles and have marked benefits in exercise capacity, obesity indexes, behavioral characteristics (including depression), and quality of life following formal, outpatient phase II cardiac rehabilitation and exercise training programs.

Prognostic value of stress echocardiography in the evaluation of atypical chest pain patients without known coronary artery disease.

Patients with atypical chest pain frequently lack significant coronary artery disease (CAD) and are, therefore, at low risk for future adverse cardiovascular events. We hypothesized that in this group of patients, stress echocardiography could identify those at risk for cardiac events. We retrospectively reviewed (mean follow-up 23.0 +/- 7.2 months) the prognostic value of stress echocardiography for major (cardiac death, myocardial infarction, congestive heart failure, and unstable angina) and total (major events plus coronary revascularization) cardiac events in 661 patients with atypical chest pain, normal global left ventricular (LV) systolic function, and no history of CAD. A positive stress echocardiogram was defined as the development of new or worsening wall motion abnormalities with exercise stress (80%) or dobutamine (20%). A total of 41 cardiac and 16 major events were noted. The event-free survival for total cardiac events was 97% for a normal stress echocardiogram and 93% for a normal stress electrocardiogram (ECG) at 30 months. A positive stress ECG predicted an event-free rate of 86% compared with 74% for stress-induced wall motion abnormalities and 42% if stress-induced LV dysfunction accompanied the wall motion abnormalities. A strategy recommending invasive studies based on positive stress echocardiogram results increased the per-patient cost, but led to greater savings per cardiac event predicted and provided incremental prognostic value for future cardiac events beyond clinical and stress electrocardiographic data. Thus, stress echocardiography in low-risk patients for CAD appears to be more cost effective than a stress ECG.

Cytokines, neuropeptides, and reperfusion injury during magnesium deficiency.

In summary, hypomagnesemia enhances reperfusion injury. We postulate that neurogenic inflammation, which occurs very early during hypomagnesemia, predisposes the myocardium to reperfusion injury by depleting endogenous antioxidants and recruiting inflammatory cells, which can participate in enhanced free radical production during postischemic reperfusion. Vitamin E supplements can prevent the occurrence of this enhanced injury possibly through the restoration of endogenous antioxidant defenses.

Evidence for cigarette smoke-induced calcitonin secretion from lungs of man and hamster.

In hamsters, acute cigarette smoke inhalation increased serum levels of the hormone calcitonin; and, in humans, smoking of two high-nicotine content cigarettes increased serum and urine levels of this hormone. The source of this immunoreactive calcitonin (iCT) does not appear to be the thyroid gland, since previously thyroidectomized patients demonstrated a similar response. In the hamster, the increased serum iCT levels were accompanied by a decreased lung tissue iCT content and hypocalcemia. It is suggested that the source of the cigarette smoke-induced hypercalcitonemia is the lung, possibly from the iCT-containing pulmonary neuroendocrine (PNE) cells. Moreover, this response appears to be dependent on the nicotine content of the cigarettes.

Dietary fibers: V. Binding of bile salts, phospholipids and cholesterol from mixed micelles by bile acid sequestrants and dietary fibers.

Mixed micelles were prepared containing combinations of either taurocholate or taurochenodeoxycholate, monoolein, oleic acid, dioleylphosphatidylcholine (lecithin) and cholesterol. These were incubated with commercial bile-acid-sequestering resins, cholestyramine and DEAE-Sephadex, or various dietary fibers and fiber components including wheat bran, cellulose, alfalfa, lignin and 2 viscosity grades of guar gum. Binding was determined as the difference between the radioactivity of each micellar component added and that recovered in the centrifugal supernatant after incubation. In general, the extent of bile salt sequestration was characteristic and reproducible for each bile salt, and was largely unaffected by the presence of one or more additional components of the micellar mixture, including the other bile salt. Cholestyramine bound 81-92% of the bile salts and 86-99% of the phospholipid and cholesterol present in micelles. DEAE-Sephadex sequestered only 49% of the taurocholate and 84% of the taurochenodeoxycholate, but completely removed all of the phospholipid and cholesterol from micelles containing either bile salt. Among the dietary fibers, guar gum of either viscosity bound between 20-38% of each micellar component, whereas lignin, alfalfa, wheat bran and cellulose were progressively less effective in sequestration of individual components of mixed micelles. The extent of sequestration of micellar components by these resins and fibers is reasonably correlated with the effects of these same materials on lymphatic absorption of lipids and to their suggested hypocholesteremic properties.

Dietary fiber supplements: effects on serum and liver lipids and on liver phospholipid composition in rats.

Rats (6 per group) were fed semipurified diets containing either particulate fibers (alfalfa, 10%; cellulose, 10%; bran, 10%), a soluble ionic fiber (pectin 5%), soluble, nonionic fibers (guar gum, 5%; Metamucil, 10%), a mixed fiber preparation (Fibyrax, 10%, or an insoluble, ionic bile acid-binding resin (cholestyramine, 2%). The control group was fed the unsupplemented diet. The feeding period, during which diet and water were provided ad libitum, was 28 days. Compared with the control group, serum total cholesterol levels were increased by more than 10% in rats fed alfalfa and decreased by more than 10% in rats fed cellulose, guar gum, Fibyrax and cholestyramine. There were no significant differences in percentage of plasma HDL cholesterol. Serum triglycerides were elevated in the groups fed alfalfa, pectin, guar gum or Fibyrax and reduced in the group fed Metamucil. Plasma phospholipids were elevated in rats fed alfalfa or bran, unaffected in rats fed pectin or Metamucil and reduced in the other groups. Liver total cholesterol was elevated in all groups but those fed wheat bran and cholestyramine. The percentage of liver cholesterol present as ester was elevated in every group except that fed cholestyramine. Liver triglycerides were reduced in rats fed guar gum or Metamucil and elevated in those fed alfalfa. Liver phospholipids were lowered in the group fed cellulose. Liver phospholipids were fractionated by thin layer chromatography to give phosphatidylcholine (PC), phosphatidylethanolamine (PE), sphingomyelin (Sph), lysophosphatidylcholine (LPC) and phosphatidylinositol plus phosphatidylserine (PI + PS).(ABSTRACT TRUNCATED AT 250 WORDS)

Comparison of the clearances of serum chylomicron triglycerides enriched with eicosapentaenoic acid or oleic acid.

Rat mesenteric lymph chylomicrons containing triglycerides enriched with either [14C]oleic acid (OA) or [14C]-eicosapentaenoic acid (EPA) were prepared by ultracentrifugation of lymph samples collected for 6 hr after a single duodenal infusion of an emulsion containing either fatty acid. These chylomicrons were injected into the jugular vein of recipient rats and, at various time intervals, blood was drawn and serum was assayed for radioactivity. In separate animals, serum lipoprotein fractions were separated by ultracentrifugation, and the redistribution of labeled fatty acid among circulating lipoproteins was determined by liquid scintillation spectrometry. When the early disappearance rates (10 min) of either total serum radioactivity or specifically the chylomicron fraction were compared, there were no differences between the groups receiving OA- or EPA-enriched chylomicrons. However, disappearance rates of EPA-enriched chylomicrons were slower than those of OA-enriched chylomicrons from 25 to 90 min. The small but significant differences in the disappearance rates for the longer time periods cannot be ascertained without further studies. At 5 min after injection of either type of chylomicron, the d less than 1.006 g/ml lipoprotein fraction of serum chylomicrons and very low density lipoproteins contained almost 90% of the original radioactivity. By 240 min, when less than 2% of the radioactivity remained, this radioactivity in the d less than 1.006 g/ml fraction was 43-46%, with concomitant increases in the low and high density lipoprotein fractions and in the lipoprotein-free serum.

Magnesium-deficient myocardium demonstrates an increased susceptibility to an in vivo oxidative stress.

Previous studies in our laboratory have indicated free radical participation in magnesium deficiency cardiomyopathy. In this study, we examined the capacity of the magnesium-deficient animals to withstand an in vivo oxidative stress. Syrian hamsters were placed on either magnesium-deficient diet or a magnesium-supplemented control diet. Animals from each group also received vitamin E. After 14 days some of animals were given the catecholamine isoprenaline; 2 days later the animals were killed. The severity of the isoprenaline-induced injury was assessed by a morphometric analysis. The isoprenaline-induced injury was dramatically increased in the magnesium-deficient animals. The addition of vitamin E reduced the severity of the injury by 81% in these animals, indicating that the injury process was primarily due to an oxidative mechanism. These data show that magnesium deficiency increases the susceptibility of the cardiovascular system to oxidative stress.