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Biomedical subjects

S Grinstein

Publications and source records attributed to S Grinstein.

At least 343 records · Page 19Linked to original sources
PubMed1981

Isolation of chromaffin cell plasma membranes on polycationic beads.

We have tried to define which proteins of chromaffin cell plasma membranes are facing the cytoplasm by surface labelling a selectively oriented membrane preparation. Viable chromaffin cells were isolated by collagenase treatment of bovine adrenals. Plasma membranes from these cells were isolated on polycationic beads by the method of Jacobson and Branton (Jacobson, B.S. and Branton, D. (1977) Science 195, 302--304). The purity and orientation of the membranes were defined by biochemical and morphological criteria. The membranes, with their external side apposed to the bead surface, were enriched about 10-fold with respect to a whole cell homogenate, and contained only small amounts of contaminating organelles. Surface specific iodination of membranes on beads with 1,3,4,6-tetrachloro-3 alpha, 6 alpha-diphenylglycoluril (Iodogen), followed by polyacrylamide gel electrophoresis, allowed the identification of cytoplasmically exposed proteins. A different pattern was observed when intact cells were labelled prior to membrane isolation. The advantages and possible uses of this immobilized membrane preparation are discussed.

Adrenal Glands↗
PubMed1981

Infections in children with malignant disease in Argentina.

During six-month period, 102 consecutive episodes of fever in 68 children (ranging from 1 month to 14 years of age) with malignant diseases were prospectively evaluated. Sixty-five had acute lymphoblastic leukemia, nine had acute myeloblastic leukemia, nine had malignant lymphoma (four Hodgkin and five non-Hodgkin), five had chronic myeloid leukemia, four had rhabdomyosarcoma, three had CNS tumors, two had neuroblastoma, one had Wilms, and four had other malignant tumors. Forty cases (39.2%) showed severe neutropenia (500 neutrophil/m3) during the episode. S. aureus, E. coli, and S. pyogenes were in 53% of the 75 microbiologic isolates. Twenty-two percent of the viral studies were positive. Mycologic studies were all negative, except one case with C. Albicans. Pneumonia (33 cases), cellulitis (15 cases), pharyngitis (12 cases), and varicella (11 cases) were the most common final diagnosis. Seventy-one percent of the episodes were etiologically documented (by bacterial isolate, characteristic serology, and/or typical clinic picture); 19% of the febrile episodes were probable infections, and 10% were fever of uncertain cause. Ninety percent of the cases responded well to therapy, and mortality of this series was 7%. Gentamicin, Carbenicillin, and Methicilin were the more common antibiotics employed. We conclude that in our population 1) infection is a frequent cause of morbidity in children with malignant diseases; 2) the most common cause of the febrile episodes is bacterial infection; 3) S. aureus, E. coli and S. pyrogenes are the most frequent bacterial isolates, and P. aeruginosa is infrequent; 4)viral infections are relatively frequent in this group of children; and 5) with adequate management, the mortality is low.

Adolescent↗
PubMed1981

Duration of the immune response in subjects inoculated with antimeningococcal A and C vaccines kept in storage at -20 degrees C and at 4 degrees C: influence of pre-vaccination titres on the vaccinal response.

The antibody titres in 250 subjects, aged 5 to 22 years, who were vaccinated with a mannitol-lyophilized antimeningococcal A + C vaccine, stable only when stored at -20 degrees C, were followed for two years. As measured by indirect haemagglutination (IHA) and indirect immunofluorescence (IF) techniques, titres for both A and C Neisseria meningitidis antibodies remained high. Two years after vaccination titres of antibodies against type A showed fourfold increase over the initial titres in from 46% to 100% of groups of subjects and against type C in from 42% to 80%. For 130 subjects vaccinated with a new lactose-lyophilized antimeningcoccal A + C vaccine (presumed stable at 4 degrees C) antibody titres were measured up to 16 months after inoculation with this vaccine stored at -20 degrees C and also after storage for several periods at 4 degrees C. Antibody titres in all these subjects had fallen to their initial titres by 16 months. The importance of evaluating the results on subjects showing low initial titres (less than or equal to 1/8 as measured by IHA) is discussed, as inclusion of high initial titres influences the extent of the response.

Adolescent↗
PubMed1981

Rotavirus infection in children hospitalized for diarrhoea in Argentina.

An epidemiological survey was undertaken to establish the importance of rotavirus as a cause of admission of children to hospital in Buenos Aires, Argentina. Between August 1978 and January 1980 rotavirus infection was detected in 62 (34%) of 180 infants and children admitted to hospital with acute gastro-enteritis. In identification of the virus, electron microscopy (EM) was found to be as sensitive as counterimmunoelectrophoresis (CIE). There was agreement between EM and CIE diagnosis in 90.5% cases. Adenovirus and coronavirus were detected by electron microscopy less often. A serological response to rotavirus was revealed by complement fixation test in 38 (72%) of 53 infected patients. Rotaviruses were detected throughout the year, but peak incidence occurred during the cooler months, May to August. Infection was commoner among children under six months of age. Diarrhoea was generally mild in these children and respiratory illness showed no significant association with rotavirus gastroenteritis. These results indicate that rotaviruses may be a major cause of infantile acute gastroenteritis in Buenos Aires, Argentina especially during the winter.

Argentina↗
PubMed1980

Intrinsic segments of band 3 that are associated with anion transport across red blood cell membranes.

After treatment of red cell ghosts with chymotrypsin, the predominant intrinsic peptides remaining in the membrane fraction are 15,000 and 9,000 daltons mol wt. After partial extraction with Triton X-100, the residual membrane vesicles have almost no other stained peptides and such vesicles are reported to carry out anion transport activities sensitive to specific inhibitors. In vesicles derived from cells treated with DIDS(4,4'-diisothiocyano-2,2'-stilbene disulfonic acid), an irreversible inhibitor of anion transport that is highly localized in an abundant intrinsic protein known as band 3, the probe is largely recovered in the 15,000 dalton peptide. The part of band 3 from which it is derived is a previously reported 17,000 transmembrane segment (Steck, T.L., Ramos, R., Strapazon, E., 1976, Biochemistry 15:1154). The 9,000-dalton peptide is present in the vesicles in a one-to-one mole ratio with the 15,000-dalton peptide, suggesting that both are derived from the same protein. This conclusion is supported by the finding that the 35,000-dalton C-terminal end of band 3, derived by chymotrypsin treatment of cells, is further proteolysed if the cells are converted to ghosts and its disappearance coincides with the appearance of the 9,000-dalton fragment. Evidence is presented that the 9,000-dalton fragment crosses the bilayer and that it is closely associated with the 15,000-dalton peptide.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1980

Interaction of the sugar carrier of intestinal brush-border membranes with HgCl2.

HgCl2 was used as an inhibitor and potential label for the glucose carrier of intestinal brush-border membranes. Half-maximal inhibition of Na+-dependent D-glucose uptake was reached with micromolar concentrations of HgCl2 when the protein concentration was 1.2 mg/ml. Similar concentrations were found to inhibit the binding of [3H]phlorizin, a reversible competitive inhibitor of sugar transport. Inhibition was reversed by dithioerythritol but only marginally by EDTA. The data support the involvement of a sulfhydryl group in the inhibitory process. Deoxycholate-extracted membranes, which are enriched in specific phlorizin binding activity, were used for labeling studies using 203HgCl2. The polypeptides were separated by gel electrophoresis and analyzed by protein staining and autoradiography. Non-specific 203HgCl2 labeling was minimized by pre-treatment with sulfhydryl reagents which do not inhibit phlorizin binding. Several bands, which are lost from the autoradiographic pattern during a negative purification of the phlorizin binding sites, could be ruled out as essential components of the sugar carrier. The polypeptide profile was also analyzed following proteolysis, which abolished phlorizin binding. Those radioactive bands of which apparent Mr values were alterd by the treatment were considered as possible candidates. Finally, samples in which inhibition was reversed by thiols were also studied. The possible identity of the polypeptide(s) involved in glucose translocation is disussed in the light of these observations.

Animals↗
PubMed1980

Inorganic anion transport in kidney and intestinal brush border and basolateral membranes.

The efflux of inorganic anions from purified brush border and basolateral membrane vesicles from dog kidney cortex was measured under equilibrium exchange conditions. Marked differences in temperature sensitivity and effects of inhibitors were found between the Cl and SO4 transport pathways and between the two types of membranes. SO4 transport in both brush border and basolateral membranes was markedly reduced by cooling, but significant inhibition by 4,4'-diisothiocyano-2,2'-disulfonic stilbene (DIDS) was only observed in basolateral vesicles. In contrast, Cl efflux from both types of vesicles was neither substantially inhibited by DIDS nor by lowering the temperature to 0 degrees C. Phosphate efflux from basolateral membrane vesicles was found to be only partially sensitive to DIDS. Attempts to label the stilbene-sensitive SO4 pathway in basolateral vesicles using [3H2]DIDS as a marker were unsuccessful due to the nonspecific labeling of many membrane components. The asymmetry in inorganic anion transport behavior exhibited by brush border and basolateral membrane vesicles from dog renal proximal tubule was also observed in equivalent vesicles prepared from rat small intestine.

Alkaline Phosphatase↗
PubMed1980

Rotavirus in Buenos Aires, Argentina.

Rotavirus was studied over a 1-year period in 141 hospitalized children suffering from gastroenteritis and in 120 control children. Electron microscopy and discontinuous counterimmunoelectrophoresis techniques were employed with equal success. Rotavirus was detected with high frequency in children suffering from diarrhea (45 cases; 32%) and in only 2 cases (1.6%) of the control group. A marked seasonal distribution, with 80% in July (winter) and a decrease (0%) in December (summer), was observed. The frequency of infection with rotavirus was highest in infants between 0 and 6 months of age. Seroconversion, studied by complement fixation, was obtained in 34 of 42 (80%) virus-positive children, and antibody without conversion was observed in 8 (19%). Of 58 virus-negative children, 51 (96.2%) were negative for antibody and only 2 (3.7%) developed antibody.

Antibodies, Viral↗
PubMed1979

Partial purification of the sugar carrier of intestinal brush border membranes. Enrichment of the phlorizin-binding component by selective extractions.

The [3H] phlorizin-binding component of brush border vesicles was enriched in situ by negative purification. Several procedures, known to effect selective solubilization of membrane components, were used separately or in combination to remove proteins unrelated to the binding. Deoxycholate ruptured the vesicles and released 67% of their protein, thereby increasing the specific [3H] phlorizin-binding activity of the pellet three-to fourfold. Extracting the deoxycholate-pellets with either NaI or alkaline solutions released up to 38% of the deoxycholate-insoluble protein without significantly affecting phlorizin binding. The polypeptide composition of the membranes at the different stages was analyzed by NaDodSO4-polyacrylamide gel electrophoresis. A number of polypeptides present in the original vesicles could be ruled out as essential components of the [3H] phlorizin binding entity. Intact and deoxycholate-treated vesicles were subjected to proteolytic attack. Papain liberated sucrase and isomaltase from intact vesicles, but affected neither other Coomassie-stained bands nor phlorizin binding. Neither the protein composition nor the binding properties of sealed vesicles were influenced by trypsin or chymotrypsin. However, all the proteolytic enzymes tested on deoxycholate-treated membranes substantially reduced [3H] phlorizin binding and produced concomitantly the disappearance of several bands from the electrophoretic profile. Pretreatment of vesicles with papain, followed by deoxycholate extraction and incubation in alkaline media, increased the specific binding activity of the membranes up to ninefold by removing close to 90% of the protein. A limited number of polypeptides are suggested as possible candidates for the glycoside-binding site of intestinal brush borders.

Animals↗
PubMed1979

Distribution of sulfhydryl groups in intestinal brush border membranes. Localization of side-chains essential for glucose transport and phlorizin binding.

1. Brush border membrane vesicles from rabbit small intestine were found to contain 46 nmol SH groups/mg protein, 52% of which could react with 4,4'-dithiodipyridine, a membrane permeating probe. Only 18% of the total SH-groups reacted with the impermeant probe 5,5'-dithiobis(2-nitrobenzoic acid), indicating that only this fraction is externally located. 2. Brush border membrane vesicles could be disrupted by a gentle treatment with deoxycholate, releasing most of their electron-dense core material. In deoxycholate-treated vesicles most of the SH groups that reacted with 4,4'-dithiodipyridine react with 5,5'-dibiobis(2-nitrobenzoic acid), suggesting that both membrane surfaces became exposed to the extravesicular medium. 3. In intact vesicles (1.2 mg protein/ml), the binding of phlorizin (a competitive inhibitor of the monosaccharide transport system) was 50% inhibited by 67 microM of the penetrating organomercurial p-chloromercuribenzoate, but was about ten times less sensitive to the poorly permeating p-chloromercuriphenylsulfonate. In contrast, binding of phlorizin to leaky (deoxycholate-treated) membranes was equally susceptible to either reagent. 4. Mercurial inhibition of phlorizin binding could be reversed by dithioerythritol in both sealed and leaky membranes, whereas the less permeant thiol L-glutathione (reduced form) could only revert the inhibition in leaky membranes.

Animals↗
PubMed1979

Transmembrane effects of irreversible inhibitors of anion transport in red blood cells. Evidence for mobile transport sites.

Experiments were designed to determine whether band 3, the anion transport protein of the red cell membrane, contains a mobile element that acts as a carrier to move the anions across a permeability barrier. The transport site-specific, nonpenetrating irreversible inhibitor 4,4'-diisothiocyano-2,2'-stilbene disulfonate (DIDS) was found to be effective only when applied extracellularly. It was used to sequester transport sites on the extracellular side of the membrane in intact cells. The membranes were then coverted into inside-out vesicles. The number of anion transport sites available on the cytoplasmic side of the vesicle membranes was then estimated by measuring the binding of N-(-4-azido-2-nitrophenyl)-2-aminoethyl-sulfonate (NAP-taurine), a photoreactive probe. Pretreatment with DIDS from the extracullular side substantially reduced the binding of NAP-taurine at the cytoplasmic side. Since NAP-taurine does not appear to penetrate into the intravesicular (normally extracellular) space, a transmembrane effect is apparently involved. About 70% of the DIDS-sensitive NAP-taurine binding sites are located in band 3, with the remainder largely in a lower molecular weight (band 4) region. A similar pattern of reduction in NAP-taurine binding is produced by high concentrations of Cl-, but this anion has little or no effect in vesicles from cells pretreated with DIDS. Thus the DIDS-modulated sites seem to be capable of binding either NAP-taurine or Cl. It is suggested that band 3 contains a mobile transport element that can be recruited to the extracellular surface by DIDS, thus becoming unavailable to NAP-taurine at the cytoplasmic face of the membrane. The results are consistent with a model of carrier-mediated transport in which the movement of the transport site is associated with a local conformational change in band 3 protein.

Binding Sites↗