Safety of oral robenacoxib in the cat.

The safety of robenacoxib, a nonsteroidal anti-inflammatory drug with high selectivity for inhibition of the cyclooxygenase (COX)-2 isoform of COX, was investigated in the cat in two randomized, blinded, placebo-controlled, parallel-group studies. Robenacoxib was administered orally to healthy young domestic short-hair cats at dosages of 0 (placebo), 5 and 10 mg/kg once daily for 28 days (study 1) and at 0 (placebo), 2, 6 and 10 mg/kg twice daily for 42 days (study 2). The recommended minimum dosage for robenacoxib tablets in cats is 1 mg/kg once daily (range 1-2.4 mg/kg). Relative to placebo treatment, no toxicologically significant effects of robenacoxib were recorded in either study, based on general observations of health, haematological and clinical chemistry variables and urinalyses in life, and by post mortem organ weight, gross pathology and histopathology assessments. Pharmacokinetic-pharmacodynamic simulations indicated that all dosages of robenacoxib were associated with marked inhibition of COX-2 at peak effect (median I(max) 97.8-99.4% inhibition) with lesser inhibition of COX-1 (median I(max) 26.8-58.3% inhibition). Inhibition of the COXs was short lasting, with >10% median inhibition persisting for 4.0 h for COX-2 and 1.5 h for COX-1. These levels of inhibition of COX-1 and COX-2 twice daily with robenacoxib were not associated with any detectable toxicity, suggesting that, as previously described in dogs, the high safety index of robenacoxib in cats may be related to a combination of its high COX-2 selectivity and short residence time in the central compartment.

[Vaccination against bluetongue: safety and immune response in the field]. / Impfung gegen die Blauzungenkrankheit: Verträglichkeit und Immunantwort in der Praxis.

Bluetongue, caused by the bluetongue virus serotype 8 has rapidly spread through Europe since 2006. The first cases in Switzerland were detected in October 2007. The European Union and Switzerland launched a vaccination campaign in June 2008. This study aims to demonstrate the safety and the immune response of the three vaccines used in Switzerland under practical conditions in the field. The trial was carried out in cattle, sheep and goats. Based on the results of this study recommendations for the 2009 campaign are presented.

Identification of the amino-acetonitrile derivative monepantel (AAD 1566) as a new anthelmintic drug development candidate.

Anthelmintic resistance has become a global phenomenon in gastro-intestinal nematodes of farm animals, including multi-drug resistance against the three major classes of anthelmintics. There is an urgent need for an anthelmintic with a new mode of action. The recently discovered amino-acetonitrile derivatives (AADs) offer a new class of synthetic chemicals with anthelmintic activity. The evaluation of AADs was pursued applying in vitro assays and efficacy and tolerability studies in rodents, sheep, and cattle. Amongst various suitable compounds, AAD 1566 eliminated many tested pathogenic nematode species, both at larval and adult stages, at a dose of 2.5 mg/kg bodyweight in sheep and 5.0 mg/kg bodyweight in cattle. The same doses were sufficient to cure animals infected with resistant or multi-drug-resistant nematode isolates. These findings, complemented by the good tolerability and low toxicity to mammals, suggest that AAD 1566, monepantel, would be a suitable anthelmintic drug development candidate.

Determination of Lufenuron in canine skin layers by radioluminography.

In a bioavailability study in dogs we investigated the quantity of [14C]-Lufenuron and its metabolites in the skin by radioluminography. Two Beagle dogs were orally administered 10 mg [14C]-Lufenuron per kg body weight. 21 days later they were euthanized. Skin specimens were taken at 6 different sites. Sections of these skin specimens were exposed on a phosphorus imaging plate and the radiolabeled Lufenuron content visualized. Radioluminographical analysis showed that [14C]-Lufenuron appeared in the subcutaneous layer of the skin. We did not discover any activity on the skin surface.

Pharmacokinetics of clomipramine in dogs following single-dose intravenous and oral administration.

OBJECTIVE: To determine pharmacokinetics of clomipramine and its principle metabolite (desmethylclomipramine) in the plasma of dogs after IV or oral administration of a single dose. ANIMALS: 6 male and 6 female Beagles. PROCEDURES: Clomipramine was administered IV (2 mg/kg), PO (4 mg/kg) after food was withheld for 15 hours, and PO (4 mg/kg) within 25 minutes after dogs were fed. Plasma clomipramine and desmethylclomipramine concentrations were measured by use of a gas chromatography with mass-selection method. RESULTS: Time to peak plasma concentrations of clomipramine and desmethylclomipramine following oral administration was 1.2 hours. For clomipramine, after IV administration, elimination half-life was 5 hours, mean residence time was 3 hours, and plasma clearance was 1.4 L/h/kg. Values for mean residence time and terminal half-life following oral administration were similar to values obtained following IV administration, and systemic bioavailability was approximately 20% for clomipramine and 140% for desmethylclomipramine, indicating fast absorption of clomipramine from the gastrointestinal tract and extensive first-pass metabolism. Administration of clomipramine with food did not alter the area under the concentration versus time curve for desmethylclomipramine but resulted in a 25% increase for clomipramine. Clomipramine and desmethylclomipramine were extensively bound (> 96%) to serum proteins. There were no significant differences in area under the concentration versus time curve between male and female dogs. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicate that there should not be any clinically important differences in efficacy regardless of whether clomipramine is administered with or without food.

Probable interaction between S100A7 and E-FABP in the cytosol of human keratinocytes from psoriatic scales.

The overexpression of E-FABP and S100A7 in lesional psoriatic skin suggests a possible link with this hyperproliferative skin disease. In order to investigate a role for the proteins in this disease, the purifications for both proteins were re-analyzed. Moreover, a specific antiserum directed against purified human S100A7 was generated. By SDS-PAGE immunoblotting we show that E-FABP and S100A7 are expressed in cultured human differentiating keratinocytes and confirm their overexpression in psoriatic scales. Gel filtration and non-denaturing PAGE revealed that S100A7 co-purified with E-FABP, indicating an association between the two proteins. Ion-exchange chromatography resulted in the dissociation of the complex. Finally, immunoprecipitations using antiserum against E-FABP revealed that S100A7 co-immunoprecipitated with E-FABP from protein extracts of psoriatic scales. These data indicate that E-FABP and S100A7 might form a complex in the cytosol of human keratinocytes.

Calcium-binding protein S100A7 and epidermal-type fatty acid-binding protein are associated in the cytosol of human keratinocytes.

Expression of epidermal-type fatty acid-binding protein (E-FABP) and S100A7 has previously been shown to be elevated in psoriatic skin, a disease characterized by abnormal keratinocyte differentiation. However, no causal relationship between the up-regulation of these proteins and the disease has been shown. E-FABP is thought to be involved in cytosolic fatty acid (FA) transport, whereas the role of S100A7 is still unknown. In this report, we show by overlay assays that E-FABP, immobilized on nitrocellulose, is able to capture S100A7 from cytosolic psoriatic protein extracts and vice versa, suggesting the formation of a complex between the two proteins. Using purified E-FABP and S100A7, the complex can be reconstituted only in presence of EDTA. Moreover, we show that increased EDTA concentrations in psoriatic cytosolic protein extracts enhance complex formation. Partial complex disruption was obtained by the addition of physiological concentrations of Zn2+ (0.1 mM), whereas Ca2+ at 5 mM and Mg2+ at 30 mM had no effect. On the other hand, high Ca2+ concentrations (30 mM) resulted in partial complex disruption. Oleic acid-binding properties were observed for free E-FABP and the complex E-FABP-S100A7, but not for free S100A7. By using confocal microscopy we show that S100A7 and E-FABP are co-localized in the cytoplasm of differentiating keratinocytes from lesional psoriatic skin. These data indicate that formation of the E-FABP-S100A7 complex and its FA-binding function might be regulated at least by bivalent cations.

The fatty acid-binding heterocomplex FA-p34 formed by S100A8 and S100A9 is the major fatty acid carrier in neutrophils and translocates from the cytosol to the membrane upon stimulation.

Since no data are available concerning fatty acid (FA) transport in neutrophils we studied the presence of possible FA carriers. The kFA-p34 complex, composed of S100A8 and S100A9, has been implicated in the intracellular transport of arachidonic acid and its precursors in human keratinocytes. Here, we show that FA-p34 is the major FA carrier in human neutrophils (nFA-p34). The complex is highly expressed in resting neutrophils (2.65% of cytosolic proteins) and translocates to the membrane fraction upon stimulation with opsonized zymosan. Comparison of purified nFA-p34 with kFA-p34 shows that both complexes are composed of nearly the same subunits and possess similar binding properties for oleic acid. Densitometrical analyses of 2D gels show that n and kFA-p34 contain twice as much S100A8 and S100A9 suggesting an estimated stoichiometry of (S100A8)2S100A9. A method is described allowing to distinguish n and kFA-p34 from S100A8/S100A9 homo- and heteromer complexes that are devoid of FA-binding properties. After solvent extraction, we find by GC analysis linoleic acid as major endogenous ligand of purified kFA-p34. Our results suggest that nFA-p34, might be involved in the shuttling of unsaturated FA between the cytosol and the plasma membrane of neutrophils.

[Intra-chamber lidocaine in addition to tetracaine eyedrop anesthesia in clear cornea phacoemulsification]. / Intrakamerales Lidocain als Ergänzung zur Tetracaintropfanästhesie bei der Clear Cornea Phacoemulsifikation.

BACKGROUND: We have been successfully using topical tetracaine 1% anesthesia in clear corneal phacoemulsification since 1992. Adding intracameral lidocaine to topical anesthesia in cataract surgery has been reported since 1996. PATIENTS AND METHODS: We examined the effectiveness of intracameral lidocaine 1%, which we dyed with methylene blue 2% in 600 cases. During surgery we merely inject 0.3 ccl lidocaine 1% twice over the iris under a shield of a viscoelastic substance. After only a few seconds time of action we remove it again from the anterior chamber. RESULTS: With intracameral lidocaine 1% postoperative pain was totally eliminated for one to two hours, and usually no pain was felt thereafter. Non of our cases showed any negative signs of postoperative reaction of the eye. With the use of intracameral lidocaine we were able to omit pre- and postoperative systemic pain medication completely. This has proven to be very advantageous in elderly outpatients. Staining the lidocaine with methylene blue 2% helps prevent mixups and emphasizes its significance. Yet, an intracameral blue flow was not visible enough to determine the spread of lidocaine. CONCLUSIONS: Intracameral lidocaine represents an ideal addition to topical tetracaine anesthesia in clear corneal phacoemulsification.

A heterocomplex formed by the calcium-binding proteins MRP8 (S100A8) and MRP14 (S100A9) binds unsaturated fatty acids with high affinity.

We show that unsaturated fatty acids (FAs) bind reversibly and with high affinity to a heterocomplex of 34 kDa (FA-p34) formed by the non-covalent association of two calcium-binding proteins of the S100 family: MRP8 (S100A8) and MRP14 (S100A9). Fatty acid-competition studies on the [3H]oleic acid.FA-p34-complex show that oleic, alpha-linoleic, gamma-linolenic, and arachidonic acids have IC50 values of about 1 microM, whereas palmitic and stearic acids are poor competitors. The binding of arachidonic acid is saturable with a single class of binding site per FA-p34, and a dissociation constant (Kd) of 0.13 microM is calculated. The individual subunits MRP8 and MRP14 show no binding properties for fatty acids, whereas a p34 complex reconstituted in vitro by the recombinant molecules exhibits binding properties, suggesting that the fatty acid-binding site of FA-p34 is created through heterocomplex formation. Furthermore, we demonstrate that lowering free Ca2+ levels to 16 nM results in a loss of the fatty acid-binding capacity of purified FA-p34. In calcium-induced differentiating keratinocytes, the amounts of FA-p34 are increased in the particulate (2.0 +/- 0.5 pmol of [3H]oleic acid/mg protein) and in the cytosolic (4.5 +/- 0.6 pmol of [3H]oleic acid/mg protein) fractions, whereas no FA-p34 can be detected in non-differentiated cultured keratinocytes. In abnormally differentiated keratinocytes (psoriasis) and in human polymorphonuclear leukocytes, FA-p34 is highly expressed (31.35 +/- 1.6 and 349.8 +/- 17.9 pmol of [3H]oleic acid/mg protein, respectively), pointing toward a role for this heteromer in mediating effects of unsaturated fatty acids in a calcium-dependent way during cell differentiation and/or inflammation.

[The co-evolutional focus in individual, couple and family therapy]. / Der koevolutive Fokus in der Einzel-, Paar- und Familientherapie.

A concrete formulation of focus is proposed that in structure and use is suitable for individual as well as couples and family therapy. The concept of coevolution (Willi 1985, 1991) forms a basis for the supposition that important personal developments are realised in human relationships. The development of one person is challenged, taxed, or hindered by the development of other persons. The formulation of a coevolutional focus consists of four steps: 1) The context of the relationship in which the current conflict emerges; 2) The identification of delayed stages of development; 3) The personal and interactional conditions which block this development; 4) Concrete changes which make the progress in the intended development recognizable. The focus in couples and family therapy integrates the impending development of the participants in their collusive interactions.

Purification and characterization of the human epidermal fatty acid-binding protein: localization during epidermal cell differentiation in vivo and in vitro.

Epidermal fatty acid-binding protein (E-FABP) was isolated from human skin and purified to homogeneity. Its molecular mass was estimated to be 15 kDa and the pI of non-denaturing protein was 5.6. Scatchard-plot analysis revealed one class of binding site for oleic acid with a Kd of 0.46 microM. Structure-binding relation experiments revealed a high affinity of E-FABP for stearic acid which decreased on reduction of the number of carbon atoms or introduction of double bonds into the fatty acid chain. Squalene, cholesterol and retinoic acid isomers showed no affinity, suggesting that E-FABP displays high specificity for fatty acids. E-FABP is a scarce cytosolic protein (0.065% of total protein). Only trace amounts could be detected in normal human skin but up to 42.5 +/- 3.4 pmol/mg of protein was found in a non-malignant defect of keratinocyte differentiation (psoriatic lesions). E-FABP levels were low in cultured human keratinocytes grown under proliferation-stimulating conditions but increased about 2-fold on induction of differentiation by Ca2+. Immunohistochemical localization showed cytosolic staining in differentiated cells of normal and psoriatic skin, suggesting a link between E-FABP and keratinocyte differentiation. The presence of E-FABP in tissues other than skin (heart, intestine and adipose tissue) excludes its specific role in fatty acid metabolism in epithelial cells or its involvement in skin lipid-barrier function.

Characterization and expression of a novel human fatty acid-binding protein: the epidermal type (E-FABP).

Using PAGE--Autoradioblotting technique we have characterized an E--FABP in human epidermal cells that is distinct from liver-, heart-, intestine- and adipose tissue-FABPs. FABP radiobinding analysis was performed directly on protein extracts without prior partial purification. E-FABP has a Mr of approximately 15 kDa and binds oleic acid with high affinity but does not bind all-trans-, 13-cis- and 9-cis-retinoic acid nor all-trans-retinol. Expression levels of E-FABP were low in normal epidermis, higher in human cultured keratinocytes and still higher in psoriasis, a disease characterized by abnormal epidermal differentiation. These findings suggest that epidermal cells may have a distinct fatty acid metabolism compared to other tissues.

Human embryo research: ethics and recent progress.

This review evaluates recent developments in the application as well as legalization of human embryo research. A number of European countries, including the United Kingdom and Spain, have recently enacted comprehensive legislation to regulate embryo research. Research reviewed here was conducted either with the aim to alleviate certain medical conditions or to improve the low success rates of assisted reproductive technology. Research was usually conducted on embryos that were considered unfit for immediate transfer or unsuitable for cryopreservation. Research projects were aimed at 1) studying the metabolism of the embryo, 2) promoting embryonic viability, 3) assisting fertilization in patients with fertilization disorders, and 4) determining gene disorders in embryos from couples at risk for transmitting genetic disease.

Cellular retinoic acid- but not cellular retinol-binding protein is elevated in psoriatic plaques.

Cellular retinoid binding proteins are thought to be involved in the molecular action of retinoids, a family of compounds successfully used in the treatment of psoriasis. Therefore, both cellular retinol (CRBP)- and retinoic acid (CRABP)-binding proteins were analyzed in psoriatic skin. Three facts emerged from our study: both CRABP and CRBP are detectable in the skin of psoriatic patients; qualitatively, they both appear similar to the corresponding proteins of normal human skin, in terms of their elution profile and apparent Kd; and quantitatively, only CRABP was found to be 3 times higher in psoriatic plaques as compared to either nonlesional skin of psoriatic patients or the skin of normal subjects. Since psoriatic plaques are particularly responsive to systemic retinoids, specifically to retinoic acid analogues, our results suggest for the first time a link between the levels of CRABP and the responsiveness of a nonneoplastic hyperproliferative tissue to systemic administration of retinoids in the human.

Cellular retinol- and retinoic acid-binding proteins in the epidermis and dermis of normal human skin.

The distribution of cellular retinol- and retinoic acid-binding proteins (CRBP and CRABP) in normal human epidermis and dermis was examined by gel filtration. We showed that CRBP is entirely bound to a lipid-protein aggregate and its free form is obtained through delipidation. The type of homogenization procedure appeared to be important for the recovery of CRBP in human skin. The level of CRABP was about 3.1 pmol/mg protein in the epidermis whereas it was only sporadically detectable in the dermis. In contrast, CRBP was found in both tissues at a concentration of about 1 pmol/mg protein. The difference in CRABP concentrations between epidermis and dermis might have biological and therapeutic implications. Dissociation constants (Kd) of CRABP and CRBP were respectively 2.2 X 10(-7) M and 2.51 X 10(-7) M. This method will facilitate the study of CRABP and CRBP in retinoid- responsive dermatoses and enable us to relate the therapeutic effects of retinoids to the levels of cellular retinoid-binding proteins in the skin.