Volume 50 (2004) No. 1

Volume 50 (2004) No. 1

Original Articles
Eukaryotic Operon Genes Can Define Highly Conserved Syntenies
Z. TRACHTULEC...........................................................................1
Department of Mammalian Molecular Genetics and Centre for Integrated Genomics, Institute of Molecular Genetics,
Academy of Sciences of the Czech Republic, Prague, Czech Republic
Corresponding author: Zdenek Trachtulec, Institute of Molecular Genetics, Academy of Sciences of the Czech Republic,
Vídeňská 1083, 142 20 Prague, Czech Republic. Tel.: +420 296 442 256, Fax: +420 296 442 154; e-mail: trachtul(zavináč)biomed.cas.cz .
Abstract.
Full text. 1-6

Loss of Tumorigenicity of Murine Colon Carcinoma MC38/0 Cell Line after Transduction with
a Retroviral Vector Carrying Murine IL-12 Genes
E. Pajtasz-Piasecka¹, A. Szyda¹, J. Rossowska¹, A. Krawczenko¹, M. Indrová³, P. Grabarczyk², P. Wysocki², A. Mackiewicz², D. Duś¹.....................................7
¹Ludwik Hirszfeld Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Wrocław, Poland
²Department of Cancer Immunology, Chair of Oncology, University of Medical Sciences at Great Poland Cancer Centre,
Poznań, Poland
^₃Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Prague, Czech Republic
Corresponding author: Elżbieta Pajtasz-Piasecka, Institute of Immunology and Experimental Therapy,
Polish Academy of Sciences, Weigla 12, 53-114 Wroclaw, Poland. E-mail: pajtasz@iitd.pan.wroc.pl.
Abstract.
Full text. 7-14

A Note on the Decreased Number and Loss of Fibrillar Centers in Nucleoli of Apoptotic HL-60 Leukaemic
Granulocytic Precursors Produced by 5-Aminolaevulinic Acid-Based Photodynamic Treatment
K. Smetana¹, D. Grebeňová¹, I. Jirásková¹, M. Doubek², Y. Marinov¹,
Z. Hrkal¹..........................................................................15
¹Institute of Haematology and Blood Transfusion, Prague, Czech Republic
²State Health Institute, Prague, Czech Republic
Corresponding author: Karel Smetana, Institute of Haematology and Blood Transfusion, U Nemocnice 1,
12800 Prague 2, Czech Republic. Fax: (+420) 221 977 249; email: karel.smetana@uhkt.cz.
Abstract.
Full text. 15-20

Short Communication
Fluorescence-Based Automated Fragment Analysis of Microsatellite Polymorphism within the Transmembrane
Region of the MIC-A Gene
P. NOVOTA^{1, 2}, L. KOLESAR², A. SLAVCEV², M. CERNA¹........................21
¹Department of Cell and Molecular Biology, 3^rd Faculty of Medicine, Prague, Czech Republic
²Department of Immunogenetics, Institute for Clinical and Experimental Medicine, Prague, Czech Republic
Corresponding author: Peter Novota, Department of Immunogenetics (Z4), Institute for Clinical and Experimental Medicine,
Videnska 1958/9, 140 21 Prague 4, Czech Republic.
Abstract.
Full text. 21-23

The Alterations of Immunological Reactivity in Heroin Addicts and Their Normalization in Patients
Maintained on Methadone
A. ZAJICOVA^1,2, H. WILCZEK³, V. HOLAN^1,2............................24
¹Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Prague, Czech Republic
²Faculty of Science, Charles University, Prague, Czech Republic
³3^rd Medical Clinic, 1^st Faculty of Medicine, Charles University, Prague, Czech republic
Corresponding author: Alena Zajicova, Institute of Molecular Genetics, Academy of Sciences of the Czech Republic,
Flemingovo nam. 2, 166 37 Prague 6, Czech Republic. Tel.: 420 220 183 460; Fax: 420 224 310 955;
e-mail: zajicova@img.cas.cz
Abstract.
Full text. 24-28

Monoclonal Antibody Register
Monoclonal Antibody Produced against Bovine MHC Class I Antigens
J. ANTALIKOVA, M. SIMON, L. HOROVSKA, J. VALENTOVICOVA
Institute of Animal Biochemistry and Genetics, Slovak Academy of Sciences, Ivanka pri Dunaji, Slovakia
Corresponding author: Jana Antalikova, Institute of Animal Biochemistry and Genetics, Slovak Academy of Sciences,
Ivanka pri Dunaji, Slovakia. Tel.: +421 2 4593 151; Fax: +421 2 4593932; e-mail: Jana.Antalikova@savba.sk
Background.
Full text. 29-31

Original Articles
Eukaryotic Operon Genes Can Define Highly Conserved Syntenies
Z. TRACHTULEC

The synteny conservation of the members of eukaryotic operons was investigated by mapping their
orthologues in Drosophila, human, and other eukaryotes. While the homologues of the operon members are generally
not linked, some examples of highly conserved syntenies were found. The most significant synteny involves two members
of one C. elegans operon, encoding fibrillarin and ribosomal protein S16. Their homologues are linked in human, mouse,
Drosophila, Anopheles gambiae, Saccharomyces cerevisiae, Schizosaccharomyces pombe, Plasmodium falciparum,
and Guillardia theta, but not in five other genomes. The distances between the genes are larger than in the nematode,
suggesting the prevalence of intrachromosomal rearrangements.
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Loss of Tumorigenicity of Murine Colon Carcinoma MC38/0 Cell Line after Transduction with
a Retroviral Vector Carrying Murine IL-12 Genes

E. Pajtasz-Piasecka, A. Szyda, J. Rossowska, A. Krawczenko, M. Indrová, P. Grabarczyk,
P. Wysocki, A. Mackiewicz, D. Duś

Cells of transplantable MC38 colon carcinoma of C57BL/6 mice were adapted to growth in vitro as the MC38/0 cell line.

Along the establishing process, MC38/0 cells preserved their tumorigenicity. After transduction with a retroviral vector carrying
murine interleukin 12 (mIL-12) genes and further selection, stable MC38/IL-12 transductant cells were obtained. These cells produced
IL-12 (approx. 2500 ng/ml /5x10⁵ cells /48 h) as evaluated in the optimized bioassay. After subcutaneous inoculation into syngeneic mice,
the IL-12-modified cells demonstrated reduced tumorigenicity as compared to parental MC38/0 cells. Mice that rejected
the MC38/IL-12 tumour became protected against subsequent challenge with MC38/0 cells. The obtained data indicate
that the IL-12-transduced murine colon carcinoma cells could be used both as a model tumour for the study of mechanisms
of anticancer immunity and/or as an adjuvant to cancer vaccines.
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The nuclear and nucleolar ultrastructure was studied by means of conventional transmission electron microscopy to provide
more and complementary information on nucleolar changes accompanying the apoptotic process in leukaemic granulocytic precursors
(HL-60 cells) produced by PDT without previous terminal differentiation. PDT induced the apoptotic process using BL irradiation and
ALA as a precursor of the photosensitizer protoporphyrin IX. PDT produced marked changes of the nucleolar ultrastructure in apoptotic
cells, such as reduction of the number and loss of fibrillar centers surrounding dense fibrillar components. Such nucleolar changes are
known to reflect an alteration of nucleolar biosynthetic activities, which are believed to be located at the periphery of fibrillar centers.
Some electron micrographs also indicated that fibrillar centers apparently migrated out from nucleolar bodies.
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Short Communication

Fluorescence-Based Automated Fragment Analysis of Microsatellite Polymorphism within the Transmembrane
Region of the MIC-A Gene
P. NOVOTA, L. KOLESAR, A. SLAVCEV, M. CERNA

MHC class I chain-related genes (MIC) are located within the MHC class I region of chromosome 6. Sequence analysis of the
MIC-A gene showed a trinucleotide repeat (GCT) microsatellite polymorphism within the transmembrane region. So far, six alleles
of the exon 5 of the MIC-A gene, which consist of 4, 5, 6, 9 and 10 repetitions of GCT, or five repetitions of GCT with an additional
nucleotide insertion (GGCT), have been identified. Recent works support the findings that MIC-A is associated with several
autoimmune diseases. In our work we present a modification of a method used for microsatellite polymorphism detection
within the transmembrane region of the MIC-A gene. It is the ALFexpress fluorescent-based automated fragment analysis. We also
present the frequencies of MIC-A exon 5 alleles found in the Czech population. We have identified five alleles of the transmembrane
region of MIC-A, which comprise 4, 5, 6 and 9 repetitions or five repetitions with an additional nucleotide insertion. The most frequent
allele was A5.1 (59.3%) and the less frequent was the allele A5 (20.0%). No A7, A8 or A10 alleles were identified.
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The Alterations of Immunological Reactivity in Heroin Addicts and Their Normalization in Patients
Maintained on Methadone
A. ZAJICOVA, H. WILCZEK, V. HOLAN

Drug addiction influences many physiological functions including reactions of the immune system. The higher occurence of infectious
and other diseases in drug addicts has been explained by the depression of immunity due to the harmful effects of the drug.
To test this assumption, we tested the proliferative responsiveness and cytokine production of PBL from a group of heroin addicts
(N = 19), patients maintained on methadone (N = 15) and healthy controls (N =15). The results show that Con A-induced proliferation
of PBL from heroin addicts was even enhanced in comparison with PBL from the control group. Similarly, production of IL-2, IL-10 and
IFN-gamma was higher in the group of heroin addicts than in healthy controls. The enhanced proliferation of PBL or the increased
production of cytokines observed in heroin addicts was partially or completely normalized in the group of patients maintained on methadone.
A significantly higher production of IL-6 was found in both unstimulated and stimulated PBL from heroin addicts and patients
maintained on methadone, when compared with PBL from healthy controls. The results thus showed enhanced proliferative activity
and increased production of various cytokines in heroin addicts and partial or complete adjustment of these alterations in patients maintained
on methadone.
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Monoclonal Antibody Register

Monoclonal Antibody Produced against Bovine MHC Class I Antigens
J. ANTALIKOVA, M. SIMON, L. HOROVSKA, J. VALENTOVICOVA

The major histocompatibility complex (MHC) genes encode cell glycoproteins that bind and present antigenic peptides to T cells.
The analysis of MHC gene expression, as the key molecule of the immune system, is thus an essential component of studies of immune
responses and susceptibility to diseases. The MHC class I molecules present endogenously synthetized peptides to CD8⁺ cytotoxic T cells
(York and Rock, 1996) and these molecules (MHC I) are also recognized by inhibitory receptors of natural killer cells (Lanier, 1998).
The "classical" MHC class I molecules of man and other studied species are expressed on most nucleated cells (York and Rock, 1996).
They consist of heterodimers of highly polymorphic α chains (Mr 44 kDa) non-covalently associated with the invariant β2-microglobulin
subunit (Mr 12 kDa) (Ploegh et al., 1981).
The bovine MHC is denoted as BoLA. One class I locus, BoLA-A, has been confirmed serologically (Spooner et al., 1979;
Davies et al., 1994) and the second locus has been suggested by biochemical evidence (Bensaid et al.,1988, 1991). Cytotoxic T
lymphocyte recognition of parasitic antigens associated with MHC class I has also been demonstrated (Goddeeris et al., 1986),
confirming the immunological importance of class I MHC molecules in cattle.
Knowledge about MHC class I structure and their biological function has been advanced by the extensive development of monoclonal
antibody (mAb) reagents. In cattle, most of the MHC class I molecule analyses were performed by cross-reactive mAb W6/32
(Parham et al. 1979; Kahn-Perles et al., 1987) commonly used for human MHC study. Only a few mouse mAbs generated against
bovine cells were reported (Bensaid et al., 1989). This paper describes the antibody IVA-26 formed against bovine cells detecting MHC
class I molecule of several species and showing analogical features with mAb MEM-147 (Tran et al., 2001) that recognizes MHC class I
on human cells.
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