Jolanta Krzysztoń-Russjan - Academia.edu (original) (raw)

Papers by Jolanta Krzysztoń-Russjan

Research paper thumbnail of Synergistic action of doxorubicin and sulindac in human cervix carcinoma cells - studies on possible mechanisms

Medical science monitor : international medical journal of experimental and clinical research, 2010

Epidemiologic and experimental studies have shown that cyclooxygenase-2 (COX-2) inhibitors as non... more Epidemiologic and experimental studies have shown that cyclooxygenase-2 (COX-2) inhibitors as non-steroidal anti-inflammatory drugs (NSAIDs) are effective chemopreventive agents. The mechanisms underlying the antitumor activity of COX-2 inhibitors are thought to involve inhibition of COX-2 enzyme activity and induction of apoptosis. The aim of the current work was to study the mechanisms of synergistic action noted in HeLa cervical carcinoma cells under doxorubicin (DOX) and sulindac (SUL) co-treatment. Cytotoxic activity of the drugs was defined with MTT test, apoptosis was detected with TUNEL test, DOX transmembrane efflux was measured fluorometrically, expression of MDR-1 and MRP-1 was determined with quantitative real time - PCR (QRT-PCR). It was shown that SUL at non-toxic concentrations, 10 and 50 microM, is an effective enhancer of cytotoxic action for DOX in 0.5 and 1 microM, respectively; however, only for SUL concentration equal to 50 microM potentiated apoptosis induced b...

Research paper thumbnail of Huntington' disease--imbalance of amino acid levels in plasma of patients and mutation carriers

Annals of agricultural and environmental medicine : AAEM, 2013

Determination of the plasma amino acid (AA) levels in Huntington's disease (HD) can make it p... more Determination of the plasma amino acid (AA) levels in Huntington's disease (HD) can make it possible to find the metabolic markers used in early diagnosis. The aim of the presented study was to determine the AA profile in plasma samples from HD patients and presymptomatic carriers, compared to healthy subjects. The AA profile was analyzed with HPLC. The study concerned 59 participants: 30 subjects with abnormal CAG repeats expansion (>36) in the HTT gene, and 29 healthy subjects. Each participant was analyzed with regard to the parameters characterizing the metabolic state and protein metabolism, such as: urea, creatinine, glucose, total protein, TSH (thyroid-stimulating hormone), cortisol, ESR (erythrocyte sedimentation rate), and CRP (C-reactive protein). Simple statistical comparisons showed 5 AA to be significantly lower in the HD group, compared to the control group, i.e.: Asn, His, Leu, Ser, Thr. Creatinine and creatinine clirens were found to be lower in the HD group, ...

Research paper thumbnail of A study of molecular changes relating to energy metabolism and cellular stress in people with Huntington’s disease: looking for biomarkers

Journal of Bioenergetics and Biomembranes, 2012

Huntington's disease (HD) is a neurodegenerative disorder characterized by a progressive motor an... more Huntington's disease (HD) is a neurodegenerative disorder characterized by a progressive motor and cognitive decline and the development of psychiatric symptoms. The origin of molecular and biochemical disturbances in HD is a mutation in the HTT gene, which is autosomally dominantly inherited. The altered huntingtin protein is ubiquitously expressed in the CNS, as well as in peripheral tissues. In this study we measured the metabolism changes in gene transcription in blood of HD gene carriers (premanifest and manifest combined) versus 28 healthy controls. The comparison revealed statistically significant Global Pattern Recognition Fold Change (FC) for 6 mRNA transcripts, reflecting an increase of: MAOB (FC03.07; p00.0005) which encodes an outer mitochondrial membrane-bound enzyme called monoamine oxidase type B; TGM2 (FC01.8; p00.02) encoding a transglutaminase 2 that mediates cellular stress; SLC2A4 (FC0 1.64; p00.02) solute carrier family 2 (facilitated glucose transporter) member 4; branched chain ketoacid dehydrogenase kinase (BCKDK) (FC01.34; p00.02); decrease of LDHA (FC0−1.16; p00.03) lactate dehydrogenase A; and brainderived neurotrophic factor (BDNF) (FC0−2,11; p00.03). These distinguished changes coincided with HD progress. The analyses of gene transcription levels in sub-cohorts confirmed these changes and also revealed 28 statistically significant FCs of gene transcripts involved in ATP production and BCAA metabolism.

Research paper thumbnail of First community-acquired methicillin-resistant Staphylococcus aureus (MRSA) strains in Croatia

Clinical Microbiology and Infection, 2006

received 40 mg dexamethasone over the course of 4 days. By day 20 the patient had recovered. Conf... more received 40 mg dexamethasone over the course of 4 days. By day 20 the patient had recovered. Conflicting results concerning the clinical efficacy of pristinamycin against macrolide-resistant S. pneumoniae with the MLS B phenotype have been reported [1,9,10]. In a double-blind study comparing the efficacy of pristinamycin with amoxycillin for the treatment of communityacquired pneumonia, pristinamycin was significantly less effective than amoxycillin (63.6% vs. 86.6%) in bacteraemic patients, with a 50% failure rate for macrolide-resistant S. pneumoniae [2]. Taken together, these data emphasise the need to reconsider the choice of pristinamycin or quinupristin ⁄ dalfopristin for the treatment of macrolide-resistant pneumococcal infections.

Research paper thumbnail of New Method for Typing Staphylococcus aureus Strains: Multiple-Locus Variable-Number Tandem Repeat Analysis of Polymorphism and Genetic Relationships of Clinical Isolates

Journal of Clinical Microbiology, 2003

The PCR-based methodology applied to multiple-locus variable numbers of tandem repeat (VNTR) anal... more The PCR-based methodology applied to multiple-locus variable numbers of tandem repeat (VNTR) analysis was recently shown to be a useful technique for the molecular typing of clinical isolates of several bacterial species. We have adopted this method for the molecular typing of methicillin-resistant Staphylococcus aureus . Five staphylococcal VNTR loci ( sdr , clfA , clfB , ssp , and spa ) were subjected to analysis, and it was shown that the method allows typing of S . aureus strains with the discriminatory power and reproducibility of pulsed-field gel electrophoresis while at the same time being rapid and applicable to analysis of large numbers of isolates.

Research paper thumbnail of Comparison of multiple-locus variable-number tandem repeat analysis and pulsed-field gel electrophoresis in a setting of polyclonal endemicity of vancomycin-resistant Enterococcus faecium

Clinical Microbiology and Infection, 2008

Research paper thumbnail of Huntington' disease--imbalance of amino acid levels in plasma of patients and mutation carriers

Annals of agricultural and environmental medicine : AAEM, 2013

Determination of the plasma amino acid (AA) levels in Huntington's disease (HD) can make it p... more Determination of the plasma amino acid (AA) levels in Huntington's disease (HD) can make it possible to find the metabolic markers used in early diagnosis. The aim of the presented study was to determine the AA profile in plasma samples from HD patients and presymptomatic carriers, compared to healthy subjects. The AA profile was analyzed with HPLC. The study concerned 59 participants: 30 subjects with abnormal CAG repeats expansion (>36) in the HTT gene, and 29 healthy subjects. Each participant was analyzed with regard to the parameters characterizing the metabolic state and protein metabolism, such as: urea, creatinine, glucose, total protein, TSH (thyroid-stimulating hormone), cortisol, ESR (erythrocyte sedimentation rate), and CRP (C-reactive protein). Simple statistical comparisons showed 5 AA to be significantly lower in the HD group, compared to the control group, i.e.: Asn, His, Leu, Ser, Thr. Creatinine and creatinine clirens were found to be lower in the HD group, ...

Research paper thumbnail of Characterization of coagulase-negative staphylococci isolated from cases of ostitis and osteomyelitis

Polish journal of microbiology, 2006

Coagulase-negative staphylococci (CoNS) are often responsible for cases of chronic ostitis and os... more Coagulase-negative staphylococci (CoNS) are often responsible for cases of chronic ostitis and osteomyelitis, especially in patients with orthopedic prosthesis/implants. The aim of this study was to characterize CoNS isolated from ambulatory patients with chronic ostitis/osteomyelitis and to compare them by PFGE (pulsed-field gel electrophoresis). Out of 263 bacterial strains isolated from wounds/sinuses of patients with chronic ostitis/osteomylitis, 41 were identified as CoNS. Twenty methicillin-resistant strains were selected for this study. Our results confirm the superior performance of cefoxitin disk test to detect methicillin resistance in heterogenous population of CoNS. High level of antibiotic resistance was observed among the studied strains: majority of CoNS were resistant to tetracycline and erythromycin and also to clindamycin and ciprofloxacin. Importantly, in 15 out of 20 studied CoNS different phenotypes of macrolides, lincosamides and streptogramin--MLS resistance w...

Research paper thumbnail of Modulation of apoptosis protein profiles--role of P-gp in HeLa cells exposed to doxorubicin

Cancer genomics & proteomics, 2012

As shown previously doxorubicin (1 μM) plus sulindac (50 μM) reduced the expression of ABCB1 (ATP... more As shown previously doxorubicin (1 μM) plus sulindac (50 μM) reduced the expression of ABCB1 (ATP-binding cassette, sub-family B (MDR/TAP), member 1) mRNA in HeLa cells and this effect was accompanied by increased apoptosis. The aim of this study was to define if the decrease of ABCB1 expression or blocking of P-glycoprotein (P-gp) can affect the expression of the apoptotic genes determined with use of quantitative real time polymerase chain reaction (qRT-PCR). Western blot was used for visualization of chosen pro- and antiapoptotic proteins. Doxorubicin was the main compound which affected the apoptotic genes. The effectiveness of the drugs in reducing of P-gp function has been shown as not being related to the regulation of apoptotic gene transcription. In this experimental scheme, regulation of apoptotic gene transcription depended on the kind of P-gp modulator.

Research paper thumbnail of Doxorubicin Treatment of Cancer Cells Impairs Reverse Transcription and Affects the Interpretation of RT-qPCR Results

Cancer genomics & proteomics, 2016

BACKGROUND Doxorubicin (DOX) acts in a variety of ways including DNA damage and enzyme inhibition... more BACKGROUND Doxorubicin (DOX) acts in a variety of ways including DNA damage and enzyme inhibition, which consequently causes changes in gene expression of cells treated with this agent. Practical validation of experimental results followed by appropriate normalization of the factors investigated is crucial for obtaining biologically relevant results in gene expression studies. MATERIALS AND METHODS Six candidates were evaluated regarding their validity as internal reference genes: RPS23, FLOT2, UBB, ABCF1, ACTB, HPRT1. Optimization for quantitative polymerase reaction (qPCR) included: sensitivity, specificity, amplification efficiency and linear dynamic range determination. The gene expression stability was evaluated by real-time quantitative polymerase reaction (RT-qPCR) in two human cervical cancer cell lines: HeLa and DOX-resistant KB-V1 Cells treated under various concentrations of DOX. RESULTS DOX treatment changed gene expression and led to re-optimization of the cDNA template...

Research paper thumbnail of Zmiany w epidemiologii inwazyjnych klonów MRSA w wybranych szpitalach w Polsce

Research paper thumbnail of Possible role of HSP60 in synergistic action of anthracyclines and sulindac in HeLa cells

As was observed in the earlier studies, doxorubicin (DOX) induced apoptosis in HeLa cells and tha... more As was observed in the earlier studies, doxorubicin (DOX) induced apoptosis in HeLa cells and that effect was potentiated significantly by sulindac (SUL). The aim of the current work was to study the effects of DOX and SUL on HSP60, HSF1 and HSP60 expression and the influence of DOX and SUL on HSP60 translocation. Expression of HSP60 and HSF1 was determined with QRT-PCR; the expression and localization of HSP60 were evaluated with Western blot. The 24-h cell cultures were co-incubated with DOX - 1 microM and/or SUL - 50 microM. The significant induction of HSF1 and HSP60 mRNA level was observed after exposure of the cells to DOX 1 microM. SUL 50 microM alone caused moderate increase in mRNA level. The significant decrease in expression of HSF1 and HSP60 was noted after DOX 1 microM and SUL 50 microM simultaneous treatment. HSP60 appeared in the higher levels in cytosol than in mitochondria No intracellular translocation was noted under treatment of the cells to DOX and/or SUL. In co...

Research paper thumbnail of Formation and preclinical evaluation of a new alloplastic injectable bone substitute material

Alloplastic bone substitute materials are raising some more interest as an alternative for autolo... more Alloplastic bone substitute materials are raising some more interest as an alternative for autologic transplants and xenogenic materials especially in oral surgery over the last few years. These non-immunogenic and completely resorbable biomaterials are the basis for complete and predictable guided bone regeneration. In the majority of cases, such a material is chosen because of its convenient application by surgeons. The main objective of our project was to design and fabricate an osteoconductive, injectable and readily tolerable by human tissues biomaterial for guided bone regeneration. For this purpose, a self-setting composite consisting of chitosan/tricalcium phosphate microparticles and sodium alginate was made. The material obtained was characterized by microsphere and agglomerate morphology and microstructure. Its features relating to setting time and mechanical properties were precisely investigated. Our material was also evaluated according to PN-EN ISO 10993 Biological ev...

Research paper thumbnail of Usefulness of Multiple-Locus VNTR Fingerprinting in detection of clonality of community- and hospital-acquired Staphylococcus aureus isolates

Antonie van Leeuwenhoek, 2008

Staphylococcus aureus has become a major source of hospital infections and the risk of colonisati... more Staphylococcus aureus has become a major source of hospital infections and the risk of colonisation and infection by community-acquired methicillin-resistant S. aureus (CA-MRSA) is increasingly higher. Because of the importance of S. aureus to public health, many molecular typing methods have been developed to determine its transmission routes and source of infection during epidemiological investigations. In this study we evaluated the usefulness of multiplex PCR based Multi-Locus VNTR Fingerprinting (MLVF) as the first step method for rapid differentiation of Croatian and Polish S. aureus isolates in hospital and community settings. This is a first report of the usefulness of MLVF in typing of hospital-acquired methicillin-sensitive S. aureus (HA-MSSA) and four CA-MRSA isolates. A total of 47 isolates of S. aureus recovered in Croatia in 2004 and in Poland in 2006 and 2007 were tested. The MLVF results were compared to those produced by other typing methods, such as Pulsed-Field Gel Electrophoresis (PFGE), Multi-Locus Sequence Typing (MLST) and spa typing. The MLVF analysis showed almost the same clonality results as the remaining typing methods although some differences were found. Epidemiological data about the relation among S. aureus isolates and the results produced by typing methods applied in the present study indicate that because of the advantages in ease and speed of Variable Number of Tandem Repeats (VNTR) procedure over PFGE, spa typing and MLST, MLVF can be used as a first screening method followed by additional typing.

Research paper thumbnail of Changes in epidemiology of invasive MRSA clones in selected Polish hospitals

Introduction: Staphylococcus aureus is a pathogen that can cause a wide spectrum of diseases from... more Introduction: Staphylococcus aureus is a pathogen that can cause a wide spectrum of diseases from relatively mild skin infections to life-threatening systemic disease. The study was conducted to assess the clonality of methicillin-resistant isolates of S. aureus (MRSA) in selected Polish hospitals. Material and Methods: A collection of strains consisted of 183 methicillin-resistant isolates of S. aureus (MRSA) and 4 methicillin susceptible isolates of S. aureus (MSSA) from 2002-2005. Isolates came from 29 hospitals. Purified total DNA of isolates was used to amplify the region X of the gene spa. Subsequently, the amplicons were sequenced and spa

Research paper thumbnail of F09 The study on molecular changes related to energy metabolism in Huntington's disease subjects: looking for biomarkers

Journal of Neurology, Neurosurgery & Psychiatry, 2012

Huntington's disease (HD) is a neurodegenerative disorder characterised by a progressive moto... more Huntington's disease (HD) is a neurodegenerative disorder characterised by a progressive motor and cognitive decline and psychiatric symptoms. The origin of molecular and biochemical disturbances in HD is the genetic defect in the HTT gene, autosomally dominantly inherited. The altered huntingtin protein is ubiquitously expressed in the Central Nervous System as well as in peripheral tissues. The aim of the study was to detect the metabolism changes on the transcription level in blood cells in HD subjects (n=29) with regards to the control, healthy subjects (n=28). The gene expression analysis was performed using quantitative PCR and StellAarray system. A panel of 94 genes involved in glycolysis, Krebs cycle, electron transport chain, BCAAs metabolism and other processes were selected to study. The results calculations were conducted by Global Pattern Recognition software with ΔΔCt type of analysis application. The comparison between HD and control revealed statistically significant Global Pattern Recognition fold change (FC) values for six transcripts, including a decrease of BDNF (−2.11), LDHA (−1.16) and an increase of BCKDK (1.34), MAOB (3.07), SLC2A4 (1.64) and TGM2 (1.8). Multiple analyses in sub-cohorts distinguished by gender, age, BMI, calf circumference, nutrition status, CAG number repeats, TFC, UHDRS motor and HD time from onset also showed 34 transcript FCs. The widest FC ranges decrease concerned to BDNF (from −1.66 to −3.6), BCAT2 (from 1.85 to −1.42) and increase to MAOB (from 1.87 to 7.89) transcript levels that coincided together with the length of HD duration period and the HD progress.

Research paper thumbnail of Development of New PCR Assay with SYBR Green I for Detection of Mycoplasma, Acholeplasma, and Ureaplasma sp. in Cell Cultures

Diagnostics

Mycoplasma, Acholeplasma, and Ureaplasma sp. are atypical bacteria responsible for in vitro cell ... more Mycoplasma, Acholeplasma, and Ureaplasma sp. are atypical bacteria responsible for in vitro cell culture contaminations that can warp the results. These bacteria also cause human and animal infections and may lead to chronic diseases. In developed polymerase chain reaction (PCR) in this study a quantitative PCR with SYBR Green I fluorochrome was applied to facilitate the Mycoplasma, Acholeplasma, and Ureaplasma sp. DNA detection and identification. Screening Test-1 v.1 (triplex qPCR) allowed for the detection of 11 species. Test-1 v.2 (three single qPCRs) pre-identified three subgroups, allowing for the reduction of using single qPCRs in Test-2 for species identification. The range of both tests was consistent with pharmacopeial requirements for microbial quality control of mammal cells and included detection of M. arginini, M. orale, M. hyorhinis, M. fermentans, M. genitalium, M. hominis, M. pneumoniae, M. salivarium, M. pirum, A. laidlawii, and U. urealyticum. Limit of detection v...

Research paper thumbnail of B23 Influence of mycoplasma arginini on inflammatory response and energy metabolism in human cells with and without HTT gene mutation

Journal of Neurology, Neurosurgery & Psychiatry

Research paper thumbnail of Role of thiamine in Huntington’s disease pathogenesis: In vitro studies

Advances in Clinical and Experimental Medicine

Background. Oxidative stress accompanies neurodegeneration and also causes abnormalities in thiam... more Background. Oxidative stress accompanies neurodegeneration and also causes abnormalities in thiamine-dependent processes. These processes have been reported to be diminished in the brains of patients with several neurodegenerative diseases. Objectives. The aim of this work was to conduct a comparative analysis of the impact of supplemented thiamine on the viability of human B lymphocytes with CAG abnormal expanded huntingtin gene (mHTT) (GM13509) and control, B lymphocytes without mHTT (GM14467) through the following studies: determination of the supplemented thiamine concentrations, which are effective for cell growth stimulation after incubation in thiamine deficit conditions; determination of cell capability to intake the exogenous thiamine; evaluation of exogenous thiamine influence on the profile of the genes related to thiamine and energy metabolism; determination of ATP synthesis and activities of thiamine-dependent enzymes, KGDHC and BCKDHC in the intact cells and upon the exogenous thiamine. Material and methods. The following methods were used: EZ4U test for cell growth analysis; HPLC for determination of thiamine intake and ATP synthesis, qRT-PCR for evaluation of the gene profiles and spectrophotometric method for KGDHC and BCKDHC activities determination. Results. Maximal cell growth stimulation was observed at 2.5 mM in GM14467 up to 135% of the control culture and at 5.0 mM in GM13509 cells up to 165% of the control culture. Native levels of total ATP and KGDHC and BCKDHC activities in both cell types were comparable and did not changed upon thiamine deficit or supplementation. GM13509 cells showed more of an increase in growth stimulation upon thiamine supplementation than GM14467 cells and this effect was reflected in the increase of intracellular thiamine concentration. Conclusions. The above results and reported changes in expression of GAPDH, IDH1 and SLC19A3 genes observed upon thiamine deficit conditions suggest that intracellular thiamine status and energy metabolism can have a role in HD pathogenesis.

Research paper thumbnail of Doxorubicin Treatment of Cancer Cells Impairs Reverse Transcription and Affects the Interpretation of RT-qPCR Results

Cancer genomics & proteomics

Doxorubicin (DOX) acts in a variety of ways including DNA damage and enzyme inhibition, which con... more Doxorubicin (DOX) acts in a variety of ways including DNA damage and enzyme inhibition, which consequently causes changes in gene expression of cells treated with this agent. Practical validation of experimental results followed by appropriate normalization of the factors investigated is crucial for obtaining biologically relevant results in gene expression studies. Six candidates were evaluated regarding their validity as internal reference genes: RPS23, FLOT2, UBB, ABCF1, ACTB, HPRT1. Optimization for quantitative polymerase reaction (qPCR) included: sensitivity, specificity, amplification efficiency and linear dynamic range determination. The gene expression stability was evaluated by real-time quantitative polymerase reaction (RT-qPCR) in two human cervical cancer cell lines: HeLa and DOX-resistant KB-V1 Cells treated under various concentrations of DOX. DOX treatment changed gene expression and led to re-optimization of the cDNA template amounts. ACTB, HPRT1, RPS23 and FLOT2 ar...

Research paper thumbnail of Synergistic action of doxorubicin and sulindac in human cervix carcinoma cells - studies on possible mechanisms

Medical science monitor : international medical journal of experimental and clinical research, 2010

Epidemiologic and experimental studies have shown that cyclooxygenase-2 (COX-2) inhibitors as non... more Epidemiologic and experimental studies have shown that cyclooxygenase-2 (COX-2) inhibitors as non-steroidal anti-inflammatory drugs (NSAIDs) are effective chemopreventive agents. The mechanisms underlying the antitumor activity of COX-2 inhibitors are thought to involve inhibition of COX-2 enzyme activity and induction of apoptosis. The aim of the current work was to study the mechanisms of synergistic action noted in HeLa cervical carcinoma cells under doxorubicin (DOX) and sulindac (SUL) co-treatment. Cytotoxic activity of the drugs was defined with MTT test, apoptosis was detected with TUNEL test, DOX transmembrane efflux was measured fluorometrically, expression of MDR-1 and MRP-1 was determined with quantitative real time - PCR (QRT-PCR). It was shown that SUL at non-toxic concentrations, 10 and 50 microM, is an effective enhancer of cytotoxic action for DOX in 0.5 and 1 microM, respectively; however, only for SUL concentration equal to 50 microM potentiated apoptosis induced b...

Research paper thumbnail of Huntington' disease--imbalance of amino acid levels in plasma of patients and mutation carriers

Annals of agricultural and environmental medicine : AAEM, 2013

Determination of the plasma amino acid (AA) levels in Huntington's disease (HD) can make it p... more Determination of the plasma amino acid (AA) levels in Huntington's disease (HD) can make it possible to find the metabolic markers used in early diagnosis. The aim of the presented study was to determine the AA profile in plasma samples from HD patients and presymptomatic carriers, compared to healthy subjects. The AA profile was analyzed with HPLC. The study concerned 59 participants: 30 subjects with abnormal CAG repeats expansion (>36) in the HTT gene, and 29 healthy subjects. Each participant was analyzed with regard to the parameters characterizing the metabolic state and protein metabolism, such as: urea, creatinine, glucose, total protein, TSH (thyroid-stimulating hormone), cortisol, ESR (erythrocyte sedimentation rate), and CRP (C-reactive protein). Simple statistical comparisons showed 5 AA to be significantly lower in the HD group, compared to the control group, i.e.: Asn, His, Leu, Ser, Thr. Creatinine and creatinine clirens were found to be lower in the HD group, ...

Research paper thumbnail of A study of molecular changes relating to energy metabolism and cellular stress in people with Huntington’s disease: looking for biomarkers

Journal of Bioenergetics and Biomembranes, 2012

Huntington's disease (HD) is a neurodegenerative disorder characterized by a progressive motor an... more Huntington's disease (HD) is a neurodegenerative disorder characterized by a progressive motor and cognitive decline and the development of psychiatric symptoms. The origin of molecular and biochemical disturbances in HD is a mutation in the HTT gene, which is autosomally dominantly inherited. The altered huntingtin protein is ubiquitously expressed in the CNS, as well as in peripheral tissues. In this study we measured the metabolism changes in gene transcription in blood of HD gene carriers (premanifest and manifest combined) versus 28 healthy controls. The comparison revealed statistically significant Global Pattern Recognition Fold Change (FC) for 6 mRNA transcripts, reflecting an increase of: MAOB (FC03.07; p00.0005) which encodes an outer mitochondrial membrane-bound enzyme called monoamine oxidase type B; TGM2 (FC01.8; p00.02) encoding a transglutaminase 2 that mediates cellular stress; SLC2A4 (FC0 1.64; p00.02) solute carrier family 2 (facilitated glucose transporter) member 4; branched chain ketoacid dehydrogenase kinase (BCKDK) (FC01.34; p00.02); decrease of LDHA (FC0−1.16; p00.03) lactate dehydrogenase A; and brainderived neurotrophic factor (BDNF) (FC0−2,11; p00.03). These distinguished changes coincided with HD progress. The analyses of gene transcription levels in sub-cohorts confirmed these changes and also revealed 28 statistically significant FCs of gene transcripts involved in ATP production and BCAA metabolism.

Research paper thumbnail of First community-acquired methicillin-resistant Staphylococcus aureus (MRSA) strains in Croatia

Clinical Microbiology and Infection, 2006

received 40 mg dexamethasone over the course of 4 days. By day 20 the patient had recovered. Conf... more received 40 mg dexamethasone over the course of 4 days. By day 20 the patient had recovered. Conflicting results concerning the clinical efficacy of pristinamycin against macrolide-resistant S. pneumoniae with the MLS B phenotype have been reported [1,9,10]. In a double-blind study comparing the efficacy of pristinamycin with amoxycillin for the treatment of communityacquired pneumonia, pristinamycin was significantly less effective than amoxycillin (63.6% vs. 86.6%) in bacteraemic patients, with a 50% failure rate for macrolide-resistant S. pneumoniae [2]. Taken together, these data emphasise the need to reconsider the choice of pristinamycin or quinupristin ⁄ dalfopristin for the treatment of macrolide-resistant pneumococcal infections.

Research paper thumbnail of New Method for Typing Staphylococcus aureus Strains: Multiple-Locus Variable-Number Tandem Repeat Analysis of Polymorphism and Genetic Relationships of Clinical Isolates

Journal of Clinical Microbiology, 2003

The PCR-based methodology applied to multiple-locus variable numbers of tandem repeat (VNTR) anal... more The PCR-based methodology applied to multiple-locus variable numbers of tandem repeat (VNTR) analysis was recently shown to be a useful technique for the molecular typing of clinical isolates of several bacterial species. We have adopted this method for the molecular typing of methicillin-resistant Staphylococcus aureus . Five staphylococcal VNTR loci ( sdr , clfA , clfB , ssp , and spa ) were subjected to analysis, and it was shown that the method allows typing of S . aureus strains with the discriminatory power and reproducibility of pulsed-field gel electrophoresis while at the same time being rapid and applicable to analysis of large numbers of isolates.

Research paper thumbnail of Comparison of multiple-locus variable-number tandem repeat analysis and pulsed-field gel electrophoresis in a setting of polyclonal endemicity of vancomycin-resistant Enterococcus faecium

Clinical Microbiology and Infection, 2008

Research paper thumbnail of Huntington' disease--imbalance of amino acid levels in plasma of patients and mutation carriers

Annals of agricultural and environmental medicine : AAEM, 2013

Determination of the plasma amino acid (AA) levels in Huntington's disease (HD) can make it p... more Determination of the plasma amino acid (AA) levels in Huntington's disease (HD) can make it possible to find the metabolic markers used in early diagnosis. The aim of the presented study was to determine the AA profile in plasma samples from HD patients and presymptomatic carriers, compared to healthy subjects. The AA profile was analyzed with HPLC. The study concerned 59 participants: 30 subjects with abnormal CAG repeats expansion (>36) in the HTT gene, and 29 healthy subjects. Each participant was analyzed with regard to the parameters characterizing the metabolic state and protein metabolism, such as: urea, creatinine, glucose, total protein, TSH (thyroid-stimulating hormone), cortisol, ESR (erythrocyte sedimentation rate), and CRP (C-reactive protein). Simple statistical comparisons showed 5 AA to be significantly lower in the HD group, compared to the control group, i.e.: Asn, His, Leu, Ser, Thr. Creatinine and creatinine clirens were found to be lower in the HD group, ...

Research paper thumbnail of Characterization of coagulase-negative staphylococci isolated from cases of ostitis and osteomyelitis

Polish journal of microbiology, 2006

Coagulase-negative staphylococci (CoNS) are often responsible for cases of chronic ostitis and os... more Coagulase-negative staphylococci (CoNS) are often responsible for cases of chronic ostitis and osteomyelitis, especially in patients with orthopedic prosthesis/implants. The aim of this study was to characterize CoNS isolated from ambulatory patients with chronic ostitis/osteomyelitis and to compare them by PFGE (pulsed-field gel electrophoresis). Out of 263 bacterial strains isolated from wounds/sinuses of patients with chronic ostitis/osteomylitis, 41 were identified as CoNS. Twenty methicillin-resistant strains were selected for this study. Our results confirm the superior performance of cefoxitin disk test to detect methicillin resistance in heterogenous population of CoNS. High level of antibiotic resistance was observed among the studied strains: majority of CoNS were resistant to tetracycline and erythromycin and also to clindamycin and ciprofloxacin. Importantly, in 15 out of 20 studied CoNS different phenotypes of macrolides, lincosamides and streptogramin--MLS resistance w...

Research paper thumbnail of Modulation of apoptosis protein profiles--role of P-gp in HeLa cells exposed to doxorubicin

Cancer genomics & proteomics, 2012

As shown previously doxorubicin (1 μM) plus sulindac (50 μM) reduced the expression of ABCB1 (ATP... more As shown previously doxorubicin (1 μM) plus sulindac (50 μM) reduced the expression of ABCB1 (ATP-binding cassette, sub-family B (MDR/TAP), member 1) mRNA in HeLa cells and this effect was accompanied by increased apoptosis. The aim of this study was to define if the decrease of ABCB1 expression or blocking of P-glycoprotein (P-gp) can affect the expression of the apoptotic genes determined with use of quantitative real time polymerase chain reaction (qRT-PCR). Western blot was used for visualization of chosen pro- and antiapoptotic proteins. Doxorubicin was the main compound which affected the apoptotic genes. The effectiveness of the drugs in reducing of P-gp function has been shown as not being related to the regulation of apoptotic gene transcription. In this experimental scheme, regulation of apoptotic gene transcription depended on the kind of P-gp modulator.

Research paper thumbnail of Doxorubicin Treatment of Cancer Cells Impairs Reverse Transcription and Affects the Interpretation of RT-qPCR Results

Cancer genomics & proteomics, 2016

BACKGROUND Doxorubicin (DOX) acts in a variety of ways including DNA damage and enzyme inhibition... more BACKGROUND Doxorubicin (DOX) acts in a variety of ways including DNA damage and enzyme inhibition, which consequently causes changes in gene expression of cells treated with this agent. Practical validation of experimental results followed by appropriate normalization of the factors investigated is crucial for obtaining biologically relevant results in gene expression studies. MATERIALS AND METHODS Six candidates were evaluated regarding their validity as internal reference genes: RPS23, FLOT2, UBB, ABCF1, ACTB, HPRT1. Optimization for quantitative polymerase reaction (qPCR) included: sensitivity, specificity, amplification efficiency and linear dynamic range determination. The gene expression stability was evaluated by real-time quantitative polymerase reaction (RT-qPCR) in two human cervical cancer cell lines: HeLa and DOX-resistant KB-V1 Cells treated under various concentrations of DOX. RESULTS DOX treatment changed gene expression and led to re-optimization of the cDNA template...

Research paper thumbnail of Zmiany w epidemiologii inwazyjnych klonów MRSA w wybranych szpitalach w Polsce

Research paper thumbnail of Possible role of HSP60 in synergistic action of anthracyclines and sulindac in HeLa cells

As was observed in the earlier studies, doxorubicin (DOX) induced apoptosis in HeLa cells and tha... more As was observed in the earlier studies, doxorubicin (DOX) induced apoptosis in HeLa cells and that effect was potentiated significantly by sulindac (SUL). The aim of the current work was to study the effects of DOX and SUL on HSP60, HSF1 and HSP60 expression and the influence of DOX and SUL on HSP60 translocation. Expression of HSP60 and HSF1 was determined with QRT-PCR; the expression and localization of HSP60 were evaluated with Western blot. The 24-h cell cultures were co-incubated with DOX - 1 microM and/or SUL - 50 microM. The significant induction of HSF1 and HSP60 mRNA level was observed after exposure of the cells to DOX 1 microM. SUL 50 microM alone caused moderate increase in mRNA level. The significant decrease in expression of HSF1 and HSP60 was noted after DOX 1 microM and SUL 50 microM simultaneous treatment. HSP60 appeared in the higher levels in cytosol than in mitochondria No intracellular translocation was noted under treatment of the cells to DOX and/or SUL. In co...

Research paper thumbnail of Formation and preclinical evaluation of a new alloplastic injectable bone substitute material

Alloplastic bone substitute materials are raising some more interest as an alternative for autolo... more Alloplastic bone substitute materials are raising some more interest as an alternative for autologic transplants and xenogenic materials especially in oral surgery over the last few years. These non-immunogenic and completely resorbable biomaterials are the basis for complete and predictable guided bone regeneration. In the majority of cases, such a material is chosen because of its convenient application by surgeons. The main objective of our project was to design and fabricate an osteoconductive, injectable and readily tolerable by human tissues biomaterial for guided bone regeneration. For this purpose, a self-setting composite consisting of chitosan/tricalcium phosphate microparticles and sodium alginate was made. The material obtained was characterized by microsphere and agglomerate morphology and microstructure. Its features relating to setting time and mechanical properties were precisely investigated. Our material was also evaluated according to PN-EN ISO 10993 Biological ev...

Research paper thumbnail of Usefulness of Multiple-Locus VNTR Fingerprinting in detection of clonality of community- and hospital-acquired Staphylococcus aureus isolates

Antonie van Leeuwenhoek, 2008

Staphylococcus aureus has become a major source of hospital infections and the risk of colonisati... more Staphylococcus aureus has become a major source of hospital infections and the risk of colonisation and infection by community-acquired methicillin-resistant S. aureus (CA-MRSA) is increasingly higher. Because of the importance of S. aureus to public health, many molecular typing methods have been developed to determine its transmission routes and source of infection during epidemiological investigations. In this study we evaluated the usefulness of multiplex PCR based Multi-Locus VNTR Fingerprinting (MLVF) as the first step method for rapid differentiation of Croatian and Polish S. aureus isolates in hospital and community settings. This is a first report of the usefulness of MLVF in typing of hospital-acquired methicillin-sensitive S. aureus (HA-MSSA) and four CA-MRSA isolates. A total of 47 isolates of S. aureus recovered in Croatia in 2004 and in Poland in 2006 and 2007 were tested. The MLVF results were compared to those produced by other typing methods, such as Pulsed-Field Gel Electrophoresis (PFGE), Multi-Locus Sequence Typing (MLST) and spa typing. The MLVF analysis showed almost the same clonality results as the remaining typing methods although some differences were found. Epidemiological data about the relation among S. aureus isolates and the results produced by typing methods applied in the present study indicate that because of the advantages in ease and speed of Variable Number of Tandem Repeats (VNTR) procedure over PFGE, spa typing and MLST, MLVF can be used as a first screening method followed by additional typing.

Research paper thumbnail of Changes in epidemiology of invasive MRSA clones in selected Polish hospitals

Introduction: Staphylococcus aureus is a pathogen that can cause a wide spectrum of diseases from... more Introduction: Staphylococcus aureus is a pathogen that can cause a wide spectrum of diseases from relatively mild skin infections to life-threatening systemic disease. The study was conducted to assess the clonality of methicillin-resistant isolates of S. aureus (MRSA) in selected Polish hospitals. Material and Methods: A collection of strains consisted of 183 methicillin-resistant isolates of S. aureus (MRSA) and 4 methicillin susceptible isolates of S. aureus (MSSA) from 2002-2005. Isolates came from 29 hospitals. Purified total DNA of isolates was used to amplify the region X of the gene spa. Subsequently, the amplicons were sequenced and spa

Research paper thumbnail of F09 The study on molecular changes related to energy metabolism in Huntington's disease subjects: looking for biomarkers

Journal of Neurology, Neurosurgery & Psychiatry, 2012

Huntington's disease (HD) is a neurodegenerative disorder characterised by a progressive moto... more Huntington's disease (HD) is a neurodegenerative disorder characterised by a progressive motor and cognitive decline and psychiatric symptoms. The origin of molecular and biochemical disturbances in HD is the genetic defect in the HTT gene, autosomally dominantly inherited. The altered huntingtin protein is ubiquitously expressed in the Central Nervous System as well as in peripheral tissues. The aim of the study was to detect the metabolism changes on the transcription level in blood cells in HD subjects (n=29) with regards to the control, healthy subjects (n=28). The gene expression analysis was performed using quantitative PCR and StellAarray system. A panel of 94 genes involved in glycolysis, Krebs cycle, electron transport chain, BCAAs metabolism and other processes were selected to study. The results calculations were conducted by Global Pattern Recognition software with ΔΔCt type of analysis application. The comparison between HD and control revealed statistically significant Global Pattern Recognition fold change (FC) values for six transcripts, including a decrease of BDNF (−2.11), LDHA (−1.16) and an increase of BCKDK (1.34), MAOB (3.07), SLC2A4 (1.64) and TGM2 (1.8). Multiple analyses in sub-cohorts distinguished by gender, age, BMI, calf circumference, nutrition status, CAG number repeats, TFC, UHDRS motor and HD time from onset also showed 34 transcript FCs. The widest FC ranges decrease concerned to BDNF (from −1.66 to −3.6), BCAT2 (from 1.85 to −1.42) and increase to MAOB (from 1.87 to 7.89) transcript levels that coincided together with the length of HD duration period and the HD progress.

Research paper thumbnail of Development of New PCR Assay with SYBR Green I for Detection of Mycoplasma, Acholeplasma, and Ureaplasma sp. in Cell Cultures

Diagnostics

Mycoplasma, Acholeplasma, and Ureaplasma sp. are atypical bacteria responsible for in vitro cell ... more Mycoplasma, Acholeplasma, and Ureaplasma sp. are atypical bacteria responsible for in vitro cell culture contaminations that can warp the results. These bacteria also cause human and animal infections and may lead to chronic diseases. In developed polymerase chain reaction (PCR) in this study a quantitative PCR with SYBR Green I fluorochrome was applied to facilitate the Mycoplasma, Acholeplasma, and Ureaplasma sp. DNA detection and identification. Screening Test-1 v.1 (triplex qPCR) allowed for the detection of 11 species. Test-1 v.2 (three single qPCRs) pre-identified three subgroups, allowing for the reduction of using single qPCRs in Test-2 for species identification. The range of both tests was consistent with pharmacopeial requirements for microbial quality control of mammal cells and included detection of M. arginini, M. orale, M. hyorhinis, M. fermentans, M. genitalium, M. hominis, M. pneumoniae, M. salivarium, M. pirum, A. laidlawii, and U. urealyticum. Limit of detection v...

Research paper thumbnail of B23 Influence of mycoplasma arginini on inflammatory response and energy metabolism in human cells with and without HTT gene mutation

Journal of Neurology, Neurosurgery & Psychiatry

Research paper thumbnail of Role of thiamine in Huntington’s disease pathogenesis: In vitro studies

Advances in Clinical and Experimental Medicine

Background. Oxidative stress accompanies neurodegeneration and also causes abnormalities in thiam... more Background. Oxidative stress accompanies neurodegeneration and also causes abnormalities in thiamine-dependent processes. These processes have been reported to be diminished in the brains of patients with several neurodegenerative diseases. Objectives. The aim of this work was to conduct a comparative analysis of the impact of supplemented thiamine on the viability of human B lymphocytes with CAG abnormal expanded huntingtin gene (mHTT) (GM13509) and control, B lymphocytes without mHTT (GM14467) through the following studies: determination of the supplemented thiamine concentrations, which are effective for cell growth stimulation after incubation in thiamine deficit conditions; determination of cell capability to intake the exogenous thiamine; evaluation of exogenous thiamine influence on the profile of the genes related to thiamine and energy metabolism; determination of ATP synthesis and activities of thiamine-dependent enzymes, KGDHC and BCKDHC in the intact cells and upon the exogenous thiamine. Material and methods. The following methods were used: EZ4U test for cell growth analysis; HPLC for determination of thiamine intake and ATP synthesis, qRT-PCR for evaluation of the gene profiles and spectrophotometric method for KGDHC and BCKDHC activities determination. Results. Maximal cell growth stimulation was observed at 2.5 mM in GM14467 up to 135% of the control culture and at 5.0 mM in GM13509 cells up to 165% of the control culture. Native levels of total ATP and KGDHC and BCKDHC activities in both cell types were comparable and did not changed upon thiamine deficit or supplementation. GM13509 cells showed more of an increase in growth stimulation upon thiamine supplementation than GM14467 cells and this effect was reflected in the increase of intracellular thiamine concentration. Conclusions. The above results and reported changes in expression of GAPDH, IDH1 and SLC19A3 genes observed upon thiamine deficit conditions suggest that intracellular thiamine status and energy metabolism can have a role in HD pathogenesis.

Research paper thumbnail of Doxorubicin Treatment of Cancer Cells Impairs Reverse Transcription and Affects the Interpretation of RT-qPCR Results

Cancer genomics & proteomics

Doxorubicin (DOX) acts in a variety of ways including DNA damage and enzyme inhibition, which con... more Doxorubicin (DOX) acts in a variety of ways including DNA damage and enzyme inhibition, which consequently causes changes in gene expression of cells treated with this agent. Practical validation of experimental results followed by appropriate normalization of the factors investigated is crucial for obtaining biologically relevant results in gene expression studies. Six candidates were evaluated regarding their validity as internal reference genes: RPS23, FLOT2, UBB, ABCF1, ACTB, HPRT1. Optimization for quantitative polymerase reaction (qPCR) included: sensitivity, specificity, amplification efficiency and linear dynamic range determination. The gene expression stability was evaluated by real-time quantitative polymerase reaction (RT-qPCR) in two human cervical cancer cell lines: HeLa and DOX-resistant KB-V1 Cells treated under various concentrations of DOX. DOX treatment changed gene expression and led to re-optimization of the cDNA template amounts. ACTB, HPRT1, RPS23 and FLOT2 ar...