The sensitivity of metabolomics versus classical regulatory toxicology from a NOAEL perspective.

The identification of the no observed adverse effect level (NOAEL) is the key regulatory outcome of toxicity studies. With the introduction of "omics" technologies into toxicological research, the question arises as to how sensitive these technologies are relative to classical regulatory toxicity parameters. BASF SE and metanomics developed the in vivo metabolome database MetaMap®Tox containing metabolome data for more than 500 reference compounds. For several years metabolome analysis has been routinely performed in regulatory toxicity studies (REACH mandated testing or new compound development), mostly in the context of 28 day studies in rats (OECD 407 guideline). For those chemicals for which a toxicological NOAEL level was obtained at either high or mid-dose level, we evaluated the associated metabolome to investigate the sensitivity of metabolomics versus classical toxicology with respect to the NOAEL. For the definition of a metabolomics NOAEL the ECETOC criteria (ECETOC, 2007) were used. In this context we evaluated 104 cases. Comparable sensitivity was noted in 75% of the cases, increased sensitivity of metabolomics in 8%, and decreased sensitivity in 18% of the cases. In conclusion, these data suggest that metabolomics profiling has a similar sensitivity to the classical toxicological study (e.g. OECD 407) design.

Application of in vivo metabolomics to preclinical/toxicological studies: case study on phenytoin-induced systemic toxicity.

BASF and Metanomics have built-up the database MetaMap(®)-Tox containing rat plasma metabolome data for more than 500 reference compounds. Phenytoin was administered to five Wistar rats of both sexes at dietary dose levels of 600 and 2400 ppm over 28 days and metabolome analysis was performed on days 7, 14 and 28. Clinical pathology did not indicate clear evidence for liver toxicity, whereas liver histopathology revealed slight centrilobular hepatocellular hypertrophy. The metabolome analysis of phenytoin shows metabolome changes at both dose levels and the comparison with MetaMap-Tox indicated strong evidence for liver enzyme induction, as well as liver toxicity. Moreover, evidence for kidney and indirect thyroid effects were observed. This assessment was based on the metabolite changes induced, similarities to specific toxicity patterns and the whole metabolome correlation within MetaMap-Tox. As compared with the classical read-out, a more comprehensive picture of phenytoin's effects is obtained from the metabolome analysis, demonstrating the added value of metabolome data in preclinical/ toxicological studies.

Nutritional impact on the plasma metabolome of rats.

Metabolite profiling (metabolomics) elucidates changes in biochemical pathways under various conditions, e.g., different nutrition scenarios or compound administration. BASF and metanomics have obtained plasma metabolic profiles of approximately 500 compounds (agrochemicals, chemicals and pharmaceuticals) from 28-day rat studies. With these profiles the establishment of a database (MetaMap(®)Tox) containing specific metabolic patterns associated with many toxicological modes of action was achieved. To evaluate confounding factors influencing metabolome patterns, the effect of fasting vs. non-fasting prior to blood sampling, the influence of high caloric diet and caloric restriction as well as the administration of corn oil and olive oil was studied for its influence on the metabolome. All mentioned treatments had distinct effects: triacylglycerol, phospholipids and their degradation product levels (fatty acids, glycerol, lysophosphatidylcholine) were often altered depending on the nutritional status. Also some amino acid and related compounds were changed. Some metabolites derived from food (e.g. alpha-tocopherol, ascorbic acid, beta-sitosterol, campesterol) were biomarkers related to food consumption, whereas others indicated a changed energy metabolism (e.g. hydroxybutyrate, pyruvate). Strikingly, there was a profound difference in the metabolite responses to diet restriction in male and female rats. Consequently, when evaluating the metabolic profile of a compound, the effect of nutritional status should be taken into account.

Chronomics, neuroendocrine feedsidewards and the recording and consulting of nowcasts--forecasts of geomagnetics.

A multi-center four-hourly sampling of many tissues for 7 days (00:00 on April 5-20:00 to April 11, 2004), on rats standardized for 1 month in two rooms on antiphasic lighting regimens happened to start on the day after the second extremum of a moderate double magnetic storm gauged by the planetary geomagnetic Kp index (which at each extremum reached 6.3 international [arbitrary] units) and by an equatorial index Dst falling to -112 and -81 nT, respectively, the latter on the first day of the sampling. Neuroendocrine chronomes (specifically circadian time structures) differed during magnetically affected and quiet days. The circadian melatonin rhythm had a lower MESOR and lower circadian amplitude and tended to advance in acrophase, while the MESOR and amplitude of the hypothalamic circadian melatonin rhythm were higher during the days with the storm. The circadian parameters of circulating corticosterone were more labile during the days including the storm than during the last three quiet days. Feedsidewards within the pineal-hypothalamic-adrenocortical network constitute a mechanism underlying physiological and probably also pathological associations of the brain and heart with magnetic storms. Investigators in many fields can gain from at least recording calendar dates in any publication so that freely available information on geomagnetic, solar and other physical environmental activity can be looked up. In planning studies and before starting, one may gain from consulting forecasts and the highly reliable nowcasts, respectively.

Phase I trial of intravenous aviscumine (rViscumin) in patients with solid tumors: a study of the European Organization for Research and Treatment of Cancer New Drug Development Group.

BACKGROUND: Aviscumine is an Escherichia coli-derived recombinant type II ribosome-inactivating protein with potent antitumor activity in vitro and in vivo. It is the recombinant counterpart of natural mistletoe lectin-I. The current study was performed to determine the safety profile, dose-limiting toxicity (DLT) and maximum tolerated dose (MTD) of the intravenous (i.v.) administration of aviscumine in cancer patients. Translational research included the evaluation of pharmacokinetics and monitoring of plasma cytokine and anti-aviscumine antibody induction after administration of the drug. PATIENTS AND METHODS: Aviscumine was given twice weekly as a 1 h central i.v. infusion in patients with advanced, refractory progressive, solid malignant tumors who had not been previously exposed to natural mistletoe preparations. They had histologically or cytologically verified disease, were > or =18 years old, had an Eastern Cooperative Oncology Group performance status < or =2 and adequate bone marrow, liver and renal function. DLT was defined as any non-hematological grade 3-4 toxicity (National Cancer Institute Common Toxicity Criteria version 2.0), neutrophil count <500/microl for > or =7 days, febrile neutropenia or thrombocytopenia grade 4. The MTD was defined as the dose at which >20% of patients experienced DLT during the first treatment cycle. The Continual Reassessment Method was used to determine the number of patients required per dose level. RESULTS: Forty-one fully eligible patients (19 male, 22 female) with a median age of 56 years (range 37-74) were enrolled. Colorectal, ovarian, renal cell and breast cancer were the most common tumor types. Dose levels of aviscumine ranged from 10 to 6400 ng/kg. The median number of cycles was two (range one to eight). Common clinical toxicities in cycle 1 were fatigue, fever, nausea, vomiting and allergic reactions. Fatigue grade 3 was dose limiting in one of six patients at 4000 ng/kg and reversible grade 3 liver toxicity (elevation in alkaline phosphatase, transaminases and/or gamma-glutamyltransferase) occurred in one of 10 patients at 4800 ng/kg and in two of five patients at 6400 ng/kg. The best response (RECIST criteria) was stable disease in 11 patients, lasting for two to eight cycles. The pharmacokinetic evaluation revealed a short alpha half-life of 13 min and linear kinetics on dose levels > or =1600 ng/kg. Aviscumine stimulated the immune system with a release of cytokines such as interleukin (IL)-1beta, IL-6 and interferon-gamma, and induced immunoglobulin (Ig) G- and/or IgM-anti-aviscumine antibodies of uncertain clinical relevance. CONCLUSIONS: The recommended dose for further clinical trials is 5600 ng/kg twice weekly. Based on the short half-life of the recombinant protein observed in this trial, the exploration of prolonged infusion schedules of aviscumine is warranted.

Prospective controlled evaluation of the influence of an excursion to a health resort on medical students' opinion about chances for outpatient rehabilitation at a health resort.

OBJECTIVE: To assess the influence of an excursion to a health resort on students' opinion on the chances for outpatient rehabilitation at a health resort for cardiac and rheumatic diseases. METHODS: A group of 17 medical students went on a 1-day excursion to Bad Orb (near Frankfurt/M.) where mainly patients with heart diseases are treated. Another group of 44 students went on a 5-day excursion to Bad Gastein (Austria) where mainly patients with rheumatic diseases are treated. Before and after the excursion students completed a questionnaire asking their opinion on the chances for outpatient rehabilitation at a health resort for acute and chronic cardiac and rheumatic diseases. RESULTS: Students judged the chances for beneficial effects of outpatient rehabilitation at a health resort for chronic rheumatic diseases to be significantly better after the excursion to Bad Gastein than before, whereas there was no difference after the excursion to Bad Orb. CONCLUSION: Students' opinion of chances for outpatient rehabilitation at a health resort can be influenced by an excursion to a spa, but a single 1-day excursion may not be sufficient.

[Influence of reflex zone therapy of the feet on intestinal blood flow measured by color Doppler sonography]. / Beeinflussung der Darmdurchblutung durch Fussreflexzonenmassage, gemessen mittels farbkodierter Dopplersonographie.

OBJECTIVE: An influence on organ-associated blood flow is considered as a possible mechanism of action of reflex zone massage of the feet (FRZM) therapy. In the present study we investigated whether changes in intestinal blood flow can be achieved by FRZM. MATERIAL AND METHODS: 32 healthy adults (19 women and 13 men) were randomly assigned to the treatment or the placebo group. Subjects of the treatment group received foot massage on the zones assigned to the intestines and those of the placebo group received massage on zones unrelated to the intestines. Before, during and after FRZM, the blood flow velocity, the peak systolic and the end diastolic velocities in the superior mesenteric artery as well as the resistive index as a parameter of vascular resistance were calculated. RESULTS: During FRZM, in the subjects of the treatment group there was a significant reduction in the resistive index (p = 0.021), suggesting an increase in the blood flow in the superior mesenteric artery and the subordinate vascular system. In contrast, there were no significant changes in the resistive index in the subjects of the placebo group. CONCLUSION: The reduction in the resistive index observed in the treatment group supports the assumption that FRZM improves blood flow in the organs considered to be associated with the specific foot zones, at least during the therapy process.

Circadian and infradian rhythms in mood.

The aim of this study was to assess any variation in positive, negative and total affect recorded longitudinally; to compare the results with those from prior transverse or hybrid population studies, based on the same or a different method of mood rating; and to test for any association of mood with cardiovascular, hormonal and geophysical variables monitored concomitantly. The study approach was as follows. A clinically healthy 34-year-old man filled out the positive and negative affective scale (PANAS) questionnaire five times a day for 86 days. Systolic (S) and diastolic (D) blood pressure (BP) and heart rate (HR) were also measured automatically at 30-minute intervals with an ambulatory monitor from May 19 to June 29, 2000, while different endpoints of heart rate variability (HRV) were also determined at 5-minute intervals from beat-to-beat electrocardiogram (ECG) monitoring for 42 days between May 3 and June 14, 2000, with only short interruptions while the subject took a shower and changed ECG tapes. Saliva samples were collected at the times of mood ratings for one month for later determination of melatonin and cortisol concentrations. Intervals of 24 hours of the record of each variable displaced in increments of 24 hours were analyzed by chronobiologic serial section at a trial period of 24 hours to assess the circadian characteristics as they changed from one day to another. Estimates of the midline-estimating statistic of rhythm (MESOR) and circadian amplitude and acrophase obtained on consecutive days were correlated among variables to assess any associations. The findings were as follows. Overall, a circadian rhythm was demonstrated for all variables. A positive association was noteworthy between the circadian amplitude of negative affect and the MESOR of both SBP (r= 0.363; P= 0.029) and DBP (r= 0.389; P= 0.019), suggesting that BP is raised in the presence of large swings in negative affect. Needing further validation was a weak association between the MESOR of negative affect and the circadian amplitude of SBP (r= - 0.272; P = 0.108), suggesting a lowering of the circadian SBP amplitude in the presence of a strong negative affect. Of further interest was the lack of a statistically significant relation between positive and negative affect, not only in terms of the MESOR but also in terms of the circadian amplitude.

Unconventional medicine in Central Europe: a misuse of public health insurance?

OBJECTIVE: To scrutinize the presumption maintained by critics that patients seeking medical treatment at a health resort may be more motivated by the prospect of a pleasant sojourn paid for by health insurance than by the impairment caused by a disease. DESIGN: Variables for mobility (occiput-to-wall distance, cervical rotation, chest expansion, thoracic flexion, lumbar flexion, and finger-to-floor distance) and C-reactive protein were determined in 181 patients (male 134, female 47; age 52.4 +/- SEM 0.8 years) with ankylosing spondylitis (AS) whose costs were covered by their health insurance (group A) and in 77 AS patients (male 66, female 11; age 51.6 +/- 1.2 years) who paid their own costs (group B). SUBJECTS: A group of 258 patients with AS presenting for 3- or 4-week speleotherapeutic radon treatment at the Gasteiner Heilstollen Hospital, a medical institution located at Badgastein in the Austrian Alps. RESULTS: After Bonferroni correction for multiple calculations no significant difference was seen between the two groups. CONCLUSIONS: The results suggest that patients presenting for medical treatment at a health resort suffer a like degree of disease impairment, whether they pay their own costs or not. There was no evidence that seeking treatment at a health resort may be an attempt by patients to misuse the health insurance for "sponsored" holidays.

[Changes of renal blood flow during organ-associated foot reflexology measured by color Doppler sonography]. / Anderung der nierenduchblutung durch organassoziierte reflexzonentherapie am fuss gemessen mit farbkodierter Doppler-sonographie.

Using colour Doppler sonography blood flow changes of the right kidney during foot reflexology were determined in a placebo-controlled, double-blind, randomised study. 32 healthy young adults (17 women, 15 men) were randomly assigned to the verum or placebo group. The verum group received foot reflexology at zones corresponding to the right kidney, the placebo group was treated on other foot zones. Before, during and after foot reflexology the blood flow of three vessels of the right kidney was measured using colour Doppler sonography. Systolic peak velocity and end diastolic peak velocity were measured in cm/s, and the resistive index, a parameter of the vascular resistance, was calculated. The resistive index in the verum group showed a highly significant decrease (p

Decrease of respiratory burst in neutrophils of patients with ankylosing spondylitis by combined radon-hyperthermia treatment.

OBJECTIVE: To define the respiratory burst activity of neutrophils, the total anti-oxidative status of plasma, and the parameters of systemic inflammation in patients with ankylosing spondylitis (AS) before and after a combined radon-hyperthermia treatment in the thermal tunnels of Böckstein-Bad Gastein in Austria. METHODS: In 20 patients with AS the effects of a total of 15 hours of radon-hyperthermia-treatment spread over a period of three weeks were studied. The respiratory burst activity of neutrophils was measured fluorometrically using dichlorofluorescein diacetate, the total anti-oxidant status was measured using azinodiethyl-benzthiazoline-sulphonate, and inflammation parameters were determined by routine laboratory assays. RESULTS: Before treatment, the basal neutrophil respiratory burst in patients (n = 20) was 409 +/- 62 fluorescence arbitrary units (AU; mean +/- SEM) and 359 +/- 37 AU in controls (n = 9; p > 0.5); the stimulated respiratory burst (fMet-Leu-Phe, 10(-6) M) was 1,027 +/- 133 AU in patients and 1,152 +/- 218 AU in controls (p > 0.5). After treatment, the basal neutrophil respiratory burst in patients (n = 19) was 137 +/- 16 and in controls it was 174 +/- 35 AU (n = 8; p > 0.1); the stimulated respiratory burst was 670 +/- 66 and 1,305 +/- 82 AU, in patients and controls respectively (p < 0.001). No effects of treatment on the total anti-oxidant status of the plasma or on the parameters of inflammation were detected. CONCLUSION: Combined radon-hyperthermia treatment reduces the respiratory burst activity of the blood circulating neutrophils in patients with AS. If respiratory burst activity from the neutrophils plays a role in the pathophysiology of ankylosing spondylitis, the observed reduction may be related to the beneficial effects of radon-hyperthermia treatment.

Heterologous expression of Arabidopsis phytochrome B in transgenic potato influences photosynthetic performance and tuber development.

Transgenic potato (Solanum tuberosum) plants expressing Arabidopsis phytochrome B were characterized morphologically and physiologically under white light in a greenhouse to explore their potential for improved photosynthesis and higher tuber yields. As expected, overexpression of functional phytochrome B caused pleiotropic effects such as semidwarfism, decreased apical dominance, a higher number of smaller but thicker leaves, and increased pigmentation. Because of increased numbers of chloroplasts in elongated palisade cells, photosynthesis per leaf area and in each individual plant increased. In addition, photosynthesis was less sensitive to photoinactivation under prolonged light stress. The beginning of senescence was not delayed, but deceleration of chlorophyll degradation extended the lifetime of photosynthetically active plants. Both the higher photosynthetic performance and the longer lifespan of the transgenic plants allowed greater biomass production, resulting in extended underground organs with increased tuber yields.

Modulatory potential of iron chelation therapy on nitric oxide formation in cerebral malaria.

To determine whether iron chelation modulates nitric oxide (NO) formation and cell-mediated immune effector function in children with cerebral malaria, serum concentrations were measured of the stable end products of NO, nitrite and nitrate (NO2-/NO3-), interleukin (IL)-4, -6, and -10, and neopterin in 39 Zambian children enrolled in a placebo-controlled trial of desferrioxamine B and quinine therapy. Mean concentrations of NO2-/NO3- increased significantly over 3 days in children receiving desferrioxamine plus quinine but not in those given placebo and quinine. Neopterin levels declined significantly with placebo but not with desferrioxamine. IL-4 levels increased progressively in the placebo group and ultimately decreased in the desferrioxamine group, but the trends were not statistically significant. IL-6 and IL-10 levels were elevated initially and decreased significantly in both groups over 3 days. These data are consistent with the hypothesis that iron chelation therapy in children with cerebral malaria strengthens Th1-mediated immune effector function involving increased production of NO.

In vitro stimulation of erythropoiesis by stem cell factor alone in myelodysplastic syndrome patients with elevated endogenous erythropoietin serum levels.

Anemia remains a therapeutic problem in patients with myelodysplastic syndrome (MDS). In view of the recently reported potential of stem cell factor (SCF) in restoring erythropoiesis in combination with erythropoietin (Epo), we first aimed to define a correlation between SCF serum levels and anemia in MDS. Endogenous SCF levels in 50 MDS patients were determined by using a quantitative sandwich enzyme immunoassay. Broad interindividual variations were observed, but SCF serum levels were in the normal range with no correlation to peripheral blood count. A soft agar culture system was used to further define the role of SCF for stimulation of erythroid growth. Bone marrow mononucleated cells of 20 MDS patients (4 refractory anemia, 5 refractory anemia with excess of blasts, 7 refractory anemia with excess of blasts in transition, and 4 chronic myelomonocytic leukemia) were investigated, and SCF plus Epo was able to stimulate burst-forming unit-erythroid significantly more than SCF or Epo alone independent of French-American-British group. When mononucleated cells from six MDS patients (two refractory anemia, two refractory anemia with excess of blasts, and 2 refractory anemia with excess of blasts in transition) with elevated serum Epo levels were incubated in the presence of SCF and autologous serum, a significant dose-dependent stimulation of burst-forming unit-erythroid number and cells per colony was detected. Erythroid differentiation was further enhanced by adding serum with high colony-stimulating activity obtained from patients with severe aplastic anemia. Our data suggest that in MDS patients with high endogenous Epo serum levels SCF alone might be effective in stimulating erythropoiesis in vivo.

Serum neopterin, interleukin-4, and interleukin-6 concentrations in cerebral malaria patients and the effect of iron chelation therapy.

To determine if iron chelation therapy alters immune responses in children with cerebral malaria, we retrospectively measured mean serum levels of neopterin, interleukin-4 (IL-4), and IL-6 in children who received desferrioxamine B or placebo for three days in addition to quinine-based therapy. Mean levels of neopterin, IL-4, and IL-6 were elevated above the expected normal range on admission. Neopterin correlated significantly with the degree of anemia, IL-4 with the duration of fever prior to admission, and IL-6 with parasite density. Serial measurements of cytokines and neopterin were performed over four days in 39 children, 21 randomized to receive desferrioxamine B and 18 to receive placebo. Mean concentrations of neopterin did not change significantly in either group while levels of IL-4 increased significantly in the placebo group (P = 0.04) but remained unchanged in the desferrioxamine B group. Interleukin-6 concentrations decreased markedly in both groups (P < 0.025). Stable IL-4 levels in children given desferrioxamine B may represent an inhibition of the T helper lymphocyte-2 (TH-2) response resulting from a strengthened TH-1 response associated with iron chelation therapy. Any effect of iron chelation on immunity in the setting of severe malaria will have to be confirmed in future prospective investigations.

[Research on the psychomotor performance of patients with vertebral syndromes at spas]. / Testpsychologische Leistungsuntersuchungen an Patienten mit Wirbelsäulen-Syndromen im Kurverlauf.

The influence of radon balneotherapy on the psychomotor performance of 169 male and female patients with vertebral syndromes was investigated. The speed and skill of motor response to various stimuli was measured by the Schoppe apparatus and the Viennese determination apparatus. The analysis of "steadiness" brought no results, whereas all the other parameters tested by the Schoppe and the Viennese devices showed highly significant improvements in the course of spa treatment. It was not possible to differentiate to which specific factors of spa treatment this positive influence can be ascribed.

Alterations of haemostasis after exposure to sauna.

Blood coagulation and fibrinolysis were examined in young healthy male volunteers before and after sauna baths. No consistent or periodical changes could be found. However on several days the increases of haemoglobin, platelet counts and factor VIII associated antigen and to a lesser degree of haematocrit, factor VIII, IX and XII reached statistical significance after sauna. The results suggest a tendency to an increased activity of the blood coagulation system.

Pharmacological studies on retinoic acid in Wistar and in atrichos rats.

Retinoic acid produces some changes in the skin which could be interpreted as a mechanism of action: release of acid hydrolases and phosphatases, inflammatory reactions, increase in lipid synthesis and transport as an energy dependent phenomenon, increase in the lipid flux perhaps as a lipoprotein complex, since there is a rise in nucleic acid synthesis. Stimulation of bone remodeling is observed only when high dosage of retinoic acid is given orally.