Hippo signaling instructs ectopic but not normal organ growth.

The Hippo signaling pathway is widely considered a master regulator of organ growth because of the prominent overgrowth phenotypes caused by experimental manipulation of its activity. Contrary to this model, we show here that removing Hippo transcriptional output did not impair the ability of the mouse liver and Drosophila eyes to grow to their normal size. Moreover, the transcriptional activity of the Hippo pathway effectors Yap/Taz/Yki did not correlate with cell proliferation, and hyperactivation of these effectors induced gene expression programs that did not recapitulate normal development. Concordantly, a functional screen in Drosophila identified several Hippo pathway target genes that were required for ectopic overgrowth but not normal growth. Thus, Hippo signaling does not instruct normal growth, and the Hippo-induced overgrowth phenotypes are caused by the activation of abnormal genetic programs.

Effect of the stiffness of bone substitutes on the biomechanical behaviour of femur for core decompression.

Core decompression is the most common procedure for treatment of the early stages of osteonecrosis of the femoral head. The purpose of this study was to compare the biomechanical performance of four different bone graft substitutes combined with core decompression. Subject-specific finite element models generated from computed tomography (CT) scan data were used for a comprehensive analysis. Two different contact conditions were simulated representing states of osseointegration at the interface. Our results showed that the use of a low-stiffness bone substitute did not increase the risk of femoral fracture in the early postoperative phase, but resulted in less micromotion and interfacial stresses than high-stiffness bone substitutes.

Experimental and computational studies on the femoral fracture risk for advanced core decompression.

BACKGROUND: Two questions are often addressed by orthopedists relating to core decompression procedure: 1) Is the core decompression procedure associated with a considerable lack of structural support of the bone? and 2) Is there an optimal region for the surgical entrance point for which the fracture risk would be lowest? As bioresorbable bone substitutes become more and more common and core decompression has been described in combination with them, the current study takes this into account. METHODS: Finite element model of a femur treated by core decompression with bone substitute was simulated and analyzed. In-vitro compression testing of femora was used to confirm finite element results. FINDINGS: The results showed that for core decompression with standard drilling in combination with artificial bone substitute refilling, daily activities (normal walking and walking downstairs) are not risky for femoral fracture. The femoral fracture risk increased successively when the entrance point is located further distal. The critical value of the deviation of the entrance point to a more distal part is about 20mm. INTERPRETATION: The study findings demonstrate that optimal entrance point should locate on the proximal subtrochanteric region in order to reduce the subtrochanteric fracture risk. Furthermore the consistent results of finite element and in-vitro testing imply that the simulations are sufficient.

Azole antimycotics--a highway to new drugs or a dead end?

Azole antimycotics are a well-known and important class of agents that are used in hospital practice, everyday health care, veterinary medicine and for crop protection. The era of azole fungicides began with the breakthrough of chlormidazole roughly 50 years ago. Since then, more than 20 drugs of this group, including triazoles, have been brought to the market. The specific chemical structure and mechanism of the action of azoles along with the eukaryotic character of fungal pathogens raise several serious issues. Resistance to drugs and disturbance to metabolic pathways are among the most important. On the other hand, these same features are responsible for unique and novel applications of these drugs. As a result, old and ineffective antifungal drugs can be successfully used in the treatment of parasitic diseases, bacterial infections or cancers. Are azoles getting their second wind?

Abuse liability of oxycodone as a function of pain and drug use history.

The relationship between pain and prescription opioid abuse is poorly understood. Determining whether a patient is seeking additional opioid medications in order to alleviate pain or to abuse the drugs can be difficult. The present study was designed to evaluate two variables that may influence the abuse liability of opioids: drug use history and the presence or absence of experimentally induced pain. Eighteen healthy participants completed this outpatient study. One group was abusing prescription opioids (N=9) and one group had used prescription opioids medically but did not abuse them (N=9). All participants completed twelve sessions during which the effects of orally delivered oxycodone (0, 15, 30mg/70kg, PO) were examined. One dose was tested per day under double-blind conditions and sessions were separated by at least 48h. During the first "sample" session each week, participants were given $10 and the dose that was available later that week. During the second "choice" session, participants could self-administer either money or the previously sampled dose. Six sessions involved repeated hand immersions in cold water (4 degrees C) and six sessions involved immersions in warm water (37 degrees C). Most of the positive subjective effects of oxycodone were similar between the groups, but oxycodone self-administration significantly differed between groups. Non-abusers self-administered active doses of oxycodone only when they were in pain while abusers self-administered oxycodone regardless of the pain condition. These data suggest that an assessment of the reinforcing effects of opioids may be a sensitive method for differentiating opioid abusers from non-abusers.

Sequencing batch reactor (SBR) as optimal method for production of granular activated sludge (GAS)--fluid dynamic investigations.

Fluid dynamic investigations of multiphase flow (fluid, air, granules) in a sequencing batch reactor (SBR) are presented. SBR can be considered as an attractive technology for cultivation of granular activated sludge (GAS). Granulation is a complicated process and its mechanism is not fully understood yet. Many factors influence the formation and structure of aerobic granular sludge in a bioreactor. Extracellular polymer substances (EPS) and superficial gas velocity (SGV) play a crucial role for granules formation. Additionally, it is supposed that EPS production is stimulated by mechanical forces. It is also assumed that hydrodynamic effects have a major influence on the formation, shape and size of GAS in SBR under aerobic condition. However, the influence of stress on granulation is poorly investigated. Thus, in the present paper, fluid dynamic investigations of multiphase flow in a SBR, particularly effect of normal and shear strain, are reported. In order to analyse multiphase flow in the SBR, optical in-situ techniques with particle image velocimetry (PIV) and particle tracking velocimetry (PTV) are implemented. Obtained results show a characteristic flow pattern in a SBR. It is pointed out that additional effects like particle-wall collisions, inter particle collisions, erosion can also affect significantly granules formation.

Analogues of arginine vasopressin and its agonist and antagonist modified in the N-terminal part of the molecule with l-beta-homophenylalanine.

In continuation of our efforts to elucidate the role of positions 2 and 3 in arginine vasopressin (AVP) and its analogues, we designed and synthesized peptides modified in these positions with l-beta-homophenylalanine (beta-Hph). Two of them had just this single modification, the next two peptides are analogues of the V2 agonist, namely [3-mercaptopropionic acid (Mpa)1]AVP (dAVP). The last two compounds were designed by substitution of positions 2 or 3 of a potent V(1a) antagonist, [1-mercaptocyclohexaneacetic acid (Cpa)1]AVP, with beta-Hph. All the peptides were tested for their pressor and antidiuretic and uterotonic in vitro activities in the rat. All the activities tested have been found to be significantly decreased. Three analogues, i.e. [Mpa(1),beta-Hph2]AVP, [Cpa1,beta-Hph2]AVP, [Cpa1,beta-Hph3]AVP, turned out to be uterotonic antagonists with pA2 = 6.3 +/- 0.2, 6.3 +/- 0.1, 6.0 +/- 0.3 respectively. The last one exhibited antipressor properties also (pA2 = 6.4 +/- 0.1).

Analogues of arginine vasopressin modified in the N-terminal part of the molecule with enantiomers of N-methylphenylalanine.

Four new analogues of arginine vasopressin (AVP) substituted in positions 2 and 3 with all possible combinations of enantiomers of N-methylphenylalanine were synthesized and studied to assess the influence of N-methylation of the peptide bonds between the first three amino acids on the pharmacological properties of the resulting peptides. The next three analogues were designed to learn how the shortening of the peptide chain, by removal of one of the N-methylphenylalanine residues, would affect pharmacological properties of the resulting compounds. The activity of the analogues was tested in the in vitro uterotonic, pressor and antidiuretic tests. None of the prepared analogues displayed significant biological activity with the exception of [Me-d-Phe(2), Me-Phe(3)]AVP and [Me-d-Phe(2,3)]AVP, which showed low antiuterotonic activity (pA(2) = 6.6 and pA(2) = 6.4, respectively). Our results, while not impressive in terms of biological activity, may be helpful for designing potent and selective oxytocin antagonists.

Cis/trans conformational equilibrium across the N-methylphenylalanine- N-methylphenylalanine peptide bond of arginine vasopressin analogs.

Two cyclic analogs of vasopressin, -Pro-Arg-Gly-NH(2) (1) and -Pro-Arg-Gly-NH(2) (2) were synthesized by the solid phase method. Their structure was determined in aqueous solution by two-dimensional NMR techniques and simulated annealing algorithm from an extended template in X-PLOR. The total chemical shift correlation spectroscopy and rotating-frame Overhauser enhancement spectroscopy of the peptides displayed four distinct sets of residual proton resonances. This suggests that both analogs adopt four families of conformations in H(2)O/D(2)O (9 : 1) (one major and three minor species). In further analysis only signals of major species (M) and of one minor species (m(1)) were considered. The major species of both peptides include a trans peptide bond between the first and second residue, and a cis form between the second and third residue. In the minor species, all peptide bonds were found to exist in trans geometry.

Potent bradykinin antagonists containing N-benzylglycine or N-benzyl-l-alanine in position 8.

Two new analogues of a previously designed bradykinin (BK) antagonist, d-Arg-Arg-Pro-Hyp-Gly-Thi-Ser-d-Phe-Thi-Arg, substituted in position 8 by N-benzylglycine and N-benzyl-l-alanine were designed, synthesized and bioassayed. The results show an impressive enhancement of B2 antagonistic potencies of both peptides in comparison with the model. In two further analogues these modifications were combined with acylation of the N-terminus with 1-adamantanacarboxylic acid. Acylated analogues exhibited higher antagonistic potency in comparison with the parent compounds, however, the range of effect was not as high as in previously described cases. The activity of analogues was assessed by their ability to inhibit vasodepressor response to exogenous BK (rat blood pressure test). Our results may be of value in the design of more potent BK antagonists.

Influence of 1-aminocyclohexane-1-carboxylic acid in position 2 or 3 of AVP and its analogues on their pharmacological properties.

In this study we describe the synthesis and some pharmacological properties of seven new analogues of arginine vasopressin (AVP) substituted in position 2 or 3 with 1-aminocyclohexane-1-carboxylic acid (Acc). All peptides were tested for the pressor, antidiuretic and uterotonic in vitro activities. The Acc3 modifications of AVP, dAVP, [d-Arg8]VP and [Cpa1]AVP have been found to be deleterious for interaction with all three neurohypophyseal hormone receptors, as judged from the several orders of magnitude decreased biological activities, whereas Acc2 substitution selectively altered the interaction with the receptors. Two of the new analogues, [Acc2]AVP and [Acc2, d-Arg8]AVP, are potent antidiuretic agonists. [Acc2]AVP exhibits moderate pressor agonistic activity and weak antiuterotonic properties. [Acc2, d-Arg8]AVP has been found to be a weak antagonist in the pressor and uterotonic tests. Another analogue - [Cpa1, Acc3]AVP - turned out to be a highly selective V2 agonist. This is an unexpected effect, as its parent peptide, [Cpa1]AVP is a very potent V1a receptor antagonist. This is the first Cpa1 modification to have resulted in V2 agonism enhancement. Besides providing useful information about structure-activity relationships, our results could open up new possibilities in the design of highly potent and selective V2 agonists.

Phosphorylation of acidic ribosomal proteins by ribosome-associated protein kinases of Saccharomyces cerevisiae and Schizosaccharomyces pombe.

Two proteins of 13 kDa and 38 kDa, the components of 60S ribosomal subunits, were identified as phosphorylation substrates for protein kinases tightly associated with S. cerevisiae and Schizosaccharomyces pombe ribosomes. An enzyme with properties of multifunctional casein kinase II was detected in ribosome preparations from both yeast species. In S. cerevisiae another protein kinase with high substrate specificity toward those proteins was also identified. By using isoelectric focusing, the protein band of 13 kDa from S. cerevisiae and S. pombe was resolved respectively into three and four major forms of different charge. The same protein forms were phosphorylated in the in vivo 32P-labelling experiments.

Sensitivity of the replica method in the detection of candidal infection among denture wearers with clinically healthy oral mucosa.

To ascertain the role of Candida in denture stomatitis, the practitioner must conduct a mycologic examination of the acrylic resin denture surface, because it acts as a reservoir for continuous reinfection of the palate. Twenty-two patients were examined to compare the sensitivity of the standard technique of swabbing the denture to that of a newly developed cast agar replica technique for detecting Candida albicans. The dentures were swabbed and cast replicas of the tissue-fitting surface of the dentures were made of both study populations. The majority of cultures obtained by swabbing failed to detect the presence of Candida albicans, while all cast agar replicas grew Candida albicans. The replica method for the detection of Candida albicans in edentulous patients seemed to be a more sensitive method than currently available mycologic methods.

Changes in brain bioelectrical activity (EEG) after repetitive exposure to an organo-phosphate anticholinesterase. II. Rat.

Effects of repetitive exposure (ten times in a period of two weeks) to chlorphenvinphos (CVP), at daily doses of 0.5 and 1.0 mg/kg, i.p., were studied in adult male Wistar rats of imp-DaK stock. It was found that 3 hrs after the last exposure, the cholinesterase (ChE) activity in the blood and brain was close to 50% of the control value in the 0.5 mg/kg group and less than 50% in the 1.0 mg/kg group. In both groups, normalization of ChE activity in plasma took less time than in erythrocytes, and the normalization of ChE activity in erythrocytes proceeded faster than in the majority of the brain areas studied. Electrophysiological investigations revealed a retardation of age-related epileptic-like cortical activity. This effect, however, was present only in the 1.0 mg/kg group, and only in the period of decreased ChE activity in the brain. Spectral analysis revealed an increase in 1-4 Hz activity in the cortical EEG of the 0.5 mg/kg group and heightened theta activity (4-7 and 7-9 Hz bands) in the hippocampal EEG of the 1.0 mg/kg group. The later effects were detected after a time sufficient for full normalization of ChE activity and manifested themselves most clearly in the presence of an acoustic stimulus associated with pain. The above results are in agreement with earlier observations on rabbits exposed repetitively to CVP. Data from both species suggest that, in the case of repetitive exposure to CVP, neither plasma nor erythrocyte ChE activity is a reliable indicator of toxicity, and that such exposure to this OP may lead to changes in EEG outlasting the period of lowered ChE activity in the blood and brain.

Brain electrical activity (EEG) after repetitive exposure to chlorphenvinphos an organophosphate anticholinesterase: I. Rabbit.

Cholinesterase (ChE) activity in blood and brain as well as the hippocampal and cortical EEG were investigated in rabbits exposed once a day for a period of two weeks to an organophosphate insecticide, chlorphenvinphos (CVP). The daily dose of CVP was 14.0 mg/kg i.p. ChE activity in plasma and erythrocytes decreased by 60 and 48%, respectively, by the end of exposure and returned to the preexposure level within 35 days. In the exposed animals, killed and dissected a day after, the ChE activity in blood had been found normal, the level of ChE activity in some parts of the brain was still significantly depressed. The spontaneous hippocampal EEG showed no changes as soon as 24 hours after the exposure. However, after a period sufficiently long for the normalization of ChE activity in blood, the hippocampal arousal response (theta rhythm) to click and to a tone associated with pain was found heightened in the exposed rabbits. Moreover, spectral analysis of 5 minute EEG samples revealed a decrease in the content of 7-13 Hz activity in the cortex of the exposed animals as compared to the control ones. The obtained data suggest that exposure to CVP may lead to functional changes in the brain outlasting the period of ChE depression.

Changes in physiological and electrophysiological parameters in rabbits after single exposure to chlorphenvinphos.

In order to evaluate some of the central effects of an organophosphorus pesticide--chlorphenvinphos (CVP), the time course of changes in the activity of blood cholinesterase (ChE), body temperature and hippocampal EEG were compared in rabbits after acute i.p. exposure. The pesticide was administered twice at an interval of 80-90 days. The Deichmann-LeBlanc scheme of dosing (one animal/one dose) was adopted starting with 22 mg/kg. CVP resulted in a dose-dependent decrease in plasma and erythrocyte ChE activity, a decrease in body temperature ranging from 0.7 degrees to 3.5 degrees C, and in an increase in content of the immobility-related, rhythmic slow activity (I-RSA) in the hippocampal EEG. Changes in body temperature appeared at lower doses than those in the hippocampal EEG. Contrary to the changes in ChE activity, which lasted 4-30 days, those in body temperature and in hippocampal EEG disappeared within 24 hours after the injection. CVP administered at the same dose 80-90 days after the first injection, resulted again in an inhibition of ChE activity, but the effect on the hippocampal EEG was less clear, and that on body temperature was variable; no effect, an increase or a decrease appeared. The data suggest that: i) body temperature is a more sensitive index of a central action of CVP administered for the first time than the hippocampal EEG; ii) the brain cholinergic mechanisms are relatively resistant to the acute action of the OP and undergo fast adaptive changes; iii) even single exposure to CVP may produce some long-lasting functional changes in the brain of some subjects, which has been proven by the changed response to the second exposure.