imagine (Institute of molecular genetics and genetic engineering)
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Gut Microbiota Confers Resistance of Albino Oxford Rats to the Induction of Experimental Autoimmune Encephalomyelitis
Get PDFAlbino Oxford (AO) rats are extremely resistant to induction of experimental autoimmune encephalomyelitis (EAE). EAE is an animal model of multiple sclerosis, a chronic inflammatory disease of the central nervous system (CNS), with established autoimmune pathogenesis. The autoimmune response against the antigens of the CNS is initiated in the peripheral lymphoid tissues after immunization of AO rats with CNS antigens. Subsequently, limited infiltration of the CNS occurs, yet without clinical sequels. It has recently become increasingly appreciated that gut-associated lymphoid tissues (GALT) and gut microbiota play an important role in regulation and propagation of encephalitogenic immune response. Therefore, modulation of AO gut microbiota by antibiotics was performed in this study. The treatment altered composition of gut microbiota in AO rats and led to a reduction in the proportion of regulatory T cells in Peyer's patches, mesenteric lymph nodes, and in lymph nodes draining the site of immunization. Upregulation of interferon-. and interleukin (IL)-17 production was observed in the draining lymph nodes. The treatment led to clinically manifested EAE in AO rats with more numerous infiltrates and higher production of IL-17 observed in the CNS. Importantly, transfer of AO gut microbiota into EAE-prone Dark Agouti rats ameliorated the disease. These results clearly imply that gut microbiota is an important factor in AO rat resistance to EAE and that gut microbiota transfer is an efficacious way to treat CNS autoimmunity. These findings also support the idea that gut microbiota modulation has a potential as a future treatment of multiple sclerosis
The Variants in the 3 ' Untranslated Region of the Matrix Metalloproteinase 9 Gene as Modulators of Treatment Outcome in Children with Asthma
No full textPurpose The maintaining of asthma control is difficult due to high variability in response to therapy among patients. Since matrix metalloproteinase 9 (MMP9) is implicated in inflammation and remodeling of asthmatic airways, it could be associated with adequate response to asthma therapy. The aim of this study was to investigate whether variants in 3' end of the MMP9 gene are associated with clinical phenotype and responsiveness to treatment in children with asthma. Methods The study included 127 asthmatic children from Slovenia. Variants in the 3' end of the MMP9 gene were analyzed by direct DNA sequencing and the obtained results were correlated with clinical parameters. Results Two variants were detected, rs13925 and rs20544. For the variant rs20544, statistically significant difference in airway hyperresponsiveness (p = 0.011) and asthma control (p = 0.049) between genotypes was found. Patients with TT genotype had lower airway sensitivity, and after 12 months of treatment showed significant improvement in Asthma Control Test (ACT) scores compared to CC and CT genotype. For the variant rs13925, the association with lung function was observed. The carriers of A allele showed noticeable improvement of lung function after the first 6 months of treatment in comparison to the carriers of G allele (p = 0.046). Conclusion The main finding of our study is the association of MMP9 genotypes rs20544 TT and rs13925 AA and AG with better asthma control, and indirectly better response to treatment. Based on these results, MMP9 deserves further research as a potential predictive biomarker for asthma
Streptomyces spp. in the biocatalysis toolbox
No full textAbout 20,100 research publications dated 2000-2017 were recovered searching the PubMed and Web of Science databases for Streptomyces, which are the richest known source of bioactive molecules. However, these bacteria with versatile metabolism are powerful suppliers of biocatalytic tools (enzymes) for advanced biotechnological applications such as green chemical transformations and biopharmaceutical and biofuel production. The recent technological advances, especially in DNA sequencing coupled with computational tools for protein functional and structural prediction, and the improved access to microbial diversity enabled the easier access to enzymes and the ability to engineer them to suit a wider range of biotechnological processes. The major driver behind a dramatic increase in the utilization of biocatalysis is sustainable development and the shift toward bioeconomy that will, in accordance to the UN policy agenda "Bioeconomy to 2030," become a global effort in the near future. Streptomyces spp. already play a significant role among industrial microorganisms. The intention of this minireview is to highlight the presence of Streptomyces in the toolbox of biocatalysis and to give an overview of the most important advances in novel biocatalyst discovery and applications. Judging by the steady increase in a number of recent references (228 for the 2000-2017 period), it is clear that biocatalysts from Streptomyces spp. hold promises in terms of valuable properties and applicative industrial potential
Prognostic significance of SOX2, SOX3, SOX11, SOX14 and SOX18 gene expression in adult de novo acute myeloid leukemia
No full textAberrant expression of different SOX (SRY-related high mobility group (HMG) box) genes has been observed in number of tumors but, little is known about their expression patterns in hematological malignancies, especially in acute myeloid leukemia (AML). In this study we investigated SOX2, SOX3, SOX11, SOX14 and SOX18 gene expression in 50 de novo adult AML patients and correlated our findings with known clinical and molecular prognostic markers of the disease. We have found that these genes are overexpressed in 10-22% of patients and preliminary findings suggest that high expression level of these genes may have prognostic significance in AML patients. This is the first study focused on examining the expression level of SOX2, SOX3, SOX11, SOX14 and SOX18 genes in AML patients. Although this is a relatively limited study, initial findings indicate the need for further investigation of these genes, their potential roles in leukemia pathogenesis as well as prognosis in AML patients
Regulation of gene expression and diversity of CarO i Omp33-36 porins in Acinetobacter spp.
Get PDFVrste roda Acinetobacter, pre svih Acinetobacter baumannii, su u poslednjih nekoliko decenija od komensalnih stanovnika zdravstvenih ustanova evoluirale u jednu od najozbiljnijih pretnji savremenog zdravstvenog sistema. Globalno rasprostranjenje i konstantan porast izolata Acinetobacter spp. rezistentnih na karbapeneme, antibiotike poslednjeg izbora u leĉenju infekcija izazvanih Gram-negativnim bakterijama, pozicionira ovu grupu bakterija na sam vrh liste patogena za koje je neophodan urgentan pronalazak novih efikasnih antimikrobnih terapeutika. Kontinuirani nadzor nad Acinetobacter spp. rezistenim na karbapeneme, koji podrazumeva praćenje klonalnog širenja i utvrĊivanje molekularnih mehanizama odgovornih za rezistenciju na ovu grupu antibiotika, je prvi korak u uspešnom ograniĉavanju diseminacije navedenih patogenih bakterija. Saznanja o prirodi signala iz spoljašnje sredine, koji mogu uticati na nivo rezistencije na antibiotike i virulencije, ukazuju na uslove pogodne za opstanak A. baumannii i idejno usmeravaju ka pronalasku novih terapeutika. Pored toga, formulacija efikasne vakcine, kao izuzetno znaĉajnog antiinfektivnog agensa, predstavlja jedan od imperativa u eliminaciji infekcija izazvanih višestruko rezistentnim sojevima A. baumannii. U skladu sa prethodno navedenim, ova teza je imala za cilj da molekularno okarakteriše izolate A. baumannii rezistentne na karbapeneme uzorkovane u periodu od 2012. do 2014. godine na Instituta za zdravstvenu zaštitu majke i deteta Srbije „Dr Vukan Ĉupić” (Beograd, Srbija) (n=28). Analizirani izolati A. baumannii su pripadali trima klonalnim grupama karakteristiĉnim za Evropu (Evropski klonovi I-III) sa izuzetkom jednog izolata (8778), koji nije pripadao nijednoj od ove tri grupe. Evropski klon II, najuspešniji bolniĉki Evropski klon, je takoĊe bio najzastupljeniji i meĊu izolatima iz Srbije (67,86%). PFGE metodom izolati A. baumannii rezistenti na karbapeneme su grupisani u dva klastera (I i II), koja su obuhvatala ĉetiri pulsotipa (A-D). Sliĉnost izolata se kretala od 85% do skoro 100%. Budući da je metodom zasnovanom na sekvencama tri gena i PFGE ustanovljena raznovrsnost izolata A. baumannii, može se zakljuĉiti da nije došlo do klonalnog širenja datih izolata unutar bolniĉke ustanove u kojoj su uzorkovani..
Mononuclear silver(I) complexes with 1,7-phenanthroline as potent inhibitors of Candida growth
No full textMononuclear silver(I) complexes with 1,7-phenanthroline (1,7-phen), [Ag(NO3-O,O') (1,7-phen-N7)(2)] (1) and [Ag(1,7-phen-N7)(2)]X, X = ClO4- (2), CF3SO3- (3), BF4- (4) and SbF6- (5) were synthesized and structurally characterized by NMR (H-1 and C-13), IR and UV-Vis spectroscopy and ESI mass spectrometry. The crystal structures of 1, 3 and 4 were determined by single-crystal X-ray diffraction analysis. In all these complexes, 1,7-phen coordinates to the Ag(I) ion in a monodentate fashion via the less sterically hindered N7 nitrogen atom. The investigation of the solution stability of 1-5 in DMSO revealed that they are sufficiently stable in this solvent at room temperature. Complexes 1-5 showed selectivity towards Candida spp. in comparison to bacteria, effectively inhibiting the growth of four different Candida species with minimal inhibitory concentrations (MIC) between 1.2 and 11.3 mu M. Based on the lowest MIC values and the lowest cytotoxicity against healthy human fibroblasts with selectivity index of more than 30, the antifungal potential was examined in detail for the complex 1. It had the ability to attenuate C. albicans virulence and to reduce epithelial cell damage in the cell infection model. Induction of reactive oxygen species (ROS) response has been detected in C. albicans, with fungal DNA being one of the possible target biomolecules. The toxicity profile of 1 in the zebrafish model (Danio rerio) revealed improved safety and activity in comparison to that of clinically utilized silver(I) sulfadiazine.Related to accepted version: [https://imagine.imgge.bg.ac.rs/handle/123456789/2228]Related to supplemetary material: [https://imagine.imgge.bg.ac.rs/handle/123456789/2229
Mesenchymal stem cells from periapical lesions modulate cytokine production by local immune cells
Get PDFBackground/Aim. Mesenchymal stem cells (MSCs) have been shown to suppress immune and inflammatory reactions. However, it is not known whether MSCs from inflammatory tissues, such as periapical lesions (PLs) have similar effects. This question was addressed in this study in which the aim was to examine the capacity of PL-MSCs for modulating cytokine production by local immune cells. Methods. PL-MSCs were isolated from asymptomatic (as) and symptomatic (sy) PLs. Their phenotype was analyzed by flow cytometry by detecting MSC surface markers. Anti-inflammatory and immunomodulatory properties of PL-MSCs were examined by measuring cytokine production in direct co-culture experiments with mononuclear cells (MNCs) isolated from asPLs and syPLs, respectively. The levels of cytokines in supernatants were determined by specific ELISA kits. Results. Both PL-MSCs lines were characterized by typical MSC phenotype, with the predominance of CD29, CD44, CD90, CD105 and CD166. However, the lines, independently of their similar phenotype had the same modulatory effect on cytokine production, but the response of asPL-MNCs and syPL-MNCs was different, in spite of similar composition of these MNCs. Both MSC lines inhibited the production of inflammatory cytokines, such as interleukin-10 (IL-10) and tumor necrosis factor-0 (TNF-0). However, IL-8 was only down-regulated in the co-culture of these MSC lines with syPL-MNCs. The PL-MSCs also modulated the production of immunoregulatory cytokines. Transforming growth factor-0 (TGF-0) was up-regulated by both as- and syPL-MNCs but IL-10 was up-regulated only by asPL-MNCs. Conclusion. Our results showed that PL-MSCs contribute to the restriction of local inflammatory and immune responses, but this effect is probably less efficient during the exacerbation of PL inflammation
Capacity Building for Rare Genomics
Get PDFInfrequent occurrence of rare diseases (RD) has led to prolonged time needed to make an accurate diagnosis. Research and
development of new treatments has been insufficient due to the
narrow market for affected patients. Fascinating progress and accumulation of knowledge in the field of genomics provided for
improvement of diagnosis and follow-up of numerous RD. It also
led to the design of molecular-targeted therapeutics and more effective treatment of RD.
Building up a centre for genomics of rare diseases comprises
strengthening of several components in an institution already involved in molecular genetics and biomedical research, such as, infrastructure, human resources, local and international networking
and dissemination and outreach activities.
Regarding infrastructure, facilities for Next-generation sequencing (NGS) and array-based comparative genomic hybridization (aCGH) are necessary to perform high-throughput genomewide screening. Establishment of the bioinformatics unit is also
very important. At the heart of genomics of RD is biological samples storage and database facilities. Biobanks are a critical resource
in the field of genomics of RD. Additionally, strong connections
with local medical institutions have to be made.
Reinforcement of human potential, especially through recruitment of experts in bioinformatics, is indispensable contribution
to the implementation of new equipment and high-throughput
methodologies. Trainings of researchers are continually in focus
of an expert centre for genomics of RD.
Integration of the centre into European networks in order to
intensify the transfer of knowledge is one of the cornerstones for
its capacity building. The data a nd biospecimens that are distributed to all researchers, are absolute requirements in the pathway to
develop modern science, diagnosis and cures for RD.
Finally, dissemination and outreach activities are needed to
ensure increased visibility of the centre within local and international scientific community and stakeholders and social entities
interested in exploiting the results of the centre
Importance of TLR9-IL23-1L17 axis in inflammatory bowel disease development: Gene expression profiling study
Get PDFBackground and aims: Mucosal gene expression have not been fully enlightened in inflammatory bowel disease (IBD). Aim of this study was to define IL23A, IL17A, IL17F and TLR9 expression in different IBD phenotypes. Methods: Evaluation of mRNA levels was performed in paired non-inflamed and inflamed mucosal biopsies of newly diagnosed 50 Crohn's disease (CD) and 54 ulcerative colitis (UC) patients by quantitative real-time PCR analysis. Results: IL17A and IL17F expression levels were significantly increased in inflamed IBD mucosa. Inflamed CD ileal and UC mucosa showed increased IL23A, while only inflamed CD ileal samples showed increased TLR9 mRNA level. Correlation between analysed mRNAs levels and endoscopic and clinical disease activity were found in UC, but only with clinical activity in CD. Conclusion: Both CD and UC presented expression of Th17-associated genes. Nevertheless, expression profiles between different disease forms varies which should be taken into account for future research and therapeutics strategies.This is the peer reviewed version of the paper: Dragasevic, S., Stankovic, B., Sokic-Milutinovic, A., Milosavljevic, T., Milovanovic, T., Lukic, S., Drazilov, S. S., Klaassen, K., Kotur, N., Pavlovic, S., & Popovic, D. (2018). Importance of TLR9-IL23-IL17 axis in inflammatory bowel disease development: Gene expression profiling study. Clinical Immunology, 197, 86–95. [https://doi.org/10.1016/j.clim.2018.09.001
Functional Characterization of the Lactolisterin BU Gene Cluster of Lactococcus lactis subsp. lactis BGBU1-4
Get PDFThe gene cluster responsible for the production of the aureocin A53-like bacteriocin, lactolisterin BU, is located on plasmid pBU6 in Lactococcus lactis subsp. lactis BGBU1-4. Heterologous expression of pBU6 confirmed that production and limited immunity to lactolisterin BU were provided by the plasmid. Comparative analysis of aureocin A53-like operons revealed that the structural genes shared a low level of identity, while other genes were without homology, indicating a different origin. Subcloning and expression of genes located downstream of the structural gene, IliBU, revealed that the lactolisterin BU cluster consists of four genes: the structural gene IliBU, the abcT gene encoding an ABC transporter, the accL gene encoding an accessory protein and the immL gene which provides limited immunity to lactolisterin BU. Reverse transcription analysis revealed that all genes were transcribed as one polycistronic mRNA. Attempts to split the lactolisterin BU operon, even when both parts were under control of the PlliBU promoter, were unsuccessful indicating that expression of lactolisterin BU is probably precisely regulated at the translational level by translational coupling and is possible only when all genes of the operon are in cis constellation. Two rho-independent transcription terminators were detected in the lactolisterin BU operon: the first in the intergenic region of the IliBU and abcT genes and the second at the end of operon. Deletion of the second transcription terminator did not influence production of the bacteriocin in lactococci