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Biomedical subjects

L Nilsson

Publications and source records attributed to L Nilsson.

At least 163 records · Page 9Linked to original sources
PubMed1995

Modulation of DNA-binding specificity within the nuclear receptor family by substitutions at a single amino acid position.

Regulation of gene expression involves a large number of transcription factors with unique DNA-binding properties. Many transcription factors belong to families of related proteins that bind to similar but distinct sequences. In this study we have analyzed how amino acid substitutions at a single position in the DNA-binding domain modulate the DNA-binding specificity within the nuclear receptor family of transcription factors. All possible amino acids were introduced at the first position in the DNA recognition helix, and the specificities of the mutants were analyzed using response elements containing all combinations of bases at two variable base pair positions. All mutant proteins were functional in DNA binding, and could be divided into classes of mutants with different response element specificities. By combining functional data with analysis of the structural effects of the mutations by molecular modeling, we could identify both prohibitive steric interactions as well as positive interactions, such as hydrogen bonds, that function as important determinants for specificity. Only the residues found naturally in the glucocorticoid and estrogen receptors, glycine and glutamate, produce unique binding specificities. The specificities of the other mutants overlap with each other somewhat but the substitutions clearly have potential to contribute to diversity within the nuclear receptor family.

Amino Acid Sequence↗
PubMed1995

Cumulative birth rates following cryopreservation of all embryos in stimulated in vitro fertilization (IVF) cycles.

OBJECTIVE: To cryopreserve all good-quality embryos available after IVF is one way to avoid an impending hyperstimulation and a more attractive alternative to a couple than cancellation of the cycle. At Sahlgrenska University Hospital this method has been practiced since 1991. The aim of this study was to assess the success rate (defined as childbirth per couple) after IVF treatment including one stimulated cycle, resulting in cryopreservation of all good-quality embryos available, followed by replacement of frozen/thawed embryos in one or more natural cycles. DESIGN: A cohort of 32 women undergoing one stimulation for IVF between January 1991 and December 1993 where all good-quality embryos were cryopreserved and transferred in a later spontaneous cycle were studied retrospectively. The cumulative childbirth rate per couple was calculated. RESULTS: A total of 28 women underwent a total of 45 transfers with frozen-thawed embryos. In 4 women no transfer has taken place so far. The clinical pregnancy rate per embryo transfer was 33% (15/45) and per patient 54% (15/28). Three spontaneous abortions occurred giving a cumulative childbirth rate per patient of 39% (11/28). In addition, 6 out of the 28 women still have embryos left in the freezer. CONCLUSION: Cryopreservation of all good-quality embryos available after IVF is a highly effective alternative to cancellation of a cycle when there is impending hyperstimulation. The use of this approach results in a cumulative childbirth rate per couple of at least 39%. Since only one stimulation was performed, it also seems to be cost effective.

Abortion, Spontaneous↗
PubMed1995

The effects of a mobile telephone task on driver behaviour in a car following situation.

The effects of a mobile telephone task on young and elderly drivers' choice reaction time, headway, lateral position, and workload were studied when the subjects were driving in a car-following situation, in the VTI driving simulator. It was found that a mobile telephone task had a negative effect upon the drivers' choice reaction time, and that the effect was more pronounced for the elderly drivers. Furthermore, the subjects did not compensate for their increased reaction time by increasing their headway during the phone task. The subjects' mental workload, as measured by the NASA-TLX, increased as a function of the mobile telephone task. No effect on the subjects' lateral position could be detected. Taken together, these results indicate that the accident risk can increase when a driver is using the mobile telephone in a car following situation. The reasons for the increased risk, and possible ways to eliminate it, are also discussed.

Accidents, Traffic↗
PubMed1995

Eosinophil granule proteins in cardiopulmonary bypass with and without heparin coating.

Extracorporeal circulation with exposure of blood to foreign surfaces causes activation of different defense systems, eg, white cells. Several potent mediators are released into plasma, capable of causing harmful effects to different organs, contributing to postoperative morbidity after operations using cardiopulmonary bypass. The eosinophil granulocyte has not previously been investigated in this respect. We studied two of its activation products, eosinophil cationic protein and eosinophil protein X in coronary bypass patients. In 17 control patients, plasma levels of eosinophil cationic protein and eosinophil protein X increased considerably during cardiopulmonary bypass. In 19 patients with heparin-coated cardiopulmonary bypass equipment the levels were significantly reduced, indicating improved biocompatibility of the cardiopulmonary bypass circuit. The heparin-coated surface causes less activation of eosinophils; also released eosinophil cationic protein is bound to the heparinized surface.

Blood Proteins↗
PubMed1995

Anaerobic biodegradation potentials in digested sludge, a freshwater swamp and a marine sediment.

An anaerobic gas production test was used for determining the potential biodegradation of 22 organic chemicals under methanogenic conditions. Nine of the examined chemicals were extensively mineralized (> 75%) both in sewage sludge and in a freshwater swamp indicating good agreement between biodegradation potentials in these habitats. Samples from a marine sediment showed a less extensive mineralization of most of the test chemicals, and lag periods of several weeks often preceded net gas production. As marine sediments usually contain sulfate at the time of collection, the assessment of biodegradation potentials in such environments is probably more reliable when using a method based on sulfate reduction instead of methanogenic gas production. The results of the tests indicate that the commonly recommended washing of sludge solids can be eliminated by applying a more diluted inoculum.

Anaerobiosis↗
PubMed1995

Diminution of preprosomatostatin-mRNA in cerebral cortex of the aged rat.

The aim of the present study was to examine the effect of normal aging on somatostatin neurotransmission. Somatostatin gene expression was analysed in several regions of the cerebral cortex and hippocampus in 3, 7 and 21 month-old Sprague-Dawley rats using quantitative in situ hybridization with a 48mer oligodeoxynucleotide antisense probe. Furthermore the distribution of 125I-Tyr11 somatostatin receptor binding sites was studied using quantitative receptor autoradiography. The results demonstrated a significant reduction of preprosomatostatin-mRNA in the frontal cortex of the aged (21 month) group compared with the young (3 month) and the middle-aged (7 month) groups. The receptor binding densities of the aged (21 month) group tended to be lower, compared to the other groups although no significant region-specific changes were evident. These results indicate neurochemical changes in somatostatin-containing neurons in the frontal cortex during aging.

Aging↗
PubMed1995

On the pH dependence of amide proton exchange rates in proteins.

We have analyzed the pH dependencies of published amide proton exchange rates (kex) in three proteins: bovine pancreatic trypsin inhibitor (BPTI), bull seminal plasma proteinase inhibitor IIA (BUSI IIA), and calbindin D9K. The base-catalyzed exchange rate constants (kOH) of solvent exposed amides in BPTI are lower for residues with low peptide carbonyl exposure, showing that the environment around the carbonyl oxygen influences kOH. We also examined the possible importance of an exchange mechanism that involves formations of imidic acid intermediates along chains of hydrogen-bonded peptides in the three proteins. By invoking this "relayed imidic acid exchange mechanism," which should be essentially acid-catalyzed, we can explain the surprisingly high pHmin (the pH value at which kex reaches a minimum) found for the non-hydrogen-bonded amide protons in the beta-sheet in BPTI. The successive increase of pHmin along a chain of hydrogen-bonded peptides from the free amide to the free carbonyl, observed in BPTI, can be explained as an increasing contribution of the proposed mechanism in this direction of the chain. For BUSI IIA (pH 4-5) and calbindin D9K (pH 6-7) the majority of amide protons with negative pH dependence of kex are located in chains of hydrogen-bonded peptides; this situation is shown to be consistent with the proposed mechanism.

Amides↗
PubMed1995

Potential of mean force calculations of the stacking-unstacking process in single-stranded deoxyribodinucleoside monophosphates.

The free energy of the stacking-unstacking process of deoxyribodinucleoside monophosphates in aqueous solution has been investigated by potential of mean force calculations along a reaction coordinate, defined by the distance between the glycosidic nitrogen atoms of the bases. The stacking-unstacking process of a ribodinucleoside monophosphate was observed to be well characterized by this coordinate, which has the advantage that it allows for a dynamical backbone and flexible bases. All 16 naturally occurring DNA dimers composed of the adenine, cytosine, guanine, or thymine bases in both the 5' and the 3' positions were studied. From the free-energy profiles we observed the deepest minima for the stacked states of the purine-purine dimers, but good stacking was also observed for the purine-pyrimidine and pyrimidine-purine dimers. Substantial stacking ability was found for the dimers composed of a thymine base and a purine base and also for the deoxythymidylyl-3',5'-deoxythymidine dimer. Very poor stacking was observed for the dCpdC dimer. Conformational properties and solvent accessibility are discussed for the stacked and unstacked dimers. The potential of mean force profiles of the stacking-unstacking process for the DNA dimers are compared with the RNA dimers.

DNA↗
PubMed1995

Molecular dynamics simulations of the glucocorticoid receptor DNA-binding domain in complex with DNA and free in solution.

Molecular dynamics simulations have been performed on the glucocorticoid receptor DNA binding domain (GR DBD) in aqueous solution as a dimer in complex with DNA and as a free monomer. In the simulated complex, we find a slightly increased bending of the DNA helix axis compared with the crystal structure in the spacer region of DNA between the two half-sites that are recognized by GR DBD. The bend is mainly caused by an increased number of interactions between DNA and the N-terminal extended region of the sequence specifically bound monomer. The recognition helices of GR DBD are pulled further into the DNA major groove leading to a weakening of the intrahelical hydrogen bonds in the middle of the helices. Many ordered water molecules with long residence times are found at the intermolecular interfaces of the complex. The hydrogen-bonding networks (including water bridges) on either side of the DNA major groove involve residues that are highly conserved within the family of nuclear receptors. Very similar hydrogen-bonding networks are found in the estrogen receptor (ER) DBD in complex with DNA, which suggests that this is a common feature for proper positioning of the recognition helix in ER DBD and GR DBD.

Amino Acid Sequence↗
PubMed1995

Structural basis for X-linked agammaglobulinemia (XLA): mutations at interacting Btk residues R562, W563, and A582.

It has been suggested that tryptophan 563 is sandwiched between residues R562 and A582 in Bruton's agammaglobulinemia tyrosine kinase (Btk). Mutations of the surrounding residues have been shown to cause X-linked agammaglobulinemia. Substitutions R562P and A582V were noticed to have impaired kinase activity. However, based on Western blot analysis, the mutant proteins were expressed at normal levels. Molecular modeling of the kinase domain has previously indicated that these residues presumably govern the position of the W563 side chain, which is thought to interact with the catalytic loop. W563 is inside the molecule and too far away from the catalytic center to interact directly with the substrate or cofactors. To prove these model-based conclusions, a conservative substitution with phenylalanine for W563 was made, and the resultant mutant lacked kinase activity. These results confirm our previous assumption that the side chain of W563, invariant in protein tyrosine kinases, is crucial for Btk kinase activity. Mutations in the surrounding residues seem to inactivate Btk by affecting the location of W563.

Agammaglobulinaemia Tyrosine Kinase↗
PubMed1995

Amyotrophic lateral sclerosis associated with homozygosity for an Asp90Ala mutation in CuZn-superoxide dismutase.

Recent reports have shown heterozygosity for some twenty different mutations in the CuZn-superoxide dismutase (CuZn-SOD) gene in familial amyotrophic lateral sclerosis (FALS), and analysed samples from patients have shown decreased enzymic activity. Here we report homozygosity for an exon 4 mutation, Asp90Ala in fourteen patients among four unrelated ALS families and four apparently sporadic ALS patients from Sweden and Finland. The erythrocyte CuZn-SOD activity is essentially normal. Our findings suggest that this CuZn-SOD mutation causes ALS by a gain of function rather than by loss, and that the Asp90Ala mutation is less detrimental than previously reported mutations.

Adult↗
PubMed1995

Microfertilization techniques--the Swedish experience.

Intracytoplasmic sperm injection (ICSI) has been studied in this animal research programme since 1990. In 1993, the technique was first applied clinically and up to the present time (September 1994), a total of 456 couples have been studied in 538 cycles. The principal indication for the use of ICSI has been severe male sub-fertility as judged by a semen analysis. In addition, men with high titres of antisperm antibodies, blockage of the vas deferens and neurological disorders such as spinal cord lesions have been included in the programme. Men with genetic disorders such as cystic fibrosis and acrosome-deficient spermatozoa have also been treated successfully. The overall fertilization rate using ICSI was 59%, which is similar to the conventional in vitro fertilization (IVF) programme in Göteborg, however, the pregnancy rate per embryo transfer (29%) and the ongoing pregnancy rate per transfer (22%) were slightly lower. The total number of pregnancies was 144 with 111 of the pregnancies either ongoing or already delivered. To date, 36 healthy children have been born following 29 deliveries and no major malformations have been diagnosed. Being the first programme in Scandinavia to perform ICSI, this unit has experienced long waiting lists which indicates that severe male sub-fertility will be one of the major groups for treatment with assisted reproductive technologies in the future.

Cryopreservation↗
PubMed1995

DNA-based mutation analysis of Bruton's tyrosine kinase gene in patients with X-linked agammaglobulinaemia.

The identification of the BTK (Bruton's tyrosine kinase) gene defective in human immunoglobulin deficiency X-linked agammaglobulinaemia (XLA) and characterisation of BTK exon-intron boundaries has now allowed the analysis of mutations and polymorphisms at the level of genomic DNA. Using Southern blot analysis and the polymerase chain reaction single strand conformation polymorphism (PCR-SSCP) assay, amplifying all 19 exons and the putative promoter region with a single annealling temperature, mutations have been identified in 19 out of 24 unrelated patients diagnosed as having XLA. Apart from a large deletion involving exon 19, nine missense (F25S, R288W, 1370M, M509V, R525P, N526K, R562W, A582V and G594R), two nonsense (E277X and R525X), five frameshift and two splice site mutations have been found affecting most coding exons and all major enzyme domains. No mutations or polymorphisms were detected in the putative promoter region. A single nucleotide deletion located in the last exon, resulting in a truncation of the eight C-terminal residues of Btk and a typical XLA phenotype, indicates structural and/or functional importance of Btk helix I in the catalytic domain. Although allelic heterogeneity at the BTK locus may partly explain clinical variability in families with XLA, compensatory and redundant mechanisms involved in B-cell development must play a role in the phenotypic diversity of the disease.

Adolescent↗
PubMed1995

The human I alpha 1 region contains a TGF-beta 1 responsive enhancer and a putative recombination hotspot.

It appears that the switch recombination machinery of a B lymphocyte targets preferentially unrearranged heavy chain genes that have been rendered transcriptionally active. Transcriptional activation of the 'germline' human C alpha 1 and C alpha 2 genes is triggered by TGF-beta 1 and is controlled by proximal positive and distal negative regulatory elements residing upstream of the alpha 1 and alpha 2 switch regions respectively. In this report we characterize the positive proximal regulatory elements and analyse their interaction with DNA binding proteins. Our data demonstrate that a 100 bp fragment that contains a cAMP responsive element (CRE)/activating transcription factor (ATF) motif, a putative Ets binding site and an element that is created by two previously described neighbouring direct repeats (DRE), can increase the basal level of transcription and confer TGF-beta 1 inducibility to a heterologous promoter in an orientation- and position-independent manner. Ubiquitously expressed DNA binding proteins interact specifically with the CRE/ATF, the Ets site and the DRE element. Additionally, nuclear proteins interact with sequences which are located downstream of this enhancer are not essential for transcription in the transient expression assays utilized; however, they contain motifs that have been previously implicated in regulating DNA recombination events. These motifs include a Chi motif and a Chi-like element previously found in the recombination hotspot region of the Bcl-2 proto-oncogene and close to chromosomal breakpoints in T-ALL lines. Our findings raise the possibility that the intervening region associated regulatory elements in addition to regulating the transcriptional activation of the Ig heavy chain genes could also facilitate the physical interaction of transcription and recombination controlling molecular mechanisms.

Base Sequence↗
PubMed1995

Gratuitous overexpression of genes in Escherichia coli leads to growth inhibition and ribosome destruction.

We attempted to test the idea that the relative abundance of each individual tRNA isoacceptor in Escherichia coli can be altered by varying its cognate codon concentration. In order to change the overall codon composition of the messenger pool, we have expressed in E. coli lacZ with the aid of T7 RNA polymerase so that their respective gene products individually accounted for 30% of the total bacterial protein. Unexpectedly, the maximum expression of either test gene has no specific effect on the relative rates of synthesis of the tRNA species that we studied. Instead, we find that there is a cumulative breakdown of rRNAs, which results in a loss of ribosomes and protein synthetic capacity. After either of the test genes is maximally induced, there is a growing fraction of protein synthesis invested in beta-galactosidase or delta tufB that is matched by a comparable decrease of the fraction of normal protein synthesis. We have also observed enhanced accumulation of two heat shock proteins during overexpression. Finally, after several hours of overexpression of either test protein, the bacteria are no longer viable. These results are relevant to the practical problems of obtaining high expression levels for cloned proteins.

Bacterial Proteins↗
PubMed1995

Results from in vitro fertilization and intracytoplasmic sperm injection with purified gonadotrophins (Metrodin HP).

Purified urinary follicle-stimulating hormone (uFSH-HP; Metrodin HP, Serono Ltd.) was compared with a combination of pure FSH and human menopausal gonadotrophin (hMG; Pergonal, Serono Ltd.) in patients undergoing standard in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI). In standard IVF, pure FSH gave a significantly higher pregnancy rate per started cycle than did the combination with hMG (35 vs. 18%, p < 0.05). No differences between standard IVF and ICSI were seen which could be associated with hormonal stimulation in an open non-randomized series of patients. In 11 ICSI cycles, the use of recombinant FSH (Gonal F, Serono Ltd.) resulted in 4 ongoing pregnancies.

Cytoplasm↗
PubMed1995

An improved method for determination of thiocyanate in plasma and urine.

An improved spectrophotometric method is described for the determination of thiocyanate in plasma and urine. Thiocyanate is adsorbed on a weak anion-exchange resin with strong affinity for chaotropic ions, and eluted with perchlorate. Thiocyanate is then chlorinated by hypochlorite and quantified according to the König (J Prakt Chem 1904; 69:105-37) reaction by use of isonicotinic acid and 1,3-dimethyl-barbituric acid. The method affords a simple, rapid and sensitive assay for thiocyanate and has a detection limit of 0.93 mumol/l. At thiocyanate concentrations of 107.1 and 167.4 mumol/l in plasma and urine the within-day CVs were 0.69% and 1.1% respectively, and the total imprecision measured for a period of 65 days was 0.98%. Analytical recoveries were quantitative both with urine and plasma samples.

Barbiturates↗