Good servant, bad master: sulfide influence on partial nitritation of sewage.

When applying partial nitritation (PN) to anaerobically pre-treated sewage, ammonium oxidizing bacteria (AOB) and nitrite oxidizing bacteria (NOB) will be exposed to dissolved sulfide and methane. Both sulfide and methane may inhibit nitrification. To gain knowledge necessary for sustaining PN under these conditions, we exposed an AOB enrichment and a mixed nitrifying culture to dissolved sulfide and methane. In the mixed nitrifying culture, sulfide selectively inhibited NOB activity (KI,AOB1 = 150 mg-S L-1, KI,NOB = 10 mg-S L-1) which shows that sulfide may help establish PN. The AOB enrichment showed similar KI,AOB2 (130 mg-S L-1), but nitritation activity lagged longer than the time necessary to remove sulfide from the liquid. This demonstrates that feeding of sulfide into established PN should be avoided. Methane inhibition of AOB enrichment was assessed in batch assays with 10 mg-CH4 L-1. As compared to control without methane, AOB enrichment activity was identical. Up to 51% of methane was converted to methanol, thus reducing the greenhouse gas emissions.

Immediate effects of whole body vibration on patellar tendon properties and knee extension torque.

PURPOSE: Reports about the immediate effects of whole body vibration (WBV) exposure upon torque production capacity are inconsistent. However, the changes in the torque-angle relationship observed by some authors after WBV may hinder the measurement of torque changes at a given angle. Acute changes in tendon mechanical properties do occur after certain types of exercise but this hypothesis has never been tested after a bout of WBV. The purpose of the present study was to investigate whether tendon compliance is altered immediately after WBV, effectively shifting the optimal angle of peak torque towards longer muscle length. METHODS: Twenty-eight subjects were randomly assigned to either a WBV (n = 14) or a squatting control group (n = 14). Patellar tendon CSA, stiffness and Young's modulus and knee extension torque-angle relationship were measured using ultrasonography and dynamometry 1 day before and directly after the intervention. Tendon CSA was additionally measured 24 h after the intervention to check for possible delayed onset of swelling. RESULTS: The vibration intervention had no effects on patellar tendon CSA, stiffness and Young's modulus or the torque-angle relationship. Peak torque was produced at ~70° knee angle in both groups at pre- and post-test. Additionally, the knee extension torque globally remained unaffected with the exception of a small (-6%) reduction in isometric torque at a joint angle of 60°. CONCLUSION: The present results indicate that a single bout of vibration exposure does not substantially alter patellar tendon properties or the torque-angle relationship of knee extensors.

Whole-body vibration training induces hypertrophy of the human patellar tendon.

Animal studies suggest that regular exposure to whole-body vibration (WBV) induces an anabolic response in bone and tendon. However, the effects of this type of intervention on human tendon properties and its influence on the muscle-tendon unit function have never been investigated. The aim of this study was to investigate the effect of WBV training on the patellar tendon mechanical, material and morphological properties, the quadriceps muscle architecture and the knee extension torque-angle relationship. Fifty-five subjects were randomized into either a vibration, an active control, or an inactive control group. The active control subjects performed isometric squats on a vibration platform without vibration. Muscle and tendon properties were measured using ultrasonography and dynamometry. Vibration training induced an increase in proximal (6.3%) and mean (3.8%) tendon cross-sectional area, without any appreciable change in tendon stiffness and modulus or in muscle architectural parameters. Isometric torque at a knee angle of 90° increased in active controls (6.7%) only and the torque-angle relation remained globally unchanged in all groups. The present protocol did not appreciably alter knee extension torque production or the musculo-tendinous parameters underpinning this function. Nonetheless, this study shows for the first time that WBV elicits tendon hypertrophy in humans.

Alpine Skiing With total knee ArthroPlasty (ASWAP): muscular adaptations.

This study investigated the effectiveness of recreational skiing as an intervention to improve quadriceps muscle architecture, strength, and antagonistic co-activation in patients with unilateral total knee arthroplasty (TKA). Hence, patients with TKA were assigned to either an intervention group (IG) or control group (CG). The IG completed a 12-week guided skiing program whereas the CG was instructed not to change their daily routines for the same period and was not allowed to ski. Before, after the intervention/after an 8-week retention period m. rectus femoris (RF) cross-sectional area (CSA), m. vastus lateralis muscle thickness, fascicle length, and pennation angle were measured with ultrasonography, while isometric (90° knee angle) knee extension, flexion torque and m. biceps femoris co-activation were assessed on an isokinetic dynamometer in 26 patients. There were significant and stable increases in RF CSA for the operated (10%; P < 0.05) and non-operated leg (12%; P < 0.01) after the training period in the IG whereas no changes were observed for the CG (all P > 0.05). There were no significant effects for other parameters (all P > 0.05). Overall, the skiing intervention was successful in increasing muscle mass in TKA older patients.

Alpine Skiing With total knee ArthroPlasty (ASWAP): effect on tendon properties.

The aim of this study was to investigate the effect of alpine skiing on patellar tendon properties in patients with total knee arthroplasty (TKA). Thirty-one adults (70.4 ± 4.7 years) with unilateral TKA were recruited 2.7 ± 0.9 years after surgery and assigned to an intervention (IG) or a control group (CG). The IG underwent a 12-week guided skiing program. Tendon stiffness, Young's modulus, and cross-sectional area (CSA) were measured before and after the intervention. In both groups, mean tendon CSA was 28% (P < 0.001) larger in the operated (OP) than in the non-operated (NOP) leg at baseline, without any difference in other tendon properties. After training, stiffness increased in the IG by 5.8% and 15.8%, respectively, in the OP and NOP legs. Likewise, mean CSA increased in the IG by 2.9% in the OP and 3.8% in the NOP leg, whereas no significant changes were found for the Young's modulus. None of the tendon parameters changed in the CG. Results indicate that patellar tendon structure and/or loading pattern are altered following TKA, but this tissue seems to retain its adaptation capacity. Further, alpine skiing appears to offer a suitable rehabilitation strategy for TKA patients.

Influence of loading rate on patellar tendon mechanical properties in vivo.

BACKGROUND: Rate-dependent properties of tendons have consistently been observed in vitro but in vivo studies comparing the effects of loading duration on this feature remain conflicting. The main purpose of the present study was to evaluate whether tendon loading rate per se would affect in vivo tendon mechanical properties. METHODS: Twenty-two physically active male subjects were recruited. Patellar tendon deformation was recorded with ultrasonography under voluntary isometric contractions at rates of 50, 80 and 110Nm/s, controlled via visual feedback. FINDINGS: Subjects were able to accurately generate all three loading rates (Accuracy=2% to 15%), with a greater steadiness at 50 (CV=12.4%) and 110Nm/s (CV=13.1%) than at 80Nm/s (CV=22.9%). Loading rate did not appreciably affect strain or stress. However, stiffness (ɳp(2)=0.555) and Youngs's Modulus (ɳp(2)=0.670) were significantly higher at 80Nm/s (21.4% and 21.6%, respectively) and at 110Nm/s (32.5% and 32.0%, respectively) than at 50Nm/s. Similarly, stiffness and Young's modulus were 9.9% and 8.8% higher, respectively, at 110Nm/s than at 80Nm/s. INTERPRETATION: These results indicate that in vivo measurements of patellar tendon mechanics are influenced by loading rate. Moreover, they bear important methodological implications for in vivo assessment of mechanical properties of this tendon and possibly other human tendons.

[Absolute and relative strength-endurance of the knee flexor and extensor muscles: a reliability study using the IsoMed 2000-dynamometer]. / Absolute und relative Kraftausdauer der Kniebeuge- und -streckmuskulatur: eine Reliabilitätsstudie unter Verwendung des IsoMed 2000-Dynamometers.

BACKGROUND: Isokinetic devices are highly rated in strength-related performance diagnosis. A few years ago, the broad variety of existing products was extended by the IsoMed 2000-dynamometer. In order for an isokinetic device to be clinically useful, the reliability of specific applications must be established. Although there have already been single studies on this topic for the IsoMed 2000 concerning maximum strength measurements, there has been no study regarding the assessment of strength-endurance so far. The aim of the present study was to establish the reliability for various methods of quantification of strength-endurance using the IsoMed 2000. METHODS: A sample of 33 healthy young subjects (age: 23.8 ± 2.6 years) participated in one familiarisation and two testing sessions, 3-4 days apart. Testing consisted of a series 30 full effort concentric extension-flexion cycles of the right knee muscles at an angular velocity of 180 °/s. Based on the parameters Peak, Torque and Work for each repetition, indices of absolute (KADabs) and relative (KADrel) strength-endurance were derived. KADabs was calculated as the mean value of all testing repetitions, KADrel was determined in two ways: on the one hand, as the percentage decrease between the first and the last 5 repetitions (KADrelA) and on the other, as the negative slope derived from the linear regression equitation of all repetitions (KADrelB). Detection of systematic errors was performed using paired sample t-tests, relative and absolute reliability were examined using intraclass correlation coefficient (ICC 2.1) and standard error of measurement (SEM%), respectively. RESULTS/CONCLUSION: In general, for extension measurements concerning KADabs and - in an weakened form - KADrel high ICC -values of 0.76-0.89 combined with clinically acceptable values of SEM% of 1.2-5.9 % could be found. For flexion measurements this only applies to KADabs, whereas results for KADrel turned out to be clearly weaker with ICC- and SEM% values of 0.42-0.62 and 9.6-17.7 % and leave considerable doubts on the clinical usefulness. However, if there should be after all a need to measure KADrel for flexion, it is - in view of the stronger reliability results - recommended (i) to concentrate on the calculation of KADrelB, (ii) to use the parameter Work and - in view of considerable familiariszation and learning effects of ≈10 % - (iii) to include a familiarisation period that extends exceeds the familiarisation session conducted in the present study.

Sagopilone (ZK-EPO, ZK 219477) for recurrent glioblastoma. A phase II multicenter trial by the European Organisation for Research and Treatment of Cancer (EORTC) Brain Tumor Group.

BACKGROUND: Sagopilone (ZK 219477), a lipophylic and synthetic analog of epothilone B, that crosses the blood-brain barrier has demonstrated preclinical activity in glioma models. PATIENTS AND METHODS: Patients with first recurrence/progression of glioblastoma were eligible for this early phase II and pharmacokinetic study exploring single-agent sagopilone (16 mg/m(2) over 3 h every 21 days). Primary end point was a composite of either tumor response or being alive and progression free at 6 months. Overall survival, toxicity and safety and pharmacokinetics were secondary end points. RESULTS: Thirty-eight (evaluable 37) patients were included. Treatment was well tolerated, and neuropathy occurred in 46% patients [mild (grade 1) : 32%]. No objective responses were seen. The progression-free survival (PFS) rate at 6 months was 6.7% [95% confidence interval (CI) 1.3-18.7], the median PFS was just over 6 weeks, and the median overall survival was 7.6 months (95% CI 5.3-12.3), with a 1-year survival rate of 31.6% (95% CI 17.7-46.4). Maximum plasma concentrations were reached at the end of the 3-h infusion, with rapid declines within 30 min after termination. CONCLUSIONS: No evidence of relevant clinical antitumor activity against recurrent glioblastoma could be detected. Sagopilone was well tolerated, and moderate-to-severe peripheral neuropathy was observed in despite prolonged administration.

Phase I study of the novel, fully synthetic epothilone sagopilone (ZK-EPO) in patients with solid tumors.

BACKGROUND: Sagopilone (ZK-EPO) is a fully synthetic microtubule-stabilizing agent that has demonstrated high antitumor activity in preclinical models. This first-in-human phase I study aimed to determine the maximum tolerated dose (MTD) and dose-limiting toxic effects (DLTs) of 3-weekly sagopilone treatment. PATIENTS AND METHODS: A total of 52 patients with advanced solid tumors received a 30-min infusion of escalating doses of sagopilone (0.6-29.4 mg/m(2)) every 3 weeks. Nine additional patients were recruited to a 3-h infusion arm (16.53- or 22.0-mg/m(2) dose) to assess the incidence of neuropathy with prolonged infusion. RESULTS: The MTD was established as 22.0 mg/m(2). DLTs comprised peripheral sensory neuropathy (PNP), infection, hyponatremia, diarrhea, and central ataxia. PNP was the most common grade 3 event, with a similar incidence in the 30-min and 3-h arms. Hematologic adverse events were rare and of low intensity. One confirmed partial response (PR) and one unconfirmed PR were reported in the 30-min arm, and a further unconfirmed PR was observed in the 3-h arm. Eleven patients achieved disease stabilization. Sagopilone showed high levels of tissue binding and no obvious serum accumulation in both arms. CONCLUSIONS: These data demonstrate that sagopilone therapy is feasible and well tolerated. The recommended dose for phase II studies is 16.53 mg/m(2), once every 3 weeks.

Weekly administration of sagopilone (ZK-EPO), a fully synthetic epothilone, in patients with refractory solid tumours: results of a phase I trial.

BACKGROUND: Epothilones are a novel class of microtubule-stabilising agents, and sagopilone is a fully synthetic epothilone that has shown marked in vivo and in vitro preclinical activity. METHODS: This phase I, open-label study investigated the maximum tolerated dose (MTD) and dose-limiting toxicities (DLTs) of weekly sagopilone. Twenty-three patients with malignancy resistant or refractory to standard treatment were enrolled into this study evaluating sagopilone doses from 0.6 to 7.0 mg m(-2). RESULTS: The incidence of drug-related haematological adverse events (AEs) was low, with two grade 3 events observed. Nonhaematological AEs were generally mild and reversible; increased gamma-GT was the only grade 4 event and grade 3 events comprised peripheral neuropathy (n=2), diarrhoea (n=1) and fatigue (n=1). Two grade 3 events were DLTs (diarrhoea and peripheral neuropathy at 7.0 mg m(-2)). The MTD of weekly sagopilone was therefore established as 5.3 mg m(-2). Stable disease was the best overall response (n=3). Microtubule bundle formation in peripheral blood mononuclear cells increased post-treatment, peaking after 1 h. Sagopilone disposition was similar across treatment courses and showed rapidly decreasing serum concentrations after infusion end and a long terminal disposition phase with no obvious accumulation in the serum, probably reflecting a fast uptake into tissues followed by a slow release. CONCLUSION: Weekly administration of sagopilone could represent an alternative to the 3-weekly administration currently evaluated in phase II trials.

Deep endometriosis, including intestinal involvement--the interdisciplinary approach.

Deep endometriosis is a disease which may involve all organs of the pelvis. The lesion is most often located at the backside of the uterus, involving the uterosacral ligaments and/or the rectovaginal septum. The involvement of adjacent organs, e.g. bowel, ureter, and bladder, makes an interdisciplinary approach necessary. There is a correlation between the radicalness of endometriosis resection and the postoperative improvement of complaints. In a series of 202 patients with deep endometriosis including the bowel we performed a segmental resection with anterior anastomosis including radical excision of all endometriotic lesions. The follow-up of 142 patients shows a significant improvement of pelvic pain (96%), dyschezia (88%), and dyspareunia (87%). Of 95 patients with a desire for children, 50% became pregnant. The postoperative complication rate was low. A leakage of anastomosis was seen in six cases (3%).

[Laparoscopic therapy of intestinal endometriosis and the ranking of drug treatment]. / Die laparoskopische Therapie der Darmendometriose und der Stellenwert der medikamentösen Therapie.

UNLABELLED: Patients with severe endometriosis involving the rectovaginal septum and bowel makes surgery necessary. The laparoscopic approach offers the possibility to perform the complete resection with minimal invasive techniques. Small lesions can be removed by full-thickness-resection. Large nodules make a segmental resection necessary. Deep lesions are resected with stapling devices. Nodules in the sigmoid ore removed by laparoscopic assisted hand-sewn anastomosis. RESULTS: Between 3/96 and 7/03 142 patients with severe endometriosis involving the bowel have been treated laparoscopically. The pre-operative complains have been reduced as following: dyschezio: 88%, dyspareunia 87%, chronic pelvic pain 96%, disturbance of sexuality 75%. The complication rate was minimal: leakage of the anastomosis n=4 (2.8%), paraproctial abscess 2 (1.4%), blood transfusion 1 (0,7 %), severe stenosis of the anastomosis 6 (4.2%). CONCLUSION: Laparoscopic approach is a safe and effective technique to treat deep infiltrating endometriosis with bowel involvement. The hormonal treatment of bowel endometriosis is used in patients with minimal symptoms. The postoperative treatment becomes necessary in incomplete operations or in patients with severe adenomyosis and infertility.

Neuronal and glial coexpression of argininosuccinate synthetase and inducible nitric oxide synthase in Alzheimer disease.

The enzyme argininosuccinate synthetase (ASS) is the rate limiting enzyme in the metabolic pathway leading from L-citrulline to L-arginine, the physiological substrate of all isoforms of nitric oxide synthases (NOS). ASS and inducible NOS (iNOS) expression in neurons and glia was investigated by immunohistochemistry in brains of Alzheimer disease (AD) patients and nondemented, age-matched controls. In 3 areas examined (hippocampus, frontal, and entorhinal cortex), a marked increase in neuronal ASS and iNOS expression was observed in AD brains. GFAP-positive astrocytes expressing ASS were not increased in AD brains versus controls, whereas the number of iNOS expressing GFAP-positive astrocytes was significantly higher in AD brains. Density measurements revealed that ASS expression levels were significantly higher in glial cells of AD brains. Colocalization of ASS and iNOS immunoreactivity was detectable in neurons and glia. Occasionally, both ASS-and iNOS expression was detectable in CD 68-positive activated microglia cells in close proximity to senile plaques. These results suggest that neurons and astrocytes express ASS in human brain constitutively, whereas neuronal and glial ASS expression increases parallel to iNOS expression in AD. Because an adequate supply of L-arginine is indispensable for prolonged NO generation, coinduction of ASS enables cells to sustain NO generation during AD by replenishing necessary supply of L-arginine.

Arginine metabolism and the synthesis of nitric oxide in the nervous system.

The biochemistry and physiology of L-arginine have to be reconsidered in the light of the recent discovery that the amino acid is the only substrate of all isoforms of nitric oxide synthase (NOS). Generation of nitric oxide, NO, a versatile molecule in signaling processes and unspecific immune defense, is intertwined with synthesis, catabolism and transport of arginine which thus ultimately participates in the regulation of a fine-tuned balance between normal and pathophysiological consequences of NO production. The complex composition of the brain at the cellular level is reflected in a complex differential distribution of the enzymes of arginine metabolism. Argininosuccinate synthetase (ASS) and argininosuccinate lyase which together can recycle the NOS coproduct L-citrulline to L-arginine are expressed constitutively in neurons, but hardly colocalize with each other or with NOS in the same neuron. Therefore, trafficking of citrulline and arginine between neurons necessitates transport capacities in these cells which are fulfilled by well-described carriers for cationic and neutral amino acids. The mechanism of intercellular exchange of argininosuccinate, a prerequisite also for its proposed function as a neuromodulator, remains to be elucidated. In cultured astrocytes transcription and protein expression of arginine transport system y(+) and of ASS are upregulated concomittantly with immunostimulant-mediated induction of NOS-2. In vivo ASS-immunoreactivity was found in microglial cells in a rat model of brain inflammation and in neurons and glial cells in the brains of Alzheimer patients. Any attempt to estimate the contributions of arginine transport and synthesis to substrate supply for NOS has to consider competition for arginine between NOS and arginase, the latter enzyme being expressed as mitochondrial isoform II in nervous tissue. Generation of NOS inhibitors agmatine and methylarginines is documented for the nervous system. Suboptimal supply of NOS with arginine leads to production of detrimental peroxynitrite which may result in neuronal cell death. Data have been gathered recently which point to a particular role of astrocytes in neural arginine metabolism. Arginine appears to be accumulated in astroglial cells and can be released after stimulation with a variety of signals. It is proposed that an intercellular citrulline-NO cycle is operating in brain with astrocytes storing arginine for the benefit of neighbouring cells in need of the amino acid for a proper synthesis of NO.

Metabolism of a 20-methyl substituted series of vitamin D analogs by cultured human cells: apparent reduction of 23-hydroxylation of the side chain by the 20-methyl group.

We describe here for the first time the effect of introducing a 20-methyl group on the side-chain metabolism of the vitamin D molecule. Using a series of 20-methyl-derivatives of 1alpha,25-(OH)2D3 incubated with two different cultured human cell lines, HPK1A-ras and HepG2, previously shown to metabolize vitamin D compounds, we obtained a series of metabolic products that were identified by comparison to chemically synthesized standards on HPLC and GC-MS. 24-Hydroxylated-, 24-oxo-hydroxylated-, and 24-oxo-23-hydroxylated products of 20-methyl-1alpha,25-(OH)2D3 were observed, but the efficiency of 23-hydroxylation was low as compared with that of the natural hormone and, in contrast to 1alpha,25-(OH)2D3, no truncated 23-alcohol was formed from the 20-methyl analog. These data, taken together with results from other analogs with changes in the vicinity of the C17-C20 positions, lead us to speculate that such changes must alter the accessibility of the C-23 position to the cytochrome P450 involved. Using the HepG2 cell line, we found evidence that the 24S-hydroxylated product of 20-methyl-1alpha,25-(OH)2D3 predominates, implying that the liver cytochrome involved in metabolism is a different isoform. Studies with a more metabolically resistant analog of the series, 20-methyl-Delta(23)-1alpha,25-(OH)2D3, gave the expected block in 23- and 24-hydroxylation, and evidence of an alternative pathway, namely 26-hydroxylation. 20-Methyl-Delta(23)-1alpha,25-(OH)2D3 was also more potent in biological assays, and the metabolic studies reported here help us to suggest explanations for this increased potency. We conclude that the 20-methyl series of vitamin D analogs offers new perspectives into vitamin D analog action, as well as insights into the substrate preferences of the cytochrome(s) P450 involved in vitamin D catabolism.

Synthesis and biological activities of a new series of secosteroids: vitamin D phosphonate hybrids.

By a structural combination of phosphonate and bisphosphonate moieties with the vitamin D skeleton a series of new vitamin D analogs was synthesized. Derivatives with 24beta-hydroxy- or 24-keto groups exerted considerable vitamin D activities in vitro while the hypercalcemic potentials were significantly reduced as compared to 1alpha,25-dihydroxyvitamin D(3) (calcitriol). Whereas the 24-hydroxy analogs did not influence bone formation in vivo in dosages below the hypercalcemic threshold, the 24-ketones were found to induce synthesis of new bone matrix in non-hypercalcemic doses. Vitamin D bisphosphonate hybrids, on the other hand, which did not elicit substantial vitamin D activities in vitro and tend to decrease serum calcium levels in vivo clearly induced osteoid formation in rats, indicating a mechanism of action different to calcitriol.

Acquisition of blood--tissue barrier--supporting features by hepatic stellate cells and astrocytes of myofibroblastic phenotype. Inverse dynamics of metallothionein and glial fibrillary acidic protein expression.

A number of similarities between astrocytes and hepatic stellate cells (HSC) rose the question whether or not the protective barrier features of blood-tissue interface may be provided by HSC as well. To test this hypothesis, we investigated the presence of metallothionein (MT), a functional marker of blood--brain barrier, in HSC in situ and in cell culture and compared the results with those obtained with astrocytes. The dynamics of MT expression in cultured astrocytes and HSC was investigated by simultaneous labelling of the cells with a monoclonal antibody (MAb MT) against a lysine-containing epitope of the cadmium-induced monomer of MT-I from rat liver and antiserum against glial fibrillary acidic protein (GFAP). Cell activation was estimated by the presence of smooth muscle alpha-actin (SMAA). In immunoblotting, MAb MT recognized monomeric MT protein and proteins in the 30-kDa range; both bands were pronounced in brain and barely visible in liver homogenates. In situ, MAb MT reacted with very few perivascular cells situated in the parenchyma of the liver. Double immunolabelling of brain slices with MAb MT and antiserum against GFAP showed large areas of brain containing cells expressing both MT and GFAP. However, there were also regions in the brain where the cells produced solely GFAP or MT. In liver cell culture, MT was absent from HSC and hepatocytes in early periods of cultivation, during which the cells maintained their original features; however, MT was expressed strongly in HSC during their activation under prolonged culture conditions. Inversely, in astrocytes MT was expressed during early culturing and disappeared from the cells together with SMAA in late culture when GFAP was upregulated. These results suggest that the acquisition of myofibroblastic features by perivascular cells empowers them to establish a protective blood-tissue permeability barrier. In addition, this study shows that, at least in cell culture, an enrichment of perivascular cells in GFAP results in the disappearance of protective functions.

Anti-inflammatory antioxidants attenuate the expression of inducible nitric oxide synthase mediated by advanced glycation endproducts in murine microglia.

Advanced glycation endproducts (AGEs) accumulate on long-lived protein deposits including beta-amyloid plaques in Alzheimer's disease (AD). AGE-modified amyloid deposits contain oxidized and nitrated proteins as markers of a chronic neuroinflammatory condition and are surrounded by activated microglial and astroglial cells. We show in this study that AGEs increase nitric oxide production by induction of the inducible nitric oxide synthase (iNOS) on the mRNA and protein level in the murine microglial cell line N-11. Membrane permeable antioxidants including oestrogen derivatives (e.g. 17beta-oestradiol) thiol antioxidants (e.g. (R+)-alpha-lipoic acid) and Gingko biloba extract EGb 761, but not phosphodiesterase inhibitors such as propentophylline, prevent the up-regulation of AGE-induced iNOS expression and NO production. These results indicate that oxygen free radicals serve as second messengers in AGE-induced pro-inflammatory signal transduction pathways. As this pharmacological mechanism is not only relevant for Alzheimer's disease, but also for many chronic inflammatory conditions, such membrane-permeable antioxidants could be regarded not only as antioxidant, but also as potent therapeutic anti-inflammatory drugs.

Transport of L-citrulline in neural cell cultures.

Uptake of L-[(14)C]citrulline was studied in cell culture models of the main neural cell populations, in astroglia-rich primary cultures derived from neonatal rat brain, in rat glioma cells C6-BU-1, in cells of the murine microglial clone N11 and in the glioma x neuroblastoma hybrid cell line 108CC15 with neuronal properties. For comparison, cells of the peripheral macrophage cell line RAW 264.7 were also investigated. A saturable component of uptake was found in all cases with K(M) values between 0.4 and 3.4 mM and V(max) values between 15 and 35 nmol.min(-1).(mg protein)(-1). A nonsaturable component dominated uptake at high concentrations of extracellular citrulline. Rates of uptake of L-citrulline were not affected when Na(+) or Cl(-) were omitted from the incubation medium or in the presence of depolarizing concentrations of K(+). Saturable uptake of citrulline was strongly inhibited by an excess of histidine or beta-2-aminobicyclo-(2.2.1)-heptane-2-carboxylic acid; excess amounts of arginine, creatine, glutamate, cysteic acid or N-methyl-alpha-aminoisobutyric acid did not reduce citrulline uptake. Preincubation of the cells with bacterial lipopolysaccharide and interferon gamma did not stimulate transport of citrulline. The results suggest that at physiological concentrations citrulline is taken up by neural cells with the help of transport system L for large neutral amino acids. Therefore, in the brain, effective utilization of extracellular citrulline as part of an intercellular trafficking of intermediates of an NO/citrulline cycle depends on the concentrations of all neutral amino acids present.

New vitamin D receptor agonists with decreased metabolic stability.

The aim of the study was the development of vitamin D receptor agonists with decreased metabolic stability for the topical treatment of psoriasis and related hyperproliferative skin diseases. Calcitriol analogues 1, 2, 3, all of which contain modifications in the side chain, were synthesized. The obtained analogues were full agonists when the induction of CD14 expression in HL-60 cells, the induction of 5-lipoxygenase activity in Mono Mac 6 cells, and the inhibition of phytohemagglutinin (PHA)-stimulated lymphocyte proliferation were studied. The EC(50) value of the most active compound 1 was 1.2 nM in the CD14 assay and 1 nM in the 5-lipoxygenase assay, whereas calcitriol gave EC(50) values in these assays of 3.7 and 9 nM, respectively. In the lymphocyte proliferation assay, compound 1 and calcitriol had IC(50) values of 0.3 and 2.8 nM, respectively. All three compounds had receptor binding affinities similar to that of calcitriol. The compounds showed a decreased metabolic stability in rat liver homogenates and had a 50-fold lower affinity for the vitamin D-binding protein than calcitriol, which suggests that calcitriol analogues are metabolized more rapidly after systemic uptake or application. When injected into rats, the analogues displayed an approximately 100-fold lower hypercalcemic effect than calcitriol. In summary, our study presents three new and potent vitamin D receptor agonists with interesting profiles for development as antipsoriatic drugs.