Biomedical subjects

J Taskinen

Publications and source records attributed to J Taskinen.

At least 19 recordsLinked to original sources

PubMed2006

Association between environmental stress and epidermal papillomatosis of roach Rutilus rutilus.

We studied the association between environmental stress and epidermal papillomatosis of roach Rutilus rutilus L. in Finnish waters using a 'matched pairs' design. Populations impacted by industrial and/or sewage effluents were compared to reference populations from pristine sites. We examined both the prevalence (proportion of diseased fish) and intensity (number of scales covered by tumors) of the disease. Results of Generalized Linear Mixed Models (GLMM) indicated that the risk of papillomatosis was 7.5 times higher in males than females, and increased 1.3 times for every 10 mm increment in fish length. We controlled for the possible effects of fish size, sex and temporal variation through sampling procedures and statistical analyses. Mean prevalence of epidermal papillomatosis was 16.6 and 5.8% in impact and reference populations, respectively (10 population pairs; nfish = 1714). Results of GLMM suggested that the risk of being diseased was 2.7 times higher in the impact than reference populations. Thus, the prevalence of epidermal papillomatosis in roach can be used as an indicator of environmental stress. Results of Linear Mixed Models indicated no difference in the intensity of the disease between impact and reference populations (5 population pairs; nfish = 73; mean+/-SE 10.7+/-1.8 and 11.7+/- 2.9 scales, respectively), although prevalence was higher in impact populations in those 5 population pairs. The possible relationship between environmental stress and intensity of epidermal papillomatosis in natural roach populations remains to be demonstrated.

Animals↗

PubMed2006

Burrowing behaviour affects Paraergasilus rylovi abundance in Anodonta piscinalis.

Burrowing depth may affect predation rate, feeding ability and reproduction in bivalve clams. We studied the effect of burrowing depth on the abundance of the ergasilid Paraergasilus rylovi in the freshwater bivalve clam Anodonta piscinalis. We transplanted uninfected clams to a lake where they were allowed to choose their preferred burrowing depth, and were exposed naturally to copepodids of the parasite. There was a significant positive correlation between proportionate burrowing depth (PBD) and the abundance of P. rylovi at the end of the 17-day experiment, the deeper-burrowed clams harbouring more P. rylovi. Original PBD (0%, 50%, 100%) did not influence the final PBD or parasite abundance. Clam length affected PBD, smaller clams burrowing deeper, but it did not affect parasite abundance. Infected experimental clams and naturally-burrowed uninfected clams, both originating from the same lake, did not differ in their mean PBD. This indicated that burrowing of the experimental clams affected parasitism rather than the parasites altering burrowing of the clams. In line with the experimental result, we observed a significant positive correlation between PBD and the abundance of P. rylovi also among clams collected from 2 natural A. piscinalis populations.

Animals↗

PubMed2006

Factors affecting the intensity of epidermal papillomatosis in populations of roach, Rutilus rutilus (L.), estimated as scale coverage.

Studies of epidermal papillomatosis in fish populations have only rarely focused on the intensity of the disease, i.e. the number and size of papillomas. Furthermore, the methods used to evaluate the intensity of papillomatosis have not been standardized. We tested the reliability of a method based on counting of scales covered by papilloma tumours in roach, Rutilus rutilus (L). In addition, we studied the frequency distributions of the number of scales covered by papillomas within populations, evaluated the correlation between the prevalence and mean intensity of the disease among populations and examined the intensity of papillomatosis in roach with respect to sex and size of fish. Reliability of the scale coverage method was high. Therefore, the method could offer an effective way to determine the intensity of papillomatosis in fish species with large scales. The frequency distribution of the scale coverage of papillomas was highly aggregated within all populations studied. The mean intensity of papillomatosis increased with the size of the fish and was higher in males than in females. However, there was no correlation between the mean intensity and prevalence of the disease among the 19 roach populations studied.

Animals↗

PubMed2005

Long-lasting effect of stress on susceptibility of a freshwater clam to copepod parasitism.

The question whether a stress event can have a long-lasting effect on susceptibility to parasites was studied using a freshwater bivalve clam and its crustacean parasite as a model system. Anodonta piscinalis clams were collected from 2 populations during August-September 2002. Clams were transported to the laboratory and marked. The stressed clams were subjected to low oxygen for 25 days, while the unstressed control clams were caged in their lakes of origin for the same period. Then the clams were transported to a third lake where they were exposed to natural infections by the ergasilid copepod, Paraergasilus rylovi, 11 months after the stress event. The stressed clams were more intensively parasitized. They also showed lower growth, lower reproduction and lower survival than the unstressed control clams. The results indicate that susceptibility of A. piscinalis to P. rylovi infection may be condition dependent, and that stress may have a long-lasting, increasing effect on host susceptibility to parasitism in natural populations.

Animals↗

PubMed2004

Aspects of the ecology and natural history of Paraergasilus rylovi (Copepoda, Ergasilidae) parasitic in unionids of Finland.

The distribution of Paraergasilus rylovi in 17 populations of unionids was investigated. In 1 unionid population, the parasite was studied regarding host age, size, sex, and the reproductive period (occurrence of egg sacs). Results from pooled material from the years 1987--1989 and 1996 (southern Finland, 11 populations) indicated that Anodonta piscinalis (n = 1,359) is the main host (total mean prevalence 71% and intensity +/-SE of infection 16.4+/-0.6). Pseudanodonta complanata (n = 106) was infected occasionally (3% and 1.3+/-0.3), whereas Unio pictorum (n = 108) and U. tumidus (n = 17) were not infected. Results from 17 A. piscinalis populations showed that P. rylovi occurs in southern Finland but not in northern Finland. In A. piscinalis, the mean intensity of infection was higher in lake populations than in river populations. Both host age and length had a negative relationship with the intensity of P. rylovi infection. Host sex did not affect the intensity of infection. Egg sacs of P. rylovi were found from June to August. There was a tendency for higher intensities of infection in autumn. Infection by the digenean Rhipidocotyle fennica had no effect on the intensity of P. rylovi infection.

Analysis of Variance↗

PubMed2003

Reduction in the level of infection of the bivalve Anodonta piscinalis by the copepod Paraergasilus rylovi using high temperature and low oxygen.

The aim of this study was to develop a method to kill or expel the gill-dwelling crustacean parasite Paraergasilus rylovi from a common freshwater clam, Anodonta piscinalis. Naturally infected clams were exposed to different water-quality treatments and monitoring in the laboratory. In a high-temperature treatment (26 C vs. control 18 C), the mean abundance of the parasite decreased to near zero in 7 days. Because only 2 clams of 72 died in this treatment during the 14-day experiment, the survival of the host was not seriously at risk at the high temperature. 'Low oxygen, no water change' (18 C) was the second most effective treatment, followed by a 'low-oxygen, water-flow' (18 C) treatment. At the end of the experiment, the mean parasite abundance was significantly lower in all the treatments than in the control clams (18 C). A few P. rylovi individuals abandoned the host at 26 C but died in a couple of days outside the host. However, the parasites lived on average (+/-SE) 12.7 +/- 0.9 days outside the clam, and were also shown to be capable of infecting another uninfected host individual, at 18 C. The results of the present study suggest that high temperature provides an effective, ecologically sustainable method to manipulate the intensity of P. rylovi infection.

Analysis of Variance↗

PubMed2001

Quantitation of entacapone glucuronide in rat plasma by on-line coupled restricted access media column and liquid chromatography-tandem mass spectrometry.

A column-switching liquid chromatography-electrospray ionization-tandem mass spectrometric (LC-ESI-MS-MS) method was developed for the direct analysis of entacapone glucuronide in plasma. The plasma samples (5 microl) were injected onto a C18-alkyl-diol silica (ADS) column and the matrix compounds were washed to waste with a mixture of 20 mM ammonium acetate solution at pH 4.0-acetonitrile (97:3). The retained analyte fraction containing (E)- and (Z)-isomers of glucuronides of entacapone and tolcapone glucuronide (internal standard) was backflushed to the analytical C18 column, with a mixture of 20 mM ammonium acetate-acetonitrile (85:15) for the final separation at pH 7.0. The eluate was directed to the mass spectrometer after splitting (1:100). The mass spectrometer was operated in the negative ion mode and the deprotonated molecules [M-H]- were chosen as precursor ions for the analytes and internal standard. Collisionally induced dissociation of [M-H] in MS-MS resulted in loss of the neutral glucuronide moiety and in the appearance of intensive negatively charged aglycones [M-H-Glu]-, which were chosen as the product ions for single reaction monitoring. Quantitative studies showed a wide dynamic range (0.0025-100 microg/ml) with correlation coefficients better than 0.995. The method was repeatable within-day (relative standard deviation, RSD<7%) and between-day (RSD<14%) and the recovery (78-103%) was better than with the traditional, laborious pretreatment method. The use of tandem mass spectrometry permitted low limits of detection (1 ng/ml of entacapone glucuronide). The method was applied for the quantitation of (E)- and (Z)-isomers of entacapone glucuronide in plasma of rats used in absorption studies.

Animals↗

PubMed2001

High-performance liquid chromatographic method combining radiochemical and ultraviolet detection for determination of low activities of uridine 5'-diphosphate-glucuronosyltransferase.

A novel reversed-phase high-performance liquid chromatographic method was developed to measure UDP-glucuronosyltransferase (UGT) activity. Radiochemical and UV detection were combined in this UDP-[(14)C]glucuronic acid-utilizing method which was especially aimed at determination of low activities typical of N-glucuronidation of various amines and heterocycles. 4-Nitrophenol and levomedetomidine were used as substrates to validate this method, and applicability was tested with commonly used model substrates of N-glucuronidation, 4-aminobiphenyl and amitriptyline, and several 4-arylalkyl-1H-imidazole compounds. Detection limits were very low, 0.5-10 pmol, corresponding to UGT activities from 0.04 to 0.8 pmol/min/mg protein depending on UV absorbance of the glucuronide conjugate. The sensitivity was 10- to 100-fold compared with earlier HPLC assays using radiochemical detection. This method enabled quantitation without a reference glucuronide, and its high sensitivity allows for characterization of N-glucuronidation kinetics of various substrates. Using this method, human liver microsomal UGT activity was determined for a series of 4-arylalkyl-1H-imidazoles. Of these compounds, levomedetomidine was glucuronidated at the highest rate, 1.69 nmol/min/mg protein, using a 500 microM substrate concentration. In comparison, activities for the commonly used UGT substrates, 4-nitrophenol, 4-aminobiphenyl, and amitriptyline were 18.80, 3.23, and 0.23 nmol/min/mg protein, respectively.

Aminobiphenyl Compounds↗

PubMed2001

Molecular mechanisms controlling the rate and specificity of catechol O-methylation by human soluble catechol O-methyltransferase.

Molecular mechanisms determining the turn-over rate and specificity of catechol O-methylation were studied by combining enzyme kinetic measurements, computational modeling of substrate properties and fitting ligands in a 3D model of the active site of the enzyme. Enzyme kinetic measurements were carried out for 46 compounds, including most clinically used catechol drugs, by using recombinant human soluble catechol O-methyltransferase (COMT). The most important mechanism decreasing the turnover rate and increasing affinity was the electron withdrawing effect of substituents. Several other mechanisms by which substituents affected reactivity and affinity were identified. Highest turnover rates were determined for unsubstituted catechol and pyrogallol. Pyrogallol derivatives generally seemed to be more specific substrates than catechols. Catecholestrogens were the most specific endogenous substrates, whereas catecholamines were rather poor substrates. Among the catechol drugs used in the L-DOPA treatment of Parkinson's disease, the COMT inhibitors entacapone and tolcapone were not methylated, whereas the DOPA decarboxylase inhibitor benserazide was 15 times more specific substrate than L-DOPA, the target of COMT inhibition. The structure-activity relationships found allow the prediction of reactivity, affinity, and specificity with useful accuracy for catechols with a wide range of structures and properties. The knowledge can be used in the evaluation of metabolic interactions of endogenous catechols, drugs and dietary catechols, and in the designing of drugs with the catechol pharmacophore.

Binding Sites↗

PubMed2001

Solid-phase bromination and Suzuki coupling of 2-carboxyindoles.

As part of an ongoing lead discovery project we have developed a convenient method for the modification and substitution of indole moieties at the 3-position. Selective bromination of three different 2-carboxyindoles was followed by Suzuki cross-coupling with aryl and heteroaryl boronic acids on a Merrifield resin solid-phase. After column chromatography, yields of the 3- substituted indoles ranged from 42-98%.

Bromine↗

PubMed2001

Glucuronidation of 1-hydroxypyrene by human liver microsomes and human UDP-glucuronosyltransferases UGT1A6, UGT1A7, and UGT1A9: development of a high-sensitivity glucuronidation assay for human tissue.

Human UDP-glucuronosyltransferases (UGT, EC 2.4.1.17) involved in the biotransformation of pyrene were investigated by a sensitive fluorometric high-performance liquid chromatography (HPLC)method developed for determining activities toward 1-hydroxypyrene. The endpoint metabolite of pyrene, 1-pyrenylglucuronide, is a well-known urinary biomarker for the assessment of human exposure to polycyclic aromatic hydrocarbons. 1-Pyrenylglucuronide was synthesized using rat liver microsomes as biocatalyst. The yield was satisfactory, 22%. 1-Pyrenylglucuronide, identified by (1)H NMR and by electrospray mass spectrometry, was used for method validation and calibration. The HPLC assay was very sensitive with a quantitation limit of 3 pg (8 fmol) for 1-pyrenylglucuronide. The assay was precise, showing a relative standard deviation of 5% or less at 0.1 to 300 microM 1-hydroxypyrene. Only 2 microg of microsomal protein was required for the assay in human liver. The glucuronidation of 1-hydroxypyrene was catalyzed at high rates in microsomes from pooled or three individual liver samples, showing comparable apparent K(m) values. The formation of 1-pyrenylglucuronide was catalyzed by recombinant human UGT1A6, UGT1A7, and UGT1A9, the K(m) values being 45, 12, and 1 microM, respectively. The apparent K(m) values in human liver microsomes, ranging from 6.9 to 8.6 microM, agreed well with these results. The method provides a sensitive tool for measuring extremely low UGT activities and a specific means for assessing interindividual differences in 1-hydroxypyrene-metabolizing UGT activities in human liver and other tissues.

Dimethyl Sulfoxide↗

PubMed2000

Immunocompetence, developmental stability and wingspot size in the damselfly Calopteryx splendens L.

Calopteryx splendens males exhibit a remarkable variation in wing pigmentation both within and between populations. In this study, we examined whether the wingspots of male C. splendens are related to male quality. We measured the nylon implant encapsulation rate for 85 males and found that males with larger wingspots had a faster encapsulation rate, indicating a better immunocompetence. We also found that the encapsulation rate was positively correlated with the density of haemocytes in the haemolymph. Another measurement of male quality, fluctuating asymmetry of wingspots, correlated negatively with the size of the wingspots. Males with asymmetrical wingspots also had lower encapsulation rates than more symmetrical males. Our results suggest that the size of wingspot is an indicator of male quality in C. splendens.

Animals↗

PubMed2000

Direct analysis of nitrocatechol-type glucuronides in urine by capillary electrophoresis-electrospray ionisation mass spectrometry and tandem mass spectrometry.

Direct, quantitative capillary electrophoresis-electrospray ionisation mass spectrometric (CE-ESI-MS) and tandem mass spectrometric (CE-ESI-MS-MS) methods are described for the quantitation of 3-O-glucuronides of E- and Z-entacapone isomers (EEG and EZG) and tolcapone (TG) in urine. 3-O-Glucuronide of nitecapone was used as internal standard. Good separation of glucuronides was achieved with 20 mM ammonium acetate as separation solution at pH 6.84. Stacking was used to increase the sensitivity of the method by introducing samples in 5 mM ammonium acetate. CE-ESI-MS and CE-ESI-MS-MS methods are linear with correlation coefficients better than 0.9983 and 0.9982, and repeatable with relative standard deviations below 9 and 14%, respectively. The limit of detection (LOD) in CE-ESI-MS at signal-to-noise ratio 3 is 100 ng/ml for EEG and EZG and 250 ng/ml for TG. The CE-ESI-MS-MS method was the more sensitive; LOD was 7 ng/ml for all compounds, without any concentration of the sample.

Benzophenones↗

PubMed2000

Production of human UDP-glucuronosyltransferases 1A6 and 1A9 using the Semliki Forest virus expression system.

Human UDP-glucuronosyltransferases (UGTs) 1A6 and 1A9 were expressed using Semliki Forest virus (SFV) vectors. Infection of chinese hamster lung fibroblast V79 cells with recombinant SFV-UGT viruses resulted in efficient protein expression as detected by metabolic labeling, Western blot analyses and immunofluorescence microscopy. The expression of UGT 1A6 and UGT1A9 in the SFV-infected cells was approximately two fold higher than in a stable V79 cell line. No UGT signal was detected in noninfected cells. In addition, SFV-UGT viruses also efficiently infected other mammalian cells, such as baby hamster kidney (BHK), chinese hamster ovary (CHO) and human lung (WI-26 VA4) cells leading to high production of recombinant enzyme. The measurement of enzyme activities and kinetic parameters using p-nitrophenol and nitrocatechol (entacapone) as substrates for UGT1A6 and UGT1A9, respectively, showed that the overall kinetic properties of the enzymes produced by the two systems were similar. We conclude that the SFV expression system represents an efficient, fast and versatile method for production of metabolic enzymes for in vitro assays.

Animals↗

PubMed2000

The specificity of glucuronidation of entacapone and tolcapone by recombinant human UDP-glucuronosyltransferases.

The COMT inhibitors entacapone and tolcapone are rapidly metabolized in vivo, mainly by glucuronidation. In this work, the main UGT isoforms responsible for their glucuronidation in vitro were characterized by using a subset of representative cloned and expressed human UGT isoforms. Entacapone in particular was seen to be an exceptionally good substrate for UGT1A9 with an even higher reaction velocity value at 500 microM substrate concentration compared with that of the commonly used substrate, propofol (1.3 and 0.78 nmol min(-1) mg(-1), respectively). Neither entacapone nor tolcapone was glucuronidated by UGT1A6. Tolcapone was not detectably glucuronidated by UGT1A1, and the rate of glucuronidation of entacapone was also low by this isoform. However, UGT1A1 was the only UGT capable of catalyzing the formation of two glucuronides of the catecholic entacapone. Both COMT inhibitors were glucuronidated at low rates by the representative members of the UGT2B family, UGT2B7 and UGT2B15. Michaelis-Menten parameters were determined for entacapone and tolcapone using recombinant human UGT isoforms and human liver microsomes to compare the kinetic properties of the two COMT inhibitors. The kinetic data illustrates that UGT1A9 exhibited a much greater rate of glucuronidation and a far lower K(m) value for both entacapone and tolcapone than UGT2B15 and UGT2B7 whose contribution is minor by comparison. Entacapone showed a 3 to 4 times higher V(max) value and a 4 to 6 times lower K(m) value compared with those of tolcapone both in UGT1A9 cell lysates and in human liver microsomes.

Animals↗

PubMed1999

X-ray crystal structure of human dopamine sulfotransferase, SULT1A3. Molecular modeling and quantitative structure-activity relationship analysis demonstrate a molecular basis for sulfotransferase substrate specificity.

Humans are one of the few species that produce large amounts of catecholamine sulfates, and they have evolved a specific sulfotransferase, SULT1A3 (M-PST), to catalyze the formation of these conjugates. An orthologous protein has yet to be found in other species. To further our understanding of the molecular basis for the unique substrate selectivity of this enzyme, we have solved the crystal structure of human SULT1A3, complexed with 3'-phosphoadenosine 5'-phosphate (PAP), at 2.5 A resolution and carried out quantitative structure-activity relationship (QSAR) analysis with a series of phenols and catechols. SULT1A3 adopts a similar fold to mouse estrogen sulfotransferase, with a central five-stranded beta-sheet surrounded by alpha-helices. SULT1A3 is a dimer in solution but crystallized with a monomer in the asymmetric unit of the cell, although dimer interfaces were formed by interaction across crystallographic 2-fold axes. QSAR analysis revealed that the enzyme is highly selective for catechols, and catecholamines in particular, and that hydrogen bonding groups and lipophilicity (cLogD) strongly influenced K(m). We also investigated further the role of Glu(146) in SULT1A3 using site-directed mutagenesis and showed that it plays a key role not only in defining selectivity for dopamine but also in preventing many phenolic xenobiotics from binding to the enzyme.

Alanine↗

PubMed1999

Radiochemical high-performance liquid chromatographic assay for the determination of catechol O-methyltransferase activity towards various substrates.

A new chromatographic catechol O-methyltransferase (COMT) assay based on S-adenosyl-L-[methyl-14C]methionine and on-line radioactivity detection was developed. With minor modifications in the mobile phase composition the methylation velocities for 30 structurally diverse compounds including simple catechols, neurotransmitters, catecholestrogens and catecholic drugs could be measured using human and rat recombinant soluble COMT. The enzymes showed very similar substrate selectivities. The radiochemical method was validated using 3,4-dihydroxybenzoic acid as a model substrate and it was shown that accurate and reproducible methylation velocity values could be achieved for both of the catecholic hydroxyls. The method proved to be suited for determining the enzyme kinetic parameters and can probably be further used for gathering enzyme kinetic data on differentially substituted catechols in order to construct proper structure-activity relationships for COMT.

Animals↗

PubMed1999

Effect of nitro groups and alkyl chain length on the negative ion tandem mass spectra of alkyl 3-hydroxy-5-(4'-nitrophenoxy) and alkyl 3-hydroxy-5-(2', 4'-dinitrophenoxy) benzoates

Tandem mass spectrometric behaviour was studied for a small combinatorial library of alkyl 3-hydroxy-5-(4'-nitrophenoxy) benzoates (A1-A5) and alkyl 3-hydroxy-5-(2', 4'-dinitrophenoxy) benzoates (B1-B5). The spectra were recorded by negative ion electrospray low-energy collision induced dissociation (CID) tandem mass spectrometry. The product ion spectra of [M - H](-) of the benzoates A1-A5 are similar, as are those of benzoates B1-B5. However, the spectra of the B series compounds differ significantly from those of the A series owing to the second electron-withdrawing nitro substituent in the B compounds. In addition, the length of the alkyl chain has an effect on the fragmentation. However, both series of compounds exhibit an abundant nitrophenoxy ion formed by the loss of 3-hydroxybenzoate. This is at m/z 138 in A1-A5 and at m/z 183 in B1-B5. A precursor ion scan of the nitrophenoxy ion provides a rapid method to identify the synthesised compounds in this type of combinatorial mixture. Copyright 1999 John Wiley & Sons, Ltd.

Journal Article↗