Double-blind placebo controlled trial of the anxiolytic effects of a standardized Echinacea extract.

Earlier studies suggested that specific Echinacea preparations might decrease anxiety. To further study the issue, we performed a double blind, placebo controlled trial with a standardized Echinacea angustifolia root extract. Participants were volunteers scoring above 45 points on the state or on the trait subscale of the State Trait Anxiety Inventory (STAI). They were treated with 40 mg Echinacea or with placebo tablets twice daily for 7 days followed by a 3 week-long washout period. Participants were also administered the Beck Depression Inventory (BDI) and the Perceived Stress Scale (PSS). In the Echinacea group, state anxiety scores decreased by approximately 11 points by the end of the treatment period, whereas the decrease was around 3-points in the placebo group (p< 0.01). The effect maintained over the washout period. The difference from placebo was significant from the 7th day of treatment throughout. Changes were less robust with trait anxiety scores, but the preparation performed better than placebo in patients with high baseline anxiety. Neither BDI nor PSS scores were affected by the treatments. Adverse effects were rare and mild, and all were observed in the placebo group. These findings suggest that particular Echinacea preparations have significant beneficial effects on anxiety in humans.

Individual variability of venom from the European adder (Vipera berus berus) from one locality in Eastern Hungary.

We have revealed intra-population variability among venom samples from several individual European adders (Vipera berus berus) within a defined population in Eastern Hungary. Individual differences in venom pattern were noticed, both gender-specific and age-related, by one-dimensional electrophoresis. Gelatin zymography demonstrated that these individual venoms have different degradation profiles indicating varying protease activity in the specimens from adders of different ages and genders. Some specimens shared a conserved region of substrate degradation, while others had lower or extremely low protease activity. Phospholipase A2 activity of venoms was similar but not identical. Interspecimen diversity of the venom phospholipase A2-spectra (based on the components' molecular masses) was detected by MALDI-TOF MS. The lethal toxicity of venoms (LD50) also showed differences among individual snakes. Extracted venom samples had varying neuromuscular paralysing effect on chick biventer cervicis nerve-muscle preparations. The paralysing effect of venom was lost when calcium in the physiological salt solution was replaced by strontium; indicating that the block of twitch responses to nerve stimulation is associated with the activity of a phospholipase-dependent neurotoxin. In contrast to the studied V. b. berus venoms from different geographical regions so far, this is the first V. b. berus population discovered to have predominantly neurotoxic neuromuscular activity. The relevance of varying venom yields is also discussed. This study demonstrates that individual venom variation among V. b. berus living in particular area of Eastern Hungary might contribute to a wider range of clinical manifestations of V. b. berus envenoming than elsewhere in Europe.

Computer Aided Semi-Automated Evaluation of HER2 Immunodetection--A Robust Solution for Supporting the Accuracy of Anti HER2 Therapy.

HER2-positive breast cancers usually benefit from anti-HER2 therapy, thus, HER2 evaluation became inevitable for patient selection. HER2-negative (IHC 0, 1+) and strong positive (IHC 3+) cases can easily be interpreted with immunohistochemistry, but equivocal (IHC 2+) cases require further analysis of HER2 gene amplification using in situ hybridization. Our study aimed to validate digital pathology and automated image analysis for unbiased evaluation of HER2 immunostains. We developed an image segmentation algorithm for analyzing HER2-immunostaining (4B5 clone) in tissue microarrays of breast cancers. Two pathologists assessed 309 microscopic regions of at least 100 tumor cells each--representing all HER2 positivity groups--according to international guidelines either semi-quantitatively or by using the MembraneQuant software. Scoring results were statistically correlated with each other and with FISH data, and almost perfect agreement was found (inter-method Cohen's kappa = 0.872, Spearman-rho = 0.928). When clinical relevance (scoring disagreement that may define erroneous treatment selection) was examined high agreement was found (quadratic weighted kappa = 0.967). Image analysis classified cases with excellent correlation with visual evaluation, therefore, MembraneQuant software proved to be a reliable tool for assessing HER2 immunoreactions and supporting better targeting anti-HER2 therapy. As digital analysis of immunomorphological markers allows permanent archiving, standardization and accurate reviewing of results, it supports quality assurance initiatives in diagnostic pathology--especially of equivocal cases which are hard to interpret.

Further clinical evidence for the existence of neurotoxicity in a population of the European adder (Vipera berus berus) in eastern Hungary: second authenticated case.

We report a recent case of common adder (Vipera berus) envenoming causing paralytic signs and symptoms. A 12-year-old girl was bitten by the nominate subspecies of the common adder (V. berus berus) in eastern Hungary on May 2, 2012, 22 km away from where the first neurotoxic V. berus berus envenoming was reported in 2008. The patient developed unambiguous cranial nerve disturbances, manifested in bilateral impairment characterized by oculomotor paralysis with partial ptosis, gaze paresis, and diplopia. Drowsiness and photophobia were her additional symptoms; both occurred only during the first day of envenoming. Until now among viper envenomings in Europe, photophobia has only been documented by victims of Vipera aspis. Supportive and symptomatic treatments were administered during 3 days of hospitalization. Although case reports of V. berus berus envenomings are often published, clinical experience with neurotoxicity by this subspecies still remains rare. Population-based and geographic variation of venom composition in V. berus berus seems to include neurotoxic envenomings in certain populations. This second authenticated case provides new clinical evidence for the existence of a possible neurotoxic V. berus berus population in a restricted geographical area in eastern Hungary.

The anxiolytic potential and psychotropic side effects of an echinacea preparation in laboratory animals and healthy volunteers.

We investigated the toxicity, psychotropic side effects and anxiolytic potential of an Echinacea angustifolia extract that produced promising effects in laboratory tests performed earlier. Rats were studied in the elevated plus-maze, conditioned fear, open-field, object recognition and conditioned place preference tests. Toxicity was studied in rats after intragastric administration. The preparation decreased anxiety in the elevated plus-maze and ameliorated contextual conditioned fear. No lethality or behavioural signs of discomfort were noticed in rats treated with 1000 and 3000 mg/kg Echinacea angustifolia. The extract was without effect in tests of locomotion (open-field), memory (object recognition) and rewarding potential (conditioned place preference) within a wide dose range. A pharmacological formulation based on the same E. angustifolia extract was tested in human subjects. One or two tablets per day were administered for 1 week to healthy volunteers scoring high on the State-Trait Anxiety Inventory (STAI). The tablets contained 20 mg of the plant extract. Data were collected using a structured self-assessment diary technique. The high dose (2 tablets per day) decreased STAI scores within 3 days in human subjects, an effect that remained stable for the duration of the treatment (7 days) and for the 2 weeks that followed treatment. The lower dose (1 tablet per day) did not affect anxiety significantly.

[An overview on envenomings inflicted by the Common adder (Vipera berus) and their treatment in Hungary. Facts and beliefs -- part I]. / Górcso alatt a keresztes vipera (Vipera berus) által okozott marások és kezelésük Magyarországon. Tények és hiedelmek - I. rész.

Consequences of bites by the Common adder (Vipera berus) were reviewed in this study. Patients bitten by snakes from different populations may develop variable symptoms due to geographical venom variation. The correct diagnosis of snake bites and the knowledge of the distribution of venomous snake taxa have a crucial impact on snake bite therapy. The characteristic symptoms of patients bitten by V. berus in Hungary are highlighted. The habitat characteristics, seasonal activity and the Hungarian distribution of the adder are described based on literature data, museum specimens and field observations. However, envenomings are uncommon in Hungary, the annual 3 to 4 incidents have to be taken seriously, regardless of the age and actual health condition of the patients. Contrary to beliefs persisting both among laymen and professionals, the venom of V. berus is powerful. Medical observation of the patients is necessary in the first 5 to 6 hours. Any systemic symptom or progression of the edema requires hospital admission.

Clinical picture of envenoming with the Meadow Viper (Vipera (Acridophaga) ursinii).

OBJECTIVE: The vipers in the Vipera (Acridophaga) ursinii complex are small-sized insectivorous snakes found in parts of central and southern Europe. Subspecies include Vipera ursinii ursinii, Vipera ursinii moldavica, Vipera ursinii macrops, Vipera ursinii rakosiensis, and Vipera ursinii graeca and are commonly known as the meadow vipers. These are the least known European Vipera from a clinical point of view. METHODS: We identified cases of V. ursinii envenomations through three methods, including literature search in PubMed, ISI web of Knowldge, JSTOR, Biological Abstracts, Zoological Record, using the various combination of the following terms: snakebite, envenoming, bite, venom, ursinii, meadow viper, steppe viper (in English, French, Italian, Hungarian, Croatian, Serbian, Romanian), review of paper-based medical case records of hospitals in Hungary (four) and Romania (one) covering the 1970-July 2010 period, and personal communications of professional and amateur herpetologists studying V. ursinii and snake-handlers bitten by these snakes. RESULTS: We identified 64 cases from subspecies: V. u. ursinii (14), V. u. moldavica (8), V. u. macrops (5), and V. u. rakosiensis (37). Forty-five bites were collected from the literature, 5 from hospitals, 10 cases were communicated by seven herpetologists and four cases by two snake keepers. Bites were mostly asymptomatic. Forty-five envenomings (70%) resulted in mild and moderate local symptoms, involving pain with low-intensity, pruritus, numbness, swelling with or without erythema and/or local hematoma. Bullae (n = 3, 5%), mild superficial necrosis (n = 4, 6%), cellulitis (n = 1, 2%), and moderately extended edema (n = 8, 13%) of the bitten extremity rarely develop. Massive limb edema was recorded in eight (13%) cases. The most common systemic symptoms were dizziness caused by transient hypotension and tachycardia. Gastrointestinal disorders (i.e. nausea, vomiting) were rare (n = 2, 3%) compared to other Vipera, and probably triggered only by anxiety. Symptomatic and supportive therapy was applied in the relatively severe envenomings and antivenom therapy in six cases. Complete recovery ranged from 12 h to 2 weeks. Moderate and severe envenomings required significantly longer recovery. Application of first aid was associated with significantly longer recovery times. Neither the age (i.e. juvenile or adult) of the culprit specimen, nor the anatomical location of the bite determined the severity of symptoms. CONCLUSION: Professional and amateur herpetologists, and snake keepers are mainly at risk. Most V. ursinii bites do not require first aid or medical intervention, since only local symptoms develop and resolve spontaneously. The rare hospitalized cases require symptomatic and supportive treatment only. Antivenom therapy is not indicated.

Technical note on the validation of a semi-automated image analysis software application for estrogen and progesterone receptor detection in breast cancer.

BACKGROUND: The immunohistochemical detection of estrogen (ER) and progesterone (PR) receptors in breast cancer is routinely used for prognostic and predictive testing. Whole slide digitalization supported by dedicated software tools allows quantization of the image objects (e.g. cell membrane, nuclei) and an unbiased analysis of immunostaining results. Validation studies of image analysis applications for the detection of ER and PR in breast cancer specimens provided strong concordance between the pathologist's manual assessment of slides and scoring performed using different software applications. METHODS: The effectiveness of two connected semi-automated image analysis software (NuclearQuant v. 1.13 application for Pannoramic™ Viewer v. 1.14) for determination of ER and PR status in formalin-fixed paraffin embedded breast cancer specimens immunostained with the automated Leica Bond Max system was studied. First the detection algorithm was calibrated to the scores provided an independent assessors (pathologist), using selected areas from 38 small digital slides (created from 16 cases) containing a mean number of 195 cells. Each cell was manually marked and scored according to the Allred-system combining frequency and intensity scores. The performance of the calibrated algorithm was tested on 16 cases (14 invasive ductal carcinoma, 2 invasive lobular carcinoma) against the pathologist's manual scoring of digital slides. RESULTS: The detection was calibrated to 87 percent object detection agreement and almost perfect Total Score agreement (Cohen's kappa 0.859, quadratic weighted kappa 0.986) from slight or moderate agreement at the start of the study, using the un-calibrated algorithm. The performance of the application was tested against the pathologist's manual scoring of digital slides on 53 regions of interest of 16 ER and PR slides covering all positivity ranges, and the quadratic weighted kappa provided almost perfect agreement (κ = 0.981) among the two scoring schemes. CONCLUSIONS: NuclearQuant v. 1.13 application for Pannoramic™ Viewer v. 1.14 software application proved to be a reliable image analysis tool for pathologists testing ER and PR status in breast cancer.

First clinical experiences about the neurotoxic envenomings inflicted by lowland populations of the Balkan adder, Vipera berus bosniensis.

The first overall clinical description of envenomings by the lowland populations of the Balkan adder (Vipera berus bosniensis) is provided by this study. Fifty-four incidents have been collected retrospectively from the south-western Hungarian and the northern Croatian distribution area of the taxon. There were five (9%) asymptomatic, 24 (44%) mild, 12 (22%) moderate, 12 (22%) severe, and one fatal (2%) case according to the Poisoning Severity Score. The single death is a 60-year-old Hungarian case that was caused by V.b. bosniensis. Average hospitalisation was 2.75 days. The most common systemic symptoms were gastrointestinal disorders, ECG changes, persisting hypotension and neurological disorders. The initial phase of neurotoxic manifestations was always expressed in cranial nerve disturbances: ptosis, external ophthalmoplegia, diplopia, reduced focusing capability and blurred vision. Neuromuscular paralysis progressed to dyspnoea and lower limb paralysis in the most severe cases. Unusual symptoms were fluctuating arterial hypertension, drowsiness, and hypokalaemia. Laboratory results reveal leucocytosis, while deviation in the other laboratory values is not common. Envenomings by V.b. bosniensis significantly differ from those by the European adder (Vipera berus berus) in lower manifestation rate of extensive oedema, anaemia, CNS depression, and haematuria but the development of neuromuscular paralysis is high (20%). Their bites rather resulted in mild and moderate local symptoms in envenomed patients than those inflicted by the nominate form. This study presents the evidence of the frequent neurotoxic manifestations in Balkan adder-bitten patients for the first time, which strongly suggests that the venom of the lowland populations of V.b. bosniensis has neurotoxic activity.

Snakebites in Hungary--epidemiological and clinical aspects over the past 36 years.

Epidemiological and clinical aspects of snakebites in Hungary between 1970 and 2006 were surveyed. A total of 97 cases were recorded from 21 species, including the two native vipers, Vipera berus and Vipera ursinii, and various exotic species represented by Viperidae, Elapidae, and Colubridae. Bites by native species on laymen are uncommon (17 cases) and present trivial clinical manifestations. Compared with the consequences of native Vipera cases, bites by exotic species often resulted in severe or life-threatening envenomations. These cases were treated with antivenom administration, plasmapheresis, fasciotomy, and amputation. There were two fatalities caused by V. berus and Agkistrodon contortrix. Both of these cases were inflicted in snake-handlers with a previous history of Viperidae bites and the cause of deaths are attributed to anaphylactic reactions as a consequence of hypersensitivity to the venom. Snake-handlers and their physicians face a major challenge due to the diversity and severity of signs and symptoms following exotic venomous snakebites, and the risk of anaphylaxis or anaphylactoid reactions in patients with repeated exposure to snake venom and antivenom. Highly dangerous venomous snake species continue to appear in collections of Hungarian snake-handlers.

Clinical aspects and consequences of envenoming by a captive rhinoceros viper (Bitis nasicornis) in Hungary.

A case of Rhinoceros viper (Bitis nasicornis) bite is reported. The bitten hand distended to the wrist, which was tense and painful, with only mild local livid discolouration manifested around the fang mark. Slight hypertension with moderate tachycardia and temporary coagulopathy were observed. The patient received analgesic and intravenous fluids, antibiotics and anti-tetanus therapy. Use of antiserum was not necessary. The bitten person was treated in the main centre for snakebite first aid: in the Toxicological Ward of Erzsébet Hospital of Budapest. We attach importance to the implications of this case because envenoming by B. nasicornis being relatively rare in captivity all over the world (particularly in Europe and the USA), as well as in the wilderness in Africa.