A randomized, observer-blind, vehicle-control, multi-center clinical investigation for assessing the efficacy and tolerability of a 1% ectoine and hyaluronic acid 0.1%-containing medical device in pediatric patients with mild-to-moderate atopic dermatitis.

BACKGROUND: Ectoine is a widespread osmolyte enabling halophilic bacteria to withstand high osmotic stress that has many potential applications ranging from cosmetics to its use as a therapeutic agent. OBJECTIVE: The aim of this study was to compare the efficacy and tolerability of ectoine 1% and hyaluronic acid 0.1% containing (EHA) cream with a vehicle cream in children with mild-to-moderate atopic dermatitis (AD). METHODS: A randomized, controlled, observer-blind, multicenter clinical trial was conducted in children aged 2-18 years, diagnosed with mild-to-moderate AD (SCORAD ≤20). Patients were randomized to either receiving EHA cream or vehicle cream twice daily for 4 weeks. The primary outcome measure was the mean change in objective SCORAD from baseline to the final visit. The secondary outcome measures included the mean change in Investigator's Global Assessment score, patient's judgment of efficacy and patient's assessment of pruritus. Safety of EHA cream was also assessed. RESULTS: A total of 70 patients (35 in each group) were randomized and 57 were included in the final analysis set. Based on SCORAD measurements, patients using EHA cream achieved superior clinical improvement compared to the control group at 28 days (p < .001). EHA cream was also superior to the vehicle cream regarding all secondary outcome measures. Eight (23.5%) patients receiving EHA cream and two (5.7%) patients receiving vehicle cream experienced mild cutaneous adverse events (AEs). CONCLUSIONS: In children 2-18 years old with mild-to-moderate AD, EHA cream was superior to vehicle cream, with minor AEs.

A chemogenomic approach is required for effective treatment of amyotrophic lateral sclerosis.

ALS is a fatal untreatable disease involving degeneration of motor neurons. Μultiple causative genes encoding proteins with versatile functions have been identified indicating that diverse biological pathways lead to ALS. Chemical entities still represent a promising choice to delay ALS progression, attenuate symptoms and/or increase life expectancy, but also gene-based and stem cell-based therapies are in the process of development, and some are tested in clinical trials. Various compounds proved effective in transgenic models overexpressing distinct ALS causative genes unfortunately though, they showed no efficacy in clinical trials. Notably, while animal models provide a uniform genetic background for preclinical testing, ALS patients are not stratified, and the distinct genetic forms of ALS are treated as one group, which could explain the observed discrepancies between treating genetically homogeneous mice and quite heterogeneous patient cohorts. We suggest that chemical entity-genotype correlation should be exploited to guide patient stratification for pharmacotherapy, that is administered drugs should be selected based on the ALS genetic background.

ß-Methylamino-L-alanine-induced protein aggregation in vitro and protection by L-serine.

The cyanobacterial non-protein amino acid α-amino-ß-methylaminopropionic acid, more commonly known as BMAA, was first discovered in the seeds of the ancient gymnosperm Cycad circinalis (now Cycas micronesica Hill). BMAA was linked to the high incidence of neurological disorders on the island of Guam first reported in the 1950s. BMAA still attracts interest as a possible causative factor in amyotrophic lateral sclerosis (ALS) following the identification of ALS disease clusters associated with living in proximity to lakes with regular cyanobacterial blooms. Since its discovery, BMAA toxicity has been the subject of many in vivo and in vitro studies. A number of mechanisms of toxicity have been proposed including an agonist effect at glutamate receptors, competition with cysteine for transport system xc_ and other mechanisms capable of generating cellular oxidative stress. In addition, a wide range of studies have reported effects related to disturbances in proteostasis including endoplasmic reticulum stress and activation of the unfolded protein response. In the present studies we examine the effects of BMAA on the ubiquitin-proteasome system (UPS) and on chaperone-mediated autophagy (CMA) by measuring levels of ubiquitinated proteins and lamp2a protein levels in a differentiated neuronal cell line exposed to BMAA. The BMAA induced increases in oxidised proteins and the increase in CMA activity reported could be prevented by co-administration of L-serine but not by the two antioxidants examined. These data provide further evidence of a protective role for L-serine against the deleterious effects of BMAA.

Complexes of Ectoine with the Anionic Surfactants as Active Ingredients of Cleansing Cosmetics with Reduced Irritating Potential.

For many years, an increasing number of diagnosed atopy and skin problems have been observed. For people affected by the problem of atopy, the selection of skin care products, including cosmetics, is extremely important. Cleansing cosmetics, due to their ability to cause skin irritations and disturb the hydrolipidic barrier, can increase problems with atopic skin. New solutions to reduce the effects of these products on the skin are very important. In this work, the effect of ectoine on the properties of anionic surfactants was analyzed. Based on model systems, analysis of the effect of ectoine on the irritating effect of four anionic surfactants and their ability to solubilize model sebum was performed. Antioxidant activity was also evaluated, and cytotoxic studies were performed on cell cultures. It was shown that the addition of ectoine to the anionic surfactant solutions improves its safety of use. After introducing ectoine to the surfactant solution, a decrease of irritant potential (about 20%) and a decrease in the ability to solubilize of model sebum (about 10-20%) was noted. Addition of ectoine to surfactant solutions also reduced their cytotoxicity by up to 60%. The obtained results indicate that ectoine may be a modern ingredient that improves the safety of cleansing cosmetics.

Dose-Dependent Adult Neurodegeneration in a Rat Model After Neonatal Exposure to ß-N-Methylamino-L-Alanine.

The link between neonatal BMAA exposure and neurodegeneration has recently been demonstrated in rodents. We therefore investigated the behavioral and histopathological dose response to BMAA administered as a single dose. We report here that exposure to a BMAA dose as low as 50 mg/kg on PND 3 caused mild short-term behavioral alterations as well as beta-amyloid deposition together with neuronal loss in the hippocampus of adult rats. Additionally, all histopathological abnormalities and behavioral deficits that had been observed in a previous study in the brain and spinal cord tissue of rats exposed to 400 mg/kg BMAA on PND 3 were also observed here in the brain and spinal cord tissue of male and female rats exposed to 100 mg/kg BMAA at the same age, although the proteinopathy burdens and volume losses were lower. Both behavioral deficits and histopathology increased with increasing dose, and a single neonatal BMAA exposure at a dose of 100 mg/kg was the lowest dose able to cause clinical signs of toxicity, behavioral deficits, and neuropathology that are typically observed in AD, PD, and/or ALS patients.

Genotoxic effects of neurotoxin ß-N-methylamino-l-alanine in human peripheral blood cells.

The non-proteinogenic amino acid ß-N-methylamino-l-alanine (BMAA) is associated with the development of neurodegenerative diseases such as Alzheimer's disease, amyotrophic lateral sclerosis/parkinsonism-dementia complex (ALS-PDC) and amyotrophic lateral sclerosis. BMAA is known to induce neurotoxic effects leading to neurodegeneration via multiple mechanisms including misfolded protein accumulation, glutamate induced excitotoxicity, calcium dyshomeostasis, endoplasmic reticulum stress and oxidative stress. In the present study, for the first time, genotoxic activity of BMAA (2.5, 5, 10 and 20 µg/mL) was studied in human peripheral blood cells (HPBCs) using the comet and cytokinesis-block micronucleus cytome assays. In addition, the influence of BMAA on the oxidative stress was assessed. At non-cytotoxic concentrations BMAA did not induce formation of DNA strand breaks in HPBCs after 4 and 24 h exposure; however, it significantly increased the number of micronuclei after 24 and 48 h at 20 µg/mL and nucleoplasmic bridges after 48 h at 20 µg/mL. The frequency of nuclear buds was slightly though non-significantly increased after 48 h. Altogether, this indicates that in HPBCs BMAA is clastogenic and induces complex genomic alterations including structural chromosomal rearrangements and gene amplification. No influence on oxidative stress markers was noticed. These findings provide new evidence that environmental neurotoxin BMAA, in addition to targeting common pathways involved in neurodegeneration, can also induce genomic instability in non-target HPBCs suggesting that it might be involved in cancer development. Therefore, these data are important in advancing our current knowledge and opening new questions in the understanding of the mechanisms of BMAA toxicity, particularly in the context of genotoxicity.

50 years of research on α-amino-ß-methylaminopropionic acid (ß-methylaminoalanine).

The isolation of α-amino-ß-methylaminopropionic acid from seeds of Cycas circinalis (now C. micronesica Hill) resulted from a purposeful attempt to establish the cause of the profound neurological disease, amyotrophic lateral sclerosis/parkinsonism/dementia, that existed in high frequency amongst the inhabitants of the western Pacific island of Guam (Guam ALS/PD). In the 50 years since its discovery the amino acid has been a stimulus, and sometimes a subject of mockery, for generations of scientists in a remarkably diverse range of subject areas. The number of citations of the original paper has risen in the five decades from a few to 120 within the decade 2007-2016 and continues at a high rate into the next decade. The reasons for this remarkable outcome are discussed and examples from the literature are used to illustrate the wide range of scientific interest that the original paper generated.

Reduction of neutrophilic lung inflammation by inhalation of the compatible solute ectoine: a randomized trial with elderly individuals.

BACKGROUND: Compatible solutes are natural substances that are known to stabilize cellular functions. Preliminary ex vivo and in vivo studies demonstrated that the compatible solute ectoine restores natural apoptosis rates of lung neutrophils and contributes to the resolution of lung inflammation. Due to the low toxicity and known compatibility of the substance, an inhalative application as an intervention strategy for humans suffering from diseases caused by neutrophilic inflammation, like COPD, had been suggested. As a first approach to test the feasibility and efficacy of such a treatment, we performed a population-based randomized trial. OBJECTIVE: The objective of the study was to test whether the daily inhalation of the registered ectoine-containing medical device (Ectoin® inhalation solution) leads to a reduction of neutrophilic cells and interleukin-8 (IL-8) levels in the sputum of persons with mild symptoms of airway disease due to lifelong exposure to environmental air pollution. METHODS: A double-blinded placebo-controlled trial was performed to study the efficacy and safety of an ectoine-containing therapeutic. Prior to and after both inhalation periods, lung function, inflammatory parameters in sputum, serum markers, and quality-of-life parameters were determined. RESULTS: While the other outcomes revealed no significant effects, sputum parameters were changed by the intervention. Nitrogen oxides (nitrate and nitrite) were significantly reduced after ectoine inhalation with a mean quotient of 0.65 (95% confidence interval 0.45-0.93). Extended analyses considering period effects revealed that the percentage of neutrophils in sputum was significantly lower after ectoine inhalation than in the placebo group (P=0.035) even after the washout phase. CONCLUSION: The current study is the first human trial in which the effects of inhaled ectoine on neutrophilic lung inflammation were investigated. Besides demonstrating beneficial effects on inflammatory sputum parameters, the study proves the feasibility of the therapeutic approach in an aged study group.

Cyanobacterial Neurotoxin BMAA and Mercury in Sharks.

Sharks have greater risk for bioaccumulation of marine toxins and mercury (Hg), because they are long-lived predators. Shark fins and cartilage also contain ß-N-methylamino-l-alanine (BMAA), a ubiquitous cyanobacterial toxin linked to neurodegenerative diseases. Today, a significant number of shark species have found their way onto the International Union for Conservation of Nature (IUCN) Red List of Threatened Species. Many species of large sharks are threatened with extinction due in part to the growing high demand for shark fin soup and, to a lesser extent, for shark meat and cartilage products. Recent studies suggest that the consumption of shark parts may be a route to human exposure of marine toxins. Here, we investigated BMAA and Hg concentrations in fins and muscles sampled in ten species of sharks from the South Atlantic and Pacific Oceans. BMAA was detected in all shark species with only seven of the 55 samples analyzed testing below the limit of detection of the assay. Hg concentrations measured in fins and muscle samples from the 10 species ranged from 0.05 to 13.23 ng/mg. These analytical test results suggest restricting human consumption of shark meat and fins due to the high frequency and co-occurrence of two synergistic environmental neurotoxic compounds.

Effects of ectoine on behavioural, physiological and biochemical parameters of Daphnia magna.

Ectoine (ECT) is a compatible solute produced by soil, marine and freshwater bacteria in response to stressful factors. The purpose of our study was to determine the possible toxic influence of ECT on Daphnia magna. We determined the following endpoints: survival rate during exposure and recovery, swimming performance, heart rate, thoracic limb movement determined by image analysis, haemoglobin level by ELISA assay, catalase and nitric oxide species (NOx) by spectrophotometric methods. The results showed 80% survival of daphnids exposed to 50mg/L of ECT after 24h and 10% after 90h, however lower concentrations of ECT were well tolerated. A concentration-dependent reduction of swimming velocity was noted at 24 and 48h of the exposure. ECT (at 2.5 and 4mg/L) induced an increase of heart rate and thoracic limb movement (at 2.5, 4 and 20mg/L) after 24h. After 10h of the exposure to ECT daphnids showed a concentration-dependent increase of haemoglobin level synthesized and accumulated in the epipodite epithelia. After 24h we noted a concentration-dependent decrease of haemoglobin level and its lowest value was found after 48h of the exposure. ECT at a concentration of 20 and 25mg/L slightly stimulated catalase activity after 24h. NOx level was also increased after 10h of the exposure to 20 and 25mg/L of ECT reaching maximal activity after 24h. Our results suggest that ECT possesses some modulatory potential on the behaviour, physiology and biochemical parameters in daphnids.

A prospective, controlled study of SNS01 (ectoine nasal spray) compared to BNO-101 (phytotherapeutic dragées) in patients with acute rhinosinusitis.

OBJECTIVES: In this observational study, data on the efficacy, effects on quality of life and tolerability of the topical formulation SNS01 (Ectoin Rhinitis nasal spray) were compared to those of BNO-101 (Sinupret forte dragées) in patients with acute rhinosinusitis in the ear, nose, and throat (ENT) clinical setting. DESIGN AND METHODS: Patients with the diagnosis of acute rhinosinusitis were included in this non-interventional study with a treatment duration of 14-16 days. They received either a herbal phytotherapeutic dragée (control) or an ectoine-based nasal spray (investigational product), each to be taken according to the instructions for use (IFU) and summary of product characteristics (SPC). At each visit, the physician performed a nasal endoscopy, recorded the Sinusitis Symptom Score and checked for adverse events. During the entire treatment period, patients recorded the Sinusitis Symptom Score in patient diaries. In addition, patients receiving the nasal spray filled out a questionnaire to assess the tolerability of the treatment. To investigate effects on quality of life patients were asked to fill out the German version of a sinusitis-specific HRQL (health related quality of life) questionnaire. CLINICAL TRIAL REGISTRATION: NCT01684540. RESULTS: Patient diary entries, the assessment of the Sinusitis Symptom Score and the HRQL questionnaire demonstrated that the ectoine nasal spray was as effective as the phytotherapeutic dragées in treating acute rhinosinusitis. After two weeks of treatment, the assessments of both the patients' diaries and physicians' record forms indicated statistically significant improvement (p ≤ 0.001) in the symptom scores of the two groups (57.8% improvement for ectoine and 49.3% improvement for the phytotherapeutic dragées compared to baseline). Also, overall scores of 80 in the sensory questionnaire confirmed the good tolerability of the nasal spray. Correspondingly, HRQL improved significantly over the course of the treatment in both groups. CONCLUSION: SNS01 and BNO-101 demonstrated comparable effects in the treatment of acute rhinosinusitis. LIMITATIONS: Following German regulations, this trial was set up as an observational 'non-interventional' study, which does not allow for a placebo group or randomization of patients. Although the grade of evidence delivered by the study data is thus reduced from Ib to IIa, it does, however, reflect a realistic view of the most common clinical practice.

[The Parkinson puzzle]. / A Parkinson-puzzle.

Parkinson's disease is one of the most frequent progressive degenerative disorders with unknown origin of the nervous system. The commutation of the disease on Guam led to the discovery of a neurotoxin which was also found in other continents. This neurotoxin was identified in the common cyanobacteria (blue-green algae). Early clinical observations suggested some loose correlations with gastric and duodenal ulcer and Parkinson's disease, while recent studies revealed a toxin, almost identical to that found in cyanobacteria in one strain of Helicobacter pylori, which proved to cause Parkinson like symptoms in animals. Therefore, it cannot be ruled out that there is a slowly progressive poisoning in Parkinson's disease. The disease specific alpha-sinuclein inclusions can be found in nerve cells of the intestinal mucosa far before the appearance of clinical symptoms indicating that the disease may start in the intestines. These results are strengthened by the results of Borody's fecal transplants, after which in Parkinson patients showed a symptomatic improvement. Based on these observations the Parkinson puzzle is getting complete. Although these observations are not evidence based, they may indicate a new way for basic clinical research, as well as a new way of thinking for clinicians. These new observations in psycho-neuro-immunology strengthen the fact that immunological factors may also play a critical factor facilitating local cell necrosis which may be influenced easily.

[Revisit to Kii ALS--the innovated concept of ALS-Parkinsonism-dementia complex, clinicopathological features, epidemiology and etiology].

The high incidence of amyotrophic lateral sclerosis (ALS) in the residents of Hohara and Kozagawa in the Kii peninsula was reported to have disappeared by early 1980 with its etiology unsolved. However, we found continuous high incidence in Hohara that was neuropathologically characterized by ALS pathology associated with many neurofibrillar tangles (NFTs) similar to Guam ALS. We confirmed existence of neuropathologically-verified parkinsonism-dementia complex (PDC) identical to Guamanian PDC clinically and neuropathologically. The core clinical features consisted of motor neuron signs, parkinsonism and dementia, and patients presented with clinical manifestations of ALS, PDC or PDC followed by ALS. PDC predominated over ALS in incidence. Approximately 70% of patients had family history of ALS/PDC. Neuropathological findings of 12 cases revealed that they were very similar to each others, consisting of many NFTs, no or scanty amyloid plaques, and ALS pathology affecting the upper and lower motor neurons. These findings suggest that ALS and PDC may be different clinical manifestations of a single entity "ALS-parkinsonism-dementia complex". TDP-43 positive inclusions were seen in the neurons of the dentate gyrus and spinal cord in all 6 cases examined. A comparison of age-adjusted prevalence rates in 1967 and 1998 revealed moderate decline of ALS and marked increase of PDC in the latter. The age-adjusted 5-year average incidence rates during 1950 and 2000 showed gradual decline of ALS for 50 years and dramatic increase of PDC after 1990. These findings suggest that the clinical manifestations may have changed in Kii ALS/PDC as in ALS/PDC on Guam, partly because of rapid aging of the population. Gene analyses have so far failed to demonstrate mutations of SOD1, parkin, alpha-synuclein, tau, progranulin, TDP-43 and other genes related to dementia, parkinsonism and motor neuron disease. There have been no differences in drinking water and food between the residents in the high incidence area and those in the neighboring low incidence areas, and none of the patients had habits of eating the cycad, flying fox or any other odd materials. These findings suggest that genetic factors may be etiologically primary and environmental factors may modify the clinical phenotypes.

Cycad neurotoxins, consumption of flying foxes, and ALS-PDC disease in Guam.

The Chamorro people of Guam have been afflicted with a complex of neurodegenerative diseases (now known as ALS-PDC) with similarities to ALS, AD, and PD at a far higher rate than other populations throughout the world. Chamorro consumption of flying foxes may have generated sufficiently high cumulative doses of plant neurotoxins to result in ALS-PDC neuropathologies, since the flying foxes forage on neurotoxic cycad seeds.

From soil to brain: zinc deficiency increases the neurotoxicity of Lathyrus sativus and may affect the susceptibility for the motorneurone disease neurolathyrism.

Zinc deficiency and oversupply of iron to the roots of grass pea (Lathyrus sativus) induce increases in the content of the neurotoxin beta-L-ODAP (3-oxalyl-L-2,3-diaminopropanoic acid) in the ripe seeds. The transport of zinc to the shoots is enhanced by the addition of beta-L-ODAP. The neurotoxin of L. sativus is proposed to function as a carrier molecule for zinc ions. Soils, depleted in micronutrients from flooding by monsoon rains (Indian subcontinent) or otherwise poor in available zinc and with high iron content (Ethiopian vertisols), may be responsible for higher incidence of human lathyrism, one of the oldest neurotoxic diseases known to man. A role for brain zinc deficiency in the susceptibility for lathyrism is postulated.

2-Amino-3-(methylamino)-propanoic acid (BMAA) in cycad flour: an unlikely cause of amyotrophic lateral sclerosis and parkinsonism-dementia of Guam.

We conducted an investigation of the levels of the neurotoxin 2-amino-3-(methylamino)-propanoic acid (BMAA) in cycad flour. Analysis of 30 flour samples processed from the endosperm of Cycas circinalis seeds collected on Guam indicated that more than 87% of the total BMAA content was removed during processing. Furthermore, in 1/2 the samples almost all (greater than 99%) of the total BMAA was removed. We found no significant regional differences in the BMAA content of flour prepared from cycad seeds collected from several villages on Guam. Testing of different samples prepared by the same Chamorro woman over 2 years suggests that the washing procedure probably varies in thoroughness from preparation to preparation but is routinely efficient in removing at least 85% of the total BMAA from all batches. Analysis of a flour sample that had undergone only 24 hours of soaking indicated that this single wash removed 90% of the total BMAA. We conclude that processed cycad flour as prepared by the Chamorros of Guam and Rota contains extremely low levels of BMAA, which are in the order of only 0.005% by weight (mean values for all samples). Thus, even when cycad flour is a dietary staple and eaten regularly, it seems unlikely that these low levels could cause the delayed and widespread neurofibrillary degeneration of nerve cells observed in amyotrophic lateral sclerosis and the parkinsonism-dementia complex of Guam (ALS-PD).