imagine (Institute of molecular genetics and genetic engineering)
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Genetic characterization of the Yugoslavian shepherd dog - sharplanina, a livestock guard dog from the Western Balkans
Get PDFYugoslavian Shepherd Dog - Sharplanina (YSD) is a livestock guard dog from the Western Balkans present in this region over a long time, but recognized by the Federation Cynologique Internationale as a distinct breed as late as 1957. However, the information regarding the origin and the size of the breed's foundation stock is still lacking. In order to contribute towards better understanding of the genetic make-up of the YSD and its foundation stock, we re-analyzed previously generated genetic profiles of 94 registered YSD dogs assessed with nine nuclear microsatellites. Studied individuals comprised 90 unrelated dogs and two pairs of full-sibs, sampled at four sampling sites: three dog shows and at a military training centre for dogs in Serbia. We supported earlier findings on high levels of genetic diversity in YSD (H-E=0.728 +/- 0.027) and lack of inbreeding, and revealed substructure of the breed because we found two distinct gene pools in the Bayesian clustering analysis, indicated also by the excess of homozygotes (i.e., Wahlund effect) and outcomes of other analyses: linkage disequilibrium tests, Neighbour-Joining tree, principal coordinates and two-dimensional scaling analyses. The two gene pools were almost equally represented at each sampling site. One gene pool was composed of individuals with high genetic integrity, while the other gene pool was characterized with admixed ancestry, developed possibly via hybridization with native breeding stock outside the registry system, other breeds, such as the Caucasian Shepherd, and/or individuals admixed with wolves. Thus, we demonstrate rather complex and diverse ancestry implying a genetically heterogeneous foundation stock of the YSD
Validation of a novel perfusion bioreactor system in cancer research
Get PDFDevelopment of drugs is a complex, time- and cost-consuming process due to the lack of standardized and reliable characterization techniques and models. Traditionally, drug screening is based on in vitro analysis using two-dimensional (2D) cell cultures followed by in vivo animal testing. Unfortunately, application of the obtained results to humans in about 90 % of cases fails. Therefore, it is important to develop and improve cell-based systems that can mimic the in vivo-like conditions to provide more reliable results. In this paper, we present development and validation of a novel, user-friendly perfusion bioreactor system for single use aimed for cancer research, drug screening, anti-cancer drug response studies, biomaterial characterization, and tissue engineering. Simple design of the perfusion bioreactor provides direct medium flow at physiological velocities (100-250 mu m s(-1)) through samples of different sizes and shapes. Biocompatibility of the bioreactor was confirmed in short term cultivation studies of cervical carcinoma SiHa cells immobilized in alginate microfibers under continuous medium flow. The results have shown preserved cell viability indicating that the perfusion bioreactor in conjunction with alginate hydrogels as cell carriers could be potentially used as a tool for controlled anti-cancer drug screening in a 3D environment
Immunomodulatory activity and protective effects of chokeberry fruit extract onListeria monocytogenesinfection in mice
No full textChokeberry (Aronia melanocarpa) fruit extracts (CE) are rich in polyphenols and usually exhibit immunomodulatory, anti-viral and anti-bacterial effects. We have previously shown that the CE used in this study activated macrophages and stimulated effector T cell differentiationin vitro. When applied orally to healthy mice, CE increased the proportion of CD11c(+)dendritic cells in the gut-associated lymphoid tissue. CE-pretreated BALB/c mice readily eradicated orally ingestedListeria monocytogenesas evidenced by a slighter decrease in body weight and number of bacteria recovered from the spleen and reduced spleen size compared to the control infected mice. CE pretreatment in infected mice resulted in higher proportions of CD11b(+)macrophages and CD8(+)cytotoxic T cells both in the gut and the spleen. Phagocytosis, reactive oxygen species production and the proportions of activated CD86(+)macrophages (CD11b(+)) and dendritic cells (CD11c(+)) were also enhanced in CE-pretreated infected mice. Furthermore, the expression of inducible nitric oxide synthase and IL-6 was increased in CE-pretreated infected mice and similar results were obtained in peritoneal macrophagesin vitro. This effect of CE was associated with increased phosphorylation of I kappa B and Notch1 production. Finally, CE pretreatment elevated the proportion of perforin-producing cells in the spleen compared to control infected mice. This study demonstrates that prophylactic treatment with CE leads to more rapid eradication of bacterial infection withL. monocytogenespredominantly through increased activity of myeloid cells in the gut and in the spleen.Peer-reviewed manuscript: [https://imagine.imgge.bg.ac.rs/handle/123456789/1638
Benzimidazole-based dual dipeptidyl peptidase-4 and xanthine oxidase inhibitors
No full textMultiple-targeting compounds might reduce complex polypharmacy of multifactorial diseases, such as diabetes, and contribute to the greater therapeutic success. Targeting reactive oxygen species-producing enzymes, as xanthine oxidase (XO), might suppress progression of diabetes-associated vascular complications. In this study a small series of benzimidazole derivatives (1-9) was evaluated for inhibitory activity against dipeptidyl peptidase-4 (DPP-4) and XO. One 1,3-disubstituted-benzimidazole-2-imine (5) and 1,3-thiazolo [3,2-a] benzimidazolone derivative (8) were shown as effective dual DPP-4 and XO inhibitors, with IC50 values lower than 200 mu M, and predicted binding modes with both target enzymes. Both selected dual inhibitors (compounds 5 and 8) did not show cytotoxicity to a greater extent on Caco-2 cells even at concentration of 250 mu M. These structures represent new non-purine scaffolds bearing two therapeutic functionalities, being DPP-4 and XO inhibitors, more favorable in comparison to DPP-4 inhibitors with DPP-4 as a single target due to pleiotropic effects of XO inhibition.Peer-reviewed version:[https://imagine.imgge.bg.ac.rs/handle/123456789/2817
Expression Pattern and Prognostic Significance of EVI1 Gene in Adult Acute Myeloid Leukemia Patients with Normal Karyotype
No full textAccording to current criteria, patients with acute myeloid leukemia with normal karyotype (AML-NK) are classified as intermediate risk patients. There is a constant need for additional molecular markers that will help in substratification into more precise prognostic groups. One of the potential new markers is Ecotropic viral integration 1 site (EVI1) transcriptional factor, whose expression is dissregulated in abnormal hematopoietic process. The purpose of this study was to examine EVI1 gene expression in 104 adult AML-NK patients and on 10 healthy bone marrow donors using real-time polymerase chain reaction method, and to evaluate association between EVI1 expression level and other molecular and clinical features, and to examine its potential influence on the prognosis of the disease. Overexpression of EVI1 gene (EVI1(+) status) was present in 17% of patients. Increased EVI1 expression was predominantly found in patients with lower WBC count (P = 0.003) and lower bone marrow blast percentage (P = 0.005). EVI1(+) patients had lower WT1 expression level (P = 0.041), and were negative for FLT3-ITD and NPM1 mutations (P = 0.036 and P = 0.003). Patients with EVI1(+) status had higher complete remission rate (P = 0.047), but EVI1 expression didn't influence overall and disease free survival. EVI1 expression status alone, cannot be used as a new marker for more precise substratification of AML-NK patients. Further investigations conducted on larger number of patients may indicate how EVI1 expression could influence the prognosis and outcome of AML-NK patients, by itself, or in the context of other molecular and clinical parameters
Laboratory reporting on the clinical spectrum of CFTR p.Arg117His: Still room for improvement
No full textThe clinical spectrum associated with cystic fibrosis transmembrane conductance regulator ( CFTR ) variant p.Arg117His is highly variable, ranging from full-blown cystic fibrosis (CF) in a small num- ber of cases to CFTR-related disorders (CFTR-RDs) or no symptoms at all. Therefore, taking into account phenotype variability is essential for interpretation. External quality assessment (EQA) schemes can help laboratories to objectively assess the quality of genotyping and reporting by the laboratory. Methods: We performed a retrospective longitudinal data analysis on laboratory performance regarding the interpretation of p.Arg117His during CF EQA scheme participation. Completeness and accuracy of re- porting on two mock clinical cases were each compared over time (case 1: 20 05, 20 07 and 2012; case 2: 2015 and 2018). These cases concerned subjects compound heterozygous for p.Phe508del and p.Arg117His in cis with 7T, but with different clinical backgrounds (family planning (case 1) versus diagnostic testing for a child (case 2)). Furthermore, we analyzed the influence of previous participations, annual test vol- ume, accreditation status and laboratory setting on overall performance. Results: Overall performance improved over time, except during the 2007 CF EQA scheme. In addition, previous participations had a beneficial effect on laboratory performance. Accreditation status, annual test volume and laboratory setting did not significantly influence total interpretation scores. Conclusions: In general, laboratories performed well on both cases, although reporting on the variable clinical spectrum of p.Arg117His in cis with 7T and on the disease liability of individual CFTR variants can still improve. Moreover, this study underlined the educational role of CF EQA schemes
Improvement of antifungal activity and therapeutic profile of fluconazole by its complexation with copper(II) and zinc(II) ions. Complex characterization and antimicrobial activity studies
Get PDFOver the last few decades, invasive fungal infections represent a serious problem for modern-day healthcare. Aspergillus, Candida and Cryptococcus species are the most common pathogens causing life-threatening infections. Therapeutic options for the treatment of fungal infections are presently limited to only four classes of compounds and each of these drug classes has significant therapeutic limitations, including serious toxic-side effects, resistance development and limited routes of administration. In order to overcome resistance of the clinically used antifungal triazole agents, we synthesized zinc(II) and copper(II) complexes with fluconazole (flz), {[ZnCl2(flz)2]·2C2H5OH}n (1) and {[CuCl2(flz)2].5H2O}n (2). These complexes were obtained from the reactions between ZnCl2 or CuCl2·2H2O with this antifungal agent in 1 : 2 molar ratio in ethanol at room temperature. The compounds were characterized by elemental analysis, NMR, IR and UV-Vis spectroscopy and mass spectrometry. The crystal structure of complex 1 was determined by a single-crystal X-ray diffraction analysis. The antimicrobial effect of both complexes and fluconazole was evaluated against different Candida species as well as Gram-positive and Gram-negative bacteria by means of minimal inhibitory concentrations (MICs). The obtained results have shown that, in most cases, the coordination of fluconazole to Zn(II) and Cu(II) ions leads to the enhancement of its antifungal activity. Both complexes showed strong inhibitory activity against C. albicans biofilm formation at concentrations lower than MIC values, as well as strong inhibition of C. albicans filamentation.6th International Electronic Conference on Medicinal Chemistr
Preliminarno ispitivanje sastava masnih kiselina u semenkama šipurka: primena ultrazvučne ekstrakcije
Get PDFRosehip seeds represent waste material which is obtained duringthe production of jams, marmalades, juices, teas, etc. Two methods were used forthe extraction of fatty acids from rosehip seeds: ultrasound-assisted extractioncombined with organic solvent extraction (UAE/OSCE) and organic solventconventional extraction (OSCE). Different solvent-to-sample ratio was used. Theidentification and quantitation of fatty acids was done by gas chromatography witha flame ionization detector (GC/FID). Applied methods yielded different types offatty acids, with stearic acid and linoleic acid, as the most abundant ones.Semenke šipurka predstavljaju otpadni materijal, bogat uljem, koji nastaje tokom proizvodnje džemova, marmelada, sokova, čajeva itd. Za ekstrakciju masnih kiselina iz semenki šipurka korišćene su dve metode: ultrazvučna ekstrakcija u kombinaciji sa ekstrakcijom organskim rastvaračem (UZE/KEOR)i konvencionalna ekstrakcija organskim rastvaračem (KEOR). Korišćeni su različiti odnosi zapremine rastvarača i mase uzoraka. Identifikacija i kvantifikacija masnih kiselina izvršena je metodom gasne hromatografije sa plameno-jonizujućim detektorom (GC/FID). Primenjenim ekstrakcionim metodama dobijene su različite masne kiseline, od kojih su stearinska i linolna bile najzastupljenije
Zinc(II) complexes with aromatic nitrogen-containing heterocycles as antifungal agents: Synergistic activity with clinically used drug nystatin
No full textThree novel Zn(II) complexes, [ZnCl2(qz)(2)] (1), [ZnCl2(1,5-naph)](n) (2) and [ZnCl2(4,7-phen)(2)] (3), where qz is quinazoline, 1,5-naph is 1,5-naphthyridine and 4,7-phen is 4,7-phenanthroline, were synthesized by the reactions of ZnCl2 and the corresponding N-heterocyclic ligand in 1:2 molar ratio in ethanol at ambient temperature. The characterization of these complexes was done by NMR, IR and UV-Vis spectroscopy, and their crystal structures were determined by single-crystal X-ray diffraction analysis. Complexes 1 and 3 are mononuclear species, in which Zn(II) ion is tetrahedrally coordinated by two nitrogen atoms belonging to two qz or 4,7-phen ligands, respectively, and by two chloride anions, while complex 2 is a 1D coordination polymer that contains 1,5-naph as bridging ligand between two metal ions. In agar disc-diffusion assay, complexes 1-3 manifested good inhibitory activity against two investigated Candida strains (C. albicans and C. parapsilosis), while not inducing toxic effects on the healthy human fibroblast cell line (MRC-5). This activity was not fungicidal, as revealed by the broth microdilution assay, however complex 3 showed the ability to modulate Candida hyphae formation, which is an important process during infection and showed significant synergistic effect with clinically used antifungal polyene nystatin