Biomedical subjects

M Nilsson

Publications and source records attributed to M Nilsson.

At least 145 records · Page 8Linked to original sources

PubMed1997

Pharmacodynamic effects of antibiotics and acid pump inhibitors on Helicobacter pylori.

Pharmacodynamic studies of Helicobacter pylori exposed to amoxicillin, clarithromycin, metronidazole, omeprazole, and lansoprazole were performed with microscopy, viable count determination, and bioluminescence assay of intracellular ATP. The pharmacodynamic parameters determined were change in morphology, change in cell density, postantibiotic effect (PAE), and control-related effective regrowth time (CERT). The PAE is delayed regrowth after brief exposure to antibiotics or acid pump inhibitors. CERT was defined as the time required for the bacteria to resume logarithmic growth and return to the pre-exposure inoculum in the test culture minus the corresponding time for the control culture. CERT measures the combined effect of initial killing and PAE. There was a good concordance between the bioluminescence assay and viable counts for determining CERT, which makes this parameter useful for pharmacodynamic studies of the effects of antibiotics and acid pump inhibitors on H. pylori. Amoxicillin and metronidazole produced a strong, concentration-dependent initial decrease in CFU per milliliter, but there was a less prominent initial change in intracellular ATP in these cultures. Amoxicillin caused a long PAE when assayed by the bioluminescence assay but no PAE or a negative PAE when assayed by viable count determination. However, amoxicillin showed similar long CERTs with both methods. The pharmacodynamic effects of amoxicillin were concentration dependent up to a maximum response, indicating that concentrations above this level do not increase the antibiotic effect. The PAEs and CERTs of clarithromycin and metronidazole were concentration dependent with no maximum response. With omeprazole and lanzoprazole, there was no PAE or CERT.

Adenosine Triphosphate↗

PubMed1997

Variation in microbial community structure in two boreal peatlands as determined by analysis of phospholipid Fatty Acid profiles.

Analyses of phospholipid fatty acids (PLFAs) were used to assess variation in community structure and total microbial biomass in two boreal peatlands in Sweden. The total PLFA concentration in peat ranged from 0.16 to 7.0 nmol g of wet peat(sup-1) (median, 0.70 nmol g of wet peat(sup-1)). Principal-component analysis of PLFA data revealed that the degree of depth-related variation in PLFA composition was high among peatland habitats, with general differences between wet sites, with water tables within a few centimeters of the moss surface, and dry sites, with water tables >10 cm below the moss surface. However, variation in PLFA composition over the growing season was negligible. In the principal-component analyses, most PLFAs were determined to be parts of clusters of covarying fatty acids, suggesting that they originated in the same functional groups of microorganisms. Major clusters were formed by monounsaturated (typical of gram-negative eubacteria), terminally branched (gram-positive or anaerobic gram-negative eubacteria), methyl-branched and branched unsaturated (sulfate-reducing bacteria and/or actinomycetes), (omega)8 monounsaturated (methane-oxidizing bacteria), and polyunsaturated (eucaryotes) PLFAs. Within the clusters, PLFAs had rather distinct concentration-depth distributions. For example, PLFAs from sulfate-reducing bacteria and/or actinomycetes and those from methane-oxidizing bacteria had maximum concentrations slightly below and at the average water table depth, respectively.

Journal Article↗

PubMed1997

Ca2+ ion permeability properties of (R,S) alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptors in isolated interneurons from the olfactory bulb of the rat.

The aim of the study was to investigate the divalent cation permeability of native alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate (AMPA) receptors expressed in interneurons of the olfactory bulb. Kainic acid (KA) was used as agonist to activate AMPA-receptor-mediated currents, which were recorded with the use of the patch-clamp technique. In interneurons acutely isolated from the olfactory bulb, the current responses to KA showed linear/outwardly rectifying current-voltage (I-V) relationships with a positive average reversal potential of +7 mV in normal external medium (1 mM Ca2+, 1 mM Mg2+). Raising the external Ca2+ concentration to 10 mM suppressed the amplitude, whereas omission of Ca2+ enhanced the amplitude of the current. Spectral analysis of the increase in current variance produced by KA indicated that the decreased amplitude observed in 10 mM Ca2+ was accompanied by a reduction in the apparent single-channel conductance. Raising the concentration of Mg2+ from 1 to 10 mM had a weak depressant effect on the KA-evoked current amplitude. No shift in the reversal potential was observed when the concentration of Ca2+ or Mg2+ was changed from 1 to 10 mM. Increasing the external medium concentration of Ca2+ to 60 mM not only further depressed the amplitudes of the KA-evoked currents but also gave a pronounced leftward shift in the average reversal potential to -32 +/- 9 (SE) mV (N = 7). For neurons in primary culture, current responses to KA also showed linear/outwardly rectifying I-V relationships with a positive average reversal potential in normal external medium. Substituting N-methylglucamine for Na+ and increasing the Ca2+ concentration to 10 mM gave a leftward shift in the average reversal potential from +9 +/- 3 mV to -47 +/- 4 mV (N = 11) and caused a marked reduction in the amplitude of the KA-evoked currents at negative potentials. The permeability properties of the studied AMPA receptors were well predicted by the Eyring rate model (symmetrical, 2 barriers, 1 site). The model gave a pCa2+/pK+ permeability ratio of 0.06 for acutely isolated interneurons and 0.14 for interneurons in primary culture. The constant field theory, which failed to successfully reproduce all the experimental data, gave corresponding low permeability ratios of 0.18 and 0.40 for acutely isolated cells and cells in primary culture, respectively. Thus it is concluded that interneurons in the olfactory bulb mainly express AMPA receptors with low permeability to Ca2+ ions.

Animals↗

PubMed1997

Locked on target: strategies for future gene diagnostics.

A rapidly expanding number of genetic mechanisms contributing to disease can be accessed via molecular analysis. In order to take full advantage of this potential for genetic diagnostics, and to monitor treatment by gene therapy and other emerging methods, a new generation of diagnostic techniques will be required. Of central importance in future analyses is the possibility to perform very large numbers of simultaneous genetic analyses in analytical devices of small linear dimensions. We have developed a new molecular probe design that is amenable to highly multiplex, specific analyses of total genomic DNA or of RNA molecules expressed in tissue samples. Here, a brief history of gene analytical procedures is presented, followed by a discussion of the properties, means of synthesis and applications of this new class of gene analytic reagents, padlock probes.

DNA↗

PubMed1997

Marked effect of beta-lactoglobulin polymorphism on the ratio of casein to total protein in milk.

The relationship between genetic variants for milk protein and the composition of milk was analyzed on 4475 repeated milk samples from individual cows; 371 dairy cows of the Swedish Red and White breed and 204 cows of the Swedish Holstein breed were used. The registrations included percentages of casein, protein, fat, and lactose in combination with milk yield and SCC. The genotype of individual cows for alpha(s1)-CN, beta-CN, kappa-CN, and beta-LG was determined by alkaline and acidic PAGE. A mixed animal model was used for the analysis; beta-LG and aggregate casein genotypes were included simultaneously as separate fixed effects in the statistical model. The results suggest a positive additive effect of the beta-LG B allele on casein content and on the ratio of casein to total protein. For the latter trait, the beta-LG genotype accounted for a relatively large part of the phenotypic variance, corresponding to a reduction in residual variance of 11% when included in the model. The corresponding value for casein content was 0.5%. The lack of unfavorable associations between milk protein variants and the traits included in this study makes the beta-LG gene an obvious candidate when the breeding objective is improved conversion of milk protein into cheese.

Animals↗

PubMed1997

Characterization of a nuclear factor that binds juxtaposed with ATF3/Jun on a composite response element specifically mediating induced transcription in response to an epidermal growth factor/Ras/Raf signaling pathway.

We have previously identified a 20-bp sequence that mediates induced transcription in response to EGF, Ras, and Raf but not after TPA or UV stimulation. This composite response element, present in a long terminal repeat of a member within the VL30 family of retrotransposons, contains an AP-1-like site that cooperates in function with a juxtaposed sequence unrelated to known transcription factor-binding sites. Using in vitro translated proteins, we here demonstrate that the AP-1-like site preferentially binds ATF3/c-Jun and ATF3/JunD heterodimers. Results from a functional analysis indicate that the ATF3/c-Jun heterodimer, together with factors interacting with the 3' element, are most likely the important mediators of the response because overexpression of JunD, alone or in combination with ATF3, abolishes Ras-induced transcription. Partial purification by phosphocellulose and DNA-affinity chromatography in combination with Southwestern analyses reveals a 52-kDa protein that specifically binds to the sequence juxtaposed to the AP-1-like site. Scatchard analyses show that this sequence, TTAGTTAC, forms two different complexes with K(d)s of 1.9 x 10(-10) and 2.3 x 10(-9) M, respectively. Together, these results suggest that EGF/Ras/Raf induces transcription via combined activation of ATF3/c-Jun and a 52-kDa nuclear factor, whereas JunD acts as a repressor of this response.

Activating Transcription Factor 3↗

PubMed1996

Synthesis of full-length oligonucleotides: cleavage of apurinic molecules on a novel support.

The synthesis of oligodeoxynucleotides is marred by several problems that contribute to the formation of defective molecules. This in turn seriously limits the usefulness of such reagents in DNA diagnostics, molecular cloning, DNA structural analysis and in antisense therapy. In particular, depurination reactions during the cyclical steps of synthesis lead to strand scission during cleavage of the completed molecules from the support. Here we present a remedy to this problem: a novel disiloxyl linkage that connects oligonucleotides to the support withstands reaction conditions that allow the removal of the 5' parts of any depurinated molecules. This ensures that all molecules that preserve the 5' protecting group when cleaved from the support will have both correct 3'- and 5'-ends. We demonstrate the application of the support for synthesis of padlock probe molecules.

Base Sequence↗

PubMed1996

Conformational changes of three periplasmic receptors for bacterial chemotaxis and transport: the maltose-, glucose/galactose- and ribose-binding proteins.

Small-angle X-ray scattering experiments were carried out for the maltose-, glucose/galactose- and ribose-binding proteins of Gram negative bacteria. All were shown to be monomers that decrease in radius of gyration on ligand binding. The results obtained for the maltose-binding protein agree well with crystal structures of the closed, ligand-bound, and open, ligand-free protein, suggesting that these are indeed the primary forms in solution. The closed form is stabilized by protein-sugar interactions, while the open conformation is stabilized by close contacts between the two domains. Since it is the proper special relationship of the domains in the closed form that is most important for interaction with chemotaxis and transport partners, the stabilization of the open form would help keep ligand-free molecules from interfering in function. The scattering results also provide evidence that a large conformational change takes place in association with ligand binding to the glucose/galactose- and ribose-binding proteins, and that the two changes are similar. Modeling suggests that the open forms resemble those found in the related leucine and leucine/isoleucine/valine-binding proteins, but are different from those observed for the maltose-binding protein and the related purine repressor.

Bacterial Proteins↗

PubMed1996

Determination of relative amounts of ribosome and subunits in Escherichia coli using asymmetrical flow field-flow fractionation.

The 30S and 50S subunits and the 70S ribosome of Escherichia coli were separated in 6 minutes by using asymmetrical flow field-flow fractionation (FFF). The total analysis time for determination of the relative amounts of ribosomes and free subunits in a preparation from a cell suspension was 8 min. The method can detect a change in the mass fraction of ribosomes if it exceeds approx, 10%. The separation is based on differences in diffusion coefficients, i.e., hydrodynamic diameters, and these can be determined from observed retention times. The hydrodynamic diameters were in good agreement with literature values obtained from electron microscopy. The mass fraction of ribosomes changed as a function of the magnesium ion concentration which confirms previous knowledge and shows the accuracy of the method. The method appears as an alternative to ultracentrifugation analysis and avoids some of its drawbacks and artefacts. An obvious application can be the optimisation of cell design in metabolic engineering in order to maximise translation and protein production.

Chemical Fractionation↗

PubMed1996

PDGF-A signaling is a critical event in lung alveolar myofibroblast development and alveogenesis.

A mouse platelet-derived growth factor A chain (PDGF-A) null allele is shown to be homozygous lethal, with two distinct restriction points, one prenatally before E10 and one postnatally. Postnatally surviving PDGF-A-deficient mice develop lung emphysema secondary to the failure of alveolar septation. This is apparently caused by the loss of alveolar myofibroblasts and associated elastin fiber deposits. PDGF alpha receptor-positive cells in the lung having the location of putative alveolar myofibroblast progenitors were specifically absent in PDGF-A null mutants. We conclude that PDGF-A is crucial for alveolar myofibroblast ontogeny. We have previously shown that PDGF-B is required in the ontogeny of kidney mesangial cells. The PDGFs therefore appear to regulate the generation of specific populations of myofibroblasts during mammalian development. The two PDGF null phenotypes also reveal analogous morphogenetic functions for myofibroblast-type cells in lung and kidney organogenesis.

Actins↗

PubMed1996

Ca2+-dependent and Ca2+-independent regulation of the thyroid epithelial junction complex by protein kinases.

The integrity of epithelial cell junctions is controlled by E-cadherin-mediated (Ca2+-dependent) cell-cell adhesion. In thyroid follicular cells the dissociation of junctions induced by transfer to low Ca2+ medium (Ca2+ switch) is prevented by thyrotropin acting via cyclic AMP/protein kinase A (cAMP/PKA) (Nilsson et al., Eur. J. Cell Biol. 56, 308-318, 1991). In MDCK kidney epithelial cells protein kinase inhibitors elicit a similar response which, however, is cadherin-independent (Citi, J. Cell Biol. 117,169-178,1992; Citi et al., J. Cell Sci. 107, 683-692, 1994). As such inhibitors also may interfere with PKA, we examined in a single cell type, filter-cultured pig thyrocytes, the effects and possible interactions of the cAMP/PKA agonist forskolin (or thyrotropin) and the kinase inhibitor H-7 in Ca2+ switch experiments. We found that the epithelial barrier dysfunction, comprising loss of transepithelial resistance, increased transepithelial flux of [3H]inulin and redistribution of junction proteins (cadherin and ZO-1), which follows Ca2+ removal were inhibited by TSH, forskolin, and H-7. All agents were also able to induce recovery of resistance in low Ca2+. The maximal recovery effects of forskolin and H-7 were additive when given simultaneous with Ca2+ chelator. In contrast, forskolin-induced recovery initiated 10 min after Ca2+ removal was antagonized by H-7. The protection of junctions by forskolin in low Ca2+ was rapidly abolished by light trypsinization (0.001%), whereas the same concentration of trypsin had little or no effect on the corresponding action of H-7 or staurosporine, another potent kinase inhibitor. In H-7-treated cells kept in low Ca2+, trypsin caused redistribution of ZO-1 from the plasma membrane to the cytoplasm while the transepithelial resistance remained high. Taken together, the data indicate that TSH via cAMP/PKA and the protein kinase inhibitor H-7 reinforce the thyroid epithelial barrier under low Ca2+ conditions by distinct although interacting mechanisms. The high sensitivity to proteolysis in the absence of Ca2+ suggests that the cAMP-regulated mechanism is cadherin-dependent. H-7 promotes or inhibits the cAMP/PKA-mediated recovery of transepithelial resistance depending on the duration of the preceding low Ca2+ period. The trypsin-induced displacement of ZO-1 in H-7-treated cells in low Ca2+ suggests that the localization of ZO-1 to the tight junction is not necessary for the maintenance of junctional tightness.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed1996

Ras-independent activation of Rel-family transcription factors by UVB and TPA in cultured keratinocytes.

UV irradiation of mammalian cells results in the activation of transcription factors which mediate induction of early response genes designed to repair and minimise the damage sustained by the cell. Evidence from studies in HeLa cells suggest that UVC regulates NF-kappa B activity via tyrosine kinases and activation of Ras and Raf kinase. In this study we have used a previously characterized TPA-responsive element (VLTRE) that binds Rel/NF-kappa B proteins and a Ras-responsive element (B10 RRE) to analyse the signalling pathway in UVB-stimulated gene transcription in cultured keratinocytes. We demonstrate that the tumour promoters TPA and UVB use different signalling intermediates to activate different sets of Rel/NF-kappa B proteins. UVB transactivation is independent of PKC activity but dependent on tyrosine kinase activity where was TPA stimulation requires PKC but not tyrosine kinase activity. Furthermore, neither UVB- nor TPA-transactivation is mediated through p21 Ras but both stimuli are dependent on a functional Raf protein. A constitutively active Raf-1 kinase however, was unable to induce transactivation through VLTRE. Thus, Raf has an essential but permissive role in UVB activation of Rel proteins. These findings demonstrate that keratinocytes contain a novel Ras-independent pathway for induction of Rel mediated transcription.

Animals↗

PubMed1996

Penetration of antibiotics in diseased antral mucosa.

OBJECTIVES: To investigate extracellular concentrations of antibiotics in inflammatory diseased sinus mucosa in critically ill patients and to find out whether there were any correlations between the concentrations of antibiotics in extracellular tissue and the presence of bacterial infection. DESIGN: Prospective case series. Sinoscopic findings were used as the criterion standard. SETTING: The general and the neurosurgical intensive care units of a tertiary care hospital. PATIENTS: Critically ill patients, nasotracheally intubated or tracheotomized, who received ventilator treatment (> 7 days) and antibiotic therapy. INTERVENTIONS: Bilateral sinoscopies were performed; antral secretions were collected for bacterial cultures; and serum and tissue samples were obtained for bioassay of antibiotic concentrations. RESULTS: The concentrations of all antibiotics found in extracellular fluid were lower than those found in serum samples. CONCLUSION: Measurable extracellular tissue concentrations of antibiotics were found in the antral mucosa, demonstrating that inflammatory maxillary sinus disease does not impede the antibiotic penetration of the mucosa or antibiotic activity in the extracellular tissue.

Adult↗

PubMed1996

Detecting Genes with Ligases

The combination of synthetic oligonucleotide probes and DNA ligases is central to several recently developed genetic assays. Among the advantages of ligase-mediated gene detection is that ligation of probe pairs provides highly specific detection of unique DNA sequences in genomic samples. The technique also allows for convenient distinction between sequence variants, since mismatched bases at the junction of the probe pair prevent ligation. Moreover, the circumstance that two probes are joined into one molecule can be exploited for detection in several ways, for instance by observing the change in probe size upon ligation. Alternatively, a detectable function on one probe can be demonstrated to become linked to a retrievable function on another one through ligation. Ligation products can also be recruited as templates for subsequent ligation reactions in powerful amplification schemes. So-called padlock probes lock to their targets by encircling them, remaining in place even after denaturing washes. Here, we will describe two ligase-mediated assays: one that serves to monitor the presence of common sequence variants in amplified samples of genomic DNA and another that is suitable to detect localized gene sequences.

Journal Article↗

PubMed1996

Morphologic conversion of Helicobacter pylori from bacillary to coccoid form.

The morphologic conversion of Helicobacter pylori from bacillary to coccoid form was studied by microscopy, viable count on agar plates, and bioluminescence assay of bacterial ATP. When morphologic conversion from bacillary to coccoid form was detected by microscopy, the viable counts and the bacterial ATP decreased. No viable count was found after nine days of incubation, but bacterial ATP was still present. In these cultures in which only the coccoid form of Helicobacter pylori was present, there was no accumulation of extracellular ATP, indicating no leaky cells. During the transition phase from the bacillary to the coccoid form of Helicobacter pylori, the addition of fresh medium increased the intracellular ATP 26-fold. The coccoid form of Helicobacter pylori had a 1000-fold lower ATP level per cell compared to the bacillary form, which indicates a decreased metabolic activity in the coccoid form. Addition of fresh medium to the coccoid cultures from days 9 and 10 increased the ATP level twofold. However, no conversion from coccoid to bacillary form was found in these cultures during prolonged incubation in fresh broth for four weeks. Such conversion needs to be demonstrated before it is proven that the coccoid form of Helicobacter pylori is responsible for transmission and relapse of infection.

Adenosine Triphosphate↗

PubMed1996

Diagnostic accuracy of direct digital dental radiography for the detection of periapical bone lesions. II. Effects on diagnostic accuracy after application of image processing.

OBJECTIVES: The diagnostic accuracy of direct digital radiography for the detection of small experimentally made periapical lesions was evaluated. A comparison was made between original digital images and images processed with different enhancement facilities of the digital system. STUDY DESIGN: Seven observers assessed the digital images in original mode and after individual image treatment. The diagnostic accuracy was calculated. The processing functions used by the observers were recorded, and the effect of processing was evaluated. RESULTS: The overall diagnostic accuracy was not different for the two image modes. Neither were the individual results of the observers different. The image processing was most effective when alterings of contrast and brightness were used. More complicated processing procedures had less effect on the diagnostic accuracy. CONCLUSIONS: Image processing of direct digital images of high quality has a limited effect on the diagnostic accuracy. Basic processing functions, that is, alterings of contrast and brightness, were preferred for the detection of periapical lesions.

Alveolar Bone Loss↗

PubMed1996

Diagnostic accuracy of direct digital dental radiography for the detection of periapical bone lesions: overall comparison between conventional and direct digital radiography.

OBJECTIVES: The diagnostic accuracy of direct digital radiography for the detection of small, experimentally made, periapical lesions was compared with that of E speed film. The high- and low-contrast resolutions of the imaging systems were also compared. STUDY DESIGN: The high-contrast resolution was evaluated with a line pair plate. The low-contrast resolution was determined with a contrast-detail plexiglass phantom. To evaluate the diagnostic accuracy, the periapical region of dry human mandibles was examined. Receiver operating characteristic based on the readings of seven observers were generated. RESULTS: The high-contrast resolution of the digital system was inferior, but the low-contrast resolution was comparable to that of E-speed film. For diagnostic accuracy no significant difference was seen between the areas under the receiver operating characteristic curves. CONCLUSIONS: The quality of the direct digital images is comparable to that of E-speed film for the detection of periapical bone lesions.

Alveolar Bone Loss↗

PubMed1996

Simple methods for the estimation of dose distributions, organ doses and energy imparted in paediatric radiology.

The energy imparted and the effective dose can both be used to describe the risk to the patient in diagnostic radiology. Simple methods must be employed to determine these quantities in clinical situations. Methods using measured relative depth-dose distributions are presented and evaluated here. Measurements of depth-dose distributions for x-ray beams were performed with an ionization chamber, a diode and a number of TL dosimeters. The energy imparted was calculated from measurements with both phantoms and patients. The method of calculating the mean absorbed dose to organs was applied to pelvis and lumbar spine examinations. TL dosimeters were found to be an appropriate detector for measuring depth-dose distributions. When calculating the energy imparted the entrance beam area must be accurately known. The mean absorbed dose to organs can be derived from measured relative depth-dose curves if accurate information on entrance beam position and area is available for the particular examination technique used. The advantage of these methods is that the dose distribution is measured for the photon beam used for the examination of the patients.

Adolescent↗