Biomedical subjects

E Altman

Publications and source records attributed to E Altman.

At least 37 records · Page 2Linked to original sources

PubMed2001

Galectin-3-mediated adherence of Proteus mirabilis to Madin-Darby canine kidney cells.

Proteus mirabilis is an important cause of urinary tract infections (UTIs) and can result in acute pyelonephritis. Proteus mirabilis expresses several, morphologically distinct, fimbrial species, and previous studies have shown that the nonagglutinating fimbriae (NAF) mediate bacterial adherence to a number of cell lines, including Madin-Darby canine kidney (MDCK) cells. Immunoblot overlay analysis of the plasma membrane fraction from MDCK cells with purified NAF revealed a 34-kDa band, which has been analyzed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). Database search identified galectin-3 as a potential protein candidate. Immunocytochemical assay of MDCK cells with a galectin-3-specific monoclonal antibody, anti-Mac-2, confirmed its presence on the plasma membrane extracellular surface. Preincubation of P. mirabilis with anti-Mac-2 monoclonal antibodies, specific for galectin-3, resulted in the inhibition of bacterial binding to MDCK cells. These data suggest a role for galectin-3, interacting with appropriately glycosylated surface receptors and P. mirabilis fimbriae, as a mediator of bacterial adherence in vitro.

Animals↗

PubMed2000

Andreev peaks and massive magnons in cuprate superconductor-normal-superconductor junctions

The projected SO(5) theory [pSO(5)] is used to resolve the puzzle of two distinct energy gaps in high T(c) superconductor-normal-superconductor junctions. Counter to the conventional theory of multiple Andreev reflections (MAR), the differential resistance peaks are associated with the antiferromagnetic resonance observed in neutron scattering, and not with Cooper pair breaking. The pSO(5) and MAR theories differ by the expected tunneling charges at the peaks. We propose that shot noise experiments could discriminate against the conventional interpretation.

Journal Article↗

PubMed2000

Construction and characterization of a highly regulable expression vector, pLAC11, and its multipurpose derivatives, pLAC22 and pLAC33.

A number of different expression vectors have been developed to facilitate the regulated overproduction of proteins in Escherichia coli and related bacteria. Some of the more popular ones include pKK223-3, pKK233-2, pTrc99A, and the pET family of expression vectors. These vectors were designed to be regulable and can be grown under conditions that repress protein production or under conditions that induce protein production. Unfortunately, however, numerous researchers have found that these vectors produce significant amounts of protein even when grown under repressed conditions. We describe here a new expression vector, pLAC11, which was designed to be more regulable and thus more tightly repressible when grown under repressed conditions. The tight regulation of pLAC11 was achieved by utilizing the O3 auxiliary operator, CAP binding site, promoter, and O1 operator that occur in the wild-type lac control region. The pLAC11 vector can be used to conduct physiologically relevant studies in which the cloned gene is expressed at levels comparable to that obtainable from the chromosomal copy of the gene in question. In experiments in which a bacterial cell contained both a null allele in the chromosome and a second copy of the wild-type allele on pLAC11, we observed that cells grown under repressed conditions exhibited the null phenotype while cells grown under induced conditions exhibited the wild-type phenotype. Two multipurpose derivatives of pLAC11, pLAC22, and pLAC33 have also been constructed to fulfill different experimental needs.

Base Sequence↗

PubMed2000

The ABC rule for clinical detection of subungual melanoma.

BACKGROUND: Subungual melanoma is a relatively rare disease with reported incidence between 0.7% to 3.5% of all melanoma cases in the general population. Unlike the significant improvement in the diagnosis of cutaneous melanoma, the diagnosis of subungual melanoma has shown little, if any, improvement over the years. The widespread adoption of the ABCDs of cutaneous melanoma has helped increase public and physician awareness, and thus helped increase the early detection of cutaneous melanoma; the same criteria cannot be applied to the examination of the nail pigmentation. OBJECTIVE: We reviewed the world literature on subungual melanoma and arranged the available information into a system for the identification of subungual melanoma. This system has to be thorough, easy to remember, and easy to apply by both physician and lay public. A case to illustrate the delayed diagnosis often encountered in the current evaluation of nail melanoma is presented. METHODS: A thorough review of the world literature on subungual melanoma was undertaken. The important findings of various studies and case reports were compared among themselves and the salient features were summarized. The information was then categorized under the easily recalled letters of the alphabet, ABCD, that have already become associated with melanoma. RESULTS: The most salient features of subungual melanoma can be summarized according to the newly devised criteria that may be categorized under the first letters of the alphabet, namely ABCDEF of subungual melanoma. In this system A stands for a ge (peak incidence being in the 5th to 7th decades of life and African Americans, Asians, and native Americans in whom subungual melanoma accounts for up to one third of all melanoma cases. B stands for brown to black b and with breadth of 3 mm or more and variegated borders. C stands for change in the nail band or lack of change in the nail morphology despite, presumably, adequate treatment. D stands for the digit most commonly involved; E stands for extension of the pigment onto the proximal and/or lateral nailfold (ie, Hutchinson's sign); and F stands for family or personal history of dysplastic nevus or melanoma. CONCLUSION: Although each letter of the alphabet of subungual melanoma is important, one must use all the letters together to improve early detection and thus survival of subungual melanoma. Still, as with cutaneous melanoma, the absolute diagnosis of subungual melanoma is made by means of a biopsy.

Adult↗

PubMed2000

Functional genomics of Helicobacter pylori: identification of a beta-1,4 galactosyltransferase and generation of mutants with altered lipopolysaccharide.

A previously annotated open reading frame (ORF) (HP0826) from Helicobacter pylori was cloned and expressed in Escherichia coli cells and determined to be a beta-1,4-galactosyltransferase that used GlcNAc as an acceptor. Mutational analysis in H. pylori strains demonstrated that this enzyme plays a key role in the biosynthesis of the type 2 N-acetyl-lactosamine (LacNAc) polysaccharide O-chain backbone, by catalysing the addition of Gal to GlcNAc. To examine the potential role of this O-chain structure in bacterial colonization of the host stomach, the mutation was introduced into H. pylori strain SS1 which is known to be capable of colonizing the gastric mucosa of mice. Compared with the parental strain, mutated SS1 was less efficient at colonizing the murine stomach.

Alleles↗

PubMed2000

Structural characterization of the outer core and the O-chain linkage region of lipopolysaccharide from Pseudomonas aeruginosa serotype O5.

The point of attachment of the O-chain in the outer core region of Pseudomonas aeruginosa serotype O5 lipopolysaccharide (LPS) was determined following a detailed analysis of the extended core oligosaccharide, containing one trisaccharide O-chain repeating unit, present in both the wild-type strain PAO1 and O-chain deficient mutant strains AK1401 and PAO-rfc. The structure of the extended core oligosaccharide was determined by various mass spectrometric methods as well as one-dimensional and two-dimensional NMR spectroscopy. Furthermore, the one-dimensional analogues of NOESY and TOCSY experiments were applied to confirm the structure of the outer core region in the O-chain polysaccharide. In both the extended core oligosaccharide and the core of the smooth LPS, a loss of one of the beta-glucosyl residues and the translocation of the alpha-rhamnosyl residue, followed by the attachment of the first O-chain repeating unit was observed. This process is complicated and could involve two distinct rhamnosyltransferases, one with alpha-1, 6-linkage specificity and another with alpha-1,3-linkage specificity. It is also plausible that an alpha-1,3 rhamnosyltransferase facilitates the addition of the 'new' alpha-rhamnosyl residue that will act as a receptor for the attachment of the single O-antigen repeating unit in the LPS of the semi-rough mutant. The 2-amino-2-deoxy-fucosyl residue of the first O-chain repeating unit directly attached to the core was found to have a beta-anomeric configuration instead of an alpha configuration, characteristic for this residue as a component of the O-chain polysaccharide. The results of this study provide the first example of the mechanistic implications of the structure of the outer core region in a fully assembled O-chain containing LPS, differing from the O-chain deficient rough LPS.

Bacterial Capsules↗

PubMed2000

Metabolic analysis of Escherichia coli in the presence and absence of the carboxylating enzymes phosphoenolpyruvate carboxylase and pyruvate carboxylase.

Fermentation patterns of Escherichia coli with and without the phosphoenolpyruvate carboxylase (PPC) and pyruvate carboxylase (PYC) enzymes were compared under anaerobic conditions with glucose as a carbon source. Time profiles of glucose and fermentation product concentrations were determined and used to calculate metabolic fluxes through central carbon pathways during exponential cell growth. The presence of the Rhizobium etli pyc gene in E. coli (JCL1242/pTrc99A-pyc) restored the succinate producing ability of E. coli ppc null mutants (JCL1242), with PYC competing favorably with both pyruvate formate lyase and lactate dehydrogenase. Succinate formation was slightly greater by JCL1242/pTrc99A-pyc than by cells which overproduced PPC (JCL1242/pPC201, ppc(+)), even though PPC activity in cell extracts of JCL1242/pPC201 (ppc(+)) was 40-fold greater than PYC activity in extracts of JCL1242/pTrc99a-pyc. Flux calculations indicate that during anaerobic metabolism the pyc(+) strain had a 34% greater specific glucose consumption rate, a 37% greater specific rate of ATP formation, and a 6% greater specific growth rate compared to the ppc(+) strain. In light of the important position of pyruvate at the juncture of NADH-generating pathways and NADH-dissimilating branches, the results show that when PPC or PYC is expressed, the metabolic network adapts by altering the flux to lactate and the molar ratio of ethanol to acetate formation.

Anaerobiosis↗

PubMed2000

[Treatment of sputum retention by minitracheostomy].

Maintenance of bronchopulmonary hygiene is mandatory for preventing complications of respiratory therapy in the hospitalized patient. Removal of secretions from the tracheobronchial tree is crucial. Conventional therapy, designed to assist in dislodging airway secretions, includes chest physical therapy, incentive spirometry, transnasal endotracheal suctioning and bronchoscopy. Minitracheostomy was first described by Matthews and Hopkinson for recurrent endotracheal suctioning in 1984. Since then there have been few papers about it, but they report good results with low morbidity. Despite this, its use is not popular in routine clinical work. We report our experience with minitracheostomy in the prevention of sputum retention. We conclude that its use is easy, safe and very effective in preventing postoperative and post-traumatic respiratory complications.

Adult↗

PubMed2000

The binding of Proteus mirabilis nonagglutinating fimbriae to ganglio-series asialoglycolipids and lactosyl ceramide.

Proteus mirabilis is a common opportunistic Gram-negative uropathogen that infects the upper urinary tract. We have examined the role of the nonagglutinating fimbriae (NAF) of P. mirabilis in mediating bacterial adhesion to cell surface receptors. Purified NAF of P. mirabilis were demonstrated to bind to a number of glycolipids, including asialo-GM1, asialo-GM2, and lactosyl ceramide (LacCer) in solid-phase binding assays and in thin layer chromatography (TLC) overlay assays. Furthermore, preincubation of the biotinylated NAF (Bt-NAF) with anti-NAF monoclonal antibodies resulted in inhibition of NAF binding to immobilized asialo-GM1, asialo-GM2, and LacCer. In adherence assays, P. mirabilis binding to Madin-Darby canine kidney (MDCK) cells was inhibited by murine anti-asialo-GM1 monoclonal antibodies H2G10 to about 50% of the binding level in the absence of the antibody, specific for the terminal beta-galactopyranosyl residue of asialo-GM1 (Harrison et al. 1998). The results of this study suggest that NAF of P. mirabilis recognize a GalNAc beta 1-4Gal moiety present in the ganglio-series of asialoglycolipids, and that the terminal beta-galactopyranosyl-containing glycoconjugates play a role in NAF-mediated adherence of P. mirabilis to MDCK cells. Similarly to other bacteria, P. mirabilis NAF was also shown to express the LacCer specificity.

Agglutination↗

PubMed1999

Isolation and chemical analysis of a lipopolysaccharide from the outer membrane of the oral anaerobic spirochete Treponema pectinovorum.

Isolation of a putative lipopolysaccharide from the surface of the oral treponeme, Treponema pectinovorum, revealed it to contain larger amounts of 3-deoxy-D-manno-octulosonic acid compared with other oral Treponema species. This molecule was isolated from the outer membrane of T. pectinovorum and had chemical characteristics of a putative lipopolysaccharide. The yield of lipopolysaccharide was between 0.6% and to 1.1% of the bacterial dry weight. The purified molecule was resistant to the action of proteinases and consisted of both sugars and lipids. 3-Deoxy-D-manno-octulosonic acid and hexoses accounted for 6.1-8.7% and 17.6-20.2%, respectively of the dry weight. Carbohydrate compositional analysis revealed the presence of glucose, galactose, 2-acetamido-2-deoxy-glucose, rhamnose and 6-deoxy-talose in the molar ratio of 1.00:0.96:0.19:0.88:0.98, respectively. No heptose was detected. The fatty acid analysis determined the presence of straight chain, C13:00, C14:00, C15:00 and C17:00 acids, as well as branched chain, C13:00, C14:00 and two species of C15:00, acids. Electrophoretic analysis indicated that the lipopolysaccharide was present as two major species.

Cell Membrane↗

PubMed1998

Structural elucidation of the lipopolysaccharide core regions of the wild-type strain PAO1 and O-chain-deficient mutant strains AK1401 and AK1012 from Pseudomonas aeruginosa serotype O5.

Lipopolysaccharide (LPS) of the Pseudomonas aeruginosa serotype O5 wild-type strain PAO1 and derived rough-type mutant strains AK1401 and AK1012 was isolated by a modified phenol/chloroform/petroleum-ether extraction method. Deoxycholate/PAGE of the LPS from the rough mutant AK1401 indicated two bands near the dye front with mobilities similar to those of the parent strain, indicating that both LPS contain a complete core and a species comprising a core and one repeating unit. Composition analysis of the LPS from strains PAO1 and AK1401 indicated that the complete core oligosaccharide was composed of D-glucose (four units), L-rhamnose (one unit), 2-amino-2-deoxy-D-galactose (one unit), L-glycero-D-manno-heptose (Hep; two units), 3-deoxy-D-manno-octulosonic acid (Kdo; two units), L-alanine (one unit) and phosphate (three units). The glycan structure of the LPS was determined by one-dimensional and two-dimensional (2D) NMR techniques in combination with MS-based methods on oligosaccharide samples obtained from the LPS by delipidation procedures. The locations of three phosphomonoester groups on the first heptose residue were established by a two-dimensional 31P (omega1)-half-filtered COSY experiment on the reduced core oligosaccharide sample of the LPS from the wild-type strain. The presence of a 7-O-carbamoyl substituent was observed on the second heptose. The structure of the core region of the O-chain-deficient LPS from P. aeruginosa serotype 05 is as follows: [structure: see text] where R1 is beta-D-Glcp-(1-->2)-alpha-L-Rhap-(1-->6)-alpha-D-Glcp-(1--> and R2 is alpha-D-Glcp-(1-->6)-beta-D-Glcp-(1->. A structural model is presented that is also representative of that for P. aeruginosa serotype O6 LPS. A revised structure for the serotype O6 mutant strain A28 is presented.

Lipopolysaccharides↗

PubMed1998

Structural characterization of the lipopolysaccharide O-antigen and capsular polysaccharide of Vibrio ordalii serotype O:2.

Structures of the capsular and O-chain polysaccharides of Vibrio ordalii serotype O:2, the causative agent of vibriosis in salmonid fish, were determined by high-field NMR techniques, mass spectrometric methods and partial hydrolysis. Both polymers were shown to be composed of linear tetrasaccharide repeating units, having the structure: carbohydrate sequence [see text].

Animals↗

PubMed1998

A kinetics approach to the characterization of an IgM specific for the glycolipid asialo-GM1.

The unique features of protein recognition of membrane-anchored glycolipids were investigated by surface plasmon resonance (SPR) monitoring of antibody interactions with glycolipids contained in liposomes. Several positive hybridomas belonging to the IgM and IgG classes were identified when tested for binding to the glycosphingolipid asialo-GM1 (Gal beta1-3GalNAcl beta1-4Gal beta1-4Glc beta1-1-Ceramide). Preliminary screening by enzyme immunoassay and thin layer chromatography (TLC) followed by immunostaining indicated that only those of the IgM type showed specificity for this glycosphingolipid. One of the IgMs, H2G10, was purified and further characterized using a SPR technique that involved antibody binding to liposomal asialo-GM1. This method generated kinetic and affinity constants for the interaction and confirmed the specificity of H2G10 for the terminal galactose of asialo-GM1. Interestingly, inhibition of antibody binding to asialo-GM1 liposomes by the asialo-GM1 tetrasaccharide reduced the total amount of bound antibody but increased the affinity of the antigen-antibody interaction due to an inverse relationship between tetrasaccharide concentration and the H2G10 dissociation rate constant. We believe that this effect is due to the selective inhibition of lower valency binding by the tetrasaccharide which, in turn, promotes higher avidity antibody-carbohydrate interactions. The observation that slower dissociation rate constants were also observed at high antigen to antibody ratios supports this interpretation. These results highlight the insight that kinetic data can provide in efforts to promote and inhibit high avidity interactions such as those involving proteins and carbohydrates.

Antibodies, Monoclonal↗

PubMed1998

Enhancement of sample loadings for the analysis of oligosaccharides isolated from Pseudomonas aeruginosa using transient isotachophoresis and capillary zone electrophoresis - electrospray - mass spectrometry.

The analysis of underivatized core oligosaccharides arising from mild acid hydrolysis of lipopolysaccharides from Pseudomonas aeruginosa serotype 05 was achieved using a transient isotachophoretic preconcentration method coupled to capillary zone electrophoresis-electrospray-mass spectrometry (tCITP-CZE-ES-MS). The combination of a tCITP preconcentration step provided a 10- to 50-fold enhancement of sample loading and a corresponding improvement in sensitivity compared to the conventional zone electrophoresis format. Electrophoretic conditions, enabling the separation of these anionic analytes, were developed to determine possible sites of heterogeneity on either the core or the O-chain structures. The tCITP-CZE-ES-MS technique provided unparalleled resolution of the different core glycoforms and oligosaccharides obtained from the acid cleavage of the native endotoxins whether isolated following conventional gel permeation chromatography or obtained from direct hydrolysis of the bacterial isolates. These investigations also highlighted the highly phosphorylated nature of these complex cell membrane components, where the heptose residues of the core oligosaccharide can bear up to six phosphate groups.

Carbohydrate Sequence↗

PubMed1998

Rating scales for mania: is self-rating reliable?

This commentary addresses the issue of whether self-rating mania scales are reliable. After a brief introduction, several newly developed self-rating mania scales are described and critically reviewed. Among the issues discussed are: the practical utility of self-assessment in patients with mania; the influence of severity of illness on ability to self-report; and, the extent to which the presence of psychosis or the absence of insight significantly interferes with a patients' ability to reliably complete self-rating measures. Data from recent studies suggests that self-rating mania scales are both reliable and valid for patients with manic symptoms, including those with psychotic features and those having little or no insight into their illness. The merits of including self-rating measures in the assessment of mania are discussed in terms of their value for both clinical and research purposes.

Bipolar Disorder↗

PubMed1998

The effect of growth conditions on in vitro adherence, invasion, and NAF expression by Proteus mirabilis 7570.

Proteus mirabilis is a common cause of upper urinary tract infections. Fimbriae-mediated adherence of this organism to urinary tract epithelium and invasion of host cells are factors thought to be important in its pathogenesis. We have assessed the effect of growth in serum, blood, and urine on the ability of P. mirabilis 7570 to adhere to and invade in vitro the cell line EJ/28, derived from a human urinary tract tumour, and to express nonagglutinating fimbriae (NAF). Proteus mirabilis was capable of adhering to EJ/28 cells to varying degrees depending upon the growth conditions used. It was invasive under all conditions, except when grown in urine, and was found to be particularly so when serum or blood was present in the media. Expression of NAF occurred under all growth conditions examined and was limited only by a decrease in temperature.

Bacterial Adhesion↗

PubMed1998

Efficiency and validity of commonly used substance abuse screening instruments in public psychiatric patients.

BACKGROUND: The more commonly used screening instruments for substance abuse were largely developed for addictive populations. We compared several alcohol and drug abuse scales to determine their efficiency and validity for psychiatric patients. METHOD: The subjects were 100 consecutively admitted patients to a public psychiatric facility. DSM-III-R diagnoses, obtained from the alcohol and drug scales of the SCID-P, were the criterion measure. Methods of reliability included inter-rater agreement, estimates of internal consistency, and repeat test administration. Sensitivity, specificity and more infrequently used accuracy indices, such as likelihood ratios and Receiver Operating Characteristic (ROC) analysis, were utilized to assess scale validity. RESULTS: First, the reliability of all scales was high. Second, the instruments generally demonstrated highly acceptable levels of screening accuracy. Third, the intake evaluation was as reliable and valid as screening after admission on the unit. Finally, instruments were least discriminating for current problems (past 30 days). CONCLUSIONS: Lifetime measures were found to be reliable and valid for public psychiatric patients but further research is need on increasing the accuracy of screening for current substance abuse problems and the effectiveness of multiple screening approaches.

Adult↗

PubMed1997

Quantitative analysis of bacterial toxin affinity and specificity for glycolipid receptors by surface plasmon resonance.

The primary virulence factors of many pathogenic bacteria are secreted protein toxins which bind to glycolipid receptors on host cell surfaces. The binding specificities of three such toxins for different glycolipids, mainly from the ganglioside series, were determined by surface plasmon resonance (SPR) using a liposome capture method. Unlike microtiter plate and thin layer chromatography overlay assays, the SPR/liposome methodology allows for real time analysis of toxin binding under conditions that mimic the natural cell surface venue of these interactions and without any requirement for labeling of toxin or receptor. Compared to conventional assays, the liposome technique showed more restricted oligosaccharide specificities for toxin binding. Cholera toxin demonstrated an absolute requirement for terminal galactose and internal sialic acid residues (as in GM1) with tolerance for substitution with a second internal sialic acid (as in GD1b). Escherichia coli heat-labile enterotoxin bound to GM1 and tolerated removal or extension of the internal sialic acid residue (as in asialo-GM1 and GD1b, respectively) but not substitution of the terminal galactose of GM1. Tetanus toxin showed a requirement for two internal sialic acid residues as in GD1b. Extension of terminal galactose with a single sialic acid was tolerated to some extent. The SPR analyses also yielded rate and affinity constants which are not attainable by conventional assays. Complex binding profiles were observed in that the association and dissociation rate constants varied with toxin:receptor ratios. The sub-nanomolar affinities of cholera toxin and heat-labile enterotoxin for liposome-anchored gangliosides were attributable largely to very slow dissociation rate constants. The SPR/liposome technology should have general applicability in the study of glycolipid-protein interactions and in the evaluation of reagents designed to interfere with these interactions.

Biosensing Techniques↗