Jerzy Rabczyński

Melatonin stimulates the activity of protective antioxidative enzymes in myocardial cells of rats in the course of doxorubicin intoxication.

Abstract: The study aimed at determining the effect of melatonin on the activity of protective antioxidative enzymes in the heart and of lipid peroxidation products in the course of intoxication with doxorubicin (DOX). The rats were categorized into four groups, receiving: 0.9% NaCl i.p. (NaCl control); melatonin [20 mg/kg body weight (b.w.)] s.c. (control Mel); DOX (2.5 mg/kg b.w.) i.p.; melatonin plus DOX in doses as above. All the substances were administered once in a week for four consecutive weeks. Homogenates of heart tissue were examined for activities of glutathione peroxidase (GPx), superoxide dismutase (SOD), catalase (CAT), levels of reduced glutathione (GSH) and of lipid peroxidation indices (MDA + 4‐HDA). Administration of melatonin alone did not induce alterations in levels of MDA + 4‐HDA, GSH, or in activity of GPx, SOD or CAT, as compared to the group receiving 0.9% NaCl. GSH levels decreased following DOX but remained at normal levels following DOX and melatonin. The level of MDA + 4‐HDA increased following DOX, as compared with the control, a change prevented by the combination of DOX+melatonin. Activities of GPx, SOD and CAT were higher in groups receiving DOX and/or DOX plus melatonin than in control groups. Activity of CAT and the level of GSH in the group receiving DOX plus melatonin were significantly higher than in the group intoxicated with DOX alone. The obtained results demonstrate that, when given in parallel with DOX, melatonin protects cardiomyocytes from damaging effects of the cytostatic drug (reflected by the levels of MDA + 4‐HDA). The protective effect resulted, in part from the augmented levels of GSH and from stimulation of CAT activity by melatonin in cardiomyocytes subjected to the action of DOX.

Cytokine gene expression in kidney allograft biopsies after donor brain death and ischemia-reperfusion injury using in situ reverse-transcription polymerase chain reaction analysis.

Background. This study focuses on the cytokine genes expression after brain-death, ischemia-reperfusion injury, and during allograft rejection. Methods. A total of 49 needle core biopsies from kidney transplant recipients, performed before and during transplantation procedures were studied. The first biopsy was taken during procurement of the organ, the second after cold ischemia, and the third after approximately 30 min of reperfusion. We also assessed 34 allograft biopsies obtained during acute rejection. Tubular and glomerular expression of interferon (IFN)-&ggr;, transforming growth factor (TGF)-&bgr;1, platelet-desired growth factor-B (PDGF-B), interleukin (IL)-2, IL-6, IL-10 mRNA was analyzed with reverse-transcription polymerase chain reaction (RT-PCR) in situ technique, which allows to detect a few copies of the target gene without destruction of the tissue architecture. Results. Compared with normal kidney tissue from living donor, high gene expression of IFN-&ggr;, TGF-&bgr;1, PDGF-B, IL-2, IL-6, and IL-10 was detected in all procurement specimens. After reperfusion gene expressions of IL-2, IL-6, and IL-10 were significantly upregulated in renal tubules compared to biopsies taken after cold ischemia. The gene expression of IFN-&ggr;, TGF-&bgr;1, and PDGF-B remained stable after organ procurement, during cold ischemia, and after reperfusion. Gene expression of IFN-&ggr;, IL-2, IL-6, IL-10, and PDGF-B in procurement biopsies, as well as in those taken after cold ischemia and reperfusion, were significantly higher than during the period of acute rejection. Conclusion. The data presented herein strongly point out the importance of the immunological and morphological injury that occurs before and during transplantation. The increase of inflammatory response after brain death is important for further stimulation of the immune response and long-term kidney survival.

Field effect of human colon carcinoma on normal mucosa : Relevance of carcinoembryonic antigen expression

Human colon cancer usually develops on a mucosa which has already undergone multiple steps of genetic change. These multiple steps create a field effect characterized by the presence of morphologically normal, but biologically altered epithelial cells. This aims of this study were to evaluate whether the expression of carcinoembryonic antigen (CEA) can act as a phenotypic marker of the field effect, and to map its topography in relation to the presence of colorectal adenocarcinoma. The expression of CEA was tested by immunohistochemistry on morphologically normal mucosa at 4 increasing distances from 14 autologous cases of colorectal adenocarcinoma. CEA expression in the normal mucosa was compared to the tumor. The results show that in the mucosa adjacent to the edge of the autologous tumor, CEA is expressed to the same level as displayed in the carcinoma; there is a decrease in CEA expression in normal mucosa located at 1 cm or more from the edge of carcinoma. Mucosa sampled at 5 and 10 cm from the tumor expresses CEA at the same low level as in mucosa of control subjects with no colorectal neoplasm. In conclusion, this study demonstrates a gradient of CEA expression in the peritumoral area, supporting the concept of field effect, and maps its extent. These data are relevant to the biology of human colorectal cancer, and more practically, to the optimal location of surgical resection.

Role of exogenous melatonin in reducing the nephrotoxic effect of daunorubicin and doxorubicin in the rat

Abstract:  The aim of these studies was to examine the nephroprotective effect of melatonin following the anthracycline administration [daunorubicin (DNR); doxorubicin (DOX)] in rats. Application of these drugs in chemotherapy is limited because of their cardiotoxicity and nephrotoxicity. Rats of the Buffalo strain were divided into groups according to the cytostatic drug used, its dose and sequence of administration [DNR or DOX single (i.v.) dose of 10 mg/kg b.w., i.e. acute intoxication and 3 mg/kg b.w. (i.v.) weekly for 3 wk, subchronic intoxication]. Melatonin was administered subcutaneously before and after every injection of a cytostatic drug at a dose of 10 mg/kg b.w. The severity of renal alterations was examined both biochemically [levels of lipid peroxidation markers, malonyldialdehyde (MDA) and 4‐hydroxyalkenals (4‐HDA)], or histologically. A statistically significant decrease in renal damage was noted after melatonin administration to acutely or subchronically intoxicated DNR‐treated and DOX‐treated rats. Biochemical assays revealed significant decreases in MDA and 4‐HDA levels following application of melatonin during subchronic DNR or DOX intoxication. In summary, melatonin was found to exert a protective effect on the kidney, which was particularly evident after subchronic DOX and DNR intoxication, using both histological or biochemical methods.

Role of exogenous melatonin in reducing the cardiotoxic effect of daunorubicin and doxorubicin in the rat

The aim of the studies was to examine the cardioprotective effect of melatonin during the anthracycline administration (daunorubicin, doxorubicin) in rats. Application of these drugs in chemotherapy is limited because of their cardiotoxicity. Rats of Buffalo strain were divided into groups according to the cytostatic drug used, its dose and sequence of administration (single intravenous [i.v.] dose of 10 mg/kg b.w., i.e., acute intoxication; 3 mg/kg b.w. weekly for 3 weeks, subchronic intoxication). Melatonin was administered subcutaneously before and after every injection of a cytostatic drug at a dose of 10 mg/kg b.w. The degree of cardiac muscle cell alterations was examined either histologically (Mean Total Score technique and the Billingham scale), or biochemically (levels of lipid peroxidation markers, malonyldialdehyde, and 4-hydroxyalkenals). Statistically significant decrease in cardiac muscle cell damage was noted with an aid of the Billingham scale after melatonin administration in acutely intoxicated doxorubicin-treated rats (p < 0.001). The similar phenomenon was observed using the Mean Total Score technique in case of acute daunorubicin or doxorubicin (p < 0.01 and p < 0.001, respectively) intoxications. A significant reduction in cardiac muscle cell lesions was detected either by the Billingham scale or by the Mean Total Score technique during subchronic intoxication with either of the anthracyclines when melatonin was given. Biochemical assays revealed significant decreases in malonyldialdehyde and 4-hydroxyalkenals levels following application of melatonin during either acute doxorubicin (p < 0.05) or subchronic daunorubicin (p < 0.01) intoxication. In summary, melatonin was found to exert a protective effect on the cardiac muscle cells, which was particularly evident after acute doxorubicin or subchronic daunorubicin intoxication, using either histological or biochemical methods.

Dysfunction of the pancreas in healthy smoking persons and patients with chronic pancreatitis.

Objectives: The aim of the study was to assess the effect of cigarette smoking on the endocrine pancreatic function by determining the levels of serum glucose and plasma insulin as well as by defining immunohistochemical localization of insulin and glucagon in tissue specimens of the pancreata derived from healthy persons and smoking and nonsmoking patients with diagnosed chronic pancreatitis (CP). Methods: The oxidative method was used to measure fasting glycemia in blood plasma and the method enzyme-linked immunoassay to determine the level of insulin in plasma. Immunohistochemical localization of hormones in paraffin tissue specimens of the pancreas was performed using the LSAB2-HRP visual test with polyclonal insulin and glucagon antibodies. The intensity of immunohistochemical reaction was calculated with digital imaging methodology. Results: The study revealed a substantially higher level of serum glucose in smoking CP patients and in healthy persons compared with nonsmoking patients and healthy persons, whereas insulin concentration in smoking patients was statistically lower than in nonsmokers. Smoking patients showed significantly lower expression of insulin and glucagon in the pancreas compared with nonsmoking patients and healthy persons. Conclusions: Impairment of the endocrine function of &bgr; and &agr; cells in the pancreatic islets is frequently manifested by complications in pancreatitis resulting among others from long-term smoking.

Expression of Decorin in Esophageal Cancer in Relation to the Expression of Three Isoforms of Transforming Growth Factor-Beta (TGF-β1, -β2, and -β3) and Matrix Metalloproteinase-2 Activity

To determine the role of the reactive stroma in cancer progression, we investigated decorin (DCN) and transforming growth factor-beta (TGF-β expression, and matrix metalloproteinase-2 (MMP-2) activity in the tumorous esophagus. We found statistically insignificantly decreased levels of DCN expression in the pathological tissues. No obvious alterations in TGF-β expression were noticed. The highly significant increase in MMP-2 activity in cancers did not result in elevated levels of TGF-β dimers. Therefore, the system of TGF-β liberation from its complex with DCN by activated MMP-2 does not seem to contribute to esophageal cancerogenesis, although this hypothesis should be reevaluated with a larger study group.

Cytokine gene expression in kidney allograft donor biopsies after cold ischemia and reperfusion using in situ RT-PCR analysis.

It was previously reported that ischemia-reperfusion injury initiates an inflammatory response and may significantly affect the transplanted organ function. The aim of this study was to assess changes of intragraft cytokine mRNA expression in kidneys after cold ischemia (CI) and following reperfusion. We examined mRNA of a product of activated T lymphocytes (IFN-gamma) and a monocyte product (IL-6). Eleven kidneys were transplanted after CI time ranging from 16 to 39 hours. Renal needle core biopsies were obtained from donors after cold ischemia and approximately after 20 minutes of reperfusion. Tubular and glomerular expression of IFN-gamma and IL-6 mRNA were assessed using semiquantitative evaluation of the RT-PCR in situ. After reperfusion an intense increase of IL-6 mRNA expression was observed in four specimens, a slight increase was noticed in five specimens, and a very slight decrease in two specimens. Changes in IL-6 mRNA expression were limited only to tubules. In contrast, the glomerular and tubular mRNA expression of IFN-gamma and glomerular of IL-6 remained stable. Mean CI time for patients with an intense increase was higher than for patients with a slight increase and with the decrease of IL-6 mRNA expression (32.0 +/- 6.8 vs 25.2 +/- 7.3 and 26.0 +/- 5.7 hours). Our results suggest that early inflammatory changes at the time of implantation of renal allografts depends mainly on monocyte/macrophage-associated products. The observed intensity of their expression in tubules was connected to longer CI time.

MRP2 (ABCC2, cMOAT) expression in nuclear envelope of primary fallopian tube cancer cells is a new unfavorable prognostic factor.

ObjectiveTo determine the prognostic value of the immunohistochemical evaluation of the multidrug resistance-associated protein 2 (MRP2) expression, together with its subcellular localization in primary fallopian tube carcinomas (PFTCs).MethodsThe immunohistochemical analysis was performed using samples originating from 70 patients with PFTCs.Results(1) We documented that MRP2 can be localized in the plasma membrane (MRP2c), as well as in the nuclear envelope (MRP2n) of the PFTC cells. (2) Patients with more advanced stage, with progression of the disease and patients who died, showed significantly higher expression of the MRP2n. (3) Univariate and multivariate analyses showed that MRP2n is an unfavorable prognostic factor in PFTCs. (4) The analysis of the classic clinicopathological data revealed that only the FIGO stage had prognostic value, both in the univariate, as well as in multivariate analysis.Conclusions(1) This study suggests that MRP2n is a new disadvantageous prognostic factor in PFTCs and (2) that expression in nuclear envelope can be associated with lower differentiation of cancer cells and their resistance to the cisplatin. (3) We have also confirmed independent prognostic value of FIGO stage in PFTCs.

Immunohistochemical localization of metallothionein in chronic pancreatitis.

Metallothionein (MT) is a low-molecular weight intracellular protein, rich in sulfhydryl residues, and able to bind bivalent metals. MT, like Zn, is a component of the diversified elements of antioxidant system. Recent studies have shown that reactive oxygen species play a role in the pathogenesis and development of chronic pancreatitis. The aim of the study was to identify immunohistochemically (LSAB2-HRP; DAKOCytomation) the localization of metallothionein and to determine MT expression in 9 patients with chronic pancreatitis. Our studies confirm that MT is present in exocrine and endocrine cells of patients with chronic pancreatitis and chronic pancreatitis with concomitant diabetes. They also indicate increased expression of MT, particularly in acinar cells of the pancreas. This suggests that MT is greatly involved in homeostasis of the pancreas and synthesis of pancreatic hormones.