Expression and activity profiling of the steroidogenic enzymes of glucocorticoid biosynthesis and the fdx1 co-factors in zebrafish.

The spatial and temporal expression of steroidogenic genes in zebrafish has not been fully characterised. Because zebrafish are increasingly employed in endocrine and stress research, a better characterisation of steroidogenic pathways is required to target specific steps in the biosynthetic pathways. In the present study, we have systematically defined the temporal and spatial expression of steroidogenic enzymes involved in glucocorticoid biosynthesis (cyp21a2, cyp11c1, cyp11a1, cyp11a2, cyp17a1, cyp17a2, hsd3b1, hsd3b2), as well as the mitochondrial electron-providing ferredoxin co-factors (fdx1, fdx1b), during zebrafish development. Our studies showed an early expression of all these genes during embryogenesis. In larvae, expression of cyp11a2, cyp11c1, cyp17a2, cyp21a2, hsd3b1 and fdx1b can be detected in the interrenal gland, which is the zebrafish counterpart of the mammalian adrenal gland, whereas the fdx1 transcript is mainly found in the digestive system. Gene expression studies using quantitative reverse transcriptase-PCR and whole-mount in situ hybridisation in the adult zebrafish brain revealed a wide expression of these genes throughout the encephalon, including neurogenic regions. Using ultra-high-performance liquid chromatography tandem mass spectrometry, we were able to demonstrate the presence of the glucocorticoid cortisol in the adult zebrafish brain. Moreover, we demonstrate de novo biosynthesis of cortisol and the neurosteroid tetrahydrodeoxycorticosterone in the adult zebrafish brain from radiolabelled pregnenolone. Taken together, the present study comprises a comprehensive characterisation of the steroidogenic genes and the fdx co-factors facilitating glucocorticoid biosynthesis in zebrafish. Furthermore, we provide additional evidence of de novo neurosteroid biosynthesising in the brain of adult zebrafish facilitated by enzymes involved in glucocorticoid biosynthesis. Our study provides a valuable source for establishing the zebrafish as a translational model with respect to understanding the roles of the genes for glucocorticoid biosynthesis and fdx co-factors during embryonic development and stress, as well as in brain homeostasis and function.

Versatile attosecond beamline in a two-foci configuration for simultaneous time-resolved measurements.

We present our attoline which is a versatile attosecond beamline at the Ultrafast Laser Physics Group at ETH Zurich for attosecond spectroscopy in a variety of targets. High-harmonic generation (HHG) in noble gases with an infrared (IR) driving field is employed to generate pulses in the extreme ultraviolet (XUV) spectral regime for XUV-IR cross-correlation measurements. The IR pulse driving the HHG and the pulse involved in the measurements are used in a non-collinear set-up that gives independent access to the different beams. Single attosecond pulses are generated with the polarization gating technique and temporally characterized with attosecond streaking. This attoline contains two target chambers that can be operated simultaneously. A toroidal mirror relay-images the focus from the first chamber into the second one. In the first interaction region a dedicated double-target allows for a simple change between photoelectron/photoion measurements with a time-of-flight spectrometer and transient absorption experiments. Any end station can occupy the second interaction chamber. A surface analysis chamber containing a hemispherical electron analyzer was employed to demonstrate successful operation. Simultaneous RABBITT measurements in two argon jets were recorded for this purpose.

Probing nonadiabatic effects in strong-field tunnel ionization.

We investigate experimentally the validity of proposed theories extending the tunneling approximation towards the multiphoton regime in strong-field ionization of helium. We employ elliptically polarized laser pulses and demonstrate how the influence of the ion potential on the released electron encoded in the measured observable provides the desired sensitivity to detect nonadiabatic effects in tunnel ionization. Our results show that for a large intensity range the proposed nonadiabatic theories contradict the experimental trends of the data, while adiabatic assumptions are confirmed.

Spatial fingerprint of quantum path interferences in high order harmonic generation.

We have spatially and spectrally resolved the high order harmonic emission from an argon gas target. Under proper phase matching conditions we were able to observe for the first time the spatial fine structure originating from the interference of the two shortest quantum paths in the harmonic beam. The structure can be explained by the intensity-dependent harmonic phase of the contributions from the two paths. The spatially and spectrally resolved measurements are consistent with previous spatially integrated results. Our measurement method represents a new tool to clearly distinguish between different interference effects and to potentially observe higher order trajectories in the future with improved detection sensitivity. Here, we demonstrate additional experimental evidence that the observed interference pattern is only due to quantum-path interferences and cannot be explained by a phase modulation effect. Our experimental results are fully supported by simulations using the strong field approximation and including propagation.

Intravitreal VEGF levels in uveitis patients and treatment of uveitic macular oedema with intravitreal bevacizumab.

PURPOSE: Clinical data suggest a role for VEGF in uveitic cystoid macular oedema (CME), even though the data on intravitreal VEGF levels in these eyes is still inconclusive. We determined intravitreal VEGF levels and treated uveitis patients with intravitreal bevacizumab. METHODS: Intravitreal VEGF levels were measured in eight uveitis patients and 10 controls using cytometric bead array technology. In 11 eyes of a second group of uveitis patients, CME was treated using 1.25 mg bevacizumab intravitreally. Re-injections of bevacizumab were given in patients showing a transient positive effect, defined as an increase of the best-corrected vision of at least two lines on a snellen chart. Alternatively, triamcinolone was given in patients, not responding to bevacizumab. RESULTS: Mean intravitreal VEGF concentration was 82.75+/-171.71 pg/ml (+/-SD) (range, 0.0-502.1 pg/ml), and below the detection levels in controls. A significant reduction of retinal thickness was seen at weeks 2 (P=0.001) and 4 (P=0.007). A significant improvement in VA was seen at week 2 (P=0.02). Patients presenting with a CME in baseline fluorescein-angiogram responded well towards bevacizumab treatment, unless an extensive leakage from the choroid or a leakage of the optic disk was detectable. In these patients, only intravitreally administered triamcinolone led to a reduction of the CME. CONCLUSIONS: Our data suggest that patients presenting with a diffuse leakage from the choroid in the fluorescein angiogram or an extensive leakage of the optic disk should be treated with intravitreal triamcinolone, whereas in patients presenting only a cystoid macular oedema bevacizumab treatment seems like a good choice.

Adverse skin reactions to infliximab in the treatment of intraocular inflammation.

TNFalpha inhibitors are more widely used in the treatment of intraocular inflammation, thus ophthalmologists should become aware of possible adverse events, associated with this form of treatment. Herein we report two cases of cutaneous adverse events in uveitis patients treated with infliximab. In one patient, the primary outbreak of pustular psoriasis was observed after her third infusion. A second patient developed impetigo contagiosa induced by Staphylococcus aureus. Thus patients undergoing treatment with infliximab should be monitored carefully since dermal infections and pustular psoriasis, which may be triggered by streptococcal infection, may occur under this regime.

[Gaze-evoked amaurosis fugax]. / Blickinduzierte Amaurosis fugax.

Gaze-evoked amaurosis is a quite unique symptom. We present a 37-year-old patient with unilateral gaze-evoked amaurosis caused by an orbital tumor. Possible mechanisms include transient ischemia of the optic nerve and the retina or inhibition of the axonal impulses. While not common as a cause of amaurosis fugax, an orbital mass should be considered in the differential diagnosis of gaze-evoked atypical monocular amaurosis fugax.

TNF-alpha promoter polymorphisms and primary open-angle glaucoma.

PURPOSE: Primary open-angle glaucoma (POAG) is a multifactorial optic neuropathy with a strong hereditary component. Recent studies suggested a role for tumour necrosis factor-alpha(TNF-alpha) in the pathogenesis of POAG. The purpose of the present study was to investigate a hypothesized association between the TNF-alpha-308G>A and -238G>A gene polymorphisms and the presence of POAG in a Caucasian population. METHODS: The present case-control study comprised 114 unrelated patients with POAG and 228 healthy control subjects, matched for age and gender. Genotyping of the TNF-alpha-308G>A and -238G>A polymorphisms was performed using polymerase chain reaction. RESULTS: Allelic frequencies and genotype distributions of both the TNF-alpha-308G>A and -238G>A gene polymorphisms did not significantly differ between patients with POAG and control subjects. Presence of the TNF-alpha-308A-allele was associated with an odds ratio (OR) of 0.96 for POAG, whereas an OR of 0.52 was found among carriers of the TNF-alpha-238A-allele. CONCLUSION: Our data suggest that none of the investigated TNF-alphagene polymorphisms is a major risk factor among Caucasian patients with POAG.

Clinical use and rational management of homocysteine, folic acid, and B vitamins in cardiovascular and thrombotic diseases.

About half of all deaths are due to cardiovascular disease and its complications. The economic burden on society and the healthcare system from cardiovascular disability, complications, and treatments is huge and becoming larger in the rapidly aging populations of developed countries. As conventional risk factors fail to account for part of the cases, homocysteine, a "new" risk factor, is being viewed with mounting interest. Homocysteine is a sulfur-containing intermediate product in the normal metabolism of methionine, an essential amino acid. Folic acid, vitamin B(12), and vitamin B(6) deficiency and reduced enzyme activities inhibit the breakdown of homocysteine, thus increasing the intracellular homocysteine concentration. Numerous retrospective and prospective studies have consistently found an independent relationship between mild hyperhomocysteinemia and cardiovascular disease or all-cause mortality. Starting at a plasma homocysteine concentration of approximately 10 micromol/l, the risk increase follows a linear dose-response relationship with no specific threshold level. Hyperhomocysteinemia as an independent risk factor for cardiovascular disease is thought to be responsible for about 10 percent of total risk. Elevated plasma homocysteine levels (> 12 micromol/l; moderate hyperhomocysteinemia) are considered cytotoxic and are found in 5 to 10 percent of the general population and in up to 40 percent of patients with vascular disease. Additional risk factors (smoking, arterial hypertension, diabetes, and hyperlipidemia) may additively or, by interacting with homocysteine, synergistically (and hence overproportionally) increase overall risk. Hyperhomocysteinemia is associated with alterations in vascular morphology, loss of endothelial antithrombotic function, and induction of a procoagulant environment. Most known forms of damage or injury are due to homocysteine-mediated oxidative stresses. Especially when acting as direct or indirect antagonists of cofactors and enzyme activities, numerous agents, drugs, diseases, and life style factors have an impact on homocysteine metabolism. Folic acid deficiency is considered the most common cause of hyperhomocysteinemia. An adequate intake of at least 400 microg of folate per day is difficult to maintain even with a balanced diet, and high-risk groups often find it impossible to meet these folate requirements. Based on the available evidence, there is an increasing call for the diagnosis and treatment of elevated homocysteine levels in high-risk individuals in general and patients with manifest vascular disease in particular. Subjects of both populations should first have a baseline homocysteine assay. Except where manifestations are already present, intervention, if any, should be guided by the severity of hyperhomocysteinemia. Consistent with other working parties and consensus groups, we recommend a target plasma homocysteine level of < 10 micromol/l. Based on various calculation models, reduction of elevated plasma homocysteine concentrations may theoretically prevent up to 25 percent of cardiovascular events. Supplementation is inexpensive, potentially effective, and devoid of adverse effects and, therefore, has an exceptionally favorable benefit/risk ratio. The results of ongoing randomized controlled intervention trials must be available before screening for and treatment of hyperhomocysteinemia can be recommended for the apparently healthy general population.

Role of factor V Leiden and prothrombin 20210A in patients with retinal artery occlusion.

PURPOSE: Retinal artery occlusion is a common vision-threatening disease. Among other risk factors, coagulopathies leading to a hypercoagulable state have been associated with retinal artery occlusion. Numerous studies have shown that two genetic variants, factor V Leiden and prothrombin 20210A, cause a procoagulant state. However, their role in the pathogenesis of retinal artery occlusion is still unclear. The purpose of the present study was therefore to investigate a possible association between factor V Leiden, prothrombin 20210A, and retinal artery occlusion. METHODS: In the present retrospective case-control study, we studied 136 patients with retinal artery occlusion and 136 age- and gender-matched control subjects. The presence of factor V Leiden and prothrombin 20210A alleles was determined by polymerase chain reaction. RESULTS: The prevalence of heterozygosity for the prothrombin G20210A variant did not significantly differ between patients and controls (three patients vs two controls, P=0.65). Distribution of factor V Leiden genotypes revealed no significant difference among the two groups (heterozygosity: eight patients vs 11 controls, P=0.47). As for other risk factors, arterial hypertension, a history of stroke and myocardial infarction were significantly more frequent in patients than in controls. CONCLUSION: Our data suggest that factor V Leiden and prothrombin 20210A do not play a major role in patients with retinal artery occlusion.

[Transpupillary thermotherapy in exudative, age-related macular degeneration]. / Transpupilläre Thermotherapie bei exsudativer altersabhängiger Makuladegeneration.

OBJECTIVE: To assess the effectiveness of a single transpupillary thermotherapy (TTT) in patients with exudative, age-related macular degeneration (AMD). METHODS: In a prospective pilot study, 14 patients with a mean age of 78 years (range: 70-92 years) with subfoveal choroidal neovascularisation (CNV) due to exudative, age-related macular degeneration were treated with a single TTT using a diode laser (810 nm). Seven patients had a classic and seven an occult CNV. Laser beam size was 4.5 mm,the power setting was 800 mW and the exposure lasted 60 s. RESULTS: Twelve out of 14 patients could be followed for a period of 18 months. Stabilisation of the visual acuity was achieved in two patients, ten patients lost three or more Snellen lines. In none of the patients a regression of the CNV could be observed immediately after TTT. Inactive fibrotic scars developed in six patients at the end of the study. CONCLUSION: Our results suggest that TTT, according to our protocol, has no beneficial effect on the spontaneous course of the CNV in patients with AMD.

Basic care I: the effect of dementia-specific training on certified nursing assistants and other staff.

This study evaluates the effectiveness of Basic Care I, a dementia-specific training course offered by the St. Louis Alzheimer's Association. Using three standardized measures, the effects of the course on knowledge gain, stress level, and sense of work-related self-esteem are examined Findings suggest that participation in Basic Care I increases retained learning, as 76.4 percent of the sample showed improvement on dementia knowledge scores. The outcomes of stress and self-esteem measures are inconclusive but indicate areas for future study. Implications for program planning are discussed.

Search for occult secondary osteoporosis: impact of identified possible risk factors on bone mineral density.

OBJECTIVES: To determine whether the use of more elaborate diagnostic tests can identify possible risk factors for secondary osteoporosis and to evaluate the impact of these possible risk factors on the severity of bone disease in the study population. DESIGN: Cross-sectional study. SETTING AND PARTICIPANTS: We have investigated 377 subjects (285 females, 92 males) with osteoporosis (T-score less than -2.5 in dual energy X-ray absorption) or nontraumatic lumbar vertebral fractures; these patients were referred to our hospital, a secondary care centre, for evaluation and treatment of osteoporosis. RESULTS: Osteoporosis without attributable risk factor was diagnosed in 106 women (37%) and 30 men (33%). In 241 patients (179 women, 62 men) one or more possible risk factors for osteoporosis (in this paper also called subclinical disease) were revealed. The most common were lactose malabsorption, disturbed exocrine pancreatic function and renal tubular disturbances, including renal hypercalciuria, incomplete renal tubular acidosis and mild phosphate diabetes. The number of possible risk factors in the individual patient was significantly related to the severity of osteoporosis as assessed by Z-scores (Spearman correlation r = -0.43, P < 0.001, n = 172 for females; r = -0.28, P < 0.05, n = 65 for males). CONCLUSIONS: All the identified subclinical diseases would have remained undetected if the currently accepted guidelines for the investigation of patients with osteoporosis were applied. The statistically significant correlation between the number of identified possible risk factors and the severity of bone disease in the individual patient strongly suggests the pathogenetic significance of the identified subclinical diseases. It is yet to be shown, whether specific treatment of these subclinical diseases yields additional improvement of bone mass as compared with standard treatment of osteoporosis.

Role of factor XIII Val34Leu polymorphism in retinal artery occlusion.

BACKGROUND AND PURPOSE: Factor XIII (FXIII) Val34Leu, a common polymorphism in the gene for factor XIII, has been associated with a lower risk of stroke, myocardial infarction, and deep vein thrombosis. Ineffective fibrin cross-linking has been suggested to be causative. The aim of the present case-control study was to investigate the role of FXIII Val34Leu polymorphism in patients with retinal artery occlusion. METHODS: A total of 108 patients with retinal artery occlusion and 313 age- and sex-matched controls were genotyped for the FXIII Val34Leu polymorphism. Factor XIII Val34Leu genotypes were determined by use of allele-specific polymerase chain reaction. RESULTS: Homozygous Leu genotype was found significantly more often in control subjects than in patients with retinal artery occlusion (P=0.018), with an odds ratio of 0.22 (95% confidence interval 0.07 to 0.74). Distribution of the Val/Val and Val/Leu genotypes did not differ significantly between groups. CONCLUSIONS: Because prevalence of homozygous Leu genotype was significantly higher in controls, we conclude that the Leu/Leu genotype is associated with a protective effect against retinal artery occlusion.

Long-term pharmacokinetics of doxorubicin HCl stealth liposomes in patients after polychemotherapy with vinorelbine, cyclophosphamide and prednisone (CCVP).

The concentration-time profiles of Doxorubicin (DOXO) from day 0 to day 21 after i.v. infusion of 25 or 30 mg/m2 doxorubicin HCI stealth liposomes (Caelyx) were investigated in 9 patients receiving combination polychemotherapy with cyclophosphamide, vinorelbine and prednisone. Peak serum concentrations occurred from 0.04 to 4.0 days after infusion (mean tmax = 1.79 +/- 1.55 d) with a mean cmax of 4,595 +/- 2,849 ng/ml. A total amount of 12.84 +/- 2.47 mg liposomal DOXO in the plasma volume (Vp = 2,794 + 537 ml) could be estimated at tmax (= 27 % of the mean dose of 47.6 mg). Stealth liposomes were eliminated slowly from the blood with a mean t 1/2el of 1.9 + 0.5 days (MRT was 4.6 + 2.5 days). AUClast values ranged from 8,070 to 33,446 ng/ml*d (mean 10,987 +/- 9,339 ng/ml*d). The low plasma clearance (Cltot = 4,681 +/- 2,835 ml/day) and the small volume of distribution (Vz = 11.7 +/- 6.31) suggested that stealth-liposomes were stable in the blood at least for 14 days. Polychemotherapy with Hyper-CCVP schedule did not alter the stability of stealth liposomes, but peak levels of DOXO seemed to be somewhat lower compared to regression analysis of literature data (cmax versus dosage range from 20 to 60 mg/m2). Due to clast occurring between day 12 to 18, no indices for an accumulation of the drug in the blood could be found, when liposomes were given every four weeks.