Biomedical subjects

Andrzej Gamian

Publications and source records attributed to Andrzej Gamian.

At least 19 recordsLinked to original sources

PubMed2006

The effect of packed red blood cell storage on arachidonic acid and advanced glycation end-product formation.

INTRODUCTION: The transfusion of packed red blood cells (PRBCs) is a significant risk to blood recipients. Blood banking procedures permit the storage of PRBCs for up to 42 days. Storage of PRBCs can cause polymorphonuclear granulocytes (PMNs) activation and the development of neutrophil-mediated transfusion-related acute lung injury. The aim of our study was to determine if PRBC storage has an influence on the formation of arachidonic acid (AA) and advanced glycation end products (AGEs). MATERIALS AND METHODS: Twenty units of PRBCs were used to measure AA and AGE levels. The samples were taken on the 0th, 14th, 28th, and 42nd days of PRBC storage. The AA level was analyzed by gas-liquid chromatography-mass spectrometry and AGE level by an immunoenzymatic test. RESULTS: During the first 14 days of PRBC storage, the AA level significantly increased and then slowly decreased. The AGE level increased continuously during the whole time of the study. In a model experiment, the AA glycoxidation product trans-2-nonenal (T2N) formed adducts in reaction with hemoglobin which were detectable with the test for AGE. CONCLUSIONS: It is highly probable that the observed increase in AGE level is related to the decrease in AA in PRBCs, which can be associated with the formation of toxic aldehydes, especially T2N and 4-hydroxynonenal (HNE), from AA. Glucose in the PRBCs (preservative solution) can contribute to AGE formation as well. The formation of AGEs, HNE, and T2N in PRBCs, their influence on PMNs in vitro, and confirmation of our assumption need further studies.

Arachidonic Acid↗

PubMed2006

Routine clinical laboratory tests correspond to increased serum levels of 3-hydroxy fatty acids, markers of endotoxins, in cardiosurgery patients.

INTRODUCTION: Endotoxemia developing during cardiosurgery as elevated endotoxin concentrations in patient's serum may prevail over 24 h after operation. A major reason is thought to be increased gut permeability resulting in endotoxin and bacterial leakage. In this study we aimed to measure endotoxin levels on samples obtained during and after cardiovascular procedures and compare them with clinical observations and laboratory test results. MATERIALS AND METHODS: 3-Hydroxy fatty acids (3-OH FAs) of 10-18 carbon chain length, chemical markers of endotoxin (lipopolysaccharide), were determined in patient sera by gas chromatography-mass spectrometry-based analysis. RESULTS were compared with routine laboratory tests: blood morphology, urine, ALT, AST, bilirubin, kidney parameters, clotting parameters, and gasometry. RESULTS: Of a total of 16 patients, 5 patients (group I) showed increased serum 3-OH FA levels and 11 patients (group II) did not show any change in 3-OH FA levels 24 h after operation. All group I patients revealed leukocytosis, two developed post-operative anemia. Significantly different changes were observed: the initial, pre-operative 3-OH FA levels were similar for both groups, while group I patients showed increased levels of all the studied 3-OH FAs during the operation (p</=0.05), and 3-OH C14 and 3-OH C16 remained elevated 24 h after the operation. CONCLUSIONS: Cardiosurgery may strongly promote gut endotoxin translocation to the blood in some patients. Prolonged leukocytosis, deep anemia, and increased liver dysfunction markers may indicate the need for observation of possible endotoxemia development. It is recommended to monitor the endotoxin level and/or endotoxemia markers in cardiosurgery patients.

Biomarkers↗

PubMed2006

Toll-like receptor ligands reverse suppression of contact hypersensitivity reactions induced by epicutaneous immunization with protein antigen.

BACKGROUND: Epicutaneous (EC) immunization with protein antigens has been shown to induce antigen nonspecific suppression of subsequent T cell-dependent contact hypersensitivity (CS) reactions after active immunization. The aim of this work was to test if EC application of Toll-like receptor (TLR) ligands together with protein antigen could reverse suppression of CS. METHODS: Mice were EC immunized by applying gauze patches soaked with a solution of protein antigen alone or in the presence of crude bacterial material (bacterial lysates or heat-killed bacteria) or purified TLR ligands and then tested for CS response. To test if reversal of EC-induced suppression is antigen-specific, mice were patched with TNP- or OX-substituted mouse Ig alone or together with LPS and then tested for CS with corresponding or non-cross-reacting hapten. Influence of EC immunization on cytokine production by lymph node cells was measured by ELISA. RESULTS: EC immunization with protein antigen induces antigen nonspecific suppression that can be reversed by crude bacterial material as well as purified TLR-2, TLR-3, TLR-4, and TLR-9 ligands. The effect of TLR-4 ligand LPS was not observed in the Tlr-4 mutant C3H/HeJ mouse, indicating that this effect was dependent upon intact TLR-4 signaling. Unlike the antigen nonspecific suppression of CS by EC immunization with antigen alone, the reversal of suppression by TLR ligands was specific for the protein antigen applied in the EC protocol. CONCLUSIONS: Our results strongly suggest that EC immunization with protein antigen together with TLR ligands induces a particular antigen-specific cell population, akin to previously described contrasuppressor cells, which protects immune cells against the action of suppressor cells but have no direct influence on antigen nonspecific suppressor cells induced by antigen alone.

Animals↗

PubMed2006

Enterobacterial 38-kDa outer membrane protein is an age-dependent molecular marker of innate immunity and immunoglobulin deficiency as results from its reactivity with IgG and IgA antibody.

In earlier studies on an animal model we observed protective properties of outer membrane proteins (OMPs) of Shigella, Hafnia, and Escherichia coli strains. In order to investigate human sera for reactivity with OMPs we subjected these proteins to immunoblotting with umbilical cord plasma and sera from children and adults. The IgG and IgA antibodies interacted primarily with a 38-kDa protein, in similar way for several enterobacterial strains, but different for Pseudomonas aeruginosa. This observation prompted us to determine the reactivity with the purified 38-kDa OMP in the sera of several groups of children. The reactivity of the protein from Shigella flexneri serotype 3a with sera in ELISA was age dependent, increasing from low reactivity in infants to the adult antibody level. The IgG and IgA antibody specific response thus revealed the normal pattern of immunity. The level of IgA and IgG antibody was significantly low in child patients with IgA and/or IgG immunoglobulin deficiencies, but was at the healthy control level in children with recurrent respiratory tract inflammation. These data correlated with total IgA and IgG levels in immunoglobulin-deficient children. The results indicate that this protein may serve as an immunodiagnostic marker, but also as an antigen carrier in vaccines.

Adolescent↗

PubMed2006

[Glycosaminoglycans in the pathogenesis and diagnostics of Graves's ophthalmopathy].

Graves's ophthalmopathy is an extrathyroidal tissue-specific symptom of Graves's disease characterized by inflammatory infiltration of orbital tissues. In several percent of patients the course of disease is severe, leading to serious ocular complications and affecting quality of life. Progress made in understanding the pathogenesis of GO has not been followed by better outcome of treatment and it is still one of the most complex problems of clinical endocrinology. In this respect, qualification for immunosuppressive treatment based on an assessment of the disease activity is a matter of great importance. Glycosaminogycans are linear, unbranched heteropolysaccharides built of repeating disaccharide sequences. These compounds are essential for the organization and function of connective tissue. Moreover, they also take part in the initiation and regulation of immune reactions. In this paper, glycosaminogycans are described with regard to their structure, metabolism, and function, with special emphasis on the tissue specificity of the orbit. Data concerning GAG useful in the management of patients with GO are highlighted.

Glycosaminoglycans↗

PubMed2005

A novel procedure for the isolation of glycolipids from Bifidobacterium adolescentis 94 BIM using supercritical carbon dioxide.

This study describes a novel isolation procedure for major glycolipids from Bifidobacterium adolescentis 94 BIM. The procedure consists of the use of supercritical carbon dioxide (scCO(2)) with hydro-methanolic solution as co-solvent. The major glycolipids were isolated using the following operating conditions: pressure, 30 MPa, co-solvent concentration, 10% (9:1, methanol/water, v/v), CO(2) flow rate, 5 g/min, extraction time and temperature, 2h and 55 degrees C, respectively. The reference glycolipids sample was prepared by classical organic solvent extraction followed by chromatographic purification. All isolates were characterized by TLC and the major glycolipids additionally by enzyme linked immunosorbent (ELISA). Sixty milligrams of glycolipids with similar immunoreactivity as the reference glycolipids were isolated from 1g of freeze-dried biomass (6% of yield).

Bifidobacterium↗

PubMed2005

Major structural proteins of type 1 and type 3 Klebsiella fimbriae are effective protein carriers and immunogens in conjugates as revealed from their immunochemical characterization.

Fimbriae are filamentous structures present on the cell surface of many bacteria, including genus Klebsiella. The use of fimbriae as protein carriers in conjugates may allow to formulate effective multivalent vaccines and suitable diagnostics. However, the evidences have been reported that fimbriae may enhance the inflammatory response. This prompted us to examine the degree of cytokine induction by the type 1 and type 3 Klebsiella fimbriae and their conjugates. Fimbriae were assessed as carrier proteins for Escherichia coli K12 endotoxin core oligosaccharide. MALDI-MS revealed the molecular mass of fimbrial monomer major protein, which was 15,847 Da for type 1 and 18,574 Da for type 3 fimbriae of Klebsiella. These two types of fimbriae were moderate inductors of IL-6 and interferon and almost inactive with regard to the stimulation of TNF when tested in human whole blood assay. Coupling of fimbriae with E. coli K12 core oligosaccharide gave immunogenic conjugates with respect to a saccharide ligand and protein carrier, although only 10% of the pilin monomers possessed the attached oligosaccharide. Rabbit antiserum reacted with a broad spectrum of lipopolysaccharides, as measured by ELISA and immunoblotting assays. The antibodies against glycoconjugates were bactericidal for the wild, S-type bacteria of some species. Regarding the induction of cytokines by conjugates only the TNF level was noticeably elevated. These results prompt for the practical use of fimbriae, as effective protein carriers for conjugates to obtain broad-spectrum antisera for diagnostic applications.

Animals↗

PubMed2005

Structure of the O-polysaccharide of Hafnia alvei strain PCM 1189 that has hexa- to octasaccharide repeating units owing to incomplete glucosylation.

The O-polysaccharide of Hafnia alvei PCM 1189 consists of D-glucose, D-galactose, D-GalNAc and D-GlcA and lacks the strict regularity. The intact and carboxyl-reduced polysaccharides as well as oligosaccharides obtained by partial acid hydrolysis were studied by chemical and enzymatic analyses, methylation and NMR spectroscopy. The following structure was established for the O-polysaccharide, which is built up of branched hexa- to octasaccharide repeating units differing in the number of lateral glucose residues: [structure: see text] where the glucose residues shown in italics are nonstoichiometric substituents. The repeating units include also a minor O-acetyl group, whose position was not determined.

Carbohydrate Sequence↗

PubMed2005

Antibodies against human muscle enolase recognize a 45-kDa bacterial cell wall outer membrane enolase-like protein.

Enolase, is a glycolytic enzyme ubiquitous in higher organisms, where it forms tissue specific dimers of isoforms, also found in the cytoplasm of fermentative bacteria. The aim of this work was to identify enolase-like proteins in the cell wall of some Gram-negative bacteria using antibodies against human beta-enolase, an isoenzyme specific to skeletal and heart muscles. Cell wall outer membrane protein (OMP) preparations were obtained from 9 strains of Enterobacteriaceae and one of Pseudomonas aeruginosa. Specific enzymatic enolase activity was detected in the supernatant fractions of cytosolic and inner membrane material, but not in purified OMP preparations. Rabbit polyclonal antibodies specific against human beta-enolase were prepared and purified using immobilized human beta-enolase in affinity chromatography. In SDS-polyacrylamide gel electrophoresis and immunoblotting assay of purified OMP preparations, rabbit anti-enolase antibody interacted specifically with a few OMPs, of which a 45-kDa band also interacted with human sera of patients presenting Buerger disease and atherosclerosis. The most distinct interaction of human sera was observed with a 45-kDa OMP of Klebsiella pneumoniae. This protein was further isolated from K. pneumoniae cell mass in two ways, namely preparative SDS-polyacrylamide gel electrophoresis and specific affinity chromatography using immobilized affinity-purified rabbit antibody raised against human beta-enolase. The data obtained from tandem mass spectrometry tryptic peptide analysis and sequence comparison of human and bacterial enolases using protein databases, could reveal the similarity in the epitopes between membrane enolase-like protein from Klebsiella and human beta-enolase. The results show that the protein present in all studied strains has a common epitope on human beta-enolase. These data raise the question whether such a bacterial protein might be a marker for detecting and monitoring damage to skeletal and heart muscles.

Animals↗

PubMed2005

[Low-molecular antioxidant compounds of natural origin].

Vitamins C, and E, carotenoids, and phenolic compounds from plants are low-molecular substances which express strong antioxidant and antiradical activity. Free radical processes are very complicated way, and it is impossible to draw conclusions from in vitro analysis of any compound regarding the role it would play under in vivo conditions. In this study, the in vivo and in vitro antioxidant and antiradical activities of some antioxidants contained in vegetable foods as well as the possibility of their prooxidant activity are described.

Animals↗

PubMed2005

The cytotoxic and genotoxic effects of conjugated trans-2-nonenal (T2N), an off-flavor compound in beer and heat processed food arising from lipid oxidation.

This study investigates the toxic effect of E(2)nonenal (trans-2-nonenal, T2N) and its conjugate with horse muscle myoglobin (Mb) tested on murine cell line L929 and human cell line A549, as well as the genotoxic effect of these compounds assayed by measuring of micronuclei in human cells K562. It is an aldehyde, which is occurring as the substance responsible for an off flavour in aged beers, but originates also from lipid oxidation in heat processed food. T2N is an aldehyde formed from linoleic acid as a secondary oxidation product. The modification of Mb with T2N was analyzed with the use of SDS-polyacrylamide gel electrophoresis (SDS-PAGE), and electrospray ionization mass spectrometry (ESI-MS). Results from SDS-PAGE suggest that T2N substitutes Mb and additionally causes cross-linking with polymerization of Mb resulting in an insoluble fraction. The ESI-MS spectrum of the soluble fraction used in the toxicity tests, demonstrated that conjugation of T2N with Mb yielded Mb adducts with one residue of trans-2-nonenal per myoglobin molecule as the major fraction and adducts with different numbers of T2N molecules as minor fractions. In the cytotoxicity assay the T2N and its Mb conjugate causes 50% destruction of cells at the concentration 95-125 microg/ml and 200 microg/ml respectively, when L929 and A549 cell lines were used, whereas Mb control tested up to 2000 mg/ml was without any cytotoxic effect. In genotoxicity in vitro assay we have observed that the T2N and its Mb conjugate expressed the genotoxicity. The number of micronuclei in human K562 cells reached 26 +/- 2.16 promille (MN/1000 cells), comparing to 62 +/- 8.64 MN/1000 cells for the reference free T2N, whereas a control value was 10.33 +/- 1.25 MN/1000 cells. The studied compounds expressed also the apoptotic effect in K562 cells as the number of apoptotic cells increased to 44.67 +/- 4.92 promille for T2N-Mb, comparing to 168.67 +/- 37.28 promille for free T2N, whereas a control value was 30.33 +/- 1.36 promille for Mb. In these assays the T2N-Mb conjugate is several times more toxic in relation to control protein. Results indicate that T2N adducts with protein are potent to induce various cytotoxic and apoptotic effects when assayed in vitro tests. It suggests that higher level of such aldehyde might create in organism severe potential of toxicity.

Aldehydes↗

PubMed2004

Structural and serological characterization of the major glycolipid from Rothia mucilaginosa.

Structural studies on the major glycolipid isolated from Rothia mucilaginosa were carried out utilising specific chemical degradation, NMR spectroscopy and matrix-assisted laser-desorption/ionization time of flight mass spectrometry (MALDI TOF-MS). The glycolipid was found to be a dimannosylacylmonoglyceride in which the carbohydrate part was the glycerol-linked dimannoside alpha-D-Manp-(1-->3)-alpha-D-Manp-(1-->3)-sn-Gro (Man A-Man B-Gro), of which Man B was esterified at O-6 by a fatty acid residue. A second fatty acid substituted the secondary methylene position of the glycerol residue, in contrast to the glycolipid previously found in R. dentocariosa and Saccharopolyspora strains, in which the second fatty acid esterified the primary methylene position of glycerol. Results of the ELISA experiment with rabbit specific antibacterial sera indicate that these two major glycolipids are antigenic, and the patterns of serological reactivity are similar but not identical.

Animals↗

PubMed2004

Structures of two O-polysaccharides of the lipopolysaccharide of Citrobacter youngae PCM 1538 (serogroup O9).

Mild acid degradation of the lipopolysaccharide of Citrobacter youngae O9, strain PCM 1538 released a homopolysaccharide of 4-acetamido-4,6-dideoxy-D-mannose (D-Rha4NAc, N-acetyl-D-perosamine). Studies by methylation analysis and (1)H and (13)C NMR spectroscopy, using two-dimensional (1)H,(1)H COSY, TOCSY, NOESY and H-detected (1)H,(13)C HSQC experiments showed the presence of two structurally different polysaccharides consisting of the following units: -->)-alpha-D-Rhap4NAc-(1 --> and --> 3)-alpha-D-Rhap4NAc-(1 --> 3)-beta-D-Rhap4NAc-(1 -->.

Carbohydrate Sequence↗

PubMed2004

Structure of the O-polysaccharide of Citrobacter youngae O1 containing an alpha-D-ribofuranosyl group.

The lipopolysaccharide of Citrobacter youngae O1, strain PCM 1492 was degraded with acid or alkali under mild conditions, and the resultant polysaccharide was isolated by GPC and studied by sugar and methylation analyses and 1H and 13C NMR spectroscopies, including 2D COSY, TOCSY, NOESY and 1H, 13C HSQC experiments. The following structure of the branched tetrasaccharide repeating unit of the O-polysaccharide was established: [structure: see text] where substitution with the alpha-D-Ribf group is nonstoichiometric. This group occurs rarely in bacterial polysaccharides and is easily cleaved under mild acidic conditions. Studies with polyclonal rabbit antisera against whole cells of C. youngae PCM 1492 and PCM 1506 showed the serological identity of the lipopolysaccharides of C. youngae PCM 1492, PCM 1493 and PCM 1506, which are classified in serogroup O1.

Carbohydrate Sequence↗

PubMed2004

Amycolatopsis palatopharyngis sp. nov., a potentially pathogenic actinomycete isolated from a human clinical source.

The taxonomic position of an actinomycete, strain 1BDZ(T), isolated from a clinical human source was determined using a polyphasic approach. Phylogenetic analysis based on almost complete 16S rDNA sequences showed that this organism consistently formed a distinct line with the Amycolatopsis methanolica subclade within the genus Amycolatopsis, and shared moderately low 16S rDNA similarity (<96.5 %) with other species. The organism was also found to have chemical and morphological properties typical of members of the genus Amycolatopsis. A range of phenotypic characteristics readily distinguished this strain from representatives of all species of Amycolatopsis with validly published names. On the basis of these data, a novel species, Amycolatopsis palatopharyngis sp. nov., is proposed to accommodate strain 1BDZ(T) (=AS 4.1729(T)=PCM 2600(T)).

Actinomycetales↗

PubMed2004

[Bactericidal activity of normal human serum to gram-negative rods with sialic acid containing lipopolysaccharide].

The complement plays the most important role in eliminating bacterial invasion of the host, by facilitating phagocytosis of potential pathogens and by participating in the direct essential role in protecting gram-negative bacteria against bactericidal activity of serum. Sialic acids which are important constitutes of animal tissue glycoconjugates are also present in antigens of some bacterial strains. The susceptibility of gram-negative strains with sialic acid--containing lipopolysaccharides to bactericidal action sera was examined.

Blood Bactericidal Activity↗

PubMed2003

Substitution pattern of 3-deoxy-D-manno-oct-2-ulosonic acid in bacterial lipopolysaccharides investigated by methylation analysis of whole LPS.

3-Deoxy-D-manno-oct-2-ulosonic acid (Kdo) is a constituent of the inner core part of bacterial lipopolysaccharides (LPS). This sugar may contribute to biological activities of the LPS, the type of substitution of Kdo is thus of importance and this work is aimed at the evaluation of a method for monitoring the substitution of Kdo in LPS. The procedure consists of three steps, namely permethylation of the lipopolysaccharide, with iodomethane and sodium methylsulfinylmethanide or NaOH in Me(2)SO, or with methyl triflate, then the product is methanolysed with HCl in MeOH and acetylated with acetic anhydride in pyridine. The resulting partially methylated acetates of Kdo methyl glycosides were analyzed by gas-liquid chromatography-electron impact ionization mass spectrometry (GLC-MS). For several derivatives of Kdo, specific GLC retention times and MS fragmentation patterns were determined. Lipopolysaccharides from several bacterial strains were isolated and analyzed with three different methods of methylation. The complete solubilization of the LPS in the acid form allows diminishing possible undermethylation. Sodium methylsulfinylmethanide is the most efficient agent in the permethylation of the whole LPS, of all the tested procedures. Methylation with methyl triflate allows the detection of base labile substituents on Kdo residues.

Chromatography, Gas↗

PubMed2003

Structure of the O-specific polysaccharide from the lipopolysaccharide of Citrobacter gillenii O11, strain PCM 1540.

The O-specific polysaccharide of the lipopolysaccharide of Citrobacter gillenii PCM 1540 (serogroup O11) consists of D-Glc, D-Man, D-GalNAc, D-GlcNAc, 2-acetamido-2,6-dideoxy-D-galactose (D-FucNAc) and O-acetyl groups in the ratios 2:1:1:1:1:1. On the basis of sugar and methylation analyses and Smith-degradation along with 1D and 2D 1H and 13C NMR spectroscopy, the following structure of the branched hexasaccharide repeating unit was established: [structure: see text]. Citrobacter werkmanii PCM 1541 belonging to the same serogroup O11 was found to have an R-form lipopolysaccharide devoid of the O-specific polysaccharide.

Acetates↗