Breast cancer patients with hypermethylation in the promoter of BRCA1 gene exhibit favorable clinical status.

Promoter hypermethylation was shown to be involved in human cancerogenesis through silencing gene expression. Several studies were dedicated to explore the frequency and clinical significance of BRCA1 hypermethylation in sporadic breast cancer to identify a specific molecular and clinico-pathological phenotype. However the available data are limited and rather too heterogeneous. In this study we investigated the level of methylation in the promoter region of BRCA1 and its correlation with clinico-pathological and molecular characteristics in a group of 135 Bulgarian patients. Methylation specific PCR was applied to determine methylation status of tumor samples. Clinical impact of BRCA1 hypermethylation was estimated using standard statistical methods including Fisher's exact and the Chi-squared tests, the Kaplan-Meier method, the univariate and multivariate Cox proportional hazards regression model. We found that hypermethylation was present in 17.04% of the cases (23/135). Patients with hypermethylation in BRCA1 displayed favorable clinical status as their tumors were smaller in size (P = 0.066), lacked p53 gene mutations (P = 0.073) and were of lobular type (P = 0.046). The presence of hypermethylation was weakly associated with better overall survival (P = 0.2) with a hazard ratio of 0.47 (95% CI 0.14-1.54, P= 0.213). Our study provides the first data on the BRCA1 hypermethylation of Bulgarian patients and contributes to elucidation of its clinical significance in sporadic breast cancer.

[The toxicological characteristics of ammonium glycyrrhizinate (glycyram). A study of its acute and subacute toxicity]. / Toksikologicheskaia kharakteristika glitsirizinata ammoniia (glitsirama). Issledovanie ostroi i podostroi toksichnosti.

A single parenteral and oral administration of ammonium glycyrrhizinate in rat and mice experiments showed that the compound is related to practically nontoxic drugs. Its repeated administration (30 times) into the stomach in a maximum daily therapeutic dose (7 mg/kg) and in a four-fold dose (28 mg/kg) did not cause signs of intoxication, essential changes in the hematological and integral parameters, shifts in the activity of serum enzymes, morphological changes in the cell structures of the internal organs. Administration of the drug in a dose of 28 mg/kg for a second time led to changes in the activity of some enzymes in the brain, the development of parenchymatous dystrophy of the liver which changed to acidophilic necrosis attended with signs of regeneration. Under conditions of a subacute experiment the maximum daily therapeutic dose of ammonium glycyrrhizinate may be considered practically nontoxic.

[The toxicological characteristics of Gastrofenzin. IV. Its gonadotropic action]. / Toksikologichna kharakteristika na "Gastrofenzin". IV. Gonadotropno deistvie.

An evaluation of the gonadotropic effect of Gastrofenzin has been carried out in conditions of 45-day and 90-day oral application (daily, 5 times per week) in male white rats. The experiments are performed in doses 1/20, 1/100 and 1/200 LD50, respectively 33.8, 6.7 and 3.4 mg.kg-1. Functional and morphologic methods of investigation have been used-count, character and mobility, acid and osmotic resistance of the sperm cells, mean number of sperm cells in the sperm tube, spermatogenesis' indices, morphological characteristic of the testes as well as autoradiographic method for determining the proteolytic activity of the acrosome. Irrespective of the application term the highest dose (33.8 mg.kg-1) causes systemic deviations in the functional indicators and physiologic fluctuation of the morphologic parameters characterizing the spermatogenic epithelium at preserved proteolytic activity of the acrosome. The dose 1/20 LD50 (33.8 mg.kg-1) is considered to be effective on the gonads in conditions of three-month chronic oral application of Gastrofensin and the threshold of the gonadotoxic activity is about 1/100 LD50-6.7 mg.kg-1.

[The toxicological characteristics of Gastrofenzin. I. Its acute toxicity (oral, subcutaneous and intravenous)]. / Toksikologichna kharakteristika na "Gastrofenzin". I. Ostra toksichnost (oralna, podkozhna, intravenozna).

The acute toxicity of the antiulcer drug "Gastrofenzin", synthesized in Bulgaria has been studied in oral, subcutaneous and intravenous application in white rats "Wistar" and white mice "ICR". The main toxicometric parameters (LD50, LD16, LD84 and others) have been determined. The clinical picture of intoxication is characterized mainly by symptoms deriving from in the central and vegetative nerve system. According to the parameters of acute oral toxicity (LD50 for male white rats is 665.0 mg.kg-1 and for female--876 mg.kg-1) and to the classification of Hodge & Stemer Gastrofenzin refers to the group of slightly toxic drugs. The LD50 in subcutaneous application is 938.0 mg.kg-1 for the male and 891.0 mg.kg-1 for the female rats. For the intravenous application LD50 is 50,1 mg.kg-1 for the male and 43.6 mg.kg-1 for the female rats. The coefficient of lethal intoxication danger is below 0.1 in the three ways of application which confirms its status according to the upper classification. A significant sex difference in the indicators of acute oral toxicity for the white rats and white mice has not been observed. The white mice of both sexes seem to be more sensitive to the drugs' effects than the white rats.

[The toxicological characteristics of Gastrofenzin. II. Its subacute oral toxicity]. / Toksikologichna kharakteristika na "Gastrofenzin". II. Podostra oralna toksichnost.

The subacute (30 day) oral toxicity of Gastrofenzin has been investigated in white rats "Wistar" of both sexes, treated with 0.7 and 33 mg.kg-1 for the male and 0.9 and 44 mg.kg-1 for the female rats. The applied doses respond respectively to the control doses, 1/100 and 1/20 LD50 of the preparation. A complex of integral, behaviour, laboratory biochemical and histological methods has been used. The findings show that Gastrofenzin in doses 1/100 and 1/20 LD50 does not exert cumulative effect, reported by the lack of lethal effect among the animals. With the help of actograph "AIDA" an increased spontaneous activity is reported in the rats from both sexes treated with dose 1/20 LD50 of the preparation. Under the experimental conditions the doses of 7 and 9 mg.kg-1 (1/100 LD50) and 33 and 44 mg.kg-1 (1/20 LD50) do not exert toxic effect on the liver, kidneys and brain. Deviations are observed which show increasing of the oxidative-metabolic processes and trigger adaptation mechanisms in the above mentioned organs.

[The toxicological characteristics of Gastrofenzin. III. Its chronic oral toxicity]. / Toksikologichna kharakteristika na "Gastrofenzin". III. Khronichna oralna toksichnost.

A risk evaluation has been carried out under conditions of chronic three-month experiment in oral application of Gastrofenzin in doses 1/20, 1/100 and 1/200 LD50 for male and female white rats, respectively 33.2 and 43.8; 6.7 and 8.8; 3.4 and 4.4 mg.kg-1. There have been applied integral, behavioural, laboratory, biochemical (serum, liver, kidney, brain, brain mitochondria), histologic and electron microscopic methods. The highest doses cause functional and biochemical changes in the nerve system, liver and the kidneys without involving the structural elements of the respective tissues. Significant changes have not been observed in the doses 1/100 LD50 and 1/200 LD50. A dose-effect increase has been found out in the level of cytochrome-P-450 in the liver tissue, but the data are insufficient for determining the type of the caused induction. The results of the complex study show that in the three-month application in doses 1/20, 1/100 and 1/200 LD50 Gastrofenzin does not cause development of significant deviation in the organism of the mice from both sexes and its toxicological characteristic is comparatively favourable.

[The assessment of the occupational risk to workers in electric steel production]. / Otsenka na professionalniia risk na rabotnitsite ot elektrostomanodobiva.

These studies, centering on occupational environmental factors, biologic monitoring, and toxicodynamic investigations, involved a total of 105 workers distributed into eight job groups, who were 45 years, of age on the average and had from 5 to 10 years of special occupational experience at "Electrosteel" Works. Evidence was obtained for presence of unfavorable microclimate conditions, elevated equivalent levels of noise, excess of general and local vibrations, exposure to manganese aerosols, carbon monoxide, and nitrogen oxides; some of the subjects and deviations in hepatic status and connective tissue. Job groups found to be at risk were those of steel founders, crane workers, and pourers.

[The toxic hygiene problems of workers in the Dinamik automobile tire plant in Sofia]. / Toksikokhigienni problemi pri raboteshtite v zavod za avtomobilni gumi "Dinamik"--Sofiia.

Toxic hygienic investigations were carried out involving a representative group of 131 workers (50 females and 81 males) distributed into three main workshops--preparatory, confection, and vulcanization--at "Dynamic" automobile tire plant in Sofia. The majority of male and female workers were in the age range beyond 40 years, having a general occupational experience in excess of 10 years and specialized occupational experience from 10 to 20 years or more. The chemical hazard was among the leading ones in the occupational environment, differing in nature according to the technologies used. Included were various chemical substances and compounds: synthetic rubbers, fillers (soot, chalk, kaolin); softeners (mazut, paraffin, etc.); accelerants (mercaptothiazoles--captax and altax); dithiocarbamates (thiuram); vulcacides (diphenylguanidine); antiwear agents (antioxidants and antiozonators-isopropyl-phenyl-paraphenylene diamine, naphthyl-beta-naphthylamine); antiaccelerants (phthalic anhydride, ets.); organic solvents and others. The indicated chemical substances and compounds, though often found at concentrations below the mean-shift MACs, do produce health impacts by virtue of prolonged and combined exposures. Use was made of the questionnaire method. Also, hematologic, clinical laboratory, and toxic chemical testing was performed. Findings pointed to changes in hemopoiesis, deviations in hepatic functional state, while sulfate and glucuronide values confirmed the workers' high exposures.

[The cytogenetic effects (the frequency of micronuclei) in lymphocyte cultures from the peripheral blood of workers in automobile tire manufacture]. / Tsitogenetichni (chestota na mikronukleusi) v limfotsitni kulturi ot periferna kruv na rabotnitsite ot proizvodstvoto na avtomobilni gumi.

At the Preparatory Workshop of the Plant for Automobile Tires (PAT), Sofia, complex investigations were undertaken to reveal possible genotoxic exposure. The studies included chemical analyses for levels of identifiable human carcinogens in the occupational ambient air (benz(a)pyrene, mineral oils, 2-naphthylamine); special techniques--questionnaire investigations and cytogenetic analysis by cytokinesis-block micronucleus method in peripheral lymphocyte cultures from 23 workers in occupational groups at risk; and urinalysis for contents of mercapturic acids. An out-of-house control group of 13 nonexposed subjects was concurrently investigated. For contents of benz(a)pyrene and mineral oils exceeding 2.5 to 3.5 times the respective occupational environment MACs, evidence from cytogenetic analysis showed substantial, 4-fold, increase in indexes of genotoxic impairment (frequency of micronucleated-binucleated lymphocytes, number of micronuclei per 1000 binucleated lymphocytes) in the workers investigated. These results are an indicator of genotoxic exposure and point to an increased potential risk of cancer development in the group of workers studied.

[An evaluation of the biological action of carbon disulfide in clinico-laboratory studies of exposed workers]. / Otsenka na biologichnoto deistvie na serovugleroda chrez kliniko-laboratorni izsledvaniia na eksponirani rabotnitsi.

A representative group of 122 workers from shop "Predilen" with 7 to 15 years length of service in specialty is taken. The professional groups are formed on the basis of measured average-shift concentrations of carbon bisulphide in the air of the working place, which vary from 4 to 50 mg.m-3. In order to establish deviations in some of the basic links of the biochemical mechanism of the toxic compounds effect the following indices are traced: copper metabolism [copper concentration and activity of ceruloplasmin in the serum]; lipidic metabolism [content of cholesterol, cholesterol of high and low density, triglycerides, beta-lipoproteins and nonesterified lipidic acids in the serum]; intercellular substance of the connective tissue [glycoproteins, glycosaminoglycans and hexauronic acids in serum and urine]; immune status [immunoglobulins A, G and M in serum]; peripheral blood [leucocytes, DKK, morphology and lipid content in granulocytes]. Decrease of the lipids in granulocytes is registered, as well as reduction of the ceruloplasmin activity, increase of immunoglobulin M and glycosaminoglycans in urine. The already established deviations follow the "dose-effect" and "dose-response" dependence.

[A comparative evaluation of the toxic and cumulative properties of the Takal drug group]. / Sravnitelna otsenka na toksichnite i kumulativnite svoistva na lekarstvenata grupa "Takali".

The acute oral and dermal toxicity was studied as well as subacute oral toxicity [30 successive days] and skin irritating effect on experimental animals of new drug form "Takal 9", "Takal 14" and "Takal 16" with proved bactericidal and virucidal activity. A complex of toxicometric, unspecific, haematological, biochemical and histological studies were performed. The compositions "Takal 9", "Takal 14" and "Takal 16" are slightly toxic at oral and dermal application. LD50 for "Takal 9" and "Takal 16" is above 15,000 mg/kg-1 while for preparation "Takal 14" is not reached. At unrepeated dermal application LD50 for the examined group of preparations surpasses 5000 mg/kg-1. The products have no skin-irritative effect. No cumulative effect is established in the conditions of a 30-day oral treatment. On the background of no changes in the serum indices an activation of the aerobic and anaerobic oxidation in the liver is registered in animals treated with ethyl alcohol, "Takal 9" and "Takal 16". These deviations are due, most probably, to the participation of ethyl alcohol as solvent, in "Takal 9" and "Takal 16". On the basis of the experiments, as most suitable for use, is established the preparation "Takal 14".

Experimental study of subacute oral, dermal and inhalation toxicity of bulmoscide preparation.

The purpose of this study is to give the toxicological characteristics of subacute oral, dermal and inhalation exposure of microbial preparation Bulmoscide, which is based on bacterium Bacillus thuringiensis, serotype H-14. The study was performed on sexually mature Wistar rats of both sexes. No significant changes in non-specific, clinico-laboratory and biochemical parameters as well as morphological examinations in any tested groups compared to the control groups were revealed. The doses 110 mg.kg-1 (1.32 x 10(9) sp.kg-1) at oral administration in duration of 90 days, 3000 mg.kg-1 (3.6 x 10(10) sp.kg-1) in 21 day dermal application and maximum attainable concentration of dust aerosol 18 mg.kg-3 (2.2 x 10(8) sp.m-3) during a 30 day inhalation exposure are "no toxic effect" levels. On the basis of the data, the selective bacterial insecticide Bulmoscide has been determined to be of low toxic and low hazardous preparation in compliance with the Hygiene Classification of pesticides.

[The acute toxicity of a bacterial insecticidal preparation containing a beta-exotoxin]. / Prouchvane na ostrata toksichnost na bakterialen insektitsiden preparat, sudurzhasht beta-ekzotoksin.

The acute toxicity of a new Bulgarian bioinsecticide preparation is determined, produced on the basis of Bacillus thuringiensis containing 0.1% beta-exotoxin by oral, dermal, inhalational and intraperitoneal introduction in experimental animals. The evaluation of the toxicity is performed by a complex of toxicometric, integral, haematologic and pathologicoanatomic methods according to the WHO and USA criteria for studying the safety of the microbic agents for plant protection. It is already established that the preparation introduced orally in dose 10,000 mg.kg-1 (1.6.10(11) cell kg.-1 10 micrograms.kg-1 beta exotoxin), applied dermally in dose 6000 mg.kg-1 (9.6.10(10), 6 micrograms.kg-1 beta exotoxin) and in concentration 300 mg.m-3 (4.8.10(10) cell m-3, 300 micrograms.m-3 beta exotoxin) provokes no lethality, intoxication and changes in the integral, haematologic and pathologicoanatomic studies of test animals. LD50 at intraperitoneal introduction in white rats is 387.20 mg (6.2.10(10), kg-1, 387.2 micrograms kg-1 beta exotoxin) for male and 364.0 mg kg-1 (0.58.10(9) kg-1 364.0 micrograms kg-1 beta exotoxin) for female animals. The investigations point out that according to rate of acute toxicity the bacterial preparation containing 0.1% beta exotoxin is referred to the low toxic substances and reveals no danger for acute oral, dermal and inhalational poisonings when the regulations for production and use are observed.

Urinary excretion of glycosaminoglycans in patients with postmenopausal osteoporosis.

The organic bone matrix contains glycosaminoglycans (GAG) of which the precise function and importance in bone mineralisation are still unclear. We examined 85 persons--35 healthy women (25 premenopausal [preMP] mean aged 40.7 years; 10 menopausal [MP] mean aged 59.3 years) and 50 patients with postmenopausal osteoporosis [PMOP] at a mean age 60.4 years. The dynamic of urinary excretion of GAG was measured in 24-hour collected urine by precipitation with cetylpyridinum chloride and spectrophotometry at 560 nm, corrected for the level of excretion of creatinine. There was a significant increase in GAG excretion in patients with PMOP compared with healthy persons (8.25 mg/g and 9.53 mg/g vs 24.11 mg/g; p < 0.0001). A significant positive correlation was established between GAG and calcium urinary excretion and a negative one between GAG and serum estradiol levels. During the treatment with calcitonin the excretion of GAG was decreased which can be used for monitoring the changes of bone metabolism.

[An experimental evaluation of the acute and chronic oral toxicity of the antibiotic Bactericin for plant protection]. / Eksperimentalna otsenka na ostrata i khronichnata oralna toksichnost na antibiotika "Bakteritsin" za rastitelna zashtita.

In relation to the future implementation of the antibiotic bactericin in agriculture are carried out acute, subacute and chronic oral experiments on 128 male white rats and 88 female. In order to determine the parameters of acute oral toxicity, doses (200, 120, 80, 60, 50, 40 mg/kg) of the preparation dissolved in polyethylene glycol are applied. In the subacute--1/10 and 1/20 LD50, and in the chronic (four months)--1/50 and 1/100 LD50. The bactericin, meant for treatment of tomatoes before sowing, in rate of acute oral toxicity falls in class II of very toxic pesticides. The acute oral LD50 for male white rats is 73.0 (48.6-109.5) mg/kg-1 and for female--58.0 (39.9-84.0) mg/kg-1. In conditions of subacute (30 oral applications for 45 days) and chronic (four months) experiments no cumulative properties are proved as well as hepatotoxic, nephrotoxic and neurotoxic effect. On the basis of the complex study could be accepted, that the dose 1/100 LD50 (0.7 mg/kg-1) for male rats and 0.4 mg/kg-1 for female) is not active at chronic oral introduction.

[Toxicologic evaluation of propyneb on the Wistar rat]. / Evaluation toxicologique du propyneb chez le rat Wistar.

This experiment was performed in Wistar rats of both sexes exposed subchronically to 1:100, 1:500, 1:1000 and 1:1500 LD50. The evaluation was based on endpoints measured on the 30th and 90th after starting exposure and after a recovery period of 30 days: these included clinical signs, functional changes, hematological parameters, urine analysis, biochemical, histochemical, immunomorphological endpoints, electron microscopy of internal organs, chromosome examination of bone marrow. A high lethality was shown to occur with a characteristic clinical picture: interruption of weight gain, behavioural changes, leucopenia mainly involving neutrophil leucocytes, biochemical changes characteristic of liver, cardio-vascular system (myocardium and aorta) together with pathologic, biochemical, histochemical and ultrastructural changes in liver, brain, thyroid gland, myocardium, spleen and bone marrow. Endpoints were shown to be clearly dose-dependently related with small variations with the low dose, i.e. 1:1500 LD50 (5 mg/kg-1 bw).

[Biochemical and histological changes after acute oral poisoning with the acetanilide herbicide acetochlor]. / Modifications biochimiques et histologiques de l'intoxication orale aiguë par l'herbicide acétanilidique "acétochlore".

Changes in the liver of male rats were studied on the 1, 3, 5, 7, 10 and 14 days after a single oral application of 1/5 DL 50 of the chloracetanilic herbicide Acetochlor (DL50 = 1063 mg/kg-1). Two main periods in the action of acetochlor can be identified: between 1-2 days when the influence of the compound itself is predominant and between 5-7 days, when the toxic action of the slowly eliminated metabolites is most pronounced. These two periods provoke a biphasic activation/inhibition response of the liver. Remarkable is the compensatory effect of the cytochrome P-450 system responsible for the metabolism of acetochlor. The phase-specific changes were are followed by pathomorphological observations of liver tissue as well as by the time-course of the biochemical parameters studied in different liver fractions.

[The effect of the acetanilide herbicide Acetochlor on the cardiovascular system of white rats]. / Efekt na atsetanilidniia kherbitsid "Atsetokhlor" vurkhu surdechno-sudovata sistema na beli plukhove.

A study is performed on the long-term effect of the chloracetanilide herbicide "Acetochlor" in doses 21.0; 10.6: 5.5 and 2.6 mg/kg-1 in conditions of 6-month oral application and 2-month rehabilitation period on the metabolite processes and the balance of the connective-tissue components in the myocardium and aorta of male white rats. A complex of biochemical and histological methods are performed (activity of succinate dehidrogenase, lactate dehydrogenase, glucose-6-phosphate dehydrogenase, adenosin triphosphatase, cytochrome oxidase, fructoso-1,6-diphosphatase, level of the thiol groups, soluble globular, elastine, collagen fractions, insoluble collagen and elastine, general and sulphated glucosamino glycanes). The dose 21.0 mg/kg-1 leads to blocking of the thyol groups, inactivation of succinate dehydrogenase, cytochrome oxidase, adenosin triphosphatase, fructose-1,6-diphosphatase, glucose-6-phosphate dehydrogenase, activation of lactate dehydrogenase, decrease of the soluble globular, elastine, and collagen fractions and increase of the glucoseaminoglycanes in the heart muscle and aorta. The changes established in the heart muscle at 10,6 mg/kg-1 certify for stronger sensitivity of the organ of the aorta wall. The presence of single changes in the examined indices, their complete dying out after the rehabilitation period and absence of structural changes in the myocardium and aorta permit the dose of 5.5 mg/kg-1 to be accepted as not effective in the conditions of chronic experiment concerning the cardiovascular system.

[The hygienic toxicological evaluation of the viral preparation Mamestrin]. / Khigienno-toksikologichna otsenka na virusniia preparat "Mamestrin".

The preparation "Mamestrine" developed in Bulgaria on the basis of NPV virus is meant for control of the Heliothis zea. Experiments are carried out on the toxic effect of the preparation, according to the requirements of FAO/WHO for assessment of the virus preparations and the acting in this country Bulgarian State Standards. On experimental animals (white rats of both sexes, white guinea pigs and white rabbits) are studied acute oral, dermal and inhalatory toxicity, subacute oral toxicity, dermal-irritation, eye-stimulation and skin-sensitizing activity, cytotoxicity and activity of the preparation in soils and waters. Toxicometric, integral, clinical and laboratory, biochemical and histological methods are used. Unrepeated oral application of dose 2.10(9) pol.kg-1, unrepeated dermal application of dose 1.10(9) pol.kg-1, in white rats and 0,4.10(7) pol.kg-1 in guinea pigs and 4 hrs inhalation unrepeated exposure of white rats to concentration 1.10(7) pol.m-3 provoke no signs of intoxication and lethal issue and are accepted as inoperative. During a 3-month oral application of the preparation in doses 1.10(7) and 1.10(9) pol.kg-1 are established no data for toxic effect on the organism of experimental white rats of both sexes. The preparation "Mamestrine" has no skin-irritative, eye-irritative, eye-stimulative and skin-sensitizing activity. There is no cytotoxic effect on animal (kidney of monkey) and human (carcinoma of the gullet) cellular cultures.(ABSTRACT TRUNCATED AT 250 WORDS)