Long term behavior of dexamethasone-loaded cochlear implants: In vitro & in vivo.

The aim of this study was to better understand the long term behavior of silicone-based cochlear implants loaded with dexamethasone: in vitro as well as in vivo (gerbils). This type of local controlled drug delivery systems offers an interesting potential for the treatment of hearing loss. Because very long release periods are targeted (several years/decades), product optimization is highly challenging. Up to now, only little is known on the long term behavior of these systems, including their drug release patterns as well as potential swelling or shrinking upon exposure to aqueous media or living tissue. Different types of cylindrical, cochlear implants were prepared by injection molding, varying their dimensions (being suitable for use in humans or gerbils) and initial drug loading (0, 1 or 10%). Dexamethasone release was monitored in vitro upon exposure to artificial perilymph at 37 °C for >3 years. Optical microscopy, X-ray diffraction and Raman imaging were used to characterize the implants before and after exposure to the release medium in vitro, as well as after 2 years implantation in gerbils. Importantly, in all cases dexamethasone release was reliably controlled during the observation periods. Diffusional mass transport and limited drug solubility effects within the silicone matrices seem to play a major role. Initially, the dexamethasone is homogeneously distributed throughout the polymeric matrices in the form of tiny crystals. Upon exposure to aqueous media or living tissue, limited amounts of water penetrate into the implant, dissolve the drug, which subsequently diffuses out. Surface-near regions are depleted first, resulting in an increase in the apparent drug diffusivity with time. No evidence for noteworthy implant swelling or shrinkage was observed in vitro, nor in vivo. A simplified mathematical model can be used to facilitate drug product optimization, allowing the prediction of the resulting drug release rates during decades as a function of the implant's design.

Hybrid Ear Cubes for local controlled dexamethasone delivery to the inner ear.

A new type of miniaturized implants for local controlled drug delivery to the inner ear is proposed: Hybrid Ear Cubes. They are composed of two main parts: (i) a cylinder, which is placed into a tiny hole (<0.4 mm) drilled into (or close to) the oval (or round) window, and (ii) a cuboid, which is placed into the middle ear. The drug is released at a pre-programmed rate into the perilymph: (i) via the cylindrical part of the implant, which is in direct contact with this liquid, and (ii) via diffusion from the cuboid through the oval/round window. Importantly, the cylindrical part assures a reliable fixation of the drug delivery system at the site of administration. Furthermore, the cuboid provides a relatively "large" drug reservoir, without expulsing perilymph from the cochlea. The required surgery is minimized compared to the placement of an intracochlear implant. In contrast to previously proposed Ear Cubes, which are mono-block systems, Hybrid Ear Cubes consist of two halves, which can: (i) be loaded with different drugs, (ii) be loaded with the same drug at different concentrations, and/or (iii) be based on two different matrix formers. This offers a substantially increased formulation flexibility. Different types of silicone-based Hybrid Ear Cubes were prepared, loaded with 10% dexamethasone in one half and 0-60% dexamethasone in the other half. Importantly, tiny drug crystals were homogeneously distributed throughout the respective implant halves. The observed drug release rates were very low (e.g., <0.5% after 2 months), which can be attributed to the type of drug and silicone as well as to the very small surface area exposed to the release medium. Importantly, no noteworthy implant swelling was observed.

Relationship between pH of ruminal fluid during subacute ruminal acidosis and physiological response of the Polish Holstein-Friesian dairy cows.

The aim of the study was to investigate the effect of ruminal fluid pH depression on biochemical indices of blood, urine, feces, and milk, and to determine which of them may be helpful as a marker for the diagnosis of subacute ruminal acidosis (SARA). Ruminal fluid samples were obtained by rumenocentesis from 305 cows representing 13 commercial dairy herds. The herds were selected based on percentages of cows with an assigned value of ruminal fluid pH segregated into three groups as: SARA-positive herd, if at least 25% of the ruminal fluid samples indicated a pH < 5.6; SARA-risk herd, if less than 25% of ruminal fluid samples indicated a pH < 5.6, but at least 33% showed a pH ≤ 5.8; and SARA-negative herd, if less than 25% of the ruminal fluid samples indicated a pH < 5.6, but less than 33% exhibited a pH = 5.8. Moreover, the dairy cows were divided according to the ruminal fluid pH into three groups as follows: healthy cows (HC, pH>5.80, n = 196), risk cows (RC, pH 5.8 - 5.6, n = 51), and acidotic cows (AC, pH < 5.6, n = 58). Almost 19% (58/305) of the cows were classified as acidotic (pH < 5.6) and 46.2% of the herds as SARA-positive. In the AC group, higher concentrations of insulin-like growth factor-I (IGF-I), non-esterified fatty acids (NEFA), rectal temperature and lower blood pH, compared with those of the HC group, were recorded. Moreover, in the SARA-positive herds, higher concentrations of IGF-I and the lowest blood pH, compared with SARA-negative herds, were observed. The lowering of ruminal fluid pH increased the blood IGF-I and NEFA concentrations and the rectal temperature and decreased the blood pH. These measures are indicators of the physiological changes that occur as part of the pathogenesis of the condition and may be helpful for the diagnosis of the SARA syndrome when serial measurements are conducted.

Limited drug solubility can be decisive even for freely soluble drugs in highly swollen matrix tablets.

The aim of this study was to elucidate the importance of potential limited solubility effects for the control of drug release from hydrophilic matrix tablets loaded with a freely water-soluble drug. It is often assumed that the considerable amounts of water penetrating into this type of advanced drug delivery systems are sufficient to rapidly dissolve the entire drug loading, and that limited drug solubility is not playing a role for the control of drug release. Here, we show that this assumption can be erroneous. HPMC/lactose matrix tablets were loaded with 5 to 60% diprophylline (e.g. solubility in 0.1M HCl at 37°C: 235mg/mL), and drug release was measured at low and neutral pH, respectively. A mechanistically realistic mathematical theory was applied, considering drug diffusion in axial and radial direction in the cylindrical matrices and the potential co-existence of dissolved and non-dissolved drug. Importantly, only dissolved drug is available for diffusion. It is demonstrated that during major parts of the release periods, non-dissolved drug excess exists within tablets containing 30% or more diprophylline, despite of the substantial water contents of the systems. This leads to partially almost linear drug concentration distance profiles within the tablets, and reveals a major contribution of limited drug solubility effects to the control of drug release, even in the case of freely water-soluble diprophylline. It can be expected that also in other types of drug delivery systems, e.g. microparticles and implants (containing much less water), limited drug solubility effects play a much more important role than currently recognized.

Ear Cubes for local controlled drug delivery to the inner ear.

A new type of advanced drug delivery systems is proposed: Miniaturized implants, which can be placed into tiny holes drilled into (or close to) the oval window. They consist of two parts: 1) A cylinder, which is inserted into the hole crossing the oval window. The cylinder (being longer than the depth of the hole) is partly located within the inner ear and surrounded by perilymph. This provides direct access to the target site, and at the same time assures implant fixation. 2) A cuboid, which is located in the middle ear, serving as a drug reservoir. One side of the cuboid is in direct contact with the oval window. Drug release into the cochlea occurs by diffusion through the cylindrical part of the Ear Cubes and by diffusion from the cuboid into and through the oval window. High precision molds were used to prepare two differently sized Ear Cubes by injection molding. The miniaturized implants were based on silicone and loaded with different amounts of dexamethasone (10 to 30 % w/w). The systems were thoroughly characterized before and upon exposure to artificial perilymph at 37°C. Importantly, drug release can effectively be controlled and sustained during long time periods (up to several years). Furthermore, the implants did not swell or erode to a noteworthy extent during the observation period. Drug diffusion through the polymeric matrix, together with limited dexamethasone solubility effects, seem to control the resulting drug release kinetics, which can roughly be estimated using mathematical equations derived from Fick's second law. Importantly, the proposed Ear Cubes are likely to provide much more reliable local long term drug delivery to the inner ear compared to liquid or semi-solid dosage forms administered into the middle ear, due to a more secured fixation. Furthermore, they require less invasive surgeries and can accommodate higher drug amounts compared to intracochlear implants. Thus, they offer the potential to open up new horizons for innovative therapeutic strategies to treat inner ear diseases and disorders.

How to adjust dexamethasone mobility in silicone matrices: A quantitative treatment.

Silicone-based drug delivery systems offer a great potential to improve the therapeutic efficacy and safety of a large variety of medical treatments, e.g. allowing for local long-term delivery of active agents to the inner ear. Different formulation parameters can be varied to adjust desired drug release kinetics. However, often only qualitative information is available on their effects, and product optimization is cumbersome. The aim of this study was to provide a quantitative analysis, allowing also for theoretical predictions of the impact of the device design on system performance. Dexamethasone was incorporated into thin films based on different types of silicones (e.g. varying in the type of side chains and contents of amorphous silica), optionally containing different types and amounts of poly(ethylene glycol) (PEG) (5% or 10%). Furthermore, the initial drug content was altered (from 10% to 50%). In most cases, an analytical solution of Fick's second law could be used to describe the resulting drug release kinetics from the films and to determine the respective "apparent" diffusion coefficient of the drug (which varied from 2×10(-14) to 2×10(-12)cm(2)/s, depending on the system's composition). Thus, the impact of the investigated formulation parameters on drug mobility in the polymeric matrices could be quantitatively described. Importantly, the knowledge of the "apparent" drug diffusivity can be used to theoretically predict the resulting release kinetics from dosage forms of arbitrary size and shape. For instance, dexamethasone release was theoretically predicted from cylindrical extrudates based on a selection of different silicone types. Interestingly, these predictions could be confirmed by independent experiments. Hence, this type of quantitative analysis can replace time-consuming and cost-intensive series of trial-and-error experiments during product optimization. This is particularly helpful, if long-term drug release (e.g., during several weeks, months or years) is targeted.

Adiposity rebound is misclassified by BMI rebound.

BACKGROUND/OBJECTIVES: Adiposity rebound (AR) is defined as the nadir or the inflexion point of body mass index (BMI) percentiles between the age of 3 and 7 years. An early rebound is seen as a risk of obesity and, thus, AR is considered as a suitable time period for prevention. As BMI does not reflect body composition, we aimed to examine the rebounds of fat mass index (FMI) and fat-free mass index (FFMI) together with BMI. SUBJECTS/METHODS: Cross-sectional data of 19 264 children aged 3-11 years were pooled from three German studies (Kiel Obesity Prevention Study, the project 'Better diet. More exercise. KINDERLEICHT-REGIONS' and regular examinations of Jena children). Height and weight were measured. Fat mass (FM) and fat-free mass (FFM) were obtained from bioelectrical impedance analysis and analysed using a population-specific algorithm. Percentiles of BMI, FMI and FFMI were constructed by the LMS method. RESULTS: Both BMI and FMI percentiles showed a rebound, whereas FFMI percentiles steadily increased with age. On P90, FMI rebound was about 1.6-1.8 years later compared with that of BMI, that is, at ages 4.2 years (BMI) and 5.8 years (FMI) in boys and at 4.2 years (BMI) and 6.0 years (FMI) in girls. At AR, the slope of the BMI-P90 was explained by increases in FFMI rather than FMI. By contrast, at FMI rebound, the slope of BMI was strongly related to FMI. CONCLUSIONS: BMI rebound does not equal the rebound of FM. At AR, the slope in BMI is determined by the increase in FFMI. AR should be defined as FMI rebound rather than BMI rebound.

Predicting drug release from HPMC/lactose tablets.

Three mathematical models were applied to quantify drug release from HPMC/lactose-based matrix tablets loaded with varying amounts of theophylline: (i) a numerical model considering drug diffusion in axial and radial direction in cylinders as well as limited drug solubility effects, (ii) an analytical solution of Fick's second law of diffusion considering axial and radial mass transport in a cylinder, but neglecting limited drug solubility effects, and (iii) a simple early time approximation of the analytical solution, considering only radial mass transport and neglecting axial diffusion as well as limited drug solubility effects. The three models were fitted to experimentally determined drug release kinetics from various types of tablets in 0.1M HCl and phosphate buffer pH 7.4. Interestingly, the agreement between fitted theories and experimental data was similar in all cases. However, the determined system specific model parameters (apparent diffusion coefficients of theophylline in the polymeric matrices) were significantly biased when using simplified theories. Nevertheless, the reliability of theoretical predictions was similar for all three models, since the determined apparent diffusivities are partially "lumped" parameters. Thus, from a practical point of view, very simple equations can be used during product optimization, allowing estimating the effects of formulation parameters on drug release.

The effect of supportive E. coli mastitis treatment on PMN chemiluminescence and subpopulations of T lymphocytes.

The aim of this field study was to assess the impact of a single i.m. injection of lysozyme dimer and flunixin meglumine in combination with intramammary and systemic antibiotic on chemiluminescence of PMN (polymorphonuclear leucocytes) and subpopulations of lymphocyte T in blood of cows with E. coli mastitis. Examinations were performed on 30 dairy cows affected with naturally occurring acute form of E. coli mastitis. Cows were randomly divided into three groups according to the method of treatment. The first group was treated with approved intramammary antibiotic product, the same antibiotic in i.m. injection and one injection of flunixin meglumine on the first day of therapy. Next group was treated with the same antibiotic and additionally one injection of lysozyme dimer on the first day of therapy. The third one was treated only with an antibiotic and served as a control group. Blood samples were taken before treatment and on days 3 and 7. In samples haematology indices were determined, spontaneous and opsonised zymosan stimulated CL and PMA measurements were performed and the subpopulations of T lymphocyte (CD2(+), CD4(+), CD8(+)) were assayed in whole blood. There was no effect of the applied supportive treatment on the value of morphological blood indices. A significant influence of the time of sample collection on the level of CL and dynamics of lymphocytes T subpopulation was demonstrated. A single injection of flunixin meglumine or lysozyme dimer on the day of the beginning of treatment of E. coli mastitis, does not affect the level of neutrophil chemiluminescence and the percentage of T lymphocytes in the blood of mastitic cows in the analysed period of time.

[Longitudinal data of the Kiel Obesity Prevention Study (KOPS)]. / Längsschnittdaten der Kieler Adipositas-Präventionsstudie (KOPS).

The Kiel Obesity Prevention Study (KOPS) has been performed since 1996. Examinations were performed at age 6, 10 and 14 years. In addition, birth weight as well as height and weight at age 2 years were collected retrospectively. For the study 4,997, 1,671 and 748 children were recruited at baseline (at age 6 years) as well as at 4 and 8-year follow-up, respectively. In this paper we will analyze and discuss (i) the important time period for preventive measures, (ii) effect sizes needed for successful interventions and (iii) suitable approaches for preventive measures. The main results were: (i) at age 6 years persistence of overweight was 69% while at younger ages persistence was 21% only. Thus, school entry was shown to be an important period for preventive measures. (ii) Interventions have to reduce energy balance by 140 kcal/day to prevent overweight (e.g. a reduction of energy intake). (iii) Prevention programs should involve the family and focus on increasing physical activity.

Selected biochemical blood compounds in cows with abomasum displacement.

The aim of this study was to compare some blood biochemical indicators in cows with displacement of abomasum (DA) which recovered or died after treatment. Examinations were performed on 60 multiparous cows with left (L) or right (R) displacement and on 15 healthy herdmates. Diagnosis was made by experienced practitioners on the basis of clinical examination. Surgical treatment was undertaken during the first 24 hours after diagnosis. Almost all animals (55 = 91.5%) became sick in the post parturient period (21 days p.p. on average) with the exception of 5 (8.3%) that became sick later. Blood samples were taken from each cow immediately before surgical procedure. Serum nonesterified fatty acid (NEFA), glucose (Glu), cholesterol (Chol), aspartate aminotransferase (AST), total bilirubin (Bil) and blood urea nitrogen (BUN) were measured. Sick animals were characterized by low mean values of Chol (< or = 2 mmol/l) and normal level of BUN (12-15 mg/dl), higher levels of NEFA (> 600 micromol/l) and Bil (> 22 micromol/l), higher activity of AST (> 100 U/l). Seven cows (11.67%) died after surgical correction and all others recovered. No significant differences in NEFA, Chol, AST, Bil and BUN levels were observed as dependent on the efficacy of treatment (survival, deaths). It was found that cows which died after surgical treatment were characterized by significant higher levels of glucose (5.05 mmol/l) compared to surviving cows (2.93 mmol/l).

Long-acting methylphenidate has an effect on aggressive behavior in children with attention-deficit/hyperactivity disorder.

INTRODUCTION: Aggression is frequently observed in children and adolescents with attention-deficit/hyperactivity disorder (ADHD). The aim of this study was to assess the efficacy with regard to oppositional and aggressive behavior of a new long-acting methylphenidate preparation (Medikinet retard, MPH-MR), with equal portions of the immediate-release and the sustained-release active substance, and especially to look at correlations between either teacher or parent assessment of aggression and ADHD sub-symptomatology. METHODS: Eighty five children and adolescents (6-16 years) were investigated in a double-blind, randomized, clinical trial over 5 weeks under a treatment with MPH-MR using symptom checklists for ADHD, oppositional-defiant and conduct disorder according to the Diagnostic and Statistical Manual of Mental Disorders, 4th edition (DSM-IV). RESULTS: A total of 64.9% of the children showed oppositional defiant disorder/conduct disorder (ODD/CD) symptoms. A statistically significant effect was found in the group treated with MPH (verum-group). On the basis of Cohen's criteria, high effects were found for aggressive symptoms in school (d = 1.0), but not in the afternoon (d = 0.4). There were also lower effect sizes for more severe aggressive symptoms. We found characteristic correlations between ODD/CD symptoms and the ADHD subscale hyperactivity/impulsivity compared to the subscale inattention. CONCLUSIONS: Long-acting MPH is effective in the treatment of oppositional-defiant and aggressive behavior, especially concerning milder symptoms. The expected correlation between impulsivity and aggressiveness could be confirmed.

Influence of balance of protein and energy metabolism in rumen on hypomagnesaemia in cows.

This study was carried out in order to determine the influence of nitrogen excess in rumen and deficit of digestible organic matter in a feed ration for dairy cows on hypomagnesaemia at the beginning of green forage feeding. The balance of nitrogen and energy metabolism in rumen was determined using the rules of evaluation of the nutritional value of fodder (PDI system) according to the Institut National de la Recherche Agronomique (INRA), taking into consideration a potential protein synthesis from nitrogen by microorganisms in rumen (MPRN) and microbial protein synthesized from digestible organic matter (MP(RE)). It was found that an increase in the amount of rumen degradable protein (RDP) and MPRN-MPRE balance leads to a significant increase of urea nitrogen in serum (p < 0.05). An increase of total MPRN-MPRE balance in rumen results in a decrease of Mg concentration in serum. It has been demonstrated that in cows fed green forage at the end of spring (2nd half of May to 1st half of June) when the MPRN-MPRE balance is not higher than 187 g, adding 7 g or 25 g of MgO/cow/day to the same extent successfully prevents hypomagnesaemia.

High-throughput peptide synthesis and peptide purification strategy at the low micromol-scale using the 96-well format.

The increasing demand for short- and medium-sized peptides in many fields of biological, medical and pharmaceutical research requires optimized and universally applicable high-throughput synthesis and purification techniques at the low-micromol scale. Here, we describe a continuous peptide synthesis/purification approach using the 96-well format. First, a micromol scale peptide synthesis on resin beads was optimized on a novel miniaturized 96-reaction vessel block employing standard Fmoc/tBu-chemistry. Almost 90% of the synthesized peptides contained the target sequence as the main component, as judged from matrix-assisted laser desorption/ionization (MALDI) mass spectra. Impurities were mostly related to partially protected peptides. Second, we tested the applicability of ion pair reversed-phase solid-phase extraction (IP-RP-SPE) to purify individual peptides. Depending on the length and predicted hydrophobicity of the peptides, elution was performed with 25 or 35% aqueous acetonitrile in the presence of 0.1% trifluoroacetic acid (TFA). Thus, scavengers used during TFA cleavage and partially protected peptides carrying very hydrophobic protecting groups were effectively removed. Using a narrow step gradient, the target peptides were even separated from deleted sequences and protected peptides with similar hydrophobicities. Third, we combined the micromol-scale synthesis in the 96-well format with purification by IP-RP-SPE on a 96-well micro-extraction plate format. This simple, fast and parallel approach was tested on 12-mer and 15-mer peptides to map epitopes of T- and B-cell clones, respectively. Approximately 80% of all peptides were obtained at purities > 90% without purification by RP-HPLC. In summary, this novel approach has several advantages: (i) the micromol-scale reduced the cost of peptide synthesis, (ii) large numbers of peptides were purified faster, (iii) the volumes of eluents and waste were significantly reduced, and (iv) the RP-HPLC column was not contaminated with hydrophobic impurities.

A high efficiency all glass sampling and concentration device for adsorptive semivolatile organics.

A low-volume, inert sampling and enrichment device for semivolatile organic vapors is described. The device consists of two concentric fused silica capillaries. A small portion of the inter-capillary volume, cooled with a burst of compressed carbon dioxide, serves as a trap for the semivolatile organics. The low mass of the trap permits rapid sampling and desorption cycles suitable for applications requiring fast monitoring of semivolatile chemicals. The device is devoid of switching valves in the sampling train and consequently does not suffer from analyte loss due to irreversible adsorption or interference resulting from cross contamination. The device was successfully used for sampling low concentrations of highly adsorptive nitroaromatic compounds and is applicable for polychlorinated dibenzo-p-dioxins (PCDDs) and polyaromatic hydrocarbons (PAHs).

Application of electron-attachment reactions to enhance selectivity of electron-capture detector for nitroaromatic explosives.

The differences in the extent of electron-attachment reactions between thermal electrons and selected classes of organic molecules with high electron affinities were investigated. The investigations showed that interactions of thermal electrons with nitroaromatic compounds lead to the formation of neutral products with very low electron affinities. By contrast, a number of other analytes with high electron affinities such as polyhalogenated organic compounds, lead to products with high electron affinities. This difference was exploited to differentiate between nitroaromatic and polychlorinated organic compounds with a tandem arrangement consisting of two electron-capture detectors connected in series with an electron-attachment reactor.

Coping with hallucinations: exploring stress and coping framework.

This study explored application of the theoretical framework of Lazarus & Folkman (1984) to coping with hallucinations in schizophrenia. Eighty-one patients were interviewed with a structured schedule. Unprompted reports of coping indicated wide use of both 'hallucination-specific' and 'general' strategies. Factor analysis of a coping checklist produced three factors which were utilized in regression analyses. The active acceptance factor may relate to control of hallucinations, the passive coping factor predicted distress reduction but, surprisingly, the resistance coping factor, which contained hallucination-specific coping strategies, predicted poor distress reduction. Clinical and research implications are noted.

Effect of recombinant beet necrotic yellow vein virus with different RNA compositions on mechanically inoculated sugarbeets.

Beet necrotic yellow vein virus (BNYVV) inocula with different RNA compositions were prepared from infectious transcripts of RNAs 3 and 4 and the Rg 1 isolate, which has a genome consisting only of RNAs 1 and 2. The recombinant viruses were inoculated on 6- to 8-day-old sugarbeet seedlings by 'vortexing'. Inocula containing RNAs 1 and 2 or 1, 2 and 4 produced some growth reduction, but the most dramatic effects, with yield reductions of about 95% in a highly susceptible variety, were seen when RNA 3 was also present in the inoculum. Under these conditions the side roots were brown and brittle and often deteriorated, but 'root beardedness' was not observed. This might be due to the fact that our experiments were done in the absence of Polymyxa betae. Alternatively, the heavy inoculation at a very young age may either have weakened the plants to such an extent that extensive root proliferation was impaired or it may have led to rapid deterioration of the proliferating rootlets, which would therefore be lost prior to or during removal of the tap roots from the soil. In the presence of RNA 3 the virus concentrations in tap roots were markedly increased suggesting that this RNA facilitates the multiplication and/or spread of the virus in root tissues.

Complexation and medium effects on the conformation of cyclosporin A studied by NMR spectroscopy and molecular dynamics calculations.

The conformation of cyclosporin A was investigated in different environments. Whereas there was no distinct difference between the structure of CSA in CDCl3 and THF, the addition of LiCl caused a conformational change. Also, using polar solvents (DMSO, water) induced conformational changes in the backbone as well as the side chains.