Prenatal and postnatal expression of mRNA coding for rat prothrombin.

The levels of prothrombin mRNA in prenatal and postnatal rat tissues were analyzed in order to determine tissue distribution of prothrombin expression and to determine if increases in liver prothrombin mRNA during development correlated with previously documented developmental increases in plasma prothrombin levels. Maternal tissues were also analyzed in order to determine if prothrombin mRNA levels varied due to gestational or postpartum influences. Northern analysis demonstrated that rat liver prothrombin mRNA levels increased several-fold late in gestation and reached maximal levels by 13 days after birth. Prothrombin mRNA was also expressed in diaphragm, stomach, intestine, kidney, spleen and adrenal tissues during development. In maternal tissues during pregnancy, prothrombin mRNA was expressed in liver, diaphragm, stomach, uterus and placenta. Prothrombin mRNA levels in each of these tissues that were positive by Northern analysis were quantitated by solution hybridization analysis. Between gestational day 18 and postnatal day 13, liver prothrombin mRNA levels increased from approx. 600 to 2100 molecules per cell (a 3.5-fold increase). In maternal liver during pregnancy, between day 18 and day 22, prothrombin mRNA levels increased from approx. 1800 to 2100 molecules per cell. Immediately after delivery, maternal liver prothrombin mRNA levels decreased to approx. 50% of preparturition levels. Prothrombin mRNA levels in placental tissue ranged from approx. 100 to 250 molecules per cell. In other fetal, postnatal and maternal tissues, prothrombin mRNA expression was less than 100 molecules per cell. These results demonstrate that the level and tissue-type expression of prothrombin mRNA varies in response to prenatal and postnatal influences.     

Average daily metabolic rate of gerbils of two species: Gerbillus pyramidum and Gerbillus allenbyi

Gerbillus pyramidum and G. allenbyi are primarily granivorous, nocturnal rodents that are sympatric over many sandy areas of the Negev Desert. However, in their overall distribution, G. pyramidum occurs in extreme desert areas whereas G. allenbyi does not. We measured the average daily metabolic rate (ADMR) of gerbils of each species when they were offered pelleted diet. Given the difference in their distribution, we reasoned that the more xeric G. pyramidum would have lower ADMR than G. allenbyi; however, given the similarity in their diets, we reasoned that their ADMRs would be similar. The latter alternative was supported. ADMR of G. pyramidum (body mass = 31·9 ± 5·4 g) was 427·1 kJ·kg^−0.75·d⁻¹, 58% of that predicted for a rodent of its body mass; whereas ADMR of G. allenbyi (body mass = 22·3±2·3 g) was 387·7 kJ·kg^−0.75·d⁻¹, 49%) of that predicted. On the basis of these results, we suggest that factors other than their ADMRs are important in determining their geographical distribution.     

Diet selection and energy and water budgets of the common spiny mouse Acomys cahivinus

The common spiny mouse, Acomys cahirinus (body mass=47 g), is widely distributed in Israeli deserts where it inhabits natural crevices on rocky slopes. This omnivorous rodent consumes a varied diet, and in particular snails. We determined diet selection and energy and water balances of spiny mice when they were offered snails and seeds. The spiny mice maintained steady state body mass. Dry matter consumption of snails was 0.014 g* g⁻¹.-d⁻¹ and of seeds was 0.049g*.g⁻¹d⁻¹ for a total of 0.063 g*g⁻¹*d⁻¹. Total water intake was 0.101ml-g⁻¹.d⁻¹ and metabolizable energy intakewas 0.990 kJ. gxs^-1.d⁻¹ for a ratio (ml: kJ) of 0.102. This ratio was similar to that reported in a previously published study on free-living spiny mice. We concluded that snails and seeds allowed spiny mice to fulfil their energy and water requirements with minimal dry matter and fresh matter intakes. Furthermore, spiny mice selected a diet that provided them with a water (ml) to energy (kJ) ratio of approximately 0.1, although it appeared that they are able to survive on a much drier diet.     

Interaction of phytoestrogens and other environmental estrogens with prostaglandin synthase in vitro

The phytoestrogens daidzein, genistein, equol and coumestrol were found to stimulate microsomal prostaglandin H synthase (PHS) in vitro in a concentration-dependent manner when PHS-activity was measured by arachidonic acid-dependent oxygen uptake. These compounds were co-oxidized by PHS and the conversion of parent compounds was measured by HPLC analysis. The stimulation of PHS-cyclooxygenase by these compounds was partially reversed at high concentrations probably due to their antioxidant properties causing inhibition. In contrast, the monomethyl ethers of daidzein and genistein, formononetin and biochanin A, had little or weakly inhibitory effect on PHS, and appear to be no or poor co-substrates for PHS. Compared to the equine estrogen equilin, its metabolite d-equilenin was poorly metabolized by PHS and inhibited rather than stimulated PHS-cyclooxygenase activity in vitro. The resorcylic acid lactones zearalenone and zeranol, on the other hand, were surprisingly good inhibitors of PHS-cyclooxygenase. Furthermore, zeranol inhibited both the arachidonic acid and the hydrogen-peroxide-dependent oxidation of DES in contrast to indomethacin which inhibited only cyclooxygenase-dependent co-oxidation of DES. The results of this in vitro study are discussed in the context of data on synthetic and steroidal estrogens and support the idea that PHS-activity may be modulated by interaction with certain estrogenic compounds.     

Enzymatic conversion of N carbamoyl-d-amino acids to d-amino acids

An enzyme isolated from Agrobacterium radiobacter was shown to catalyse the following reaction: H₂O + N-carbamoyl-d-amino acid→ d-amino acid + NH₃ + CO₂ Some properties of this new enzyme, N-carbamoyl-d-amino acid amidohydrolase, are presented in this paper. The potential application of this enzyme for the preparation of some d-amino acids used as pharmaceutical intermediates is discussed.     

Marking otoliths of Baltic cod (Gadus morhua Linnaeus, 1758) with tetracycline and strontium chloride

Identified were suitable dosages of tetracycline hydrochloride (TET) in three single treatments and three combined treatments of TET with 2 mg/kg strontium chloride (STR) in wild western Baltic cod (Gadus morhua), in terms of (a) obtainable mark qualities (visibility of fluorescent bands), (b) growth assessment, and (c) induced mortality rates. Isotonic NaCl solution was injected in a control group (25 cod per treatment). The results provide the basis for imperative age validation studies of Baltic Sea cod. Cod originating from pound nets near Fehmarn Island were kept in swimming net cages at the harbor of Warnemünde for 1.5 months. Mean initial total length was 28(±3) cm (salinity: 13, water temperature: 13 to 8°C). Overall average growth of surviving cod was 0.8 mm/day. In single TET treatments, lowest mortality rates and best mark quality were observed for TET concentrations of 100 compared to 50 and 25 mg/kg wet mass. Mortality rates of the 100 mg/kg treatment group were remarkably lower than in the control group emphasizing the antibiotic effect of TET. By contrast, the double treatment in the TET‐STR groups resulted in a binding interaction between both markers in the fish body causing either the antibiotic potency being inhibited or TET and STR forming a non‐beneficial chelate (increased mortality), and decreased incorporation of TET in the otolith (reduced visibility of TET bands). Consequently, TET (short‐term marker) and STR (long‐term marker) should not be injected together. Our results demonstrate that the binding interactions between these substances known from homoiotherm animals also apply for poikilotherms such as fish.     

Characterization of connexin30.3-deficient mice suggests a possible role of connexin30.3 in olfaction

We have generated connexin30.3-deficient mice in which the coding region of the connexin30.3 gene was replaced by the lacZ reporter gene. The expression pattern of this connexin was characterized using beta-galactosidase staining and immunoblot analyses. In skin, beta-galactosidase/connexin30.3 protein was expressed in the spinous and granulous layers of the epidermis. Specific beta-galactosidase/connexin30.3 expression was also detected in the thin ascending limb of Henle's loop in the kidney. In addition, we found beta-galactosidase/connexin30.3 in progenitor cells of the olfactory epithelium and in a subpopulation of cells in the apical layer of the vomeronasal organ. Connexin30.3-deficient mice were fertile and displayed no abnormalities in the skin or in the chemosensory systems. Furthermore, they showed normal auditory thresholds as measured by brain stem evoked potentials. These mice did, however, exhibit reduced behavioural responses to a vanilla scent.     

Automated quantitative determination of a new polyamine biosynthesis inhibitor (CGP 48 664) and a potential metabolite in human and animal plasma by high-performance liquid chromatography

A specific and sensitive liquid chromatographic assay for the determination of 4-amidino-1-indanone-2′-amidinohydrazone (CGP 48 664, I) and a potential metabolite, 2-(4-carbamoyl-2,3-dihydro-1H-inden-1-yliden) hydrazine carboximidamide (CGP 53 391, II), in human and animal plasma was developed. CGP 51 467, a structural analog, was added to the plasma samples (up to 1 ml) as an internal standard. After mixing, the samples were processed automatically using an ASPEC solid-phase extraction system. The final extracts were chromatographed on a 5 μm Purospher RP-18 HPLC column. Chromatography was performed using a gradient system and UV detection. The described HPLC method is suitable for specific and quantitative measurement of concentrations of I, as well as its potential metabolite II down to 5–10 ng/ml in human and animal (dog, rat) plasma with acceptable reproducibility and accuracy.     

Binding of Efb from Staphylococcus aureus to fibrinogen blocks neutrophil adherence

In addition to its pivotal role in hemostasis, fibrinogen (Fg) and provisional fibrin matrices play important roles in inflammation and regulate innate immune responses by interacting with leukocytes. Efb (the extracellular fibrinogen-binding protein) is a secreted Staphylococcus aureus protein that engages host Fg and complement C3. However, the molecular details underlying the Efb-Fg interaction and the biological relevance of this interaction have not been determined. In the present study, we characterize the interaction of Efb with Fg. We demonstrate that the Fg binding activity is located within the intrinsically disordered N-terminal half of Efb (Efb-N) and that the D fragment of Fg is the region that mediates Efb-N binding. More detailed studies of the Efb-N-Fg interactions using ELISA and surface plasmon resonance analyses revealed that Efb-N exhibits a much higher affinity for Fg than typically observed with Fg-binding MSCRAMMs (microbial surface components recognizing adhesive matrix molecules), and data obtained from ELISA analyses using truncated Efb-N constructs demonstrate that Efb-N contains two binding sites located within residues 30-67 and 68-98, respectively. Efb-N inhibits neutrophil adhesion to immobilized Fg by binding to Fg and blocking the interaction of the protein with the leukocyte integrin receptor, α(M)β(2). A motif in the Fg γ chain previously shown to be central to the α(M)β(2) interaction was shown to be functionally distinguishable from the Efb-N binding site, suggesting that the Fg-Efb interaction indirectly impedes Fg engagement by α(M)β(2). Taken together, these studies provide insights into how Efb interacts with Fg and suggest that Efb may support bacterial virulence at least in part by impeding Fg-driven leukocyte adhesion events.