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Biomedical subjects

A Karlsson

Publications and source records attributed to A Karlsson.

At least 109 records · Page 6Linked to original sources
PubMed1998

Intracellular metabolism of the N7-substituted acyclic nucleoside analog 2-amino-7-(1,3-dihydroxy-2-propoxymethyl)purine, a potent inhibitor of herpesvirus replication.

We investigated the intracellular metabolism of S2242 (2-amino-7-(1,3-dihydroxy-2-propoxymethyl)purine), the only known antivirally active acyclic nucleoside analogue with the side chain substituted at the N7 position of the purine ring. Uptake of S2242 by CEM cells increased linearly with increasing extracellular concentrations of the compound and was blocked by inhibitors of nucleoside transport. S2242 was phosphorylated in a time- and concentration-dependent manner to its monophosphates, diphosphates, and triphosphates. Intracellular half-life of the diphosphates and triphosphates in CEM cells was approximately 3-6 hr. A strong correlation was found between the cytostatic action of the compound and its phosphorylation in different cell lines. In accord with the findings that (1) the cytostatic potential of S2242 is reversed by deoxycytidine (dCyd) and (2) the growth of deoxycytidine kinase-deficient (dCK-) cells is refractory to the inhibitory effect of S2242, the amount of metabolites formed from S2242 in the dCK- cell line was approximately one hundredth of that in the wild-type cells. The observation that purified dCK phosphorylates S2242 to its monophosphate further corroborates these results. The activity of S2242 against herpes simplex virus, varicella-zoster virus, and human herpesvirus type 6 was reversed by 50-100-fold on the addition of exogenous dCyd. Compound S2242 was not preferentially phosphorylated in herpes simplex virus 1-, varicella-zoster virus-, or human herpesvirus type 6-infected cells (Vero, human embryonic lung, and HSB-2 cells, respectively), and exogenously added dCyd reduced substantially the formation of S2242 metabolites in these cells. In human cytomegalovirus (HCMV)-infected human embryonic lung cells, a 5-25-fold increase in S2242 metabolite formation was observed compared with the noninfected cells, suggesting that an HCMV-encoded or -induced enzyme causes the specific phosphorylation of S2242. Exogenously added dCyd had little effect on the activity of S2242 against HCMV and on the phosphorylation of the compound in HCMV-infected cells. S2242 was not specifically phosphorylated by the HCMV-encoded UL-97 kinase in cells infected with a vaccinia/UL-97 recombinant. S2242 was found to be a substrate (K(m) = 90 microM) for purified human deoxyguanosine kinase; the latter enzyme was stimulated 3-4-fold in HCMV-infected cells.

Alkaline Phosphatase↗
PubMed1998

Evaluation of the Etest method for determining fluconazole susceptibilities of 402 clinical yeast isolates by using three different agar media.

The performance of the Etest for fluconazole susceptibility testing of 402 yeast isolates was assessed against the National Committee for Clinical Laboratory Standards (NCCLS) microdilution broth method. The NCCLS method employed RPMI 1640 broth medium, and MICs were read after incubation for 48 h at 35 degrees C. Etest MICs were determined with RPMI agar containing 2% glucose (RPG), Casitone agar (CAS), and Mueller-Hinton agar (MHA) and were read after incubation for 48 h at 35 degrees C. The yeast isolates included Candida albicans (n = 161), Candida glabrata (n = 41), Candida tropicalis (n = 35), Candida parapsilosis (n = 29), Candida krusei (n = 32), Candida lusitaniae (n = 31), Candida species (n = 19), Cryptococcus neoformans (n = 40), and miscellaneous yeast species (n = 14). The Etest results correlated well with reference MICs. Overall agreement was 94% with RPG, 97% with CAS, and 53% with MHA. When RPG was used, agreement ranged from 89% for Candida spp. to 100% for C. krusei. When CAS was utilized, agreement ranged from 93% for Cryptococcus neoformans to 100% for C. tropicalis, C. parapsilosis, C. lusitaniae, Candida spp., and miscellaneous yeast species. With MHA, agreement ranged from 17% for C. parapsilosis to 90% for C. krusei. Both RPG and CAS supported growth of all yeast species, whereas growth on MHA was comparatively weaker. Etest results were somewhat easier to read on CAS. The Etest method using either RPG or CAS, but not MHA, appears to be a viable alternative to the NCCLS reference method for determining fluconazole susceptibilities of yeasts.

Agar↗
PubMed1998

Differential intracellular compartmentalization of herpetic thymidine kinases (TKs) in TK gene-transfected tumor cells: molecular characterization of the nuclear localization signal of herpes simplex virus type 1 TK.

The thymidine kinases (TKs) of herpes simplex virus type 1 (HSV-1), HSV-2, and varicella-zoster virus (VZV) were expressed in human osteosarcoma cells as fusion proteins with the green fluorescent protein (GFP), and their intracellular localizations were determined. The three TK-GFP fusion products were localized in different subcellular compartments of the transfected tumor cells. HSV-1 TK-GFP was localized exclusively in the nucleus, HSV-2 TK-GFP was predominantly found in the cytosol, while VZV TK-GFP was localized in both the nucleus and the cytosol. In support of these findings, we identified a nuclear localization signal (NLS) in the N-terminal arginine-rich region of HSV-1 TK that was absent in HSV-2 and VZV TK. The first 34 amino acids proved necessary for the specific nuclear localization of HSV-1 TK and, when added to the VZV TK-GFP gene construct, also sufficed to specifically target VZV TK-GFP to the nucleus. Further analysis of this NLS through site-directed mutagenesis revealed that the basic amino acid-rich nonapeptide 25R-R-T-A-L-R-P-R-R33 is of crucial importance in the nuclear targeting of HSV-1 TK. In particular, we revealed that the presence of the arginine residues at positions 25, 26, 30, 32, and 33 is obligatory for efficient NLS functioning, whereas arginine and histidine residues outside of the nonapeptide (i.e., residues R18, R20, and H22) did not change the functional properties of the NLS.

Amino Acid Sequence↗
PubMed1998

Thermal and mechanical characterization of cellulose acetate phthalate films for pharmaceutical tablet coating: effect of humidity during measurements.

Films from a polymer used in pharmaceutical coating (cellulose acetate phthalate) were analyzed by thermomechanical techniques including dynamic mechanical thermal analysis (DMTA) and tensile tests. Emphasis was placed on relative humidity (RH) at the measurement site (as opposed to storage or conditioning RH). The films were plasticized with either triethyl citrate or diethyl phthalate. The results show that the films respond rapidly to changes in the environmental humidity. This in turn influences the data obtained from DMTA and tensile testing; thus, good control of the humidity is essential. Absorption isotherms have been obtained for the two types of films, and the results were interpreted in terms of the equilibrium moisture content which is determined by the polar nature of the plasticizer. This factor must be considered when formulating a film composition because moisture can, apart from its influence on mechanical properties, also speed the chemical degradation processes. When the stability of a film composition during storage is studied, the actual measurements on the films should preferably be done at 0% RH, irrespective of the storage RH. At 0% RH, the mechanical testing results will reflect the chemical stability correctly, without interference from plasticizing effect of absorbed water. A functionality test for films has been suggested: DMTA under isothermal conditions using either step or continuous humidity scans. Isothermal dynamic humidity scans on the DMTA were performed for the first time, and the utility of these measurements is discussed.

Cellulose↗
PubMed1998

Characterization of cellulose acetate phthalate (CAP).

Cellulose acetate phthalate (CAP) is a commonly used enteric coating polymer. CAP powder has been studied by various methods to determine characteristics that have an influence on its functionality. While some of the parameters are well known, such as free-acid content and substituent composition, new methods have been developed to examine them. Other characteristics, such as the molecular mass distribution, have not been reported earlier. Fourier transform infrared spectroscopy (FTIR), nuclear magnetic resonance (NMR), and thermal analysis have also been performed on fresh samples, as well as samples stored under various temperature and humidity conditions. Humidity is by far a more critical storage parameter than temperature, although the two act in conjunction; high humidity is more deleterious to the functionality of the polymer than high temperature. Functionality in this case is taken to be determined by the substituents and by the molecular mass distribution. Mass-average molecular mass of a number of batches of the polymer has been measured and ranges around 48 kg/mol with a degree of polydispersity of 1.6. A method to perform a rough estimation of the molecular mass of CAP has also been suggested based on knowledge of the substituent content. It may be possible to use the values of and obtained here for any other batch of the same viscosity grade of CAP. NMR has been employed to determine the fraction substituents in the polymer. However, an attempt to obtain the pattern of substitution of the CAP molecule by NMR was unsuccessful. Glass transition temperatures of CAP samples were measured. However, this characteristic of the polymer is judged not as sensitive to the loss of substituents as the molecular mass. Thermal treatment of the polymer in oxygen and inert atmospheres gave slightly different degradation products.

Acetic Acid↗
PubMed1997

Human deoxycytidine kinase is located in the cell nucleus.

Human deoxyribonucleoside kinases are required for the pharmacological activity of several clinically important anticancer and antiviral nucleoside analogs. Human deoxycytidine kinase and thymidine kinase 1 are described as cytosolic enzymes in the literature, whereas human deoxyguanosine kinase and thymidine kinase 2 are believed to be located in the mitochondria. We expressed the four human deoxyribonucleoside kinases as fusion proteins with the green fluorescent protein to study their intracellular locations in vivo. Our data showed that the human deoxycytidine kinase is located in the cell nucleus and the human deoxyguanosine kinase is located in the mitochondria. The fusion proteins between green fluorescent protein and thymidine kinases 1 and 2 were both predominantly located in the cytosol. Site-directed mutagenesis of a putative nuclear targeting signal, identified in the primary structure of deoxycytidine kinase, completely abolished nuclear import of the protein. Reconstitution of a deoxycytidine kinase-deficient cell line with the wild-type nuclear or the mutant cytosolic enzymes both restored sensitivity toward anticancer nucleoside analogs. This paper reports that a deoxyribonucleoside kinase is located in the cell nucleus and we discuss the implications for deoxyribonucleotide synthesis and phosphorylation of nucleoside analogs.

Amino Acid Sequence↗
PubMed1997

Cloning of the cDNA and chromosome localization of the gene for human thymidine kinase 2.

Human thymidine kinase 2 (TK2) is a deoxyribonucleoside kinase that phosphorylates thymidine, deoxycytidine, and deoxyuridine. The enzyme also phosphorylates anti-viral and anti-cancer nucleoside analogs. We have identified an expressed sequence tag cDNA that encoded a 27.5-kDa protein approximately 30% similar to the human deoxycytidine kinase and deoxyguanosine kinase. The protein was expressed in Escherichia coli and shown to have similar substrate specificity as reported for purified native human TK2. The recombinant TK2 was shown to phosphorylate the anti-cancer nucleoside analog 2',2'-difluorodeoxycytidine. Northern blot analysis showed two mRNA species at 2.4 and 4.0 kilobases predominantly expressed in liver, pancreas, muscle, and brain. We identified a sequence-tagged site designed from the 3' region of the TK2 cDNA. The sequence-tagged site has been mapped to 81-84 centimorgans from the top linkage group of chromosome 16, which corresponds to the 16q22 region. Our data show that deoxycytidine kinase, deoxyguanosine kinase, and TK2 belong to a family of closely related enzymes. At the time of this report all four of the known human deoxyribonucleoside kinases have been cloned. This provides the opportunity to characterize their individual contribution to therapeutic and toxic effects of nucleoside analogs.

Amino Acid Sequence↗
PubMed1997

Desensitization of the fMLP-induced NADPH-oxidase response in human neutrophils is lacking in okadaic acid-treated cells.

The chemoattractant N-formyl-methionyl-leucyl-phenylalanine (fMLP) interacts with neutrophils, generating signals that induce activation of the superoxide anion/hydrogen peroxide-producing NADPH-oxidase. Low temperature binding of fMLP to its neutrophil surface receptors is associated with a desensitization of the cells with respect to activation of the oxidase. Other stimuli can still activate the oxidase (in fact even induce a primed response), indicating that the observed phenomenon is stimulus specific and could not be accounted for by an effect on the oxidase itself. Furthermore, no desensitization is obtained in the presence of cytochalasin B, suggesting that the cytoskeleton is involved in the process leading to desensitization. Okadaic acid is a toxin produced by dinoflagellates and exerts its effects by an inhibition of cellular phosphatases. To investigate the role of phosphorylation/dephosphorylation events in the desensitization process we used okadaic acid as a scientific tool. We show that neutrophils treated with okadaic acid are primed with respect to the fMLP-induced production of superoxide anion, and that no desensitization is obtained in toxin-treated cells. Because the recovery of ligand-receptor complexes in a Triton X-100-insoluble fraction is very low in the cells treated with okadaic acid, we suggest that protein dephosphorylation is required to obtain binding to the cytoskeleton of occupied fMLP receptors; binding of the occupied receptors to the cell cytoskeleton being the mechanism behind desensitization.

Adult↗
PubMed1997

Symptoms, lung and liver function, blood counts, and genotoxic effects in coastal tanker crews.

OBJECTIVE: The deck crew on tankers can be exposed to high concentrations of benzene and other chemicals during loading, unloading and tank-cleaning operations. The objective of this study was to investigate whether genotoxic or other early health effects of cargo vapour exposure could be detected in coastal tanker crews. METHODS: The association between exposure to cargo vapours and clinical symptoms and signs, spirometry, blood cell count, blood test for liver function, and the frequency of micronuclei and sister chromatid exchanges in peripheral lymphocytes was studied in a cross-sectional investigation of 107 male crew members (66 deck crew and 41 others) on ten coastal tankers. RESULTS: Seven of the tankers had automatic cargo level gauging systems but some of the ships still had open hatches during loading and unloading operations. Acute symptoms such as headache, nausea, vertigo, fatigue and dizziness after loading or tank-cleaning operations were reported by 56 of the 66 deck crew members (85%). Irritation of the mucous membrane in eyes and upper respiratory tract by cargo vapours were also common in this group. Obstructive symptoms were more common in the group with the highest exposure to cargo vapours but persistent effects on lung function (vital capacity and forced expiratory volume in 1 s), nervous system, liver enzymes or blood counts were not found. The frequency of micronuclei after mitotic stimulation with phytohaemagglutinin was higher among the deck crew (mean 4.2 SEM 0.40) than in other crew members (mean 3.6, SEM 0.35). although the difference was not statistically significant. We found no association between exposure and the frequency of sister chromatid exchanges or micronuclei after stimulation with pokeweed mitogen. CONCLUSION: This study indicates that exposure to cargo vapours in coastal tanker crews may cause symptoms in the respiratory and nervous systems.

Adolescent↗
PubMed1997

The ankle strategy for postural control--a comparison between a model-based and a marker-based method.

When analysing postural control statistical characteristics of the centre of pressure or the ground reaction force are often used. A complement would be to analyse movement strategies as well. The purpose of this study was to determine to what extent the ankle strategy is used to maintain the balance in standing. One important question is if it is sufficient to use only force plate data in this determination, or if markers have to be used. To answer this question we studied two different methods--a marker-based method and a model-based method. The latter needs force plate data only as input to an inverted pendulum model. Different measures were calculated and used to draw conclusions of to what extent the ankle strategy was used. Experiments demonstrated that the data from the two methods provided the same general conclusions, although there were some differences in the estimated measures. By using the model-based method instead of tracking markers we can increase the clinical effectiveness considerably, get a lesser amount of data, and still get results comparable to the marker-based method.

Ankle↗
PubMed1997

Estimation of the knee joint location using colour video.

Analysis of the knee joint moment might be useful for predicting the risk of prosthetic loosening. The moment can be estimated if the location of the centre of pressure in the knee joint and the ground reaction force vector are known. Usually, skin mounted markers are used to locate the knee joint centre. Since marker based methods have several drawbacks investigations of alternative methods are needed. A new method for location of the knee joint is outlined in this paper. The subject is video filmed during gait with an elastic blue band wrapped around the knee. The band is located by splitting the video images into the RGB-components, and the centroid of the band is used as an estimate of the knee joint centre location. The band is easier to mount than markers, and it is not sensitive for axial leg rotation like markers are, which simplifies the analysis. If desired, the axial leg rotation can be determined by attaching a circular cyan marker on the band. The centroid of the marker relative to the band centroid gives an approximation of the leg rotation. In this study, the band could easily be located in the images and the leg axial rotation estimated in the frontal and the sagittal view showed the same trend. Further, the band-based results were found to be comparable to results where a marker was used to locate the knee joint centre.

Adult↗
PubMed1997

Surface-related triggering of the neutrophil respiratory burst. Characterization of the response induced by IgG adsorbed to hydrophilic and hydrophobic glass surfaces.

Hydrophilic and hydrophobic glass surfaces precoated with human albumin, fibrinogen, or IgG were investigated with respect to their ability to activate the neutrophil NADPH-oxidase. We found that IgG-coated surfaces induced a substantial and prolonged neutrophil production of reactive oxygen species (ROS). When a hydrophilic surface was used to support protein binding, a somewhat lower neutrophil response (around 35%) was obtained, compared with the response induced by IgG on a hydrophobic surface. The production of ROS was completely eliminated when cytochalasin B was added to the measuring system, suggesting the involvement of the cell cytoskeleton in the activation process. The relation between the intra- and extracellular generation of ROS was further assessed, and we found that most of the ROS produced were released from the cells, in agreement with a model in which the activating surfaces induce a 'frustrated' phagocytic response. Serum totally inhibited 'frustrated' phagocytosis provided that the IgG molecules were sticking to a hydrophilic surface.

Albumins↗
PubMed1997

Coreceptor usage of primary human immunodeficiency virus type 1 isolates varies according to biological phenotype.

The biological phenotype of primary human immunodeficiency virus type 1 (HIV-1) isolates varies according to the severity of the HIV infection. Here we show that the two previously described groups of rapid/high, syncytium-inducing (SI) and slow/low, non-syncytium-inducing (NSI) isolates are distinguished by their ability to utilize different chemokine receptors for entry into target cells. Recent studies have identified the C-X-C chemokine receptor CXCR4 (also named fusin or Lestr) and the C-C chemokine receptor CCR5 as the principal entry cofactors for T-cell-line-tropic and non-T-cell-line-tropic HIV-1, respectively. Using U87.CD4 glioma cell lines, stably expressing the chemokine receptor CCR1, CCR2b, CCR3, CCR5, or CXCR4, we have tested chemokine receptor specificity for a panel of genetically diverse envelope glycoprotein genes cloned from primary HIV-1 isolates and have found that receptor usage was closely associated with the biological phenotype of the virus isolate but not the genetic subtype. We have also analyzed a panel of 36 well-characterized primary HIV-1 isolates for syncytium induction and replication in the same series of cell lines. Infection by slow/low viruses was restricted to cells expressing CCR5, whereas rapid/high viruses could use a variety of chemokine receptors. In addition to the regular use of CXCR4, many rapid/high viruses used CCR5 and some also used CCR3 and CCR2b. Progressive HIV-1 infection is characterized by the emergence of viruses resistant to inhibition by beta-chemokines, which corresponded to changes in coreceptor usage. The broadening of the host range may even enable the use of uncharacterized coreceptors, in that two isolates from immunodeficient patients infected the parental U87.CD4 cell line lacking any engineered coreceptor. Two primary isolates with multiple coreceptor usage were shown to consist of mixed populations, one with a narrow host range using CCR5 only and the other with a broad host range using CCR3, CCR5, or CXCR4, similar to the original population. The results show that all 36 primary HIV-1 isolates induce syncytia, provided that target cells carry the particular coreceptor required by the virus.

Acquired Immunodeficiency Syndrome↗
PubMed1997

Characteristics of the Pro225His mutation in human immunodeficiency virus type 1 (HIV-1) reverse transcriptase that appears under selective pressure of dose-escalating quinoxaline treatment of HIV-1.

Treatment of human immunodeficiency virus type 1 (HIV-1)-infected CEM cell cultures with escalating concentrations of the quinoxaline S-2720 resulted in an ordered appearance of single and multiple mutant virus strains that gradually became resistant to the quinoxaline and other nonnucleoside reverse transcriptase (RT) inhibitors (NNRTIs). A novel mutation, Pro225His, consistently appeared in a Val106Ala RT-mutated genetic background. The contribution of this mutation to the resistance of the mutant HIV-1 RT to NNRTIs was additive to the resistance caused by the Val106Ala mutation. Interestingly, site-directed mutagenesis studies revealed that the Pro225His-mutated RT had acquired markedly greater sensitivity to bis(heteroaryl)piperazine (BHAP U-90152) (delavirdine) but not to any of the other NNRTIs. The kinetics of inhibition of the Pro225His mutant RT by the NNRTIs (including BHAP U-90152) was not substantially different from that observed for the wild-type RT. The hypersensitivity of the mutant enzyme and virus to BHAP U-90152 could be rationally explained by the molecular-structural determinants of the RT-BHAP complex, which has recently been resolved by X-ray crystallography.

Amino Acid Substitution↗
PubMed1997

Estimated frequency of the RN- allele in Swedish Hampshire pigs and comparison of glycolytic potential, carcass composition, and technological meat quality among Swedish Hampshire, Landrace, and Yorkshire pigs.

The frequency of the dominant RN- allele (Rendement Napole) was estimated in purebred Swedish Hampshire pigs (n = 208) by using the estimated glycogen content in the longissimus muscle at slaughter (glycolytic potential; GP). Carriers (n = 177) and noncarriers (n = 31) of the RN- allele were compared with purebred Swedish Yorkshire (Y, n = 208) and Swedish Landrace (L, n = 114) pigs with respect to GP and carcass composition. Technological meat quality was compared between the RN phenotypes from the Hampshire breed and Yorkshire pigs. The distribution of GP deviated from a normal distribution in all three breeds. Hampshire pigs with GP > or = 183 micromol lactate equivalents per gram wet weight were regarded as carriers of the RN- allele (RN-/RN-, or RN-/rn+), and those constituted 85% of all Hampshire pigs evaluated, giving a frequency of .61 for the dominant allele (RN-). The RN- carriers had higher GP than noncarriers, Landrace, and Yorkshire pigs, but noncarriers and Landrace did not differ regarding GP. The two RN phenotypes did not differ in carcass composition, but the carriers were leaner than Landrace and Yorkshire, with larger proportions of meat plus bone in ham and back. Noncarriers and Yorkshire did not differ in leanness, but both these groups were leaner than Landrace. The RN- carriers had lower Napole yield (cured cooked muscle) and higher drip loss than noncarriers and Yorkshire, but in comparison with noncarriers they also had lower shear force values. In conclusion, the frequency of the RN- allele is high in purebred Swedish Hampshire. Most of the effects of the allele on technological meat quality (i.e., ultimate pH, water-holding capacity, and technological yield) found in crossbred pigs seem also to be consistent for purebred pigs. However, the RN- allele exerted less influence on meat content in purebred pigs.

Alleles↗
PubMed1997

Enterobius vermicularis and finger sucking in young Swedish children.

OBJECTIVE: To study the prevalence of Enterobius vermicularis and its association with finger sucking in young Swedish children. DESIGN: Cross-sectional survey with a questionnaire for symptoms of infestation with Enterobius vermicularis, and the children's habit of finger sucking (including fingernail biting). Perianal tape-test for identification of eggs of Enterobius vermicularis. SETTING: Primary care, day-care centres, and schools in a Swedish middle-sized town (approx. 80,000 inhabitants). PARTICIPANTS: 172 children of both sexes, 4-10 years old. MAIN OUTCOME MEASURES: The prevalence of Enterobius vermicularis and its association with finger sucking. RESULTS: 21% of the children were symptom-free carriers of Enterobius vermicularis, and finger sucking was strongly associated with a positive tape-test (p = 0.01). CONCLUSION: More children than previously known seemed to be symptom-free carriers of Enterobius vermicularis. Finger sucking should be considered when treating infested children and especially those with relapsing symptoms.

Animals↗