Janusz A. Madej

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.

Toxicity and antineoplastic effect of (24R)-1,24-dihydroxyvitamin D3 (PRI-2191)

Many efforts have been made to obtain active and less toxic Vitamin D analogs for new clinical applications. The results of previous studies demonstrated the efficacy and safety of topical treatment of psoriasis with one of these analogs, 1,24-dihydroxyvitamin D(3), tacalcitol (1,24-(OH)(2)D(3)). In the present study, we evaluated the toxicity and antitumor effect of this analog. Lethal toxicity of 1,24-(OH)(2)D(3) after s.c. injection was significantly lower than that of calcitriol. No significant differences were observed in the toxicity of the analogs when administered p.o. Calcium levels in the serum of mice treated with calcitriol were significantly higher (111%) than those in mice treated with 1,24-(OH)(2)D(3) (89%) at 5 day after the first s.c. (10 microg/kg/day) administration in comparison to the control (healthy, untreated animals). Oral administration increased the calcium level by 78% for calcitriol and only to 47% over the control for 1,24-(OH)(2)D(3). Parallel administration of clodronate prevented the calcitriol- and 1,24-(OH)(2)D(3)-induced lethal toxicity and also prevented increase in calcium levels. Single therapy with calcitriol did not affect tumor growth in the 16/C mouse mammary cancer model. In contrary, 1,24-(OH)(2)D(3) alone reduced tumor volume to 41% of control. Cisplatin alone did not affect growth of 16/C tumor in these conditions. The growth of tumors in the presence of cisplatin was inhibited by 1,24-(OH)(2)D(3) but not by calcitriol. Interestingly, the inhibition of tumor growth in cisplatin-treated mice by 1,24-(OH)(2)D(3) was greater, than that observed in mice treated with this analog alone. In conclusion, 1,24-(OH)(2)D(3) revealed higher antitumor and lower calcemic activity and toxicity than calcitriol. Application of biphosphonates along with Vitamin D analogs is sufficient to overcome the calcemic and toxic side effects of the proposed treatment.

Liposomal formulation of 5-fluorocytosine in suicide gene therapy with cytosine deaminase – for colorectal cancer

It is generally accepted that successful gene therapy depends on two major factors: tumor-specific expression of a therapeutic gene and the efficient transfer of a therapeutic gene to tumor cells. For gene-directed enzyme prodrug therapy (GDEPT) involving Escherichia coli cytosine deaminase (CD) and 5-fluorocytosine (5-FC), several tumor-specific promoters and virus-based vectors were used. No attention whatsoever was paid to the way of 5-FC delivery to solid tumors, despite the fact that the delivery of drugs to such tumors is generally low because of their insufficient transfer from the blood. To compare the effectiveness of GDEPT with free and liposomal 5-FC, the prodrug was encapsulated in liposomes composed of dipalmitoylphosphatidylcholine (DPPC) and cholesterol (1:1). When the liposomal form of 5-FC was administered i.v., mice treated with a dose of 5mg of liposomal 5-FC/kg body weight for 10 days, showed complete regression of transplanted tumors and complete cure was observed, whereas in animals treated with the same amounts of the free prodrug, 50% tumor regression and only insignificantly prolonged median survival were found. In summary, these results showed a remarkable enhancement of the antitumor effects of the liposomal form of 5-FC in comparison with the free prodrug. Therapy with liposomal 5-FC thus represents a new approach to achieving a high local concentration of the prodrug for suicide gene therapy using E. coli CD.

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.

Prevalence of tumours in domestic animals in the Lower Silesia (Poland) in 2009-2011.

Abstract The study aimed at the analysis of prevalence frequency and localisation of tumours in domestic animals. The research material comprised 4,212 tumours developed in dogs, cats, horses, and exotic animals, isolated during surgery, autopsy or biopsy, performed for the purpose of histopathological diagnosis. The most numerous group involved canine tumours, including 3,585 cases (85.1%), followed by tumours in cats (532 cases, 12.6%), ferrets (34 cases, 0.81%), rats (19 cases, 0.45%), horses (15 cases, 0.36%), and rabbits (14 cases, 0.33%). A significant increase in incidence of tumours was noted, as compared to studies performed in the same region of Poland in 1957-1995 and 2000-2004 or in 2005-2008. This was particularly evident in exotic animals, in which 80 cases (1.9%) were detected in 2009-2011 but only 20 cases (1.2%) in 2005-2008. The most frequent localisation of neoplasia involved the skin, subcutaneous tissue, and mammary gland.

SEMINOMA, SERTOLIOMA, AND LEYDIGOMA IN DOGS: CLINICAL AND MORPHOLOGICAL CORRELATIONS

Abstract The study aimed at presenting the most frequent male gonadal tumours in dogs, their clinical and histopathological aspects, at outlining aetiopathogenesis and differential diagnosis of the tumours. As examples of the most frequently manifested testicular tumours, three clinical cases were presented, involving tumour of interstitial (Leydig) cells, tumour of Sertoli cells, and seminoma. Respective clinical diagnosis employed USG, X-ray patterns, and morphological and biochemical tests. The surgically sampled material was stained with H+E and an attempt was made to establish expression of E-cadherin, calretinin, and Ki-67. It was shown that histopathological diagnosis of testicular tumours in dogs is frequently very difficult and complex and requires multidirectional studies.

Comparative analysis of markers of cell proliferation in canine mast cell tumours according to current classifications

The study aimed at immunohistochemical analysis of various markers of cell proliferation and comparison of the results with canine mast cell tumours grading systems according to the Patnaik and Kiupel. Tissue sections were stained using classical technique with haematoxylin and eosin, and immunohistochemical studies were performed with Ki-67, PCNA and MCM-3 antibodies. Additionally the mitotic index was assessed. Statistical analysis including rank correlation Spearmans and ANOVA Friedman analysis was performed. The significance was set at p<0.05. Expression of all examined antigens was detected. The results obtained allow concluding that there is a strong relationship between all the cell markers. However, due to the very strong response and positive reaction in the majority of tumours PCNA is not recommended as a prognostic indicator. Ki-67 and MCM-3 can be successfully used in the evaluation of canine mast cell tumours.

Ceramide galactosyltransferase (UGT8) as a molecular marker of canine mammary tumor malignancy.

Thirty-two canine mammary tubulopapillary carcinomas and 14 simple adenomas were studied by immunohistochemistry for the expression of UDP-galactose:ceramide galactosyltransferase (UGT8). The majority of tissue specimens (57%) representing adenomas had no or weak reaction with anti-UGT8 antibodies (0-2 pts according to IRS scale) in comparison to the majority of carcinomas (90%) which stained with high intensities (3-9 pts according to IRS scale). When the average values of the reaction intensities (IRS) for malignant and benign tumors were compared, using the Mann-Whitney U-test, significant differences in UGT8 expression between them were found (P < 0.001). Mammary tubulopapillary carcinomas were further analyzed by IHC and the same rabbit polyclonal antibody directed against UGT8 according to their malignancy grade. It was found that the level of UGT8 increased in tumor specimens together with their grading. A comparison of the average values of the reaction intensity (IRS scale) revealed a significant difference (Mann-Whitney U-test, P < 0.05) in UGT8 expression between tumors representing malignancy grades G3 and G1. Based on the obtained results, it is proposed that UGT8 is associated with malignancy of canine mammary gland cells and may have a potential value as a diagnostic marker.

Pathomorphological Changes of the Myocardium in Canine Dilated Cardiomyopathy (DCM)

Abstract The study was conducted on ventricular and atrial wall preparations from 11 dogs with clinically diagnosed dilated cardiomyopathy. After fixation, the specimens were stained with haematoxylin and eosin and Masson-Goldner trichrome technique. Parenchymal changes (fibrosis and fatty infiltration), vascular changes (congestion and coronary vessel wall hypertrophy), degenerative changes (loss of striation, changes in cardiomycyte and nuclei structure), and presence of inflammatory infiltrates (mononuclear and polynuclear) were estimated. Complex histological changes in both ventricular and atrial muscles were shown. It was not determined whether the processes occurring in the myocardium have a primary character, or are a consequence of developing heart failure. Such issues will be put under further and more detailed examination.

Histopathological pattern of gonads in cases of sex abnormalities in dogs: An attempt of morphological evaluation involving potential for neoplasia

Disturbances in sex differentiation (DSD - disorder of sexual development) may result from disturbances in sex chromosomes or a disturbed development of gonads, or from genotypic disturbances. The objective of this article is to describe the histological structure of gonads in dogs showing sexual disturbances and a case of a cancer resembling gonadoblastoma in one of the animals. Among the 10 examined dogs with disturbances of sex development only a single case of a gonadoblastoma was observed. In animals with sex disturbances, similarly to humans, there exists a potential tendency for neoplastic lesions in dysgenetic gonads. As a rule, its frequency in population is confined due to the early procedure of castration of non-breeding dogs. In the present study dogs demonstrated phenotypical traits of bitches with developmental anomalies such as hyperplastic clitoris with vestigial os penis (baculum), or abnormalities in the location and structure of the vulva. The material for the study included canine gonads of various breeds, sampled from phenotypical bitches, aged 7 months to 4 years - patients of the Department of Reproduction and Clinic of Farm Animals, Faculty of Veterinary Medicine, University of Environmental and Life Sciences in Wroclaw (Poland) in years 2006-2013. The organs were surgically removed from the abdomen and sent for histopathological examination for the purpose of determining their histological structure. The 10 examined cases of altered gonads included 6 bilateral cases of testes (60%), 2 cases of bilateral ovotestis (20%), one case of co-manifestation of testis and ovotestis (10%), and a single case of a testis and a neoplastically altered gonad (gonadoblastoma) (10%).