Prevalence of subclinical uterine pathologies diagnosed by biopsy and cytological and bacteriological findings in cyclic bitches.

The study was performed on 45 bitches in different cycle phases that were divided into the following groups: anoestrus (I, n=15), heat (pro-oestrus (n=7) or estrous (n=8) (II, n=15) and metestrus (III, n=15). Moreover, all experimental dogs were grouped according to their age: younger than 5 years (Y, n=35) and older than 5 years (O, n=10). The endometrial status was evaluated using cytological, bacteriological and biopsy samples obtained after ovariohysterectomy. The main uterine pathology diagnosed by biopsy was endometritis, since 40%-66% of bitches, independent of the experimental group, developed this condition. No significant differences were found among the cycle phase groups (p>0.05). By contrast, significant differences were found in the age groups; the prevalence of this pathology was higher in older bitches (p=0.0019). The general prevalence of cystic endometrial hyperplasia (CEH) and a normal endometrium (NE) was lower (6.7-26.7% vs 26.7-53.3%) in all groups, and no statistically significant differences were found between certain groups (p>0.05). The percentage of polymorphonuclear neutrophilic leukocytes (PMNs) in endometrial cytology was generally low (⟨ 2%) and did not differ significantly among the experimental groups (p=0.142). In general, a low degree of correlation was found between the diagnostic results by endometrial cytology and biopsy (Kappa Coefficient= 0.046). Positive bacteriological findings were found in approximately 50% of the bitches, independent of the cycle phase and health status of the endometrium. No correlation was found between the bacteriological and histopathological findings (p=0.883). In conclusion, uterine cytology is not a reliable diagnostic method to detect the subclinical inflammatory and degenerative uterine pathologies in cyclic bitches.

Equine Chorionic Gonadotropin and Human Chorionic Gonadotropin Stimulation Increase the Number of Luteinized Follicles and the Progesterone Level Compared with Cabergoline Stimulation in Anoestrus Bitches.

In this study, ovarian morphologies and blood progesterone concentrations following oestrous induction in bitches were examined. Fifty-three clinically healthy anoestrus bitches received cabergoline at a daily dose of 5 µg/kg of body weight per os for 21 days (group I) or subcutaneous equine chorionic gonadotropin at a dose of 20 IU/kg of body weight for five consecutive days with an additional 500 IU s.c. per bitch of human chorionic gonadotropin on the last day of treatment (group II). Twenty bitches that spontaneously displayed oestrous signs were left untreated and served as controls (group III). The induced oestrous rates and ovulation rates in groups I and II were 60.0% vs 64.3% and 86.7% vs 83.3%, respectively. Morphological assessments of the ovarian structures after ovariohysterectomy revealed an increase in the number of luteinized follicles and cysts in group II compared with the two other groups (p < 0.001). In contrast, the numbers of corpora lutea and follicles were similar in all groups. In accordance with the above-mentioned alteration, the progesterone concentration in the gonadotropin group (II) was increased (p < 0.001) in the periovulatory period compared with the other two groups. During the entire sampling period, the progesterone profiles in the cabergoline (I) and control (III) groups were similar and typical of normally cycling bitches. In conclusion, gonadotropin treatment is associated with an increased progesterone level during the periovulatory period that probably originates from luteinized follicles, whereas cabergoline treatment induces cycles with both physiological progesterone concentrations and ovarian morphologies.

Evaluation of the effect of 2,4-dichlorophenol on oxidative parameters and viability of human blood mononuclear cells (in vitro).

2,4-Dichlorophenol (2,4-DCP) is formed in drinking water as a result of its chlorination, and it is created in the environment during transformation of various xenobiotics such as triclosan or herbicide 2,4-dichlorophenoxyacetic acid (2,4-D). The molecular mechanism depicting the action of phenolic compounds on nucleated blood cells has been insufficiently studied, and therefore, we have assessed the effect of 2,4-DCP on the structure and viability of human peripheral blood mononuclear cells (PBMCs). We have evaluated necrotic, apoptotic, and morphological changes (alterations in the size and granulation) in PBMCs incubated with 2,4-DCP in the concentration ranging from 10 to 500 µg mL(-1) for 4 h at 37°C. Moreover, we have estimated changes in reactive oxygen species (ROS) formation, lipid peroxidation, and protein carbonylation in the incubated cells. We have noted that 2,4-DCP increased ROS formation and lipid peroxidation (from 10 µg mL(-1)) and oxidized proteins (from 50 µg mL(-1)) in PBMCs. The compound studied also provoked apoptotic (from 50 µg mL(-1)), necrotic (from 100 µg mL(-1)) and alterations in the size and granulation (from 50 µg mL(-1)) in the incubated cells. The analysis of quinolinium 4-[(3-methyl-2(3H)-benzoxazolylidene)methyl]-1-[3-(trimethyl-ammonio)-propyl]-diiodide/propidium iodide staining revealed that 2,4-DCP (50-250 µg mL(-1)) more strongly increased the number of apoptotic than necrotic cells, which suggests that this cell death type is mainly provoked by this compound in PBMCs. The observed changes were caused by relatively high concentrations of 2,4-DCP, which cannot influence human organism during environmental exposure and thus may only occur as a result of acute or subacute poisoning with this compound.

Extremely long time stability study of selected antibiotic standards.

A study was conducted to investigate the quality of reference substances which were produced long before the introduction of advanced analysis and purification techniques. The quality and antimicrobial activity of WHO and EP oxytetracycline, doxycycline, colistin and spiramycin reference standards were assessed. The oldest substance was stored for 54 years in a freezer. Assay and purity tests were conducted according to Ph. Eur. 6.0. Additionally, antibacterial activity was tested with the microbiological method according to Ph. Eur. 4.0 (agar diffusion method). The results of the study show that several of the tested substances remained stable for over 40 years and one for over 50 years of storage. In most cases, the determined potency is close to the declared one, regardless of the method used (HPLC or microbiological). Composition analysis of multi-substance antibiotics (colistin and spiramycin) showed important differences compared with new reference substances. Results also indicate that no excessive degradation occurred during the entire storage period and impurity levels have not changed significantly.

Comparison of the effect of phenol and its derivatives on protein and free radical formation in human erythrocytes (in vitro).

The effect of phenolic compounds: phenol, 2,4-dichlorophenol (2,4-DCP), 2,4-dimethylphenol (2,4-DMP) and catechol on human erythrocytes was studied. The level of fluorescent label - 6-carboxy-2',7'-dichlorodihydrofluorescein diacetate (H(2)DCFDA) oxidation by phenolic compounds in erythrocytes as well as the carbonyl group content and hemoglobin denaturation were monitored. H(2)DCFDA has been utilized extensively as a marker for studies of oxidative stress at the cellular level. We noted that 2,4-DCP, 2,4-DMP and catechol induced an increase in the concentration- and time-dependent H(2)DCFDA oxidation. We also observed an increase in carbonyl group content and the changes in parameter T (denaturation of hemoglobin) in erythrocytes incubated with 2,4-DCP, catechol and 2,4-DMP. The highest level of H(2)DCFDA oxidation was provoked by 2,4-DCP. The biggest changes of proteins in erythrocytes measured as the carbonyl group content were induced by 2,4-DMP, but measured as parameter T they were induced by catechol. It was observed that phenol did not oxidize H(2)DCFDA up to the concentration of 2.5 mM after 3 h of incubation. Phenol did not affect the carbonyl group content but decreased parameter T (induced denaturation of hemoglobin). To sum up, the kind of the substituent in a phenolic ring determines the molecular mechanism of action of the individual compound and the capacity of reactive oxygen species generation and thus damages the specified structures in human erythrocytes.

The influence of 2-chlorodeoxyadenosine (2-CdA) on the adenine energy charge and glutathione content of human erythrocytes.

We have examined the effect of exposure of human erythrocytes to the new chemotherapy drug 2-chlorodeoxyadenosine (2-CdA, cladribine), focusing on the glutathione (GSH and GSSG) content and the adenine energy charge (AEC). Incubation of erythrocytes with 0.1-5 microg/ml 2-CdA induced no significant change in the reduced or total glutathione level or in the AMP and ATP concentrations. The ADP concentration increased slightly and the AEC value is in the range typical of healthy organisms. Incubation of erythrocytes with 2-CdA also caused cell shape changes, converting most of the cells to echinocytes.

Catalase activity in human erythrocytes: effect of phenoxyherbicides and their metabolites.

The effects of exposure to different concentrations of phenoxyherbicides and their metabolites were studied in human erythrocytes, with particular attention to catalase (CAT-EC. 1.11.1. 6- hydrogen peroxide: hydrogen peroxide oxidoreductase). 4-chloro-2-methylphenoxyacetic acid (MCPA), 2,4-dimethylphenol (2, 4-DMP) and 2,4-dichlorophenoxyacetic acid (2,4-D) did not affect CAT activity, but 2,4-dichlorophenol (2,4-DCP) and 2,4,5-trichlorophenol (2,4,5-TCP) decrease its activity, the latter being the more inhibitory.

Influence of phenoxyherbicides and their metabolites on the form of oxy- and deoxyhemoglobin of vertebrates.

The effect of phenoxyherbicides and their metabolites on the structure of oxy- and deoxyhemoglobin was studied by using different doses and times of incubation of hemoglobin with the herbicide. It was ascertained that among the investigated hemoglobins the most sensitive was carp oxyhemoglobin incubated with 2,4-D (2,4-dichlorophenoxyacetic acid) and the least sensitive was human hemoglobin. Comparing the toxicity of 2,4-D, MCPA (2-methyl-4-chlorophenoxyacetic acid), 2,4-DCP (2,4-dichlorophenol), 2,4-DMP (2,4-dimethylphenol) it was found that the highest decrease occurred in bovine hemoglobin incubated with 2,4-DMP. The phenoxyherbicides caused stabilization of the structure of T-deoxyhemoglobin in vitro, in that they decreased the oxygen affinity with a simultaneous increase in methemoglobin concentration.

Changes in peripheral blood composition following single and repeated courses of intermittent cancer chemotherapy.

In fifteen patients treated for lung cancer with a combination of vincristine (VCR) (2 x 0.325 mg/kg), amethopterin (MTX) (3 x 0.4 mg/kg) and cyclophosphamide (CY) (30-50 mg/kg) at monthly intervals the peripheral blood parameters were determined every 2-4 days. The values of these parameters including neutrophils, lymphocytes, platelets, erythrocytes and reticulocytes decreased uniformly reaching the lowest level about the 11th day post each course of chemotherapy. Subsequently, by the 17th day, the values of all these parameters except the red cell count returned to the pretreatment levels. The only parameter whose nadir value was associated with known risk to the health was the absolute neutrophil count. The period of dangerous decrease of this count began on the 6th day and ended approximately on the 17th day after cyclophosphamide infusion. Repeated courses had no additive toxicity since the time interval between courses allowed for hematopoietic recovery. However, the gradual decrease in the red cell count was observed during intermittent chemotherapy.

Studies on isolation of a toxic substance from human and animal skin burned in vitro.

Attempts to obtain burn toxin from animal and human skin are reported. Mouse skin was burned with a metal stamp by the method of Allgöwer. The human and the skin of various experimental animals was scalded in NaCl solution by the method of Rosenthal. Using the latter method it was possible to obtain the so-called "crude burn toxin" from the human skin. It was highly lethal to mice, rats, rabbits and guinea pigs after toxic. No toxic preparations were obtained from the skin of various experimental animals using the method of burning and processing described by Rosenthal.

Comparative investigations of in vitro and in vivo scalding of mouse skin.

Mouse skin was scalded in vitro by the wet (immersion) method and dry (metal stamp) method. For in vivo scalding only the dry method was used. The content of dry mass of skin was investigated. In extracts of scalded skin the content of soluble components: protein, collagen, albumin and IgG was determined. The analyses and statistic estimation revealed that with respect to the observed effects the in vivo scalding of skin is similar to the ion vitro dry scalding but different from the in vitro wet method. Moreover, it is suggested that protein substances which are products of thermic denaturation of skin can penetrate from scalded skin into the surrounding tissues.

Isolation and immunochemical properties of burn toxin.

Human skin and the skin of some laboratory animals was minced, scalded and homogenized. The preparation from scalded human skin (the "crude burn toxin") revealed to be highly toxic to mice after i.v. administration. It was found by immunodiffusion and immunoelectrophoresis, that the urea and buthanol extracts of the crude "toxin" contain tissue antigens which were absent in a control preparation from a native (non scalded) skin.