M.W. Łuczak, P.P. Jagodziński: The role of DNA methylation in cancer development pp. 143-154
M. Podhorska-Okołów,
P. Dzięgiel, B. Dolińska-Krajewska, M. Dumańska, M. Cegielski, Z.
Jethon, K. Rossini, U. Carraro, M. Zabel: Expression of metallothionein
in renal tubules of rats exposed to acute and endurance exercise
pp.
195-200
P. Szeparowicz, J. Popko, B. Sawicki, S. Wołczyński:
Is the
repair of articular cartilage lesion by costal chondrocyte
transplantation donor age-dependent? An experimental study in rabbits
pp. 201-206
J. Patera,
J.
Chorostowska-Wynimko, J. Słodkowska, A. Borowska, P. Skopiński, E.
Sommer, A.
Wasiutyński, E. Skopińska-Różewska: Morphometric and functional
abnormalities of kidneys in the progeny of mice fed chocolate during
pregnancy and lactation pp. 207-211
The role of DNA methylation in cancer development
Michał W. Łuczak and Paweł P. JagodzińskiDepartment of Biochemistry and Molecular Biology, University of Medical Sciences,
Poznań, Poland
Abstract: Epigenetic modifications include DNA methylation and covalent modification of histones. These alterations are reversible but very stable and exert a significant impact on the regulation of gene expression. Changes in methylation of promoter or first exon may mimic the effect of mutations of various tumor suppressor genes (TSGs) or protooncogenes. Carcinogenesis can also result from aberrations in genomic DNA methylation that include hypermethylation and hypomethylation of promoter or first exon of cancer-related genes. Hypermethylation of promoter of various TSGs causes their transcriptional silencing. However, hypomethylation of regulatory DNA sequences activates transcription of protooncogenes, retrotransposons, as well as genes encoding proteins involved in genomic instability and malignant cell metastasis. The methylation of genomic DNA in malignant cells is catalyzed by DNA methyltransferases DNMT1 and DNMT3B, revealing significantly elevated expression in different types of cancers. The reversibility of hypermethylation can be used as target of therapeutic treatment in cancer. DNMT1 and DNMT3B inhibitors including 5-Aza-2’-deoxycytidine and antisense oligonucleotides have been applied in clinical trials of such treatment. Identification of aberrations of DNA methylation in cancer cells is new field of investigation in carcinogenesis. We believe that epigenetic cancer diagnostic and therapy will be achieved in the next decades.
Author’s e-mail: pjagodzi@am.poznan.pl
SDF-1 alone and in co-operation with HGF regulates biology of human cervical carcinoma cells
Marcin Majka1, Justyna Drukala2, Ewa Lesko1, Marcin Wysoczynski4, Alfred B. Jenson3, Mariusz Z. Ratajczak1,4
1Department of Transplantation, Polish-American Institute
of Pediatrics, Jagiellonian University Medical College, Cracow, Poland
2Department of Cell Biology, Faculty of Biotechnology,
Jagiellonian University, Cracow, Poland,
3James Graham Brown Cancer Center, University of Louisville,
Louisville, Kentycky, USA,
4Stem Cell Biology Program at James Graham Brow Cancer
Center, Louisville, Kentycky, USA
Abstract: Stromal Derived Factor-1 (SDF-1)-CXCR4 axis plays a pivotal role in biology and metastasis of several tumors. The aim of this study was to see if SDF-1 alone or in combination with Hepatocyte Growth Factor (HGF) affects biology of human cervical carcinoma (HCC) cells. We found that HCC cell lines investigated in our study highly express CXCR4 on their surface. CXCR4 was also expressed on tumor cells in tissue sections derived from cervical cancer patients. At the same time normal cervical epithelium was negative for CXCR4 expression what suggests a strong correlation between CXCR4 and malignant cell phenotype. Subsequently, we studied a potential role of the SDF-1-CXCR4 axis in HCC and noticed that SDF-1 (1) chemoattracted HCC cells, (2) enhanced their scattering, (3) stimulated nuclear localization of b-catenins and upregulated their target gene cyclin D1 and I(4) at the molecular level induced calcium flux and activated RAS-MAPK, PI3-AKT and JAK-STAT pathways. SDF-1-mediated functions were additionally enhanced in the presence of HGF. Thus, our data show that the SDF-1-CXCR4 axis affects biology of HCC cells. Furthermore, we postulate that this axis might become a potential target to prevent progression of cervical cancer.
Author’s e-mail: mmajka@cm-uj.krakow.pl
Immunohistochemical expression of androgen receptor and prostate-specific antigen in breast cancer
Diana Narita1, Marius Raica2, Cristian Suciu2, Anca Cimpean2 and Andrei Anghel1
1Department of Biochemistry and 2Department of Histology, “Victor Babes” University of Medicine and Pharmacy, Timisoara, Romania
Abstract: AR (androgen receptor) and PSA (prostate-specific antigen) are involved in the pathogenesis of breast cancer, but their role is not clearly defined. The purpose of this study was to analyze by immunohistochemistry the AR and PSA (prostate-specific antigen) expression in 156 female breast carcinomas and to correlate the results with some histopathological parameters, like ER (estrogen receptor), PR (progesterone receptor), HER2/neu, nodal and metastasis status, histological type and grade. ARs and PSA were expressed in 112/156 (72%) and respectively in 61/156 (39%) of cases and we found a positive correlation between AR and PSA expression in breast carcinomas (p<0.0002). We also found an association between the histological type of the tumor and AR (p<0.001), respectively PSA (p=0.01) and between AR and the grade of differentiation (p=0.007) and the nodal status (p=0.02). No correlations were found between the metastasis status and AR or PSA. 47.3% (53/112) of AR-positive cases and 46% (28/61) of PSA-positive cases were ER-negative. High frequency of AR (87.5%) and PSA (75%) expression was found in medullary carcinomas and 53% of lobular invasive carcinomas co-expressed AR and PSA. We found an inverse correlation between HER2/neu and PSA (p=0.05). Although most of the PSA-positive carcinomas were lymph node negative, well and moderately differentiated, we did not find any statistically significant correlations between these parameters and PSA expression. Our study confirms that ARs are commonly expressed in breast cancer and the expression of PSA and AR are highly correlated. Moreover, all the lobular carcinomas and the majority of medullary carcinomas co-expressed AR and PSA, the majority of AR-positive carcinomas were lymph node-negative, well and moderately differentiated, and large number of ER-negative carcinomas expressed AR and PSA.
Author’s e-mail: diana_narita@yahoo.com
Immunohistochemical detection of angiotensin receptors AT1 and AT2 in normal rat pituitary gland, estrogen-induced rat pituitary tumor and human pituitary adenomas
Marek Pawlikowski
Department of Neuroendocrinology, Chair of Endocrinology, Medical University, Łódź, Poland
Abstract: Male rat pituitary glands, diethylstilbestrol (DES)-induced rat pituitary tumors and 12 human pituitary adenomas were immunostained with antibodies raised against AT1 and AT2 angiotensin receptor proteins. Positive immunostaining of AT1 was observed in a subpopulation of anterior and intermediate pituitary lobe cells as well as in some nerve endings of the neurohypophysis. In the DES-induced rat pituiary tumors, the subpopulation of AT1-immunnopositive cells was smaller than in the non-tumoral anterior pituitary. In human pituitary adenomas, weak AT1 immunostaining was found in 5 tumors. In the remaining adenomas, the AT1 immunostaining was trace (doubtful) or absent. The AT1 immunostaining in the peritumoral non-neoplastic pituitary tissue was stronger than that observed in the tumors. The normal rat pituitaries and rat tumors did not show immunostaining with anti-AT2 antibody. In human pituitary adenomas, the tumoral cells were AT2- negative but moderate to strong AT2 immunostaining was observed in intratumoral blood vessel walls. The data suggest that the experimental (in rat) and spontaneous (in man) pituitary tumorigenesis is associated with the down-regulation of AT1 receptors. The expression of AT2 receptors, in turn, may be connected with the process of tumoral neo-angiogenesis.
Author’s e-mail: pawlikowski.m@wp.pl
A false expression of CD8 antigens on CD4+ T cells in a routine flow cytometry analysis
Karolina Bukowska-Straková, Jarosław Baran, Małgorzata Gawlicka and Danuta Kowalczyk
Department of Clinical Immunology, Polish-American Institute of Paediatrics, Jagiellonian University Medical College, Kraków, Poland
Abstract: The two-colour flow cytometry method applied in routine enumeration of peripheral blood T lymphocyte subsets reveals that in some patients the entire population of CD4+ lymphocytes seems to express CD8 determinants as well. However, expression of the CD8 antigens on the cell surface is much lower in comparison with typical CD8+ cells. Moreover, in one-colour staining with an anti-CD8 antibody, cells with weak CD8 expression are not observed and only one typical population of CD8+ lymphocytes is seen. Investigating this phenomenon, we showed that after washing patient cells in RPMI before CD4/CD8 staining, the CD4+ T cell population did not show CD8 "co-expression". These results suggest that CD4+ lymphocytes, which seem to co-express CD8 antigen, in fact do not have this antigen on their surface. Moreover, after the addition of patient plasma to healthy donor cells prior to CD4/CD8 staining, a weak CD8 expression on normal CD4+ cells was noticed. Therefore we can assume that the agent(s) causing this phenomenon is/are present in the plasma of some patients. Altogether, these observations suggest that this phenomenon is nonspecific and probably results from cross-linking of anti-CD8 mAbs with anti-CD4 mAbs caused by factor(s) present in plasma of some patient. However, identification of that/these factor(s) requires further research.
Author’s e-mail: dkowalcz@cm-uj.krakow.pl
Androgen receptor in apoptotic follicles in the porcine ovary at pregnancy
Maria Słomczyńska1, Zbigniew Tabarowski2, Małgorzata Duda1, Małgorzata Burek1 and Katarzyna Knapczyk1
1Laboratory of Endocrinology and Tissue Culture, and 2Laboratory
of Experimental Hematology, Institute of Zoology, Jagiellonian
University, Kraków, Poland
Abstract: Localization of androgen receptor (AR) was investigated in ovarian follicles developing and undergoing atresia during pregnancy in the pig. Immunohistochemical staining was conducted on ovarian antral follicles isolated on different days of gestation: 10, 18, 32, 50, 70, and 90. Paraffin sections were also subjected to in situ DNA labeling. TUNEL staining revealed the presence of positive follicles on all days of pregnancy but the amount of atretic follicles increased with time. However, even on day 90 of gestation many follicles were normal, with no signs of atresia. In atretic follicles, apoptotic cells were localized predominantly in the granulosa while theca was much less affected. Atretic follicles with many apoptotic cells were negative for AR. Nuclear immunostaining for AR was positive in follicles with limited amount of apoptotic cells. The same relationship was observed in ovarian follicles isolated at various days of pregnancy.
Author’s e-mail: slom@zuk.iz.uj.edu.pl
Sources of porcine longissimus dorsi muscle (LDM) innervation as revealed by retrograde neuronal tract-tracing
Mariusz Chyczewski1, Joanna Wojtkiewicz2, Agnieszka Bossowska2, Marek Jałyński1, Wojciech Brzeski1, Ireneusz M. Kowalski3 and Mariusz Majewski2
1Division of Surgery and Rentgenology, Department of
Clinical Sciences, Faculty of Veterinary Medicine, Warmia and Mazury
University, Olsztyn;
2Division of Clinical Physiology, Department of Functional
Morphology, Faculty of Veterinary Medicine, Warmia and Mazury
University, Olsztyn;
3Provincial Childrens’ Hospital and Rehabilitation Centre,
Ameryka, Poland
Abstract: The aim of the present study was to establish the origin of the motor, autonomic and sensory innervation of the L1-L2 segment of the porcine longissimus dorsi muscle (LDM), in order to provide morphological basis for further studies focusing on this neural pathway under experimental conditions, e.g. phototerapy and/or lateral electrical surface stimulation. To reach the goal of the study, multiple injections of the fluorescent neuronal tracer Fast Blue (FB) were made into the LDM region between the spinal processes of the vertebrae L1 and L2. The spinal cord (Th13-S1 segments) as well as the sensory and autonomic ganglia of interest, i.e., dorsal root (DRG) and sympathetic chain ganglia from corresponding spinal cord levels were collected three weeks later. FB-positive (FB+) motoneurons were observed exclusively within the nucleus ventromedialis at L1 and L2 spinal cord level, forming the most ventro-medially arranged cell column within this nucleus. Primary sensory and sympathetic chain neurons were found in appropriate ipsilateral ganglia at Th15-L3 levels. The vast majority of retrogradely traced neurons (virtually all motoneurons, approximately 76% of sensory and 99.4% of sympathetic chain ganglia neurons) was found at the L1 and L2 levels. The morphometric evaluation of FB-labeled DRG neurons showed that the majority of them (approximately 66%) belonged to the class of small-diameter perikarya (10-30 μm in diameter), whereas those of medium size (30-80 μm in diameter) and of large diameter (more than 80 μm) constituted 22.6% and 11.5% of all DRG neurons, respectively. The results of the present study demonstrated that the nerve terminals supplying porcine LDM originated from different levels of the spinal cord, dorsal root and sympathetic chain ganglia. Thus, the study has revealed sources and morphological characteristic of somatic, autonomic and spinal afferent neurons supplying porcine LDM, simultaneously pointing out the characteristic features of their distribution pattern.
Author’s e-mail: Mariusz.Majewski@uwm.edu.pl
Full
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Expression of metallothionein in renal tubules of rats exposed to acute and endurance exercise
Marzena Podhorska-Okołów1, Piotr Dzięgiel1, Barbara Dolińska-Krajewska1, Małgorzata Dumańska1, Marek Cegielski1, Zbigniew Jethon2, Katia Rossini3, Ugo Carraro3 and Maciej Zabel1, 4
1Department
of Histology and Embryology, Medical University, Wrocław, Poland
2Department of
Physiology, University of Physical Education, Wrocław, Poland
3Department of Biomedical Science, University of Padua, Italy
4Department of Histology
and Embryology, University of Medical Sciences, Poznań, Poland
Abstract: The induction of exercise-induced apoptosis in not actively involved in exercise organs, such as kidney could be a result of oxidative stress. Metallothionein (MT) exerts a protective effect in the cell against oxidative stress and apoptosis. We have previously demonstrated an increased incidence of apoptosis in distal tubular cells and collecting ducts in rat kidney after acute exercise. The present study was designed to test the hypothesis that MT may play a protective role in rat renal tubules against exercise-induced apoptosis after the acute exercise and regular training. Male Wistar rats were divided into control, acute exercised and 8-wk regularly trained groups. The kidneys were removed after a rest period of 6 h and 96 h. The ultrastructure of renal tubular cells was examined by electron microscopy. Apoptosis was detected in paraffin sections by the TUNEL technique. Expression of MT was examined by immunohistochemistry. The level of lipid peroxidation (thiobarbituric acid reactive substances – TBARS) was assayed in renal tissue homogenates. After acute exercise the occurrence of apoptosis was restricted to distal tubules and collecting ducts of rat kidney, whereas the proximal tubules remained unaffected. The 8-wk training did not result in increased apoptosis in tubular cell. MT expression was confined exclusively to proximal tubules in all groups. However, it was significantly increased in acutely exercised animals, as compared to control and trained rats. After the 8-wk training, MT expression remained unaltered as compared to the control group. TBARS levels were significantly increased after acute exercise, while after regular training they remained unchanged. A significant correlation between TBARS level and MT expression was demonstrated. The findings could suggest a protective role of MT against oxidative stress and apoptosis in proximal tubular cells.
Author’s e-mail: mapod@hist.am.wroc.pl
Is the repair of articular cartilage lesion by costal chondrocyte transplantation donor age-dependent? An experimental study in rabbits
Paweł Szeparowicz1, Janusz Popko1, Bogusław Sawicki2 and Sławomir Wołczyński3
1Department of Pediatric Orthopaedics and Traumatology, 2Department
of Histology and Embryology, and 3Department of Gynecological
Endocrinology, Medical University, Białystok, Poland
Abstract: The repair of chondral injuries is a very important problem and a subject of many experimental and clinical studies. Different techniques to induce articular cartilage repair are under investigation. In the present study we have investigated whether the repair of articular cartilage folowing costal chondrocyte transplantation is donor age-dependent. Transplantation of costal chondrocytes from 4- and 24-week old donors, with artificially induced femoral cartilage lesion, was performed on fourteen 20-week-old New Zealand White male rabbits. In the control group, the lesion was left without chondrocyte transplantation. The evaluation of the cartilage repair was performed after 12 weeks of transplantation. We analyzed the macroscopic and histological appearance of the newly formed tissue. Immunohistochemistry was also performed using monoclonal antibodies against rabbit collagen type II. The newly formed tissue had a hyaline-like appearance in most of the lesions after chondrocyte transplantation. Positive immunohistochemical reaction for collagen II was also observed in both groups with transplanted chondrocytes. Cartilage from adult donors required longer isolation time and induced slightly poorer repair. However, hyaline-like cartilage was observed in most specimens from this group, in contrary to the control group, where fibrous connective tissue filled the lesions. Rabbit costal chondrocytes seem to be a potentially useful material for inducing articular cartilage repair and, even more important, they can also be derived from adult, sexually mature animals.
Author’s e-mail: szeparowicz@poczta.onet.pl
Morphometric and functional abnormalities of kidneys in the progeny of mice fed chocolate during pregnancy and lactation
Janusz Patera1, Joanna Chorostowska-Wynimko2, Janina Słodkowska3, Adamina Borowska4, Piotr Skopiński5, Ewa Sommer6, Aleksander Wasiutyński6 and Ewa Skopińska-Różewska6
1Department of Pathology, Military Institute of Medicine,
2Laboratory of Molecular Diagnostics, National Institute of
Tuberculosis and Lung Diseases, 3Laboratory of
Telepathology, National Institute of Tuberculosis and Lung Diseases, 4Department
of Laboratory Diagnostics, National Institute of Tuberculosis and Lung
Diseases, 5Department of Histology and Embryology, Medical
University, 6Department of Pathology, Medical University,
Warsaw, Poland
Abstract: Even most commonly consumed beverages like tea, coffee, chocolate and cocoa contain methylxanthines, biogenic amines and polyphenols, among them catechins, that exhibit significant biological activity and might profoundly affect the organism homeostasis. We have previously shown that 400 mg of bitter chocolate or 6 mg of theobromine added to the daily diet of pregnant and afterwards lactating mice affected embryonic angiogenesis and caused bone mineralization disturbances as well as limb shortening in 4-weeks old offspring. The aim of the present study was the morphometric and functional evaluation of kidneys in the 4-weeks old progeny mice fed according to the protocol mentioned above. Progeny from the mice fed chocolate presented considerable morphometric abnormalities in the kidney structure, with the lower number of glomeruli per mm2 and their increased diameter. Moreover, higher serum creatinine concentration was observed in that group of offspring. No morphometric or functional irregularities were found in the progeny of mice fed theobromine.Abnormalities demonstrated in the offspring of mice fed chocolate are not related to its theobromine content. Consequently, identification of active compound(s) responsible for the observed effects is of vital importance.
Author’s e-mail: ewaskop@hotmail.com