Cellular & Molecular Biology Letters
International Scientific Journal
Established 1996
Volume 10 (2005) No. 2
| Volume 10 (2005) pp 203-215 | |
| Title | THE ROLE OF PLASMID CONSTRUCTS CONTAINING THE SV40 DNA NUCLEAR-TARGETING SEQUENCE IN CATIONIC LIPIDMEDIATED DNA DELIVERY |
| Authors | Tekkatte Krishnamurthy Prasad and Nalam Madhusudhana Rao* |
| Abstract | One of the steps that limit transfection efficiency in non-viral gene delivery is inefficient nuclear import of plasmid DNA, once it has been delivered into the cytoplasm. Recently, via microinjection into the cytoplasm and in situ hybridizations into a few cell types, it was shown that a region of Simian virus 40(SV40), specifically a c. 372-bp fragment of SV40 genomic DNA encompassing the SV40 promoter-enhancer-origin of replication (SV40 DTS), could enable the nuclear import of a plasmid carrying these sequences (Dean D.A. Exp. Cell Res. 230 (1997) 293). In this report, we address the issue of the suitability of the SV40 DTS for cationic lipid-mediated gene delivery, and its capacity to improve the efficiency of the transfection process. For this study, we used transient reporter gene expression assays on various cell types. The gene expression from the plasmid constructs carrying the SV40 DTS varied with cell type and plasmid construct used. Such cell-type and plasmid-construct dependency on gene expression from plasmids containing the SV40 DTS suggests that the gene expression from plasmids is not entirely dependent on its ability to enhance the nuclear import of said plasmids. |
| Adress and Contact Informations | Centre for Cellular and Molecular Biology, Uppal Road, Hyderabad (AP),
India 500007 * Corresponding author, tel: +91-40-7192552, fax: +91-40-7160591/ 7160311 e-mail: madhu@ccmb.res.in |
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| Volume 10 (2005) pp 217-226 | |
| Title | THE PURINE NUCLEOTIDE CONTENT IN HUMAN LEUKEMIA CELL LINES |
| Authors | Irena Baranowska-Bosiacka1*, Bogus³aw Machaliñski2 and Jolanta Tarasiuk1** |
| Abstract | The HPLC method was used to determine the purine nucleotide (ATP, ADP, AMP, GTP, GDP, GMP, NAD+) contents and the values of the adenylate energy charge (AEC) and guanylate energy charge (GEC) for three human acute myelogenous leukemia (AML) cell lines: HL60 (M3 subtype of AML), THP1 (M5 subtype of AML), and HEL (M6 subtype of AML) in French- American-British classification (FAB) and for one chronic myelogenous leukemia (CML) cell line: K562. The results showed that the examined leukemic cells had some significant changes in their purine nucleotide concentrations relative to healthy cells. On the basis of the obtained results, it seems that two of the tested acute myelogenous leukemia cell lines, HL60 and HEL, have similar purine nucleotide metabolisms, while the third AML cell line, THP1, has a purine nucleotide metabolism like that of the chronic myelogenous leukemia cell line, K562. |
| Adress and Contact Informations | 1Department of Biochemistry, University of Szczecin, ul. Felczaka 3a, 71-412
Szczecin, Poland, 2Department of General Pathology, Pomeranian Medical University, Al. Powstañców Wlkp. 72, 70-111 Szczecin, Poland * Present address: Pomeranian Medical University, Department of Biochemistry and Chemistry, Al. Wielkopolska 72, 70-111 Szczecin, Poland ** Corresponding author; e-mail: tarasiuk@univ.szczecin.pl, tel/fax: (+48) (91) 4441550 |
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| Volume 10 (2005) pp 227-235 | |
| Title | THE BINDING ACTIVITY OF YEAST RNAs TO YEAST hek2p AND MAMMALIAN hnRNP K PROTEINS, DETERMINED USING THE THREE-HYBRID SYSTEM |
| Authors | Agnieszka Paziewska1, Lucjan S. Wyrwicz2 and Jerzy Ostrowski1* |
| Abstract | K homology (KH) domains are scaffolds for the binding of RNAs by the heterogeneous nuclear ribonucleoprotein (hnRNP) K protein and its yeast ortholog, Hek2p. KH domains are remarkably conserved between mammals and yeast. To assess the binding activity for yeast RNA of the two proteins, we used full-length K protein and Hek2p as baits in the yeast three-hybrid system. All the unique RNA sequences bound by Hek2p and all but two bound by K protein represented different fragments of only two transcripts, encoded by the 18S and 25S ribosomal RNA genes. Most of them were transcribed from the antisense strand. The RNA-binding activity of K protein was significantly higher than that of Hek2p. These results and those from our previously published reports demonstrate that the specificity of target RNA recognition by both the K protein and Hek2p depends on both RNA-specific sequences and the structure of the protein. Both mammalian K protein and its yeast ortholog may be involved in the regulation of gene expression. |
| Adress and Contact Informations | 1Department of Gastroenterology, Medical Center for Postgraduate Education at
Maria Sk³odowska-Curie Memorial Cancer Center and Institute of Oncology,
Roentgena 5, 02-781 Warsaw, Poland, 2BioInfoBank Institute, 60-744 Poznañ, Poland * Corresponding author; tel: (4822) 6440102, fax: (4822) 6447601, e-mail: jostrow@warman.com.pl |
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| Volume 10 (2005) pp 237-245 | |
| Title | EFFICIENT SNP ANALYSIS ENABLED BY JOINT APPLICATION OF THE μTGGE AND HETERODUPLEX METHODS |
| Authors | Md. Salimullah1, Keiichi Hamano2,5, Masayoshi Tachibana3,5, Kinji Inoue4,5 and Koichi Nishigaki1,5* |
| Abstract | Gene science-based diagnoses have become an increasingly realistic option as the state of knowledge has improved regarding the genetic basis of disease. To facilitate the creation of this potential diagnostic tool, researchers have made large-scale detection of point mutations a key issue. Here, we propose an inexpensive and convenient method with a high performance level for this purpose: micro temperature gradient gel electrophoresis (mTGGE)- empowered heteroduplex analysis (mTG-HD). First, mTGGE was shown to separate double-stranded DNA containing single nucleotide polymorphism (SNP) with sufficiently high resolution when used in the mode of perpendicular TGGE. Using human c-Ki-ras and rat p53 DNA, point mutations could be unequivocally detected by mTG-HD when parallel TGGE was employed. The mutation type (such as G/C to A/T), the position of the point mutation (centre or not) and the DNA size (around 100 or 200 bp) were examined and found to be detectable. Thus, μTG-HD could detect point mutations efficiently at a much lower cost by having multiple lanes per gel. |
| Adress and Contact Informations | 1Department of Functional Materials Science and 4Department of Regulation
Biology, Saitama University, 255 Shimo-Okubo, Sakura-ku, Saitama-shi,
Saitama 338-8570, Japan, 2TAITEC Corporation, 2693-1 Nishikata, Koshigayashi, Saitama, 343-0822, Japan, 3Brain Research Institute, Niigata University, 1 Asahimachi, Niigata 951-8510, Japan, 5REDS Group, Saitama Small Enterprise Promotion Corporation, SKIP City, Kawaguchi, 333-0844, Japan * Corresponding author: tel/fax: +81-48-858-3533, e-mail:koichi@fms.saitama-u.ac.jp |
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| Volume 10 (2005) pp 247-253 | |
| Title | THE EFFECT OF DNA LABELING WITH THE FLUORESCENT DYES R110 AND R6G ON GENOTYPE ANALYSIS USING CAPILLARY ELECTROPHORESIS |
| Authors | Haseeb Ahmad Khan* |
| Abstract | We investigated the mobility of DNA fragments labeled with the fluorescent dyes R110 and R6G, specifically for use in genotyping using capillary electrophoresis. Genomic DNA was isolated from blood, and a highly polymorphic region of the HLA-C gene was amplified by PCR with the use of either [F]-dNTP-[R110] or [F]-dNTP-[R6G]. Pre-diluted (30-fold) PCR products were mixed with formamide, denatured (at 95ºC for 2 min.), and rapidly cooled on ice before being subjected to electrophoresis. The results showed that the number and mobility of allele-specific DNA fragments were independent of the two dyes used. Both dyes were equally efficient at differentiating homozygous or heterozygous allelic presentation. An additional dye-specific peak of 132- base-pair mobility was observed with the use of [F]-dNTP-[R110]; it significantly impaired the resolution of one allele-specific peak. The electropherograms obtained with [F]-dNTP-[R6G] were free from any interfering peaks within the target region, thus the [R6G]-based procedure is more preferable for genotype analysis. As this procedure does not involve any post-PCR cleanup, it is simple, rapid and cost-effective. |
| Adress and Contact Informations | Department of Biochemistry, College of Science, King Saud University, Riyadh
11451, Saudi Arabia * Tel: +966-1-4676039, fax: +966-1-4675791, e-mail: haseeb@ksu.edu.sa |
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| Volume 10 (2005) pp 255-264 | |
| Title | LIPID PEROXIDATION AND ANTIOXIDANT STATUS IN PATIENTS WITH PERIODONTITIS |
| Authors | Kuppusamy Panjamurthy1, Shanmugam Manoharan1* and Cinnamanoor Rajamani Ramachandran2 |
| Abstract | Our aim was to assess the degree of oxidative stress in patients with periodontitis by measuring their levels of thiobarbituric acid reactive substances (TBARS), enzymatic antioxidants (superoxide dismutase (SOD), catalase (CAT), glutathione peroxide (GSHPx)), and non-enzymatic antioxidants (vitamins –E and C, reduced glutathione (GSH)). This study was conducted on 25 adult chronic periodontitis sufferers who were patients in Rajah Muthiah Dental College and Hospital, Annamalai University. The levels of TBARS and non-enzymatic antioxidants, and the activities of enzymatic antioxidants in the patients' plasma, erythrocytes and gingival tissues were assayed using specific colorimetric methods. The periodontitis sufferers had a significantly higher TBARS level than the healthy subjects. In the plasma, erythrocytes, erythrocyte membranes and gingival tissues of the periodontitis sufferers, enzymatic antioxidant activities were found to be significantly higher, whereas the levels of non-enzymatic antioxidants were significantly lower (except for reduced glutathione in the gingival tissues) relative to the parameters found for healthy subjects. The disturbance in the endogenous antioxidant defense system due to over-production of lipid peroxidation products at inflammatory sites can be related to a higher level of oxidative stress in patients with periodontitis. |
| Adress and Contact Informations | 1Department of Biochemistry, Faculty of Science, Annamalai University,
Annamalai Nagar – 608 002, India, 2Dean, Rajah Muthiah Dental College and Hospital Annamalai University, Annamalai Nagar – 608 002,. India * Corresponding author: e-mail: manshisak@yahoo.com; tel: + 91-4144-238343 (Off.) + 91-4144-232788 (Res.); fax : + 91-4144-238145 |
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| Volume 10 (2005) pp265-280 | |
| Title | A BIOLOGICAL AND MOLECULAR CHARACTERIZATION OF SOME EGYPTIAN BARLEY GENOTYPES WHICH ARE RESISTANT TO NET BLOTCH DISEASE |
| Authors | Mahmoud M. Saker* |
| Abstract | A survey for resistance against net blotch disease (caused by Pyrenophora teres) was performed on some Egyptian barley landraces and some selected resistance and susceptible standard German barley genotypes. The results indicated that most of the Egyptian barley landraces are extremely resistant to the disease. Molecular analysis using RAPD and AFLP showed unique banding profiles for the different genotypes, and specific AFLP markers for the Egyptian genotypes were identified. The effectiveness of RAPD and AFLP for identifying different barley genotypes of different origins and with different reactions against P. teres was discussed. The results of the biological evaluation and molecular characterization done in this study can be seen as the starting point needed to identify the valuable net blotch resistant Egyptian barley germplasm at both the phenotype and genotype levels and draw the attention of breeders and banks of natural plant genetic resources towards this valuable yet neglected germplasm. |
| Adress and Contact Informations | Plant Biotechnology Dept., National Research Centre, Dokki, Cairo, Egypt * fax: 002-02-3370931, tel. 002-02-7617186, e-mail: Sakrmahmoud@yahoo.com |
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| Volume 10 (2005) pp 281-285 | |
| Title | PARALLELISATION OF THE BLAST ALGORITHM |
| Authors | Yutao Qi1* and Feng Lin2 |
| Abstract | Retrieving homologous DNA and protein sequences from existing databases is a fundamental routine in bioinformatics research. Programs of the NCBI BLAST family are widely used for this purpose. We evaluated paraBLAST, a parallelised version of the NCBI BLAST algorithm, using a Message Passing Interface (MPI) on a multi-node compute cluster. Here, we propose static and dynamic database-partitioning schemes based on the availability of the cluster. We evaluated the application of the algorithm in querying nucleotide sequences against a large-scale sequence database with different numbers of database partitions, and hence, different numbers of CPUs. Since the program's tasks are performed independently of each other, each available CPU can run its own copy of BLAST queries, resulting in reduced interference between processes and leading to a highly scalable solution. |
| Adress and Contact Informations | 1,2BioInformatics Research Center, School of Computer Engineering
Nanyang Technological University, Nanyang Avenue, Singapore 639798 * Corresponding author, e-mail: antonioqi@pmail.ntu.edu.sg (Yutao Qi), tel.: +65 67906609, fax: +65 63162780 |
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| Volume 10 (2005) pp 287-303 | |
| Title | THE EXPRESSION OF CONNEXIN 43 IN CHILDREN WITH TETRALOGY OF FALLOT |
| Authors | Jacek Ko³cz1, Justyna Druka³a2, Ma³gorzata Bzowska3, Bart³omiej Rajwa4, W³odzimierz Korohoda2 and Edward Malec1* |
| Abstract | Abnormalities in the expression and distribution of Connexin 43 (Cx43) in cardiomyocytes may lead to anomalous conotruncal embryogenesis and disturbances in the maturation and function of the heart. Tetralogy of Fallot (TOF) is the most frequent, cyanotic congenital heart defect in which conotruncal anomalies, right ventricle dysfunction and life-threatening arrhythmias occur. In this study, age-related changes in the expression and spatial distribution of Cx43 in cardiomyocytes from TOF children compared to patients without right ventricular outflow tract pathology were determined Confocal microscopy and flow cytometry were used to assess the changes. Disturbances in both the expression and distribution of Cx43 were found. In the group of infants with TOF, a lower level of expression of the protein was determined. Cardiomyocytes from TOF hearts were found to have Cx43 distributed over their entire surface, which is the pattern seen in immature tissue. In the controls, Cx43 was located within the intercalated disks. Expression of Cx43 in TOF hearts increases with the age of the subject, whereas its spatial distribution remains the same in both infants and older children. Disturbances in Cx43 expression and localization may influence heart embryogenesis and maturation, contribute to hypertrophy and dysfunction of the right ventricle and induce arrhythmias in children with TOF. Early redistribution of Cx43 and functional maturation of the heart muscle support a strategy of early total correction of the defect. |
| Adress and Contact Informations | 1Department of Pediatric Cardiac Surgery, Polish-American Children's Hospital,
Collegium Medicum, Jagiellonian University, Wielicka 265, 30-663 Kraków,
Poland, 2Department of Cell Biology, Faculty of Biotechnology, Jagiellonian University, Gronostajowa 7, 30-387 Kraków, Poland, 3Department of Immunology, Faculty of Biotechnology, Jagiellonian University, Gronostajowa 7, 30-387 Kraków, Poland, 4Department of Biophysics, Laboratory of Confocal Microscopy and Image Analysis, Jagiellonian University, Gronostajowa 7, 30-387 Kraków, Poland * Corresponding author, tel: 0048 12 658 10 23, fax: 0048 12 657 39 47, e-mail: mimalec@cyf-kr.edu.pl |
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| Volume 10 (2005) pp 305-319 | |
| Title | CYCLIC NUCLEOTIDE PHOSPHODIESTERASES (PDEs) IN HUMAN OSTEOBLASTIC CELLS; THE EFFECT OF PDE INHIBITION ON cAMP ACCUMULATION |
| Authors | Mikael Ahlström*, Minna Pekkinen, Minna Huttunen and Christel Lamberg-Allardt |
| Abstract | The regulation of the secondary messengers, cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), is crucial in the hormonal regulation of bone metabolism. Both cAMP and cGMP are inactivated by cyclic nucleotide phosphodiesterases (PDEs), a superfamily of enzymes divided into 11 families (PDE1-11). We compared the PDEs of cultured human osteoblasts (NHOst) and SaOS-2 osteosarcoma cells. The PDE activity of NHOst cells consisted of PDE1, PDE3 and PDE7, whereas PDE1, PDE7 and PDE4, but no PDE3 activity was detected in SaOS-2 cells. In line with the difference in the PDE profiles, rolipram, a PDE4 inhibitor, increased the accumulation of cAMP in SaOS-2, but not in NHOst cells. Expression of PDE subtypes PDE1C, PDE3A, PDE4A, PDE4B, PDE7A and PDE7B was detected in both cell types. NHOst cells additionally expressed PDE1A. |
| Adress and Contact Informations | Department of Applied Chemistry and Microbiology, University of Helsinki,
P.O. Box 66, 00014 University of Helsinki, Finland *Corresponding author, tel: +358 (9) 1915 8276, fax: +358 (9) 1915 8269, e-mail: mikael.eb.ahlstrom@helsinki.fi |
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| Volume 10 (2005) pp 321-329 | |
| Title | THE EFFECT OF MELATONIN ON LIPID PEROXIDATION AND NITRITE/NITRATE LEVELS, AND ON SUPEROXIDE DISMUTASE AND CATALASE ACTIVITIES IN KAINIC ACID-INDUCED INJURY |
| Authors | Yasemin Delen Akcay1, Ayfer Yalcin2 and Eser Yildirim Sozmen1 |
| Abstract | Kainic acid (KA) initiates neuronal injury and death by inducing oxidative stress and nitric oxide release from various regions of the brain. It was recently shown that melatonin has free radical-scavenging action and may protect against kainate-induced toxicity. In order to assess the possible supportive effect of melatonin treatment in KA-induced injury in the rat brain cortex, we determined malondialdehyde (MDA) levels as an index of lipid peroxidation, and assessed the activities of catalase (CAT) and superoxide dismutase (SOD) and the levels of nitrite/nitrate 35 male rats were divided into five groups, each receiving a different intraperitoneal treatment: saline solution (0.2 ml), kainic acid (15 mg/kg), melatonin (20 mg/kg), KA then melatonin (each as above, 15 min apart), or melatonin then KA (each as above, 30 min apart). Administration of KA caused an about five-fold increase in the catalase activity and an increase in the SOD activity in the cortex relative to the activities for the controls. Treatment with melatonin 15 min after KA injection kept malondialdehyde levels and catalase and superoxide dismutase activities at the normal levels, and led to an increase in the levels of nitrite/nitrate. Our data suggests that melatonin treatment following KA administration has a protective effect on antioxidant enzyme activities and thus supports the role of melatonin and oxidative stress in the regulation of antioxidative enzyme activity. |
| Adress and Contact Informations | 11Department of Biochemistry, Faculty of Medicine, Ege University, 35100
Bornova, Izmir, Turkey, 2Department of Biochemistry, Faculty of Pharmacy, Ege University, 35100 Bornova, Izmir, Turkey |
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| Volume 10 (2005) pp 331-344 | |
| Title | CONSTRUCTION OF A GENETIC LINKAGE MAP USING MFLP AND IDENTIFICATION OF MOLECULAR MARKERS LINKED TO DOMESTICATION GENES IN NARROW-LEAFED LUPIN (Lupinus angustifolius L.) |
| Authors | Jeffrey G. Boersma1,2*, Margaret Pallotta3, Chengdao Li1, Bevan J. Buirchell1, Krishnapillai Sivasithamparam2 and Huaan Yang1 |
| Abstract | A mapping population of F8 derived recombinant inbred lines (RILs) was established from a cross between a domesticated breeding line 83A:476 and a wild type P27255 in narrow-leaf lupin (Lupinus angustifolius L.). The parents together with the 89 RILs were subjected to DNA fingerprinting using microsatellite-anchored fragment length polymorphism (MFLP) to rapidly generate DNA markers to construct a linkage map. Five hundred and twenty two unique markers of which 21% were co-dominant, were generated and mapped. Phenotypic data for the domestication traits: mollis (soft seeds), leucospermus (white flower and seed colour); Lentus (reduced pod-shattering), iucundis (low alkaloid), Ku (early flowering) and moustache pattern on seed coats; were included. Three to 7 molecular markers were identified within 5cM of each of these domestication genes. The anthracnose resistance gene Lanr1 was also mapped. Linkage groups were constructed using MapManager version QTXb20, resulting in 21linkage groups consisting of 7 or more markers. The total map length was 1543cM, with an average distance of 3.4cM between adjacent markers. This is the first published map for a lupin species. The map can be exploited for marker assisted selection for genetic improvement in lupin breeding programs.. |
| Adress and Contact Informations | 1Department of Agriculture, Western Australia, 3 Baron-Hay Court, South Perth,
W.A. 6151, Australia, 2School of Earth and Geographical Sciences, The University of Western Australia, Nedlands, W.A. 6009, Australia, 3Australian Centre for Plant Functional Genomics, Hartley Grove, Waite Campus, Urrbrae, S.A. 5064, Australia * Corresponding author, e-mail: jboersma@agric.wa.gov.au |
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| Volume 10 (2005) pp 345-362 | |
| Title | In-vitro ANALYSIS OF THE EXPRESSION OF TGFb-SUPERFAMILYMEMBERS DURING CHONDROGENIC DIFFERENTIATION OF MESENCHYMAL STEM CELLS AND CHONDROCYTES DURING DEDIFFERENTIATION IN CELL CULTURE |
| Authors | Ulrich Reinhart Goessler1*, Peter Bugert2, Karen Bieback2, Moritz Deml1, Haneen Sadick1, Karl Hormann1 and Frank Riedel1 |
| Abstract | Traditional surgical methods for the reconstruction of cartilage defects rely on the transplantation of autologous and allogenous tissues. The disadvantages of these techniques are the limited availability of suitable tissues and the donor site morbidity of transplants. In addition, in cultured chondrocytes, the dedifferentiation of cells seems unavoidable during multiplication. In this study, we investigated the expression of distinct markers during the dedifferentiation of human chondrocytes (HC) and human mesenchymal stem cells (MSC) in cell culture using microarray technique, immunohistochemistry and RT-PCR. Transforming growth factor b (TGFb) is a multifunctional peptide that plays play a crucial role in inducing and maintaining chondrogenic differentiation. In dedifferentiating chondrocytes, the gene for TGFb1 was constantly expressed, while the gene for TGFb2 was never expressed. The genes for TGF a, TGFb4 and TGFbi were activated with ongoing dedifferentiation. TGFb-receptor 3 was constantly expressed, while the genes for the TGFb-receptors 1 and 2 were never expressed. Immunohistochemical staining for TGFb3 revealed upregulation in the course of dedifferentiation. The genes for LTBP1 and LTBP2 were activated with ongoing dedifferentiation, whereas the gene for LTBP3 was constantly expressed, and negative results were obtained for the gene for LTBP4. The genes for LTBP1 and LTBP2 were activated with ongoing dedifferentiation. During chondrogenic differentiation, the MSCs constantly expressed TGFb1, b2, b3 and b4. LTBP1, LTBP2 and TGFb-R3 were downregulated. In conclusion, TGFb3, TGFb4, TGFbi, LTBP1 and LTBP2 may assist the process of dedifferentiation, while TGFb1 and b2 might not be involved in this process. Of the TGFb-receptors, only type 3 might be involved in dedifferentiation. |
| Adress and Contact Informations | 1Department of Otolaryngology, Head and Neck Surgery, 2Institute of Transfusion Medicine and Immunology, Red Cross Blood Service of Baden- Württemberg/Hessen, Ruprecht-Karls-University Heidelberg, Faculty of Clinical Medicine Mannheim, Germany * Corresponding author; tel: (+49) 621 383 1600, fax: (+49) 621 383 3827, e-mail: ugoessler@web.de |
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Office:
Cellular & Molecular Biology Letters
Faculty of Biotechnology
University of Wroc³aw
Przybyszewskiego 63/77
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Faculty of Biotechnology
University of Wroc³aw
Przybyszewskiego 63/77
51-148 Wroc³aw, Poland
fax:
+48 71 375 6208 or
+48 71 375 6234
+48 71 375 6234
e-mail:
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cmbl@ibmb.uni.wroc.pl