Biomedical subjects

H H Meyer

Publications and source records attributed to H H Meyer.

At least 37 records · Page 2Linked to original sources

PubMed2000

Transrectal Doppler sonography of uterine blood flow.

Transrectal Doppler ultrasound was used for the noninvasive investigation of uterine blood flow in cows. Both the left and right Aa. uterinae were scanned to obtain blood flow velocity waveforms over 2 consecutive estrous cycles. Blood flow was reflected by the resistance index (RI) and the time-averaged maximum velocity (TAMV). Intra-observer reproducibility of Doppler measurements was evaluated. The intra-class correlation coefficient (Intra-CC) was 0.97 for the RI and 0.95 for TAMV. While RI values did not differ between the left and right A. uterina (P > 0.05), differences in TAMV occurred between both vessels in 2 cows. These differences were not related to the ovary bearing the dominant follicle or to the corpus luteum (P < 0.001). As in all cows, changes of RI and TAMV values between the left and right artery during the estrous cycle were correlated (correlation coefficient r > 0.72; P < 0.0001); the mean values of both sides were used for subsequent analyses. Variance component estimates for the effect of cow on RI and TAMV were 8 and 13% and for the influence of day of estrous cycle they were 70 and 47%, respectively (P <0.0001). Between estrous cycles no significant differences could be measured within cows (P > 0.05). The highest RI and lowest TAMV values occurred on Day 0 (= day of ovulation) and Day 1, while the lowest RI and highest TAMV values were measured between Days -3 and -1 of the estrous cycle, respectively. There was a positive correlation between TAMV and estrogen concentrations and a negative correlation between RI and plasma estrogen levels. Plasma progesterone levels and TAMV were negatively correlated, but no correlation could be measured (P > 0.05) between RI values and plasma progesterone concentrations. While there were no differences in plasma concentrations of estrogens and progesterone between estrous cycles within cows, the levels of these hormones differed between cows. The results show that transrectal Doppler sonography is a useful, noninvasive method for examining uterine blood flows in cows. If there is an influence of uterine perfusion on fertility in cows its role needs further investigation.

Animals↗

PubMed2000

Characterisation of the affinity of different anabolics and synthetic hormones to the human androgen receptor, human sex hormone binding globulin and to the bovine progestin receptor.

For the steroidal growth promoters trenbolone acetate (TBA) and melengestrol acetate (MGA) neither the complete spectrum of biological activities nor the potential endocrine disrupting activity of their excreted metabolites in the environment is fully understood. The potency of these substances in [3H]dihydrotestosterone ([3H]-DHT) displacement from the recombinant human androgen receptor (rhAR) and from human sex-hormone binding globulin (hSHBG) was evaluated. In addition, the potency for [3H]-ORG2058 displacement from the bovine uterine progestin receptor (bPR) was tested. For comparison, different anabolics and synthetic hormones were also tested for their binding affinities. For 17beta-trenbolone (17beta-TbOH), the active compound after TBA administration, an affinity the rhAR similar to dihydrotestosterone (DHT) and a slightly higher affinity to the bPR than progesterone were demonstrated. The affinity of the two major metabolites, 17alpha-trenbolone and trendione, was reduced to less than 5% of the 17beta-TbOH-value. The affinity of these three compounds and of MGA to the hSHBG was much lower compared with DHT. MGA showed a 5.3-fold higher affinity than progesterone to the bPR but only a weak affinity to the rhAR. The major MGA metabolites have an affinity to the bPR between 85% and 28% of the affinity of progesterone. In consequence, MGA and TBA metabolites may be hormonally active substances, which will be present in edible tissues and in manure. We conclude that detailed investigations on biodegradation, distribution and bio-efficacy of these substances are necessary.

Anabolic Agents↗

PubMed2000

Dose-dependent effects of melengestrol acetate (MGA) on plasma levels of estradiol, progesterone and luteinizing hormone in cycling heifers and influences on oestrogen residues in edible tissues.

Melengestrol acetat (MGA) is widely used as a growth promoting feed additive in cattle breeding in the USA and several other non-European countries. To explore the physiological effects of MGA four heifers were fed during 8 weeks with 0.5 mg MGA daily as registered in the USA and two heifers each received 0, 1.5 or 5 mg/day, respectively. Plasma samples were collected twice a week and concentrations of MGA, progesterone (P4) and estradiol-17beta (E2-17beta) were quantified. The pulsatile secretion of luteinizing hormone (LH) was investigated in 6-hour profiles before and during treatment. After slaughter the reproductive organs were examined and oestrogen residues in edible tissues were measured. Four days after the beginning of MGA feeding MGA concentrations in plasma reached levels of 30 and 100-400 pg/mL depending on the dose received. Three weeks after the beginning of MGA feeding P4 plasma concentrations had dropped to base levels below 0.3 ng/mL in all three treatment groups. Mean plasma E2-17beta levels increased in physiological range from 1 to 5 pg/mL during 0.5 mg MGA/day feeding with many acyclic peaks. Overdosed MGA decreased E2 levels and suppressed cyclic peaks. Number and size of ovarian follicles were not altered by any treatment. Mean LH levels and pulse frequencies increased significantly during labelled treatment (0.5 mg/day), while higher doses had reducing effects. The development of corpus luteum was suppressed. E2-17beta residues in fat increased about 300% following labelled MGA treatment.

Administration, Oral↗

PubMed1999

Solid-state NMR and FTIR studies on bilayer membranes from 1,2- dioctadec-(14-ynoyl)-sn-glycero-3-phosphatidylcholine.

Multilamellar dispersions from a new model phospholipid, 1, 2-dioctadec-(14-ynoyl)-sn-glycero-3-phosphatidylcholine (DO(14-yne)PC), bearing a triple bond in the fatty acid chains are studied by (2)H, (31)P NMR and Fourier transform infrared (FTIR) spectroscopy. The investigations are focused on the evaluation of the molecular properties of the lipid molecules as function of temperature and sample composition. Information about the fatty acid chain conformations are obtained from FTIR measurements by analysing the CH(2) wagging and stretching modes. (2)H NMR studies are performed on two selectively deuterated compounds that provide further insights into the molecular characteristics at two specific positions along the fatty acid chains. These studies demonstrate that the introduction of the triple bond is accompanied by a reduction of fatty acid chain order which holds for both the conformational and the orientational order. Likewise, (31)P NMR spectroscopy is used for the determination of the dynamics and ordering in the head group region. Here, particular emphasis is given to the evaluation of the lipid lateral motions that are quantified over a large temperature range within the liquid crystalline phase. It is found that the lateral mobility of the lipid molecules is almost unaffected by the triple bond in the fatty acid chains. The addition of cholesterol gives rise to a reduction in lateral mobility for DO(14-yne)PC, as can be followed by spin echo, 2D-exchange NMR and stimulated echo experiments.

Crystallization↗

PubMed1999

The mouse p97 (CDC48) gene. Genomic structure, definition of transcriptional regulatory sequences, gene expression, and characterization of a pseudogene.

Here we present the first description of the genomic organization, transcriptional regulatory sequences, and adult and embryonic gene expression for the mouse p97(CDC48) AAA ATPase. Clones representing two distinct p97 genes were isolated in a genomic library screen, one of them likely representing a non-functional processed pseudogene. The coding region of the gene encoding the functional mRNA is interrupted by 16 introns and encompasses 20.4 kilobase pairs. Definition of the transcriptional initiation site and sequence analysis showed that the gene contains a TATA-less, GC-rich promoter region with an initiator element spanning the transcription start site. Cis-acting elements necessary for basal transcription activity reside within 410 base pairs of the flanking region as determined by transient transfection assays. In immunohistological analyses, p97 was widely expressed in embryos and adults, but protein levels were tightly controlled in a cell type- and cell differentiation-dependent manner. A remarkable heterogeneity in p97 immunostaining was found on a cellular level within a given tissue, and protein amounts in the cytoplasm and nucleus varied widely, suggesting a highly regulated and intermittent function for p97. This study provides the basis for a detailed analysis of the complex regulation of p97 and the reagents required for assessing its functional significance using targeted gene manipulation in the mouse.

Adenosine Triphosphatases↗

PubMed1999

Directed evolution of an esterase: screening of enzyme libraries based on pH-indicators and a growth assay.

In order to resolve a sterically hindered 3-hydroxy ethyl ester, which was not accepted as substrate by 20 wild-type hydrolases, a directed evolution of an esterase from Pseudomonas fluorescens (PFE) was performed. Mutations were introduced using the mutator strain Epicurian coli XL1-Red. Enzyme libraries derived from seven mutation cycles were assayed on minimal media agar plates supplemented with pH indicators (neutral red and crystal violet), thus allowing the identification of active esterase variants by the formation of a red color caused by a pH decrease due to the released acid. A further selection criteria was introduced by using the corresponding glycerol estar, because release of the carbon source glycerol facilitates growth on minimal media. By this strategy, one double mutant (A209D and L181V) of PFE was identified, which hydrolyzed the 3-hydroxy ethyl ester in a stereoselective manner (25% ee for the remaining ester, E approximate to 5).

Cell Division↗

PubMed1999

Detection of melengestrol acetate residues in plasma and edible tissues of heifers.

The aim of this study was to gain knowledge of residue formation after the use of melengestrol acetate (MGA) as a growth-promoting agent. Two Holstein-Friesian heifers each received a daily dose through the feed of 0, 0.5 mg (2 heifers with and without withdrawal each), 1.5 mg or 5.0 mg MGA for 8 weeks. MGA residues in plasma were screened by enzyme immuno-assay (EIA). Concentrations in kidney, liver, and muscle were quantified by liquid-chromatography-mass spectrometry (LC-MS), and in fat by gas chromatography-mass spectrometry (GC-MS). MGA levels in plasma were 40, 128, and 280 ng/L, respectively. Residues accumulated in muscle and kidney (5-fold), liver (20-to-40-fold), and fat (200-fold). After administration of 1.5 mg per day the mean MGA concentration in fat was 29 micrograms/kg and thus violated USA regulations which specify a limit of 25 ppb. Therefore the labelled use of MGA (0.5 mg per day) has to be officially controlled.

Adipose Tissue↗

PubMed1998

The p47 co-factor regulates the ATPase activity of the membrane fusion protein, p97.

The highly conserved ATPase p97, a member of the AAA-ATPases, is found in a complex with its co-factor p47 in rat liver cytosol. Previously it had been shown that p97-mediated reassembly of Golgi cisternae from mitotic Golgi fragments requires p47 which mediates the binding of p97 to a Golgi t-SNARE (soluble N-ethylmaleimide-sensitive factor attachment factor receptor), syntaxin 5. Here we show that it also suppresses the ATPase activity of p97 by up to 85% in a dose-dependent and saturable manner suggesting that it has other roles in the membrane fusion cycle.

Adenosine Triphosphatases↗

PubMed1998

Clenbuterol as a marker in baits for oral vaccination of dogs against rabies.

Clenbuterol was investigated as a potential marker of baits for the oral vaccination of dogs (Canis familiaris) against rabies in Turkey. Orally administered clenbuterol is incorporated into the hair fibre during hair growth, and the uptake of clenbuterol into the hair of 18 dogs was therefore investigated in a controlled laboratory experiment. Clenbuterol could be detected in the hair of the dogs 28 and 56 days after they had eaten a bait containing 0-5 mg clenbuterol. In a field study, 150 baits containing clenbuterol were of distributed at selected sites along roads in the suburban areas of Ferhatpasa, Istanbul; the baits incorporated a vaccine container with the live modified rabies virus vaccine SAD B19. By the following morning, 93 per cent of the baits had gone. Hair samples from nine of 31 recaptured dogs contained more than 1 ng clenbuterol/g, indicating that they had consumed a bait. However, only four of the 31 dogs had an increased antibody titre. The results of this field study indicate that the placing of baits at selected sites is not a very efficient method of vaccinating ownerless dogs.

Administration, Oral↗

PubMed1998

Directed evolution of an esterase for the stereoselective resolution of a key intermediate in the synthesis of epothilones.

The directed evolution of an esterase from Pseudomonas fluorescens using the mutator strain Epicurian coli XL1-Red was investigated. Mutants were assayed for their ability to hydrolyze a sterically hindered 3-hydroxy ester, which can serve as a building block in the synthesis of epothilones. Screening was performed by plating esterase producing colonies derived from mutation cycles onto minimal media agar plates containing indicator substances (neutral red and crystal violet). Esterase-catalyzed hydrolysis of the 3-hydroxy ester (ethyl or glycerol ester) was detected by the formation of a red color due to a pH decrease caused by the released acid. Esterases isolated from positive clones were used in preparative biotransformations of the ethyl ester. One variant containing two mutations (A209D and L181V) stereoselectively hydrolyzed the ethyl ester resulting in 25% ee for the remaining ester.

Antineoplastic Agents↗

PubMed1998

Comparative binding affinity study of progestins to the cytosol progestin receptor of endometrium in different mammals.

The relative binding affinity of 5 alpha-reduced progestins and a newly synthesized antiprogestin J912 (progesterone 100%) was determined in a competitive receptor binding assay using [3H]ORG-2058 as radiolabeled ligand for the progestin receptor. Uteri obtained from 12 different species of four mammalian orders were examined. The relative binding affinity of 75-100% and a blood prevalence of 5 alpha-pregnane-3,20-dione in horses and African elephants suggest a biological role of this particular 5 alpha-reduced progesterone. For pigs the binding affinity of 5 alpha-pregnane-3,20-dione was about 50% of progesterone, but blood levels are unknown. In all other cases the low binding affinity of investigated progestins precludes possible biological role. For 5 alpha-pregnane-3 alpha-ol-20-one, 5 alpha-pregnane-20 alpha-ol-3-one, and 5 alpha-pregnane-3 beta,20 alpha-diol the relative binding affinity was less than 1%. A rather low binding (< 15%) was observed in 5 alpha-pregnane-3,20-dione in all ruminant species investigated. The antiprogestin J912 was found to be highly efficient in displacing progesterone from its endometrial binding sites in carnivores and might therefore be used for pregnancy interruption during diapause in certain species, e.g., in captive bears.

Animals↗

PubMed1998

Accumulation of polyunsaturated fatty acids by concentrate selecting ruminants.

Depot fat samples from ruminants of different feeding type and--for comparison--fat samples from simple-stomached animals were collected within 3 months. Individual fatty acid proportions, especially the relation of essential, polyunsaturated fatty acids to nonessential, saturated plus monounsaturated fatty acids were analyzed by gas chromatography. Species can be separated into two distinct groups: Depot fat of wild boar showed by far the highest content of essential fatty acids compared with all ruminant species. The subsequent inter-ruminant comparison yielded a further separation into two distinct groups related to feeding type. Roe deer and moose, constituting the first group of concentrate selectors, showed significantly higher percentages of polyunsaturated fatty acids than the other ruminant species comprised of the grass and roughage eaters or intermediates. The data document that the mode of feeding and/or the diet affects the body composition of the species investigated and that the depot fat composition of these ruminant species is markedly related to feeding type.

Animals↗

PubMed1998

Application of an androgen receptor assay for the characterisation of the androgenic or antiandrogenic activity of various phenylurea herbicides and their derivatives.

The potency of different substances for [3H]dihydrotestosterone ([3H]DHT) displacement from the bovine androgen receptor was tested. The phenylurea herbicide linuron and its derivative 3,4-dichloroaniline (3,4-DCA), which are found in sediments and surface waters, are known to displace bound testosterone from the rat androgen receptor. Because 3,4-DCA is rapidly taken up by fish and metabolised into 3,4-dichloroacetanilide (3,4-DCAc), it was investigated whether the displacement effects are attributable to 3,4-DCA or to 3,4-DCAc. The potency of 3,4-DCAc androgen receptor binding was compared with that of several phenylurea compounds. In a radioreceptor assay with calf uterus cytosol as androgen receptor preparation, the specific binding of [3H]DHT, the endogenous ligand, was completely displaceable by increasing concentrations of 3,4-DCAc. The relative binding affinities (RBA) of the various compounds were about 1/10(4) to 1/10(5) of that of DHT. 3,4-DCAc had the relative highest affinity (1.31 x 10(-4)), followed by linuron, 3,4-dichlorophenylurea, flutamide, 3,4-DCA and diuron with the lowest RBA (2.4 x 10(-5)). Hence the metabolism of xenobiotic compounds has to be considered to estimate potential ecotoxiocological effects. This test not only can be used to screen for androgen- and antiandrogen-like substances in environmentally relevant samples such as surface waters, but might also be applied for drug testing and for residue monitoring.

Acetanilides↗

PubMed1998

Production of CTC-containing porcine reference materials.

The European Commission (EC) established the Standards, Measurements and Testing programme for the preparation of Reference Materials (RMs) as an aid to harmonise testing for veterinary drug residues throughout the European Union (EU). The production of chlortetracycline (CTC)-free and CTC-incurred pig tissues as candidate RMs is described. High performance liquid chromatography (HPLC) with fluorescence detection of CTC and 4-epi-CTC was used for all tissue analyses. A pilot study revealed that incurred CTC residues were stable in pig kidney, liver and muscle lyophilised powders during storage for 10 weeks at -70, -20 and +37 degrees C, obviating the need for addition of a stabiliser (thimerosal). In the main study, 500 vials each of CTC-free and CTC-incurred kidney, liver and muscle were produced. Target concentrations in the CTC-incurred lyophilised tissue powders were 750-1500, 500-1000 and 300-600 micrograms kg-1 for kidney, liver and muscle, respectively. Following lyophilisation, the mean +/- s concentrations of CTC in the incurred positive RMs were 1,315 +/- 56.9, 765 +/- 35.3 and 378 +/- 16.8 micrograms kg-1 for kidney, liver and muscle respectively. Residual moisture in the RMs ranged from 1.6 +/- 0.53% for muscle to 3.0 +/- 0.50% for liver. Between-vial homogeneity for incurred powders was determined for 20 vials of each material, which had been removed at regular intervals during the filling process. Relative standard deviations (RSDs) for kidney, liver and muscle were 4.3, 4.6 and 4.4% respectively, being within the interassay RSD of the method and indicating that mixing was effective. Stability of powders stored at -18, 4, 20 and 37 degrees C was assessed over a period of 79 weeks. No measurable degradation occurred over this time period at any of the storage temperatures. It is concluded that these candidate RMs are homogenous, stable and are suitable for certification.

Animals↗

PubMed1998

Quantification of insulin-like growth factor-1 (IGF-1) mRNA: development and validation of an internally standardised competitive reverse transcription-polymerase chain reaction.

To investigate the role of local IGF-1 mRNA expression in various tissues, we developed and validated a method which allows for a specific, sensitive and reliable quantification of IGF-1 mRNA: an internally standardised Reverse Transcription-Polymerase Chain Reaction (RT-PCR). A synthetic competitive template IGF-I standard cRNA (IGF-1 cRNA) was designed, which contains the same flanking primer sequences used to amplify the wild type IGF-1 mRNA, but differs by 56 bp in length. To obtain the IGF-1 mRNA concentration present in tissue RNA samples, series of 250 ng total-RNA were spiked with three known quantities of the standard IGF-1 cRNA, incubated for competitive RT-PCR reactions and the two amplificates obtained (184 bp from IGF-1 cRNA and 240 bp from the wild type IGF-I mRNA) were subsequently separated and quantified by HPLC-UV. For every individual tissue RNA sample, the ratio R (R = competitor PCR product / wild type PCR product) was plotted against the number of starting molecules of the competitor IGF-1 cRNA. The initial amount of IGF-1 mRNA present in the sample can then be read off where R = 1. The validated assay had a detection limit of 1600 IGF-1 cRNA molecules/reaction, the intra-assay variation was 7.4% (n = 5) and linearity (r = 0.997) was given between 140 ng to 840 ng total-RNA input. The present method was first applied to study the effect of long term castration on the IGF-1 expression rates in bovine tissues. The hepatic IGF-1 mRNA concentrations were well correlated (r = 0.81) with the plasma concentrations as quantified by RIA and were higher in intact than in castrated animals. In two skeletal muscles (m. splenius and m. gastrocnemius) IGF-1 mRNA concentrations were 20- and 35- times lower than in liver, respectively, without any differences between steers and bulls. In bulls, the IGF-1 mRNA expression was higher in m. splenius (p < 0.01) than m. gastrocnemius, indicating that locally produced IGF-1 might be important for sexually dimorphic muscle growth patterns.

Animals↗

PubMed1998

Viability of small preantral ovarian follicles from domestic cats after cryoprotectant exposure and cryopreservation.

About 1500 preantral follicles can be recovered from a single cat ovary by mechanical dissection. This is a potentially rich source of genetic material if ova could be preserved and grown in vitro, especially from rare or endangered species that die abruptly or are ovariectomized for medical reasons. The aims of this study were to examine cryoprotectant toxicity and then the potential of successfully cryopreserving preantral cat follicles. In the initial toxicity trial, isolated cat follicles (40-90 micron) were exposed to dimethylsulfoxide, glycerol, 1,2-propandiol or ethylene glycol at 0 degrees C for 15 min. Follicle viability was assessed by supravital staining using a combination of Trypan blue and Hoechst 33258 at O h, and after 18 h and 1 week of culture. Percentages of follicles with intact oocytes and granulosa cells were similar (P > 0.05) among control (no cryoprotectant), dimethylsulfoxide, 1,2-propandiol and ethylene glycol treatments at all time points, but were reduced (P < 0.05) after glycerol exposure. On the basis of this finding, dimethylsulfoxide and 1,2-propandiol were used to cryopreserve intact follicles, and post-thaw viability was assessed by supravital staining and 5-bromo-2'-deoxyuridine uptake into oocytes and granulosa cells during culture. Of control (noncryopreserved) follicles, 31.4% +/- 2.9%, 18.8% +/- 1.9% and 16.2% +/- 1.6% were intact after O h, 18 h and 1 week of culture, respectively. Uptake of 5-bromo-2'-deoxyuridine occurred in approximately 20% of follicles at all time points. On the basis of the presence of both a healthy oocyte and granulosa cells, cryopreservation in dimethylsulfoxide or 1,2-propandiol allowed approximately 19% of follicles to survive. Approximately 10% demonstrated clear evidence of cell activity that was sustainable for 1 week. In conclusion, the cat ovary contains a population of preantral follicles that are not adversely affected by short-term exposure to most conventional cryoprotectants. Furthermore, there is a subpopulation of these follicles capable of surviving cryopreservation, remaining structurally intact and physiologically active after thawing.

Animals↗

PubMed1998

Synthesis of an unsaturated fatty acid analogue (18-(4'-azido-2'-hydroxybenzoylamino)-oleic acid) and its interaction with lysophosphatidylcholine: acyl-CoA-O-acyltransferase.

Acylation/deacylation reactions represent a basic requirement of triglyceride as well as phospholipid metabolism, and maintenance of membrane lipid composition. In order to examine enzymes participating in these pathways, we synthesized 18-(4'-azido-2'-hydroxybenzoylamino)-oleic acid, an iodinable photoaffinity analogue of oleic acid as a new tool for analyzing enzymes, especially those binding unsaturated fatty acids or acyl-CoAs. For the synthesis of omega-amino-oleic acid, coupling two bifunctional Cg-components was used. The described synthesis scheme is also suited for the specific generation of other fatty acid analogues with distinct positions of the double bond. The functionality of 18-(4'-azido-2'-hydroxybenzoylamino)-oleic acid was investigated with the enzyme lysophosphatidylcholine:acyl-CoA-O-acyltransferase (LAT) [EC 2.3.1.23], an enzyme that shows high specificity towards (poly)unsaturated fatty acyl-CoAs. It could be shown that the photolabel, esterified with coenzyme A, acts in the dark as a reversible inhibitor of the enzyme activity, but photolysis of the label results in irreversible inactivation of LAT. This inactivation could be prevented by addition of the native substrate arachidonyl-CoA during photolysis. Several proteins could be specifically visualized using the iodinated analogue. The data indicate that this new photoaffinity label may have application to identify and characterize lipid biosynthetic enzymes using unsaturated fatty acids as well as acyl-CoA binding proteins and the active site of these proteins.

1-Acylglycerophosphocholine O-Acyltransferase↗

PubMed1998

Uterine androgen receptor mRNA expression in metestrous and anestrous bitches being healthy or suffering from Pyometra.

The importance of androgens for the female reproductive system has been investigated for decades and a number of androgen sensitive processes has now been identified in female reproductive organs. For carnivore species no data were available so far about uterine androgen sensitivity and its regulation. The present study therefore aimed to investigate whether androgen receptors (AR) are present in the dog uterus, whether they are regulated throughout the ovarian cycle and whether pyometra affects their expression rate. Uterine tissue samples were collected from 28 bitches of different ages and various breeds. The samples were grouped according to the stage of estrous cycle (metestrus ME or anestrus AE) and the pathological status of the uterus (i.e. suffering from pyometra or not). Androgen receptor mRNA (AR mRNA) was quantified from 500 ng of total RNA isolated from the tissue samples using an internally standardized reverse transcription polymerase chain reaction (RT-PCR) described previously. The amount of total RNA extractable per g tissue was elevated during pyometra. The successful amplification of the expected 172 bp fragment from canine uterine RNA together with the confirmation of the identity of this fragment by sequence analysis, demonstrates that AR is expressed in this particular tissue. Comparing the expression rates in uteri from bitches during ME or AE being healthy (H) or suffering from pyometra (P), the only significant (p < 0.01) difference was found between H and P uteri during ME with 3.5-fold lower expression rates in P. Although the same seems true for AE bitches, a significant difference could not be demonstrated due to the low number (n = 2) of diseased animals in the AE group. There was no evident effect of the stage of ovarian cycle on uterine AR mRNA levels.

Anestrus↗