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Biomedical subjects

A Fasano

Publications and source records attributed to A Fasano.

At least 19 recordsLinked to original sources
PubMed1992

Multifocal corneal topographic changes with excimer laser photorefractive keratectomy.

Excimer laser photorefractive keratectomy can flatten the central cornea, thereby eliminating myopic refractive errors; in older patients, however, presbyopia limits satisfaction. Computer-assisted topographic analysis of corneas after refractive surgery indicates that a minority of patients achieve a multifocal lens effect, such that they maintain reasonable acuity over a range of defocus. We have purposefully attempted to create a multifocal refractive effect and have analyzed the subsequent topographies quantitatively to determine if multifocality was achieved. In corneas not operated on and plastic hemispheres, a fairly small range of corneal powers is observed; the range of powers is increased after a monofocal ablation. After multifocal ablations, a greater spread of surface powers is observed, often with a bimodal distribution, indicative of an apparent multifocal effect. These observations suggest that in some patients undergoing photorefractive keratectomy for myopia, it may be possible to reduce symptoms of presbyopia, although a decrease in image contrast or monocular diplopia may complicate this approach.

Animals↗
PubMed1992

Role of Vibrio cholerae neuraminidase in the function of cholera toxin.

Vibrio cholerae neuraminidase (NANase) is hypothesized to act synergistically with cholera toxin (CT) and increase the severity of a secretory response by increasing the binding and penetration of CT to enterocytes. To test this hypothesis, the NANase gene (nanH) from V. cholerae Ogawa 395 was first cloned and sequenced. Isogenic wild-type and NANase- V. cholerae 395 strains were then constructed by using suicide vector-mediated mutagenesis. The influence of NANase on CT binding and penetration was examined in vitro by using culture filtrates from these isogenic strains. Fluorescence due to binding of fluorescein-conjugated CT to C57BL/6 and C3H mouse fibroblasts exposed to NANase+ filtrates increased five- and eightfold, respectively, relative to that with NANase- filtrates. In addition, NANase+ filtrates increased the short-circuit current measured in Ussing chambers 65% relative to that with NANase- filtrates, although this difference decreased as production of CT increased. The role of NANase in V. cholerae pathogenesis was examined in vivo by intragastric inoculation of the isogenic strains into CD1 suckling mice. No difference in fluid accumulation ratios was seen at doses of 10(4) to 10(8) CFU, but NANase+ strains produced 18% higher fluid accumulation ratios at 10(9) CFU than NANase- strains when inoculated into nonfasted suckling mice. It is concluded that NANase plays a subtle but significant role in the binding and uptake of CT by susceptible cells under defined conditions.

Amino Acid Sequence↗
PubMed1992

Cloning of a gene (zot) encoding a new toxin produced by Vibrio cholerae.

Live oral candidate cholera vaccines have previously been constructed by deletion of Vibrio cholerae sequences encoding the enzymatically active A subunit of the cholera toxin. However, volunteer studies have shown that these non-cholera toxin-producing strains still provoke mild to moderate diarrhea in some individuals. We recently reported the identification of a second toxin produced by V. cholerae which may be responsible for this residual diarrhea (A. Fasano, B. Baudry, D. W. Pumplin, S. S. Wasserman, B. D. Tall, J. M. Ketley, and J. B. Kaper, Proc. Natl. Acad. Sci. USA 88:5242-5246, 1991). This new toxigenic factor increases the permeability of rabbit ileal mucosa by affecting the structure of the intercellular tight junctions (zonula occludens). We now report the identification and cloning of the gene encoding this new toxin. This gene, named zot (for zonula occludens toxin), consists of a 1.3-kb open reading frame which could potentially encode a 44.8-kDa polypeptide. The location of the zot gene encoding the new toxin is immediately upstream of the ctx operon encoding cholera toxin.

Amino Acid Sequence↗
PubMed1992

Enterotoxigenicity of Vibrio parahaemolyticus with and without genes encoding thermostable direct hemolysin.

Vibrio parahaemolyticus produces a thermostable direct hemolysin (TDH) that has been implicated in the pathogenesis of diarrheal disease caused by this organism. However, previous studies attempting to demonstrate the contribution of the hemolysin to virulence have been inconclusive. We investigated this putative virulence factor by using an isogenic TDH-negative (TDH-) strain constructed by specifically inactivating the two copies of the tdh gene encoding TDH. The enterotoxigenicities of the parent strain (AQ3815) and the mutant strain were tested by adding sterile culture supernatants to rabbit ileal tissue mounted in Ussing chambers. The culture filtrate of the parent strain produced a significant increase in short-circuit current (Isc), compared with the change induced by the TDH- mutant. The capacity of the culture filtrate of AQ3815 to increase the Isc was reduced by neutralization with anti-TDH serum, and the return of the cloned tdh gene to the TDH- mutant restored the ability to increase the Isc. These results were corroborated by rabbit ileal loop assays in which AQ3815 caused fluid accumulation but the TDH- mutant did not. No microscopic damage was seen in mucosal tissues exposed to the culture filtrate of either strain. These results indicate that TDH has an enterotoxigenic effect on rabbit small intestine and could be responsible for the watery diarrhea seen with V. parahaemolyticus.

Animals↗
PubMed1992

Multifocal corneal topographic changes after radial keratotomy.

Quantitative analysis of the central corneal refractive powers of both eyes of three patients demonstrated that, after radial keratotomy, the corneas tended to have a greater range of refractive powers than preoperatively, and that the corneas with a multifocal effect had a wider distribution of refractive powers than the corneas of eyes without this effect. It may be possible to intentionally achieve a multifocal lens effect by designing a refractive procedure to predictably widen the distribution of central corneal refractive powers.

Cornea↗
PubMed1991

Vibrio cholerae produces a second enterotoxin, which affects intestinal tight junctions.

Attenuated Vibrio cholerae vaccine strains specifically mutated in genes encoding cholera toxin (CT) are still capable of causing mild to moderate diarrhea. Culture supernatants of V. cholerae strains, both CT-positive and CT-negative, were examined in Ussing chambers, and a toxin was found that increases the permeability of the small intestinal mucosa by affecting the structure of the intercellular tight junction, or zonula occludens. The activity of this toxin is reversible, heat-labile, sensitive to protease digestion, and found in culture supernatant fractions containing molecules between 10 and 30 kDa in size. Production of this factor (named ZOT for zonula occludens toxin) correlates with diarrheagenicity of V. cholerae strains in volunteers and may represent another virulence factor of infectious diarrhea that must be eliminated to achieve a safe and effective live oral vaccine against cholera.

Animals↗
PubMed1991

Diarrhea in ciguatera fish poisoning: preliminary evaluation of pathophysiological mechanisms.

Ciguatera fish poisoning is a clinical syndrome consisting of a combination of gastrointestinal and neurological symptoms occurring after eating toxin-containing tropical reef fish; it is a major cause of morbidity in Hawaii, the South Pacific, Australia, and the Caribbean. In an effort to define pathophysiological mechanisms responsible for the diarrheal component of the illness, we examined the effect of crude and fractionated toxin preparations on isolated rabbit ileal tissue in a Ussing chamber model. Both the crude toxin preparation (prepared from toxic Ctenochaetus strigosus) and 10% and 50% methanol-chloroform toxin fraction (prepared from a pool of toxic fish samples) gave a striking increase in transepithelial electrical potential difference and short-circuit current. Enterotoxic activity seemed to be mediated by calcium. When examined by light microscopy, the intestinal mucosa was not damaged by the toxin preparations used. Our data demonstrate that toxins involved in ciguatera fish poisoning directly stimulate intestinal fluid secretion without accompanying tissue damage and suggest that calcium is the "second messenger" mediating the process.

Animals↗
PubMed1991

Enteroaggregative Escherichia coli elaborate a heat-stable enterotoxin demonstrable in an in vitro rabbit intestinal model.

Enteroaggregative Escherichia coli (EAggEC) have been associated with persistent diarrhea in young children, but little is known about its pathogenesis. We assayed for enterotoxic activity in culture filtrates (CF) of EAggEC strains in Ussing chambers mounted with rabbit ileal mucosa. CF from strain 17-2, a prototype Chilean EAggEC strain, caused a greater rise in potential difference and short circuit current (SCC) than that seen in HB101 control, and this effect was abolished by protease pretreatment and partially stable after heat treatment. Ultrafiltration of 17-2 CF preparations localized the active moiety to the 2-5 kD Mr size range. CF from HB101 transformed with the 17-2 plasmid showed Ussing chamber activity. less than 10-kD CF fractions from five of six other EAggEC strains screened in Ussing chambers gave SCC responses of similar magnitude to 17-2. The 17-2 CF activity was not neutralized after pretreatment with polyclonal anti-STa antibody. Additionally, all of the seven EAggEC strains studied were nonreactive by heat-stable enterotoxin variant STa ELISA, were negative in the suckling mouse assay, and failed to hybridize with heat-stable enterotoxin variant STh and STp DNA probes. In summary, our data indicate that 17-2 produces a low molecular weight, partially heat-stable, protease-sensitive enterotoxin which appears to be plasmid associated, and genetically and immunologically distinct from E. coli STa. Preliminary screening suggests that this tox+ phenotype may be common among EAggEC.

Animals↗
PubMed1990

Bile acids reversible effects on small intestinal permeability. An in vitro study in the rabbit.

To define the action of deconjugated bile acids on the small intestinal permeability in an in vitro system, we investigated the effects of chenodeoxycholic acid and ursodeoxycholic acid on the rate of transmural flux of lactulose in jejunal and ileal mucosa of rabbits, stripped of their muscle layers and mounted in Ussing chambers. In a series of experiments, tissue samples from small intestinal segments either exposed to bile acids or not also were examined by scanning and transmission electron microscopy to study the integrity of the tight junctions. Results show that chenodeoxycholate, starting at the concentration of 0.1 mM, enhanced in a dose-related manner the transepithelial flux of lactulose in the ileum. Both chenodeoxycholate (0.5 mM) and ursodeoxycholate (0.5 mM) significantly increased mucosal permeability to lactulose in jejunum and ileum; the effect of chenodeoxycholate was also shown to be reversible, as it completely disappeared within 40 min after its withdrawal and it did not result in permanent changes of epithelial transport function. Finally, the tight junctions appeared loosened by the addition of 1 mM chenodeoxycholate, suggesting that this is the major site of the transient bile acid increase of small intestinal permeability to compounds such as lactulose, having a molecular radius wider than 0.5 nm.

Animals↗
PubMed1990

Enterotoxin and cytotoxin production by enteroinvasive Escherichia coli.

It has long been suspected that besides their ability to invade enterocytes, enteroinvasive Escherichia coli (EIEC) strains have the ability to elaborate an enterotoxin. We tested 35 EIEC strains for cytotoxins and 9 (1 per serogroup) for enterotoxins. All 35 strains exhibited low levels of Vero cell cytotoxins that are immunologically and genetically distinct from Shiga-like toxin I or II of enterohemorrhagic E. coli. Sterile supernatants and cell lysates of two EIEC strains were tested in rabbit ileal loops, and both stimulated moderate fluid accumulation (circa 0.5 ml/cm) without tissue damage; secretory activity was confirmed in Ussing chambers, where these two strains and the seven others tested significantly increased short circuit current without altering tissue conductance. Curing the 140-MDa invasiveness plasmid from an EIEC strain did not diminish enterotoxin production. Culture in minimal Fe2+ medium is necessary to detect expression of the enterotoxin which is circa 68 to 80 kDa in size and is distinct from the EIEC cytotoxin.

Animals↗
PubMed1989

Diarrhoea in jaundiced neonates treated with phototherapy: role of intestinal secretion.

Thirty jaundiced neonates with diarrhoea who were being treated with phototherapy and 30 matched control infants were studied to try and find out the cause of the diarrhoea. Faecal osmolality and electrolyte concentrations were measured, which gave clear evidence that the diarrhoea arose from intestinal secretion. Rectal water and electrolyte absorption in 10 jaundiced infants receiving phototherapy, in 10 jaundiced infants not receiving phototherapy, and in 10 healthy controls was measured with a rectal dialysis bag. A further group of eight jaundiced infants was also studied both during and after phototherapy to document the reversal of ion transport changes. Absorption of water, sodium chloride, and potassium was significantly impaired in the patients receiving phototherapy compared with each of the control groups. Such impairment was transient, as it was not apparent when the jaundice faded and phototherapy was stopped. These data show that the colon plays a part in the pathogenesis of secretory diarrhoea and that both hyperbilirubinaemia and phototherapy are necessary for such an effect to develop.

Diarrhea, Infantile↗
PubMed1988

Unconjugated bilirubin and the bile from light exposed Gunn rats inhibit intestinal water and electrolyte absorption.

Jaundiced babies undergoing phototherapy often develop diarrhoea. The cause of it is still uncertain. Increasing evidence supports a role of a secretory mechanism for the diarrhoea. We therefore studied the effects of bile from congenitally jaundiced rats undergoing phototherapy and of unconjugated bilirubin on rat small intestine in vivo and in vitro. Results suggest that: (1) the bile from homozygous Gunn rats under phototherapy has an anti-absorptive effect when tested in the perfused jejunum of normal Wistar rats; (2) unconjugated bilirubin has a dose dependent secretory effect on the intestinal transport of water and electrolytes, when tested in the same system. Alteration of cyclic AMP or cyclic GMP, known intracellular mediators of secretion, was not observed. We conclude that free bilirubin is an intestinal secretagogue acting by an as yet unknown mechanism, that may mediate the secretory type of diarrhoea in jaundiced neonates undergoing phototherapy.

Animals↗
PubMed1988

Pathogenesis of postantibiotic diarrhoea caused by Clostridium difficile: an in vitro study in the rabbit intestine.

To elucidate the pathophysiological changes leading to postantibiotic diarrhoea caused by Clostridium difficile and its cytotoxin, oral ampicillin was given to rabbits, and jejunal, ileal, and caecal segments of those that developed diarrhoea were investigated in vitro. The rabbits that, in response to treatment, harboured Clostridium difficile in their colonic lumen were studied, and the results expressed according to the presence or absence of Clostridium difficile and/or its cytotoxin. Thus, we refer to either CD+ or CD- segments. The influx of glucose, phenylalanine, glycylphenylalanine, and lysine across the brush border of jejunum and ileum of CD+ segments was severely impaired, while only slightly blunted in CD-. No significant change was detected in the influx of glutamic acid in the jejunum of all treated animals and in the CD- ilea. Morphologic damage in ileum and caecum of CD+ was also more evident than in CD-. Transepithelial ion transport across short circuited ileal mucosa (CD+ and CD-) revealed secretory changes in Cl net transport that were more marked in CD-. We conclude that: (1) Clostridium difficile may also colonise the upper intestinal tract, where it induces morphological and functional damage, severely impairing nutrient absorption; and (2) the ileum contributes to the diarrhoea caused by CD even when the micro-organism is confined to the more distal gut by showing moderate impairment of nutrient absorption and marked electrolyte secretion.

Ampicillin↗
PubMedDate not supplied

Effects of berberine on basal and secretagogue-modified ion transport in the rabbit ileum in vitro.

The antisecretory effect of berberine sulfate (BS) was determined in the rabbit ileal mucosa in vitro mounted in Ussing chambers. When present in the luminal side, the drug had no effect on potential difference (PD) or on the short-circuit current (Isc) changes induced by theophylline or prostaglandin E2 (PGE2). When present in the serosal medium at a concentration of 2 mM, however, berberine reduced PD and Isc and abolished residual ion flux (bicarbonate secretion) but had no significant affect on basal net Na and Cl transepithelial transport. BS effect on basal PD was not affected by the absence of Ca2+ in the medium; neither was it inhibited by the alpha-adrenergic blocker yohimbine. Serosally added BS effectively antagonized the secretory effect on Isc of agents that increase each of the known intracellular mediators of ion secretion: cyclic 3',5'-adenosine monophosphate (cAMP), cyclic 3',5'-guanosine monophosphate (cGMP), and Ca2+; it also inhibited the ion transport modifications evoked by PGE2. The drug had no effect on the concentration of cyclic nucleotides either under basal condition or in the presence of some secretagogues. Finally, BS did not impair the PD rise due to luminal addition of glucose.

Animals↗