Danuta Plewka

Ozonotherapy in an Induced Septic Shock. I. Effect of Ozonotherapy on Rat Organs in Evaluation of Free Radical Reactions and Selected Enzymatic Systems

The confirmed advantageous effects of oxygen/ozone therapy in several clinical conditions stimulated experimental studies on effects of the therapy in rats with an induced septic shock. The studies were conducted on adult male rats of Wistar strain. Four groups of the animals, each of 15 rats, included: I—control group, (C); II—animals intraperitoneally administered with O2/O3 (CO), III—rats given of Escherichia coli endotoxin (lipopolysaccharide—LPS) (CL), IV—rats administered with the lipopolysaccharide plus administered with the oxygen/ozone mixture (OL). Activities of catalase and superoxide dismutase and of free radical reactions were estimated. The exposure to LPS augmented activities of SOD and of catalase in liver, lungs and heart. In all the examined organs LPS induced significant changes in levels of free radicals. Except of the lungs, parallel administration of the rats with LPS and ozone/oxygen revoked development of the alterations. The obtained results point to a strong, stabilizing and regenerative effect of ozonotherapy.

Immunohistochemical localization of selected pro-inflammatory factors in uterine myomas and myometrium in women of various ages

Uterine myomas represent one of the most frequently manifested benign tumors in women. They originate from smooth muscle cells of myometrium or its blood vessels. Many studies suggest that inflammation and pro-inflammatory factors may play a role in the carcinogenesis with an involvement of the transcription factor NF-kappaB which activity can be controlled by various environmental factors, including many cytokines. The aim of the study was to investigate the expression of NF-B, interleukin-1β (IL-1β), tumor necrosis factor a (TNF-α), cyclooxygenase 2 (COX-2) and inducible nitric oxide synthase (iNOS) in myometrium and uterine myomas of women of various age. The expression of NF-kappaB, selected cytokines and enzymes was estimated in women of reproductive or perimenopausal age by semiquantitative immunohistochemistry. The expression of the examined proteins was higher in myomas than in control myometrium and was dependent on the size of myomas and the age of women. However, the expression of the cytoplasmic NF-kappaB observed in uterine myomas was independent on the size of myomas and no significant differences were observed in the number of stained nuclei between control and myoma groups. Thus, the expression of proinflammatory factors in myomas was not accompanied by the nuclear activation of NF-kappaB p65. The results of our study indicate that the examined factors may be involved in the pathogenesis of benign tumors and not only malignant diseases.

Intestinal tissue kallikrein–kallistatin profile in inflammatory bowel disease ☆

The profile of tissue kallikrein (TK) and its inhibitor, kallistatin was evaluated in patients with active ulcerative colitis (UC) and Crohns disease (CD). Tissue kallikrein is mainly localized to goblet cells and kallistatin to epithelial cells of human intestine. Intestinal tissue kallikrein (ITK) and kallistatin are significantly decreased in inflamed intestine compared to noninflammatory controls. TK mRNA as well as kallistatin mRNA is significantly decreased in intestinal biopsy samples from UC-active patients compared with controls. The difference in distribution and levels of ITK and kallistatin in protein and mRNA in patients with inflammatory bowel disease (IBD) compared to controls suggest a role in inflammatory state.

Ozone Therapy in Induced Endotoxemic Shock. II. The Effect of Ozone Therapy Upon Selected Histochemical Reactions in Organs of Rats in Endotoxemic Shock

Mitochondria and lysosomes were evaluated by assessment of changes in activity of selected enzymes: lactate dehydrogenase (LDH), succinate dehydrogenase (SDH), adenosinetriphosphatase (ATPase), acid phosphatase (AcPase) and β-glucuronidase (BG) in rats under profound hypoxia induced by endotoxemic shock. The study was conducted on adult male Wistar rats. The animals formed the following four groups of 15 rats each: control animals (C);—rats receiving intraperitonally O2/O3 (CO), rats receiving of Escherichia coli toxin (LPS) (CL); rats receiving LPS plus oxygen–ozone mixture (OL). Histoenzymatic examinations of liver, kidney, lungs, and heart muscle were performed. Lipopolysaccharide suppressed activities of all the enzymes except for LDH, the activity of which as high as a fourfold increase. The results demonstrated potent, stabilizing and regenerative effects of ozone therapy on body enzymatic processes in course of induced endotoxemic shock in rats, which might prove to be of clinical significance.

Immunohistochemical visualization of pro-inflammatory cytokines and enzymes in ovarian tumors

Epithelial ovarian cancer represents one of the most deadly gynaecological neoplasms in developed countries and is a highly heterogeneous disease. Epidemiological studies show that anti-inflammatory drugs reduce the incidence and mortality of several types of cancer, indicating the potential role of pro-inflammatory factors in carcinogenesis. The expression of pro-inflammatory factors in various cancer types, including ovarian cancer, was assessed in many studies, yielding in consistent results, often due to the histological heterogeneity of various cancers. The aim of the study was to investigate the expression of IL-1, IL-6, TGF-β, TNF-α, COX-2, iNOS, and NF-kB in serous and mucinous ovarian cancers. Ninety cases of ovarian tumors classified into mucous and serous type (45 patients in each group) were selected. Each group was classified into subgroups according to the three stages of tumor differentiation, i.e. into (i) benign, (ii) borderline and (iii) malignant tumors. The presence of proteins of interest in paraffin sections was analysed by immunohistochemistry. The expression of most of the studied factors depended on the histological tumor subtype and the degree of malignancy. Expression of NF-κB appears to be related to the level of the neoplastic differentiation only in the group of serous tumors, while the presence of IL-6 in the mucinous tumor subtype was observed only in the case of benign lesions. Expression of IL-1, TNF-α and COX-2 increased with the stage of the disease in both serous and mucinous tumors. The highest level of TGF-β expression was observed in serous borderline tumors. The different levels of iNOS immunoreactivity between the groups of serous and mucinous tumors were observed only in borderline tumors. The results of our study may be helpful in designing therapeutic strategies depending on the type of ovarian cancer.

Expression of selected cytochrome P450 isoforms and of cooperating enzymes in colorectal tissues in selected pathological conditions.

The current interest in CYP expression in the colon results from its uniqueness as a target organ for cancer. To date, the CYP expression profiles in the colon have not yet been subject of comprehensive research. In this study, we investigated 40 patients with Crohns disease, 40 with ulcerative colitis, and 40 healthy subjects as a control group. Colon tissues were fixed, dehydrated, cleared in xylene and embedded in paraffin. Sections were prepared from paraffin blocks for immunohistochemical staining with specific antibodies. We used antibodies to the human CYP1A1, CYP2B6, CYP2C9, CYP2E1 and CYP3A4 isoforms, as well as antibodies to the human glycoprotein P, glutathione-S transferase and antibody to the UDP-glucuronosyltransferase. The sections were stained immunohistochemically and examined using light microscopy. Cellular localization was determined, and computer image analysis was used. In all cases with Crohns disease, the proteins studied showed at least a twofold expression. Ulcerative colitis showed a much weaker influence regarding the expression of the proteins studied but in case of CYP2C9 and UDP-glucuronosyltransferase, a decrease of expression was observed.

Morphological and enzymatic changes caused by a long-term treatment of female rats with a low dose of gonadoliberin agonist and antagonist

Summary Background Long-term treatment with gonadoliberin analogs is used to block the hypothalamic-pituitary-gonadal axis. The use of these agents is generally considered to be safe; however, some observations suggest the possibility of adverse effects. Material/Methods We investigated whether a 3-months administration of a low dose (6 μg/kg b.w.) of dalarelin – a new agonist, and cetrorelix – a known antagonist of GnRH to female rats causes morphological changes in pituitary gland, ovaries, uterus and liver (HE and VG staining); effects on pituitary, hepatic and blood enzyme activities (histochemical and kinetic methods, respectively), and on the blood lipid profile (colorimetric methods); and to what extent these changes are reversible. Results Applying analogs effectively inhibited ovulation, affected the uterine endometrium and changed histological appearance of the liver (e.g., steatosis). They altered activities of marker enzymes of cellular respiration, gluconeogenesis and intracellular digestion in the liver and, partially in the pituitary gland, caused undesirable changes in the activities of aspartate aminotransferase, alanine aminotransferase, lactate dehydrogenase, and creatine kinase, and a concentration of cholesterol HDL fraction and triglycerides in the blood. Both morphological and enzymatic effects were more evident after antagonist administration; changes in the blood lipid profile were more evident after agonist administration. In both analogs histological and enzymatic changes persisted a relatively long time after the discontinuation of the treatment. Conclusions The low dose of dalarelin and cetrorelix is sufficient to cause limited damage of hepatic cells and may modify the function of pituitary, ovaries, uterus and liver as well as other organs, even after discontinuation of the treatment.

Developmental toxicity of hexachloronaphthalene in Wistar rats. A role of CYP1A1 expression.

Hexachloronaphthalene (HxCN) is one of the most toxic congeners of polychlorinated naphthalenes (PCNs). This study assesses the prenatal toxicity of HxCN after daily administration at doses of 0.1-1.0mg/kg b.w. to pregnant Wistar rats during organogenesis. We evaluated also the expression of CYP1A1 mRNA and protein in the livers of dams and fetuses, as well as the placenta. The results indicate that 0.3mg/kg b.w. was the lowest HxCN toxic dose for dams (LOAEL) while a dose of 0.1mg/kg b.w. was sufficient to impair the intrauterine development of embryos/fetuses without maternal toxicity. Regardless of the applied dose, HxCN generated embryotoxic effects. Dose-dependent fetotoxic effects were associated with HxCN exposure. HxCN was found to be a strong inducer of maternal and fetal CYP1A1. Expression of CYP1A1 mRNA in the placenta appears to be the most sensitive marker of HxCN exposure.

The relation between plasma α-synuclein level and clinical symptoms or signs of Parkinson's disease

INTRODUCTION Parkinson disease (PD) is the common neurodegenerative disease. α-Synuclein (ASN), main aggregating protein in neural cells of CNS in PD, was found in peripheral fluids. Testing ASN in plasma is potential test for diagnose PD, but previous studies are controversial. The aim of this study was to investigate if plasma ASN level may be a valuable biomarker, is the level of plasma ASN concentration different in various motor subtypes of diseases, is there a relation between the level of plasma ASN and the severity of motor symptoms. METHODS Patients with PD hospitalized in Neurology Department, Medical College were performed sequencing the 8th and 9th exon of GBA gene. Next plasma ASN level was tested in 58 patients with sequenced GBA gene and in 38 healthy volunteers (HV), matched by the age (respectively 68.43 vs. 64.57 years of age) and sex (female %, respectively: 43.10 vs.44.74). Patients were assessed with the scales: UPDRS (II, III, IV), Hoehn-Yahr (HY) and qualified to PIGD or TD subtype. For homogeneity of the group patients with GBA mutation were excluded from the analysis. RESULTS The ASN level did not differ between patients and HV (respectively: 4.53 vs. 3.73ng/ml) and between patients with different subtypes. There was inverse correlation between ASN and HY in PIGD subtype. CONCLUSIONS Plasma ASN level is not valuable marker of the disease. It does not differ in subtypes of the disease. There is relation between plasma ASN level and the severity of the disease in PIGD subtype.