M. Grzybek, A. Kozubek, P. Dubielecka, A.F. Sikorski: Rafts – the current picture pp. 3-10
Z. Tabarowski, M. Szołtys, M. Bik, M. Słomczyńska: Atresia of large ovarian follicles of the rat pp. 43-50
C. Calabrò, M.P. Albanese, E.R. Lauriano, S. Martella, A. Licata: Morphological, histochemical and immunohistochemical study of the gill epithelium in the abyssal teleost fish Coelorhynchus coelorhynchus pp. 51-56
ABSTRACTS
Michał Grzybek1, Agnieszka Kozubek2, Patrycja Dubielecka1 and Aleksander F. Sikorski1,2
1Institute of Biochemistry and Molecular Biology, University of Wrocław and 2Academic Centre for Biotechnology of Lipid Aggregates, Wrocław, Poland
Abstract: Although evidences that cell membrane contains microdomains are accumulating, the exact properties, diversity and levels of organization of small lipid patches built mainly of cholesterol and sphingomyelin, termed rafts, remain to be elucidated. Our understanding of the cell membrane is increasing with each new raft feature discovered. Nowadays rafts are suggested to act as sites of cell signaling events, to be a part of protein sorting machinery but also they are used by several pathogens as gates into the cells. It is still unclear how rafts are connected to the membrane skeleton and cytoskeleton and with how many different types of rafts are we actually dealing with. This review summarizes some of the most recent discoveries trying to make a view of the complex raft properties.
Author’s e-mail: afsbc@ibmb.uni.wroc.pl
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Capillary density and capillary-to-fibre ratio in vastus lateralis muscle of untrained and trained men
J. A. Zoladz1, D. Semik2, B. Zawadowska2, J. Majerczak1, J. Karasinski2, L. Kolodziejski3, K. Duda1,3 and W. M. Kilarski2
1Department of Muscle Physiology, Academy of Physical Education, 2Department of Cytology and Histology, Institute of Zoology, Jagiellonian University, 3Cancer Institute, Cracow, Poland
Abstract: Muscle fibre profile area (Af), volume density (Vv), capillary-to-fibre ratio (CF) and number of capillaries per fibre square millimetre (CD) were determined from needle biopsies of vastus lateralis of twenty-four male volunteers (mean ± SD: age 25.4±5.8 years, height 178.6±5.5 cm, body mass 72.1±7.7 kg) of different training background. Seven subjects were untrained students (group A), nine were national and sub-national level endurance athletes (group B) with the background of 7.8±2.9 years of specialised training, and eight subjects were sprint-power athletes (group C) with 12.8±8.7 years of specialised training. Muscle biopsies of vastus lateralis were analysed histochemically for mATPase. Capillaries were visualized and counted using CD31 antibodies against endothelial cells. There were significant differences in the Vv of type I and type II muscle fibres in both trained groups, B (51.8%; 25.6%) and C (50.5%; 26.4%). However, in untrained group A that was treated as a reference group, the difference between Vv of type I and type II fibres was less prominent, nevertheless statistically significant (42.1%; 35.1%). There was also a significant difference in CF: 1.9 in group A and 2.1 in groups B and C. The number of capillaries per mm2 (CD) was 245 (group A), 308 (group B) and 325 (group C). Significant differences (P<0.05) in CF and CD, were found only between group A (1.9; 245) and both groups of trained men, B and C (2.1; 308 and 325). However, endurance athletes (group B), such as long-distance runners, cyclists and cross country skiers, did not differ from the athletes representing short term, high power output sports (group C) such as ice hockey, karate, ski-jumping, volleyball, soccer and modern dance.
Author’s e-mail: kilar@zuk.iz.uj.edu.pl
Modification of immunocytochemical ZAP-70 assay for potential clinical application in B-cell chronic lymphocytic leukemia
Agnieszka Bojarska-Junak1, Jacek Roliński1 and Jerzy Kawiak2
1Department of Clinical Immunology, University School of Medicine, Lublin,
2Department of Clinical Cytology, Medical Center of Postgraduate Education, Warsaw, Poland
Abstract: The ZAP-70 protein is a member of the Syk/ZAP protein tyrosine kinase family, normally expressed in T cells and NK cells but not found in normal, mature B cells. The protein plays a critical role in the initiation of T-cell signaling. Leukemic cells from patients with B-cell chronic lymphocytic leukemia (B-CLL) that expressed nonmutated immunoglobulin V genes were found to express levels of ZAP-70 protein that were comparable to those detected in T cells of healthy adults. The ZAP-70 protein expression can be evaluated by flow cytometry and may be used as a prognostic marker in B-CLL patients. We modified the method of immunocytochemical assessment of ZAP-70 expression. The traditional two-step method with monoclonal anti-ZAP-70 antibody in the first step followed by FITC-conjugated goat anti-mouse IgG was changed for one-step method with monoclonal anti-ZAP-70 antibody labeled by Zenon Alexa Fluor 488. The method is simple and fast. The major advantage of Zenon labeling technique is its compatibility with simultaneous staining of surface antigens. The cells may be earlier immunostained for CD3, CD19 and/or CD5 to compare of the ZAP-70 kinase expression in B and T cells.
Author’s e-mail: agnieszkajunak@poczta.onet.pl
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Generation of dendritic cells from human peripheral blood monocytes – comparison of different culture media
Nataliya Tkachenko, Kamila Wojas, Jacek Tabarkiewicz and Jacek Rolinski
Department of Clinical Immunology, Medical University, Lublin, Poland
Abstract: Culture medium or medium supplement is one of the factors responsible for dendritic cell (DC) generation, but little is known about the influence of various media on DC culture. In our study we generated DC from adherent monocytes of human peripheral blood in the presence of GM-CSF, IL-4 and TNF-a . The following culture media were used: RPMI 1640 supplemented with 2% human serum albumin; RPMI 1640 supplemented with 2% TCH serum replacement; X-VIVO 15 and Panserin 501. Flow cytometry analysis revealed that in all media cells were CD83+ and lost CD14. Interestingly, the use of Panserin and RPMI with albumin preferentially gave rise to CD1a+ DC, whereas in X-VIVO and RPMI with TCH we observed both CD1a+ and CD1a-. Our results showed that RPMI with TCH yielded the highest percentage of cells expressing both CD80 and CD86 molecules and, in contrast to other media, the higher percentage of CD86+ cells in comparison to CD80+ cells.
Author’s e-mail: natkaukr@yahoo.com
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Prognostic evaluation of CD44 expression in correlation with bcl-2 and p53 in colorectal cancer
H.N. Zavrides1, A. Zizi-Sermpetzoglou1, D. Panousopoulos2, G. Athanasas2, I. Elemenoglou1 and G. Peros3
1Department of Pathologic Anatomy, Tzanio Hospital, Athens,
2Department of Surgery, Athens University Medical School,
3Propaedeutic Surgical Clinic, Athens University Medical School, Greece
Author’s e-mail: hzavrides@hotmail.com
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Modulation of testicular macrophage activity by collagenase
Krzysztof Bryniarski1, Marian Szczepanik2, Maria Ptak1 and W³odzimierz Ptak1
1Department of Immunology and 2Department of Human Developmental Biology, Medical College, Jagiellonian University, Cracow, Poland
Abstract: Testicular macrophages (TMs) are located in the interstitial tissue of male gonad. These phagocytic cells take part in forming the organ-specific functional blood-testis barrier and participate in the regulation of the local hormonal balance. In the present study, we isolated TMs from testicular tissues using previously described methods – mechanical (M-TMs) or enzymatic, by treatment with collagenase (E-TMs) and then we studied production by these cells of several cytokines and reactive oxygen intermediates (ROI’s). Similarly treated oil-induced peritoneal macrophages (PMs) were used as control cells. PMs had a higher baseline level of production of TNF-alfa, IL-6, IL-10 and IL-12 than M-TMs and collagenase treatment increased the production of these cytokines (except IL-12) by both cell populations. This effect was significantly more expressed in TMs. In contrast to PMs, TMs produced little ROI’s when stimulated by zymosan. We conclude that in the case of local inflammation in the testis, ROI-negative TMs do not contribute to the tissue damage and instead may direct the local immune response into humoral pathway.
Author’s e-mail: mmbrynia@cyf-kr.edu.pl
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Atresia of large ovarian follicles of the rat
Zbigniew Tabarowski1, Maria Szołtys2, Małgorzata Bik2 and Maria Słomczyńska2
1Laboratory of Experimental Hematology and 2Laboratory of Endocrinology & Tissue Culture, Institute of Zoology, Jagiellonian University, Cracow, Poland
Abstract: In the rat, at the beginning of pregnancy a cohort of antral follicles develops until the preovulatory stage. However, these follicles, differentiating in the hyperprolactinemic milieu, produce only small amount of estradiol, do not ovulate and undergo rapid degeneration. They constitute an interesting physiological model of atresia. In the present study, we analysed the development and subsequent degeneration of such follicles. The study was performed on Wistar female rats killed in succession between days 1-9 of pregnancy. Excised ovaries were submitted to a routine histological procedure. Paraffin sections were subjected to hematoxylin and eosin staining or in situ DNA labelling. Histological and TUNEL staining revealed that the investigated group of follicles grew slower than that on the corresponding days of the estrous cycle and reached a preovulatory size and morphological appearance on day 5 of pregnancy. They did not ovulate and between days 6 and 9 of pregnancy an increasing number of apoptotic cells appeared within these follicles. They were localized predominantly in the antral granulosa layer, especially near the cumulus oophorus complex (COC) and in the region linking the COC with the follicular wall. The COC and the theca layer were much less affected. In late stages of atresia, also cumulus cells became apoptotic but degenerating oocytes did not exhibit positive TUNEL staining. Only limited number of the theca cells have undergone apoptosis and generally they were not hypertrophied. Our findings indicate that much smaller than normal amount of intrafollicular estradiol was sufficient to support a normal, according to the morphological criteria, although slower development of antral follicles to the late preovulatory stage.
Author’s e-mail: tab@zuk.iz.uj.edu.pl
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Morphological, histochemical and immunohistochemical study of the gill epithelium in the abyssal teleost fish Coelorhynchus coelorhynchus
Concetta Calabrò, Maria Pia Albanese, Eugenia Rita Lauriano, Silvestro Martella and Aurelio Licata
Department of Animal Biology and Marine Ecology, Faculty of Science, University of Messina, Italy
Abstract: Histochemical and immunohistochemical study was carried out on nitrinergic innervation and neuroendocrine system in the gill epithelium of the abyssal fish Coelorhynchus coelorhynchus. The results showed that nNOS-positive nerve fibers, originating from the branchial arch were present in the subepithelial tissue of branchial primary filament. nNOS-positive neuroendocrine cells were also present in the primary filaments and secondary lamellae. Numerous mucous cells in the gill epithelium were AB/PAS-positive, while sialic acid was absent as confirmed by neuraminidase reaction and WGA lectin histochemistry. The mucus compounds in abyssal teleost fish are different from those found in pelagic species, being related to their living conditions. In abyssal species, greater numbers of chloride and neuroendocrine cells are involved in the movement of water and electrolytes. Neuroendocrine cells possess oxygen receptors which mediate the cardiovascular and ventilatory response to oxygen deficiency, as reported in teleost species. Besides, NO contributes through nervous stimulation to the regulation of vascular tone and blood circulation in the gill.
Author’s e-mail: alicata@unime.it
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Stereological analysis of mitochondria in embryos of Rana temporaria and Bufo bufo during cleavage
Marek Romek and Ewa Krzysztofowicz
Department of Cytology and Histology, Institute of Zoology, Jagiellonian University, Cracow, Poland
Abstract: Total numbers of mitochondria and their morphology have been quantitatively determined in mature oocytes and in cleaving embryos of two anuran species Rana temporaria and Bufo bufo using stereological methods. Surface densities of inner mitochondrial membranes for both studied species during cleavage ranged from 5.43 m2/cm3 to 7.53 m2/cm3, whereas volume densities of mitochondria did not exceed 1.65%. Since values of these parameters were low, thus embryos during cleavage may be considered as metabolically “silent”. Transition of ultrastructural morphology of mitochondria towards that characterising actively respiring organelles occurs at stage 9 for R. temporaria and at stage 8 for B. bufo, correlated with blastula-gastrula and mid-blastula transition, respectively. The total numbers of mitochondria N(c) in mature oocytes are as high as 114.8 and 107.2 millions for R. temporaria and B. bufo, respectively, and during cleavage at late blastula stages they increase to 300 millions for both species under study. We suggest that an undefined mechanism might eliminate during cleavage those amphibian embryos which contain small number of mitochondria and low levels of nutrient substances.
Author’s e-mail: romek@zuk.iz.uj.edu.pl
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GA3 content in young and mature antheridia of Chara tomentosa estimated by capillary electrophoresis
Andrzej Kaźmierczak and Dariusz Stępiński
Department of Cytophysiology, University of Łódź, Poland
Abstract: The content of gibberellic acid (GA3) in male sex organs of Chara tomentosa L. was estimated using capillary electrophoresis. Young antheridia contained 0.25 m g GA3 while mature ones 0.48 m g per antheridium. Although there are significant differences in GA3 content in antheridia between C. vulgaris and C. tomentosa, these values calculated per one spermatid are 2.4 and 3.3 pg, respectively. The present results compiled with the previous knowledge about regulation of GA3-dependent development of Characeae species allow an implication that the mechanisms controlling antheridia differentiation in both species can be similar.
Author’s e-mail: kazand@biol.uni.lodz.pl