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Biomedical subjects

E Marti

Publications and source records attributed to E Marti.

At least 19 recordsLinked to original sources
PubMed2001

Control of nitrogen export from watersheds by headwater streams.

A comparative (15)N-tracer study of nitrogen dynamics in headwater streams from biomes throughout North America demonstrates that streams exert control over nutrient exports to rivers, lakes, and estuaries. The most rapid uptake and transformation of inorganic nitrogen occurred in the smallest streams. Ammonium entering these streams was removed from the water within a few tens to hundreds of meters. Nitrate was also removed from stream water but traveled a distance 5 to 10 times as long, on average, as ammonium. Despite low ammonium concentration in stream water, nitrification rates were high, indicating that small streams are potentially important sources of atmospheric nitrous oxide. During seasons of high biological activity, the reaches of headwater streams typically export downstream less than half of the input of dissolved inorganic nitrogen from their watersheds.

Absorption↗
PubMed2000

Partial sequences of feline and caprine immunoglobulin epsilon heavy chain cDNA and comparative binding studies of recombinant IgE fragment-specific antibodies across different species.

Parts of the feline and caprine IgE epsilon heavy chain cDNA (third and fourth constant domains, IgEf3/4) were cloned, sequenced, and expressed to raise antibodies (Abs). The DNA and derived protein sequences of the feline recombinant IgEf (rIgEf) shared high homology with the analogous canine parts (81% at the nucleotide and 71% at the protein levels) and the caprine with the ovine ones (95%/84%), respectively. The polyclonal Abs raised in chickens against the feline and caprine rIgEf3/4 were subjected to a comparative binding study utilizing an ELISA including rIgEf and specific Abs to these rIgEf from dog and horse (rIgEf2 and rIgEf3/4) and sheep (rIgEf3/4). All but the ovine-specific rIgEf3/4 Ab were polyclonal, which had been raised in chickens, and bound to most applied rIgEf; the ovine-specific monoclonal mouse Ab recognized only in addition to ovine rIgEf3/4 the closely related caprine rIgEf3/4. Significant, positive correlations were detected between binding reactions of the polyclonal Abs in ELISA and percentage protein sequence homology (p<0.01). Thus, the newly described feline and caprine IgE nucleotide sequences and corresponding Abs represent useful tools for further species-specific and comparative allergy and disease-associated research.

Amino Acid Sequence↗
PubMed2000

First comprehensive low-density horse linkage map based on two 3-generation, full-sibling, cross-bred horse reference families.

Two 3-generation full-sibling reference families have been produced and form a unique resource for genetic linkage mapping studies in the horse. The F(2) generations, now comprising 61 individuals, consist of 28- to 32-day-old embryos removed nonsurgically from two pairs of identical twin mares. The same stallion sired all F(2)s such that the two full-sibling families are half-sibling with respect to each other. The families are crossbred to maximize levels of heterozygosity and include Arabian, Thoroughbred, Welsh Cob, and Icelandic Horse breeds. Milligram quantities of DNA have been isolated from each embryo and from blood samples of the parents and grandparents. The families have been genotyped with 353 equine microsatellites and 6 biallelic markers, and 42 linkage groups were formed. In addition, the physical location of 85 of the markers is known, and this has allowed 37 linkage groups to be anchored to the physical map. The inclusion of dams in the genotyping analysis has allowed the generation of a genetic map of the X chromosome. Markers have been assigned to all 31 autosomes and the X chromosome. The average interval between markers on the map is 10.5 cM, and the linkage groups collectively span 1780 cM. The results demonstrate the benefits for horse linkage mapping studies of genotyping on these unique full-sibling families, which comprise relatively few individuals, by the generation of a comprehensive low-density map of the horse genome.

Animals↗
PubMed2000

Allergen-specific IgE levels against crude mould and storage mite extracts and recombinant mould allergens in sera from horses affected with chronic bronchitis.

Immunoglobulin E antibody (IgE) levels against four recombinant (r) mould allergens (r-Aspergillus fumigatus [rAsp f] 7, 8 and 9; r-Alternaria alternata 1 [rAlta1]) and crude mould (Aspergillus fumigatus, Alternaria alternata, Penicillium notatum) and storage mite extracts were determined by ELISA in sera from 24 pulmonary sound control horses and 26 horses suffering from chronic bronchitis/bronchiolitis (CB), also called chronic obstructive pulmonary disease (COPD). Serum IgG and IgA titres were also determined against Aspergillus fumigatus extract and rAsp f 8.IgE against the crude extracts could be measured in all sera, but there was no significant difference between CB-affected and control horses. In contrast, only 8-30% of the horses, depending on the r-allergen tested, had detectable IgE levels in serum against the r-allergens. Horses with CB had significantly more often detectable IgE levels than controls against rAlt a 1 (10/26 and 3/24, respectively, p=0. 054), rAsp f 7 (13/26 and 2/24, respectively, p<0.01) and rAsp f 8 (11/26 and 1/24, respectively, p<0.01). Only four horses (three CB-affected and one healthy, p0.05) had detectable IgE levels against rAsp f 9. Furthermore, CB-affected horses were often sensitised against two or more r-allergens (13/26 of the CB-affected horses) while only one of the 24 healthy horses had positive IgE levels against more than one r-allergens. Similarly to IgE levels, no significant differences between CB-affected and healthy horses were found for IgG titres against the Aspergillus fumigatus extract. However, horses with CB had significantly higher serum IgG titres against rAsp f 8 than healthy controls (median=28 versus 10 relative ELISA units [REU], p<0.01). Additionally, horses with detectable IgE titres against rAsp f 8 had significantly higher IgG titres against this r-allergen than horses with undetectable IgE titres (median IgG titres=46 and 13 REU, respectively; p<0.01). For serum IgA titres, neither differences between healthy and CB-affected animals nor correlations between IgA and IgG or IgE titres could be found. These results show that horses suffering from CB are more often sensitised to some Aspergillus fumigatus and Alternaria alternata allergens than control horses and that they are partly sensitised to the same fungal proteins as mould-allergic human patients. Furthermore, this study shows that r-allergens allow a much more sensitive determination of specific serum antibody levels by ELISA than crude mould extracts.

Allergens↗
PubMed2000

Expression of caspases and their substrates in the rat model of focal cerebral ischemia.

Experimental evidence suggests that the massive release of glutamate during experimental brain ischemia both directly and indirectly regulates downstream mechanisms of cell suicide. Cerebral ischemia was produced by distal, permanent occlusion of the middle cerebral artery (MCAO) in the rat. Sets of three animals and one sham-operated for each time-point were kept alive for 0-30 min, 1, 4, 12, 24, and 48 h, and 4 days. Additional animals were treated by local administration of a 10 microM (in 10 microl) cocktail of caspase inhibitors (YVAD-cmk, DEVD-fmk, IETD). Immunohistochemistry was performed on free-floating tissue sections with goat polyclonal antibodies to procaspase-1, -2, -3, -6, and -8. Some sections were processed for double-labeling procaspase immunohistochemistry and in situ end-labeling of nuclear DNA fragmentation (TUNEL method). Both immunohistochemistry and double-labeling procaspase immunohistochemistry and TUNEL method were carried out on formalin-fixed sections. For gel electrophoresis and Western blotting, we used antibodies to poly (ADP-ribose) polymerase (PARP), lamin B, and PKC-delta, as specific cleavage substrates of caspases. There was increased immunoreactivity ipsilaterally in the areas corresponding to the infarct and surrounding penumbra with the peak of immunoreactivity between 12 and 24 h for most of the procaspases. Procaspases were present early in the infarcted tissue neurones and their dendrites and axons. Additional procaspase expression occurred in astrocytes and microglial cells at different times following ischemia. Cells with positive in situ end-labeling of nuclear DNA fragmentation appeared in high number predominantly in the infarcted areas and at the edge of the infarction and colocalized with enhanced procaspase expression. These findings suggest increased procaspase expression in dying cells at the edge of the infarction. A major product of PARP degradation of about 89 kDa was found in the samples taken from the infarcted and penumbra areas. There was no difference in the intensity of the bands corresponding to lamin B or PKC-delta. Injection of procaspase inhibitors reduced the levels of major PARP products of 89 kDa and decreased the number of TUNEL-positive cells at 12 h post-MCAO. In conclusion, these results give support to further research on the use of caspase inhibitors as add-on therapeutic agents for the treatment of ischemia.

Animals↗
PubMed2000

Expression and induction of CYP1A1/1A2, CYP2A6 and CYP3A4 in primary cultures of human hepatocytes: a 10-year follow-up.

1. The aims were to refine experimental conditions (using 76 human hepatocyte preparations) in terms of the selection of enzyme inducers and their optimal concentration, the treatment duration with inducers and the choice of specific cytochrome P450 isoform(s) probes to optimize the use of primary hepatocytes for predicting the potential induction by new chemical entities of cytochrome P450 isoforms in vivo in man. 2. In the absence of any inducer, basal cytochrome P450 isoform(s)-mediated activities decreased to 20% of their initial activity (end of the seeding period) by 72-96 h. In contrast, UGT-dependent enzyme activities remained at a constant level (+/- 20%) up to the fifth day of culture. 3. Beta-naphthoflavone, at an optimal concentration of 50 microM and after a 3-day treatment, specifically and potently induced 7-ethoxyresorufin (10.4 +/- 10.4-fold, n = 74) and phenacetin (6.6 +/- 6.4-fold, n = 60) O-deethylation processes, markers for CYP1A1 and CYP1A2 isoforms respectively. Only a 2-fold increase was noted following treatment with 2 mM phenobarbitone, whereas dexamethasone and rifampicin had no effect at all. 4. A 3-day treatment of human hepatocytes with 50 microM dexamethasone was associated with a major induction of both coumarin 7-hydroxylation (9.4 +/- 11.4-fold, n = 49) and nifedipine dehydrogenation (4.7 +/- 3.8-fold, n = 61), markers for CYP2A6 and CYP3A4 respectively. Phenobarbitone, however, exhibited a broad but moderate inducing effect on 7-ethoxyresorufin (2.2 +/- 1.5-fold, n = 55) and phenacetin (1.7 +/- 0.9-fold, n = 54) O-deethylation, coumarin 7-hydroxylation (3.9 +/- 9.2-fold, n = 50) and nifedipine dehydrogenation (2.1 +/- 2.0-fold, n = 47). 5. Km obtained for the different cytochrome P450 isoform substrates in untreated hepatocytes were in the same range of magnitude that those determined on human hepatic microsomal fractions. Enzyme induction processes were characterized by a large increase in apparent Vmax whereas apparent Km were not affected. 6. These studies demonstrate that human hepatocytes in primary culture can respond specifically and quantitatively to model inducers. This in vitro system offers a useful approach to study the regulation of human hepatic biotransformation activities and should facilitate the demand for a reproducible method for addressing cytochrome P450 induction.

Adult↗
PubMed1999

Sulfidoleukotriene generation from peripheral blood leukocytes of horses affected with insect bite dermal hypersensitivity.

Sulfidoleukotrienes (sLT) generated in vitro after incubation of equine peripheral blood leukocytes (PBL) with different inducing agents were determined in 18 healthy and 16 insect bite dermal hypersensitivity (IDH)-affected horses. PBL from these 32 horses were stimulated with Concanavalin A, Parascaris equorum, Culicoides nubeculosus and Simulium extracts, and with a six-Grass mix. The cells of all but four horses generated sLT after incubation with Concanavalin A; these four horses did also not produce sLT with the other inducing agents. Of the 28 remaining horses (12 affected with IDH and 16 healthy), all but three generated sLT with the P. equorum extract. The six-Grass mix did not induce sLT production in any of the tested horses. sLT generation with Concanavalin A and Parascaris was statistically not different between IDH-affected and healthy horses. PBL of the diseased horses, however, produced significantly more sLT with the Culicoides (p < 0.01) and Simulium (p < 0.05) extracts than those of the healthy animals. Additionally, sLT generation with the Culicoides extract was measured at different times of the year in one IDH-affected animal and remained high even in winter, when the horse was asymptomatic. sLT and histamine release were determined in 10 horses in parallel. Positive correlations of 0.81 and 0.82 for Concanavalin A and Parascaris (p < 0.01 and p < 0.05, respectively), and of 0.95 and 0.94 for Culicoides and Simulium (p < 0.01) were found between sLT and histamine release. These results indicate that, alike in humans, sLT are released in vitro from equine basophils along with histamine in response to various stimuli and that immediate type hypersensitivity reactions to Culicoides and Simulium are often involved in the pathogenesis of IDH. Thus, sLT generation from equine basophils offers an in vitro diagnostic tool for IDH even in sensitised but asymptomatic horses.

Animals↗
PubMed1999

Domain mapping and comparative binding features of eight dog IgE-specific reagents in ELISA, immunoblots, and immunohistochemistry.

Eight dog IgE-specific reagents including monoclonal and polyclonal antibodies (Ab) and a cross-reactive alpha chain of the human high affinity IgE receptor were mapped to recombinant fragments of the second (IgEf2) and third/fourth (IgEf3/4) domains of the dog IgE heavy chain. In ELISA, five out of eight reagents reacted to solid-phase bound IgEf2, of which two polyclonal Ab bound in addition to IgEf3/4. All Ab which recognized at least one recombinant IgE fragment, also bound to IgE in ELISA, immunoblots, and immunohistochemistry. In contrast, only one monoclonal Ab, that did not bind to the recombinant IgE fragments, reacted with immunoblots of serum and immunohistochemistry. The alpha chain could only be applied to ELISA with serum IgE. Furthermore, there was a wide range of heat-lability of binding reactions. Comparative analysis of available dog IgE-specific reagents enables more in-depth functional studies on IgE-mediated phenomena in dogs, and helps to further establish the dog as an animal model for allergy research.

Animals↗
PubMed1999

Modulation of early but not later stages of programmed cell death in embryonic avian spinal cord by sonic hedgehog.

Sonic hedgehog (Shh) is a secreted glycoprotein expressed by the notochord and floor plate that is involved in the induction and specification of ventral phenotypes in the vertebrate neural tube. Recently, Shh has also been shown to promote the survival of cultured rat embryo ventral brain and spinal cord cells. We have examined whether Shh can promote the survival of chick embryo neurons in vivo or in vitro. In the chick, Shh is expressed in notochord, floor plate, and ventral neural tube/spinal cord at several stages at which programmed cell death (PCD) occurs. However, the administration of exogenous Shh to embryos in vivo or to motoneuron cultures at these stages failed to promote the survival of several different neuronal populations, including spinal motoneurons, spinal interneurons, sympathetic preganglionic neurons, sensory neurons, and neuronal precursor cells. Rather, at the earliest stage of PCD examined here (embryonic day 3) Shh selectively induced the death of ventral neuronal precursors and floor-plate cells, resulting in a net loss of cells in the neural tube. Altered concentrations of Shh induce aberrant phenotypes that are removed by PCD. Accordingly, normal PCD in the early neural tube may play a role in dorsal-ventral patterning.

Animals↗
PubMed1999

Influence of sex and age on serum total immunoglobulin E concentration in Beagles.

OBJECTIVE: To establish an ELISA for detection of serum total IgE concentration in dogs and to analyze IgE values in a dog colony. ANIMALS: 147 healthy Beagles (31 males and 116 females). PROCEDURE: 2 canine IgE-specific polyclonal antibodies elicited by 2 recombinant fragments of the epsilon chain in hens were used to develop a capture ELISA specific for serum total IgE concentration. The IgE values were calculated by comparing serum dose-response curves (1:50 to 1:6,400) with a reference serum pool assigned 100 relative ELISA units (REU). Results-Mean IgE concentration in female Beagles was 51.2 REU (range, 0 to 337.8 REU; median, 31.4 REU), whereas mean IgE concentration in male dogs was only 7.5 REU (range, 0 to 32.6 REU; mean, 3.6 REU). Distribution of IgE values was skewed; approximately 80% of dogs had IgE values < 50 REU. Analysis of natural logarithmically transformed IgE values indicated that sex and age significantly (P < 0.05) influenced IgE values; mean serum IgE values increased until the age of 4 years. Heritability estimates of IgE concentration indicated a trend toward a genetic influence. CONCLUSION: A reliable capture ELISA specific for canine IgE was developed. Serum total IgE values vary with age and sex in the sample population. CLINICAL RELEVANCE: Serum total IgE concentration can now be evaluated in various dog breeds and, subsequently, in dogs with IgE-mediated diseases provided that these significant influences are accounted for. Serum total IgE values may then prove to be of diagnostic value, similar to their use in human beings.

Age Factors↗
PubMed1998

Characterization of two dog IgE-specific antibodies elicited by different recombinant fragments of the epsilon chain in hens.

Two recombinant [His]6-tagged fragments of the canine immunoglobulin E (IgE) heavy chain (second domain: IgEf2 and third and fourth domains: IgEf3/4) were cloned, expressed in Escherichia coli (E. coli) as [His]6-tagged proteins, and affinity-purified over nickel-nitrilotriacetic acid columns. The recombinant proteins were used to immunize hens. The raised and affinity-purified chicken antibodies (Ab) isolated from egg yolk exhibited specific binding to the respective recombinant canine IgE fragment (IgEf) on immunoblots and displayed high titers against the IgEf in ELISA. Immunoblotting of canine serum separated by PAGE under native conditions with the IgEf2- and IgEf3/4-specific Ab resulted in staining of a protein of approximately 180 kilodaltons (kD). The IgEf3/4-specific Ab further recognized an 80 kD protein in IgEf3/4-specific Ab affinity-enriched dog serum separated under denaturing conditions. In an ELISA for the detection of antigen-specific IgE in dog serum, reduced binding of the IgEf-specific Ab was observed after heat treatment of the dog serum. The reactivity of both of the raised chicken Ab was only present in postimmune reagents and could only be inhibited by preincubation with the IgEf used for immunization and not with dog immunoglobulin G, E. coli extract, or with a nonrelevant recombinant [His]6-tagged protein. In immunohistochemistry, the IgEf3/4-specific Ab specifically recognized cells in paraffin-embedded tissue sections of lymph nodes. Furthermore, both of the IgEf-specific Ab elicited positive immediate type 1 skin reactions in dogs. Semiquantitative assessment of total serum IgE in dogs was developed using IgEf2-specific Ab as coating reagent and the biotinylated IgEf3/4-specific Ab as developing Ab in ELISA. In conclusion, both IgEf-specific Ab recognize native dog IgE with the advantages that they are directed against different and known constant domains of the IgE molecule, and that they can be used for immunohistochemistry on paraffin-embedded tissue. The two dog IgE-specific Ab could initiate clinical research on the involvement of immediate-type hypersensitivity reactions in dogs.

Animals↗
PubMed1998

Equine sarcoids.

Sarcoids, the most common tumor of the horse, are fibroblastic, wart-like skin lesions that show variable manifestations. They are often invasive and recurrent, although they do not fulfill all criteria of malignancy. Due to their anatomic location, these tumors can sometimes cause loss of use of the horse. There is very strong evidence that sarcoids are caused by viruses closely related or identical to bovine papilloma viruses, and genetic studies have shown associations between genes in or near the equine major histocompatibility complex (MHC) and susceptibility to sarcoid. Several types of treatments have been successful in treating sarcoids, although the response to therapy is not consistent. Current treatment of sarcoids primarily involves antitumor therapy, but the development of preventative measures in the future may be directed against the causative papilloma virus. Sarcoid continues to be an important clinical entity for the equine practitioner.

Animals↗
PubMed1998

Polymorphism of Old Kladruber horses, a surviving but endangered baroque breed.

Analysis of MHC class I and class II polymorphism, as well as data from other polymorphic systems (non-MHC lymphocyte alloantigen, blood groups systems, biochemical polymorphisms and microsatellite loci), was used to characterize the extent and distribution of the genic polymorphism of Kladruber horses. A breed-characteristic distribution of the MHC polymorphism was found. The repertoire of defined MHC class I specificities was restricted, especially in the grey subpopulation and in stallions, but a high frequency of blanks suggests the possible existence of undetected specificities. Despite the small population size and a relatively high degree of inbreeding, high heterozygosity in MHC haplotypes has been conserved. The extent of polymorphism and the degree of heterozygosity in other loci were also relatively high. A comparison of the two existing subpopulations, grey and black, at all the loci tested, including RAPD markers, characterized them as genetically distinct, although clearly related. The genetic distances between them were of the same order of magnitude as between distinct breeds. The results may be useful in defining short-term and long-term breeding policy within the breed and for further studies of associations with disease and other traits.

Animals↗
PubMed1997

Diadenosine polyphosphate hydrolase from presynaptic plasma membranes of Torpedo electric organ.

The diadenosine polyphosphate hydrolase present in presynaptic plasma membranes from the Torpedo electric organ has been characterized using fluorogenic substrates of the form di-(1, N6-ethenoadenosine) 5',5'''-P1,Pn-polyphosphate. The enzyme hydrolyses diadenosine polyphosphates (ApnA, where n=3-5), producing AMP and the corresponding adenosine (n-1) 5'-phosphate, Ap(n-1). The Km values of the enzyme were 0.543+/-0.015, 0.478+/-0.043 and 0. 520+/-0.026 microM, and the Vmax values were 633+/-4, 592+/-18 and 576+/-45 pmol/min per mg of protein, for the etheno derivatives of Ap3A (adenosine 5',5'''-P1,P3-triphosphate), Ap4A (adenosine 5',5"'-P1,P4-tetraphosphate) and Ap5A (adenosine 5',5'''-P1,P5-pentaphosphate) respectively. Ca2+, Mg2+ and Mn2+ are enzyme activators, with EC50 values of 0.86+/-0.11, 1.35+/-0.24 and 0.58+/-0.10 mM respectively. The fluoride ion is an inhibitor with an IC50 value of 1.38+/-0.19 mM. The ATP analogues adenosine 5'-tetraphosphate and adenosine 5'-[gamma-thio]triphosphate are potent competitive inhibitors and adenosine 5'-[alpha,beta-methylene]diphosphate is a less potent competitive inhibitor, the Ki values being 0.29+/-0.03, 0.43+/-0.05 and 7.18+/-0.8 microM respectively. The P2-receptor antagonist pyridoxal phosphate 6-azophenyl-2',4'-disulphonic acid behaves as a non-competitive inhibitor with a Ki value of 29.7+/-3.1 microM, and also exhibits a significant inhibitory effect on Torpedo apyrase activity. The effect of pH on the Km and Vmax values, together with inhibition by diethyl pyrocarbonate, strongly suggests the presence of functional histidine residues in Torpedo diadenosine polyphosphate hydrolase. The enzyme from Torpedo shows similarities with that of neural origin from neurochromaffin cells, and significant differences compared with that from endothelial vascular cells.

Acid Anhydride Hydrolases↗