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Biomedical subjects

A A Lima

Publications and source records attributed to A A Lima.

69 records · Page 4Linked to original sources
PubMed1992

Persistent diarrhea in northeast Brazil: etiologies and interactions with malnutrition.

With the improved control of acute diarrheal illness mortality with oral rehydration therapy, persistent diarrhea is now emerging as a major cause of childhood mortality in tropical developing areas like the impoverished populations in Brazil's Northeast. "Graveyard surveillance" in the rural community of Guaiuba in northeastern Brazil revealed fully half of the 70% diarrhea mortality was due to persistent diarrheal illnesses. Furthermore, 11% of 14 or more diarrheal illnesses per child per year in an urban slum in Fortaleza persisted beyond 14 days, a definition that clearly identified the high risk children for heavy diarrhea burdens. Not only did heavy diarrhea burdens ablate the key "catch-up" growth seen in severely malnourished children and in children following previous diarrheal illnesses, but malnutrition significantly predisposed children to a greater incidence and duration of diarrhea as well as a greater incidence of persistent diarrhea. Etiologic studies of 37 children presenting with persistent diarrhea to Hospital das Clinicas in Fortaleza revealed that Cryptosporidium (in 13%) and enteroadherent E. coli (36% with aggregative, 29% with diffuse and 13% with localized adherence to HEp-2 cells) were the predominant potential pathogens found in the stool or upper small bowel. These findings suggest that persistent diarrhea is emerging as an important health problem in Brazil's Northeast, that it identifies a high risk child for heavy diarrhea burdens, that important interactions occur with malnutrition and that Cryptosporidium and enteroadherent E. coli warrant further study as potential etiologies of this major cause of morbidity and mortality.

Animals↗
PubMed1992

Pathogen and host differences in bacterial adherence to human buccal epithelial cells in a northeast Brazilian community.

The adherence of several strains of Escherichia coli to human buccal epithelial cells was studied, using cells obtained from five groups: healthy adults, healthy children, children with acute diarrhea, children with persistent diarrhea associated with cryptosporidial parasites, and children with noncryptosporidial persistent diarrhea. All groups lived or worked in an urban slum in northeastern Brazil. Samples of buccal epithelial cells from subjects in each of these groups were incubated with wild-type E. coli K-12 (strain C600), the enteroaggregative E. coli strains 17-2 and PDAS 30-5, CFA/II-positive E. coli 1392+ and its plasmid-cured derivative 1392-, and hydrophobic E. coli 132-3. Samples were evaluated microscopically to determine background contamination and the percentage of cells with more than 15% of their surface area obscured by adherent bacteria after incubation and washing. The assay was tested under field conditions and was shown to produce reliable and consistent results. Both enteroaggregative strains of E. coli were shown to adhere to a significantly higher percentage of all groups of human buccal epithelial cells than any of the other tested strains. In addition, buccal epithelial cells from children with nonparasitic persistent diarrhea showed substantially more bacterial adherence in both the native state and with all tested strains of E. coli than did cells from children with persistent cryptosporidial diarrhea or acute diarrhea or from healthy controls. This study provides evidence that enteroaggregative strains of E. coli demonstrate increased adherence to human buccal epithelial cells (as well as to cultured HEp-2 cells) and that buccal epithelial cells from children with noncryptosporidial persistent diarrhea appear to be more susceptible to bacterial adherence and colonization than buccal epithelial cells from control groups. These findings suggest that host differences as well as pathogen differences are important in the pathogenesis of persistent diarrhea and have implications for a potential role for buccal epithelial cell analysis in the diagnosis or risk stratification of children susceptible to noncryptosporidial persistent diarrhea.

Bacterial Adhesion↗
PubMed1992

Measurement of fecal lactoferrin as a marker of fecal leukocytes.

While diarrheal illnesses are extremely common in communities and hospitals throughout the world, an etiologic diagnosis may be expensive and cost-ineffective. Although the presence of fecal leukocytes are helpful in the diagnosis and specific therapy of inflammatory diarrheas, this requires prompt microscopic examination of fecal specimens (preferably obtained in a cup rather than a swab or diaper) by a trained observer. We developed a simple, sensitive test for the detection of leukocytes in fecal specimens using antilactoferrin antibody. Whereas radial immunodiffusion detected 0.02 micrograms of lactoferrin (LF) per microliter or greater than or equal to 2,000 leukocytes per microliter, latex agglutination (LA) readily detected greater than or equal to 0.001 micrograms of LF per microliter or greater than or equal to 200 leukocytes per microliter added to stool specimens. Despite the destruction or loss of morphologic leukocytes on storage for 1 to 7 days at 4 degrees C or placement of specimens on swabs, measurable LF remained stable. Initial studies of stool specimens from six patients with Salmonella or Clostridium difficile enteritis were positive and those from three controls were negative for LF by LA. Of 17 children in Brazil with inflammatory diarrhea (greater than or equal to 1 leukocyte per high-power field), 16 (94%) had LF titers of greater than 1:50 by LA, whereas only 3 of 12 fecal specimens with less than 1 leukocyte per high-power field on methylene blue examination and none of 7 normal control specimens had an LF titer of greater than 1:50 by LA. Of 16 fecal specimens from patients with C. difficile diarrhea (cytotoxin titers, >/= 1:1,000), 95% (n = 15) had detectable LF by LA (in titers of 1:100 to 1: 800). Finally, of 48 fecal specimens from healthy adult U.S. volunteers before and after experimental shigellosis and of 29 fecal specimens from children with documented shigellosis and hospitalized controls in northeastern Brazil, fecal LF titers ranged from 1:200 to >/= 1:5,000 in 96% (25 of 26) samples from patients with shigellosis (and reported positive for fecal leukocytes), while 51 controls consistently had fecal LF titers of </= 1:200. We conclude that fecal LF is a useful marker for fecal leukocytes, even when they are morphologically lost swab specimens or when they are destroyed on transport or storage or by cytotoxic fecal specimens.

Feces↗
PubMed1992

Effects of thermostable Escherichia coli enterotoxin peptide on the isolated rat kidney.

In order to compare the function of sodium transport between the intestine and renal tubule, we studied the effect of thermostable E. coli enterotoxin on rat kidneys. Isolated kidneys from adult male hooded rats weighing 240-335 g were perfused with Krebs-Henseleit solution containing 60 mg/ml dialyzed bovine serum albumin. The effects of E. coli enterotoxin (STa; molecular weight approximately 2000; 18 amino acids with three disulfide bonds) were studied on glomerular filtration rate (GFR), net urinary flow rate (UF) and fractional sodium reabsorption (% TNa+). All experiments were preceded by a 30-min control period, and in some kidneys the time course of the variables was followed without toxin infusion, for a paired control. STa (0.1 microgram/ml) infused into the perfusate 30 min after the beginning of the experimental period promoted a significant decline in % TNa+ from 78.4 +/- 1.6 (control period) to 51.6 +/- 6.8 (P < 0.001) 90 min after the administration of the toxin. This effect was followed by an increase in net urinary flow (UF) in toxin-treated kidneys (UFSTa = 0.120 +/- 0.009 vs UFcontrol = 0.056 +/- 0.011 ml g-1 min-1, P < 0.008). The GFR of control and STa-treated kidneys did not change during the total time of perfusion and after toxin infusion. Our data demonstrate that STa promotes a specific decrease in tubular sodium transport in the rat kidney.

Animals↗
PubMed1992

Cotransport of sodium with glutamine, alanine and glucose in the isolated rabbit ileal mucosa.

To investigate the effect of substrates during oral rehydration therapy, we studied intestinal cation cotransport (ICC) with glutamine (Gln), alanine (Ala) and glucose (Glu). The specific aims were to determine the biological effects of these three different cotransport systems on intestinal function. Isolated rabbit ileal mucosa preparations mounted in Ussing chambers were studied. ICC was determined by measuring short-circuit current (Isc) and potential difference (PD) while monitoring tissue resistance (TR). The data are reported as the mean +/- SEM of 4-6 experiments for each amino acid concentration. Increasing concentrations of Gln (10(-5) to 10(-2) M), Ala (10(-5) to 10(-1) M) and Glu (10(-5) to 10(-2) M) caused a significant (P < 0.05) increase in ICC. Gln (30 mM) and Ala (0.1 M) had a maximal effect (Em(Gln) = 100% and Em(Ala) = 66%, P < 0.05) which was higher than that obtained with 30 mM Glu (Em(Glu) = 35%). When sodium was replaced with choline on the mucosal side, Ringer solution completely abolished the response with Gln, Ala and Glu. The presence of all three substrates (10(-2) M Gln, 10(-1) M Ala, and 10(-2) M Glu) in Ringer solution on the mucosal side caused a significant increase in ICC (delta increase of short circuit current = 111 +/- 43 microA, P < 0.05). These results demonstrate that Gln, Ala and Glu each increased sodium-dependent cation cotransport, and that sodium-dependent intestinal cation cotransport was higher with Gln than with Ala or Glu.

Alanine↗
PubMed1992

Mathematical dose-response model for sodium cotransport with glucose and glutamine in the rabbit ileal mucosa.

Hyperbolic and allosteric mathematical equations are used extensively to describe cumulative dose-response curves in pharmacology. In 1954, using an axiom described by Clark, Ariëns suggested the following equation for cumulative dose-response curves: Em/E-1 = Kd/[D], where Em = maximum effect, E = effect, Kd = dissociation constant of the drug-receptor complex and [D] = drug concentration. To evaluate the validity of the modified Ariëns equation on the dose-response curves with a Hill coefficient statistically different from one, we used sodium intestinal cotransport (SIC) with glutamine (Gln) and glucose (Glu). Increasing doses (10 microM to 0.1 M) of Gln and Glu were added to the mucosal side of the isolated rabbit ileum mucosal membrane mounted on Ussing chambers. Sodium cotransport was recorded by measuring short-circuit current (Isc), potential difference (PD) and tissue resistance (TR) at 20-min intervals for 2 h. The Em and ED50 for SIC with glucose were 25% and 150 microM, respectively, using the method proposed by Lineweaver-Burk. The same method could not be used to estimate the Em and ED50 in SIC with glutamine, suggesting a specific limitation of the Gln cotransport kinetic curve in the equation. Therefore, we modified the Ariëns equation to Em/E-1 = (Kd/[D])p. With p = Log (S), Log (Kd) = -pD2 and Log[D] = -pDx, we obtain E = Em/(1 + S(pD x -pD2)), where S is a hyperbolic constant.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Transport of glutamine, alanine and glucose by rabbit intestinal membrane.

Malnutrition and dehydration are the immediate consequences of diarrheal diseases. To investigate the biological significance of glutamine, alanine and glucose in the intestinal mucosa, we have used Ussing chambers to determine electrolyte transport by measuring short-circuit current (Isc), potential difference (PD) and tissue resistance (TR) in rabbit intestinal mucosa. Increasing doses (10(-5) M to 10(-1) M) of glutamine, alanine and glucose cause a significant increase in intestinal cation cotransport. Although glucose had a slightly earlier effect, 30 mM glutamine and 0.1 M alanine had a maximal effect which was more than two times that caused by 30 mM glucose. The pD2 values for glucose, glutamine and alanine were 3.0, 2.5, and 2.0, respectively. The dose-response curves of these substrates suggest that the intestinal cotransport kinetics for glutamine is different from that of glucose and alanine. Our results demonstrate that all three substrates cause a significant increase in Isc or PD, suggesting an increase in the intestinal mucosa cation cotransport. Glutamine has a larger effect on cation cotransport than alanine and glucose. These combinations should be studied further for the development of an oral rehydrating solution for diarrhea treatment which could prevent the resulting malnutrition, especially in those cases of prolonged diarrheal diseases.

Alanine↗
PubMed1989

Clostridium difficile toxin A. Interactions with mucus and early sequential histopathologic effects in rabbit small intestine.

Clostridium difficile produces two toxins, A (enterotoxic) and B (cytopathic), that are implicated in the pathogenesis of pseudomembranous colitis. However, the relationship of the secretory effect and the early histopathologic events is still unclear. We examined the early histopathologic effect of purified C. difficile toxin A in rabbit ileal loops and correlated the mucosal damage with the secretory response. As early as 2 hours after inoculation, toxin A at 1.0 micrograms caused cytolysis of the basal portion of the apical epithelia cells. Electron microscopy showed the basal portions of the absorptive epithelial cells necrotic. No significant fluid accumulation occurred after 2 or 4 hours in ileal loops inoculated with either 0.1 or 1.0 micrograms toxin A. In contrast, with 3-10 micrograms toxin A there was significant hemorrhagic fluid accumulation after 2 hours that correlated with the severity of the histologic lesions. An early mucoid exudate was demonstrated not to be related to a mucus secretagogue effect of toxin A. Prior exposure of toxin A to rabbit intestinal mucus preparations did not alter its subsequent enterotoxic or cytopathic (Chinese hamster ovary cells) effects. These studies demonstrate that the early intestinal tissue damage by toxin A was dose and time dependent and involved disruption of the basal portions of the absorptive epithelial cells with marked separation of the cells along the mucosal surfaces that preceded the secretory response.

Animals↗
PubMed1988

Effects of Clostridium difficile toxins A and B in rabbit small and large intestine in vivo and on cultured cells in vitro.

Clostridium difficile is recognized as the major cause of antibiotic-associated colitis. C. difficile produces two toxins, A (enterotoxin) and B (cytotoxin), that are implicated in the pathogenesis of the colitis. We examined the dose responses, time course, and synergism of these two toxins in ligated rabbit intestinal loops and in tissue culture. In rabbit small intestinal loops, toxin A caused histologically demonstrable intestinal tissue damage as early as 2 h. The secretory response greater than or equal to 8 h was similar to that of a cholera toxin control. The effect of toxin A on tissue damage or secretion was seen even if toxin was removed after 5 min. Purified toxin A caused significant net accumulation of sodium, chloride, potassium, and total protein and slightly increased osmolality of the fluid content at 6 h; these effects were similar to those caused by crude C. difficile culture filtrates containing toxins A and B. Crude C. difficile toxin caused fluid accumulation with a delayed time course in the rabbit large intestine, and in contrast to its effect in small intestine, crude toxin caused net accumulation of bicarbonate and increased pH. In tissue culture, toxin A caused a rounding up of CHO and T-84 colonic carcinoma cells. A monoclonal antibody (PCG-4) that has no effect on tissue culture cytotoxicity with toxins A and B completely inhibited the secretory and tissue-damaging effects in the intestine. Toxins A and B were synergistic in the gut only at high doses of toxin B (greater than or equal to 10 micrograms/ml), and they were additive in tissue culture. The cytopathic effect in tissue culture was not consistently associated with trypan blue uptake. The cytopathic effect of toxin A in tissue culture did not appear to involve inhibitable Ca2+-dependent or prostaglandin synthesis pathways or intact microfilament or microtubule function for its activity and was not inhibited by reducing or lysosomotropic agents. Our results suggest that toxins A and B have independent and distinct effects in vivo and in vitro.

Animals↗
PubMed1987

Production of a unique cytotoxin by Campylobacter jejuni.

Campylobacter jejuni is an important diarrheal pathogen worldwide; the mechanisms by which it causes disease remain unclear. Because of its association with inflammatory diarrhea, we postulated that C. jejuni might produce a cytotoxin similar to that produced by Shigella sp., enterohemorrhagic Escherichia coli O157, or Clostridium difficile. Filtrates of 12 polymyxin-treated isolates of C. jejuni were placed on HeLa cells (sensitive to Shiga toxin cytotoxicity) and Chinese hamster ovary (CHO) cells. Of 12 isolates of C. jejuni tested, 5 killed 50% of the cells at greater than or equal to 1:4 dilutions of filtered suspensions of 10(9) bacteria per ml; killing was similar in HeLa and CHO cells (the CHO cells being insensitive to Shiga cytotoxin). One isolate produced a titer of 1:32 to 1:128. The relative potency in HeLa cells was comparable to that of E. coli strains that produce intermediate amounts of Shiga-like toxin. The other seven strains showed no cytotoxic effect, nor did the control diluents, polymyxin B, or supernatants of C. jejuni not treated with polymyxin B. Sonication also released active cytotoxin, but slightly less well than did polymyxin. The cytotoxic effect was dose dependent. Concentration of the C. jejuni in suspension by 10-fold before treatment with polymyxin B resulted in a 10-fold increase in the 50% cytotoxic dose. The cytotoxin effect was not neutralized by Shiga toxin immune serum against either Shiga-like toxin I or II or by anti-Clostridium difficile antiserum. The C jejuni cytotoxin was partially labile to trypsin (0.25%) and to heating to greater than or equal to 60 degrees C. Cytotoxicity was retained in Scientific Products dialysis tubing D1615-1 (Mr cutoff, 12,000 to 14,000). Some isolates of C. jejuni release a substance lethal to HeLa or CHO cells in vitro that is distinct from Shiga-like or Clostridium difficile toxin. This cytotoxin may contribute to the colonic mucosal invasive process that characterizes C. jejuni enteritis.

Animals↗
PubMed1986

Production of type II heat-labile enterotoxin by Escherichia coli isolated from food and human feces.

Escherichia coli strains isolated in Sao Paulo, Brazil, from feces of patients with diarrhea and from food samples produced toxin(s) that was shown to be related both immunologically and genetically to the recently characterized type II heat-labile enterotoxin of E. coli. The new isolates of type II heat-labile enterotoxin-producing E. coli belonged to five different serotypes and did not represent a single clone.

Bacterial Toxins↗
PubMedDate not supplied

Absence of intestinal secretion on supernatants from macrophages stimulated with Clostridium difficile toxin B on rabbit ileum.

Several studies have documented the involvement of both Clostridium difficile, toxins, A and B in the pathogenesis of antibiotic-associated diarrhea. Recently, we demonstrated that IL-1 beta is the intestinal secretory factor released by macrophages stimulated with toxin A. The aim of this study was to evaluate the importance of macrophages stimulated with toxin B on rabbit ileal ion transport. The changes in ion transport were analyzed by studying the short-circuit current of the rabbit ileal mucosa mounted in Ussing chambers. The supernatants of macrophages treated with toxin B (3.6 x 10(-7) M) had no effect on the ion transport (change in short-circuit current =28.0+/-9.2 vs. control=26.8+/-3.6 microA cm(-2)). Supernatants of macrophages stimulated with toxin A (3.2 x 10(-7) M), our positive control, induced a significant change in ileal ion transport (delta I(sc)=55.2+/-5.7 mA cm(-2)). It was also observed that, like toxin A, toxin B stimulated macrophages to produce TNF-alpha (555.0+/-37.9 pg/ml vs. control=182.0+/-39.8 pg/ml; p<0.05). Nevertheless, in contrast to toxin A, toxin B did not stimulate IL-1 beta synthesis (28.0+/-7.5 pg/ml vs. control=40. 0+/-14.4 pg/ml; p>0.05). We conclude that the supernatants of macrophages stimulated with toxin B are not able to stimulate ion transport and that both toxins stimulate the genesis of TNF-alpha, but only toxin A induces the synthesis of IL-1 beta, which, we have earlier reported, causes an electrogenic intestinal response in rabbit ileum.

Animals↗
PubMedDate not supplied

[Multiple antibiotic resistance mediated by R plasmid in Shigella flexneri strains isolated in the northeast of Brazil].

In Shigella strains were studied the molecular mechanism that mediated the multiply antibiotic-resistance. Twenty-six strains of Shigella flexneri were utilised in this investigation. These strains were submitted to disk diffusion test, mating experiments and plasmid isolation. In relation to antibiotics resistance standard it was observed that all Shigella flexneri strains were resistant to at least, three antibiotics tested. From twenty-six Shigella flexneri strains donors submitted to conjugation process, 34.6% (nine strains) resulted in variable frequency of transconjugants. From strains that conjugated, 100%, transferred the resistance factor acquainted with ampicillin. Being that, in all transconjugants which were observed, just one plasmid with 23.1 Kb was evidenced. This plasmid found in all strains was characterised as the cause of resistance to ampicillin.

Ampicillin Resistance↗
PubMedDate not supplied

[Clostridium difficile as an inducer of inflammatory diarrhea].

Clostridium difficile has been pointed out as an important agent of diarrheal diseases associated with antibiotic use. However, due to its complexity, the physiopathology of these diseases is only partially elucidated, although a series of scientific works has demonstrated the importance of toxins A and B in the pathogenesis of the inflammatory diarrhea induced by this microorganism. The inflammatory mechanisms involved in the biological activities of these toxins are complex. There are some studies demonstrating that toxin B has no enterotoxic activity in vivo. However, this toxin causes dose-dependent eletrophysiologic and morphologic modifications of human colonic mucosa in vitro. In addition, toxin B stimulates the synthesis of potent inflammatory mediators by monocytes and macrophages. The effects provoked by toxin A on the intestinal mucosa are quite evident and are characterized by intense fluid secretion and by inflammatory cell accumulation, such as macrophages, mast cells, lymphocytes and neutrophils, with the consequent release of mediators such as prostaglandins, leukotrienes, platelet activating factor, nitric oxide and cytokines.

Animals↗