imagine (Institute of molecular genetics and genetic engineering)
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Vision, challenges and opportunities for a Plant Cell Atlas
Get PDFWith growing populations and pressing environmental problems, future economies will be increasingly plant-based. Now is the time to reimagine plant science as a critical component of fundamental science, agriculture, environmental stewardship, energy, technology and healthcare. This effort requires a conceptual and technological framework to identify and map all cell types, and to comprehensively annotate the localization and organization of molecules at cellular and tissue levels. This framework, called the Plant Cell Atlas (PCA), will be critical for understanding and engineering plant development, physiology and environmental responses. A workshop was convened to discuss the purpose and utility of such an initiative, resulting in a roadmap that acknowledges the current knowledge gaps and technical challenges, and underscores how the PCA initiative can help to overcome them
A novel 9 bp deletion (c.1271_1279delGTGCCCGCG) in exon 10 of CYP21A2 gene causing severe congenital adrenal hyperplasia
No full textBackground Congenital adrenal hyperplasia (CAH) is an autosomal recessive disorder of adrenal steroidogenesis with a broad spectrum of clinical presentations, ranging from the severe classical salt-wasting (SW) and simple-virilizing (SV) form, to the mild nonclassical form. A large variety of CYP21A2 genotypes in correlation with phenotype have been described. Materials and methods DNA samples from a 14-day-old male newborn with clinical and laboratory signs of SW CAH and family members were subjected for molecular analysis of the nine most common point CYP21A2 mutations by ACRS/PCR method. Direct DNA sequencing of the whole CYP21A2 gene was performed to detect the second mutant allele in the patient. The in silico predicting analysis and the crystal structure analysis of the mutated CYP21A2 protein have been performed. Results Molecular analysis confirmed that the patient was compound heterozygote carrying p.Q318X mutation inherited from the mother and a novel c.1271_1279delGTGCCCGCG (p.G424_R426del) variant in exon 10 inherited from the father. The in silico predicting software tools classified the novel mutation as pathogenic. Crystal structure analysis showed that the three residues affected by the novel in-frame deletion form several hydrogen bonds that could lead to impaired stability and function of the CYP21A2 protein. These findings were concordant with the patient's phenotype. The need of several molecular methods to elucidate the genotype in this patient has also been discussed. Conclusions A novel 9 bp deletion in CYP21A2 gene with predicted pathogenic effect on the enzyme activity was detected in neonatal patient causing severe SW CAH
Colorectal cancer surgery in Serbia 2010-2014: An evaluation of a multicenter registry
Get PDFUvod/Cilj. Hirurški registri i baze podataka su posebno vredni za praćenje karakteristika lečenja i otkrivanje potencijalnih problema kod karcinoma. Za bolesnike sa kolorektalnim karcinomom (KRK) u Srbiji, podaci o primljenoj terapiji, faktorima koji mogu imati uticaja na ishod, odnosno da li je lečenje uspešno i odgovarajuće ili ne, trenutno nisu sistematizovano obrađeni. Cilj rada bio je uspostavljanje baze usaglašenih podataka o hirurškom lečenju KRK u različitim državnim bolnicama širom Srbije i procena ishoda operativnog lečenja obolelih od KRK u periodu od 2010. do 2014. godine. Metode. Istraživanjem su obuhvaćena 52 državna hirurška centra u Srbiji u kojima su sakupljani brojčani podaci o bolesnicima podvrgnutim hirurškoj resekciji KRK. Kreirana je elektronska baza podataka pod nadzorom Prve hirurške klinike Kliničkog centra Srbije, Beograd. Podaci su nezavisno sakupljani korišćenjem posebno sastavljenog standardizovanog upitnika koji je uključio broj operisanih bolesnika, lokalizaciju primarnog tumora, tip hirurške intervencije, urgentnost zahvata i postoperativni mortalitet. Rezultati. Ukupno 22 037 kolorektalnih hirurških procedura izvedeno je u Srbiji u periodu od 2010. do 2014. godine (približno 4 400 godišnje) od čega je bilo 78,5% elektivnih, a 21,5% urgentnih operacija. Najčešći uzrok urgentnih operacija bio je ileus (3 618 bolesnika, 76,4%), dok su manje česti uzroci bili perforacija (899 bolesnika, 18,9%) i krvarenje (216 bolesnika, 4,5%). Postoperativna smrtnost tokom perioda istraživanja, izražena kao procenat svih KRK intervencija, iznosila je prosečno 2,8% sa diskretnom tendencijom rasta. Na kraju studijskog perioda, 127 lekara je bilo obučeno za obavljanje kolonoskopije. Zaključak. Glavni ishod istraživanja bilo je uspostavljanje neophodnih preduslova za multicentrično sakupljanje podataka koji uključuje veliki broj obolelih od KRK. Studija je podržala pretpostavku da je razvoj nacionalne baze podataka hirurškog lečenja KRK dostižan i da može obezbediti značajan uvid u rutinsko hirurško lečenje KRK u Srbiji i doprineti uvećanju resursa za buduća istraživanja.Background/Aim. Surgical registries and databases are especially valuable in monitoring the performances in cancer treatment and detecting potential problems. For Serbian patients with colorectal cancer (CRC), data regarding the treatment received, the factors that may impact the outcome, and whether or not treatment is successful and appropriate are not currently captured. The aim of this study was to establish a collection of a consensus dataset capturing surgical treatment of CRC at multiple public hospitals across Serbia and estimate outcome in CRC patients subjected to surgical treatment in the period 2010-2014. Methods. The study encompassed all 52 public CRC surgical units in Serbia. Numerical data on all patients who underwent operative CRC resection were included. An electronic database was created and overseen by the First Surgical Clinic of the Clinical Center of Serbia, Belgrade. Data were collected independently using a specifically designed standardized questionnaire, including the number of operated patients, localization of the primary tumor, type of surgical intervention, type and urgency of surgical intervention, and postoperative mortality. Results. A total number of 22,037 colorectal surgical procedures was performed in Serbia in the period 2010-2014 (approximately 4,400 per year). It was shown that 78.5% of the total number of procedures were elective and 21.5% were emergency. The most common cause of emergency surgeries was ileus (3,618 cases, 76.4%), while the less common causes were perforation (899 cases, 18.9%) and bleeding (216 cases, 4.5%). Postoperative mortality during the study period expressed as a percentage of all interventions for CRC was 2.8% on average with a slight increasing tendency. At the end of the study period, 127 doctors were educated for performing colonoscopy. Conclusion. The main outcome of this study was the establishment of the necessary preconditions for the multicenter data collection involving large numbers of CRC patients. The study supported the premise that the development of the national database for surgical treatment of CRC is achievable and could provide valuable insight into the routine surgical management of CRC in Serbia, creating a significant resource for further research
Sex differences in the effects of early-life probiotic treatment on TNBS-induced colitis in rats
No full textSynthesis and Laccase-Mediated Oxidation of New Condensed 1,4-Dihydropyridine Derivatives
Get PDFWe describe herein the synthesis and laccase mediated oxidation of six novel 1,4-dihydropyridine (DHP)-based hexahydroquinolines (DHP1-DHP3) and decahydroacridines (DHP4-DHP6). We employed different laccase enzymes with varying redox potential to convert DHP1-DHP3 and DHP4-DHP6 to the corresponding pyridine-containing tetrahydroquinoline and octahydroacridine derivatives, respectively. Intensively coloured products were detected in all biocatalytic reactions using laccase from Trametes versicolor (TvLacc), possibly due to the presence of conjugated chromophores formed in products after oxidation. The NMR assessment confirmed that the oxidation product of DHP1 was its corresponding pyridine-bearing tetrahydroquinoline derivative. Laccase from Bacillus subtillis (BacillusLacc) was the most efficient enzyme for this group of substrates using HPLC assessment. Overall, it could be concluded that DHP2 and DHP5, bearing catecholic structures, were easily oxidized by all tested laccases, while DHP3 and DHP6 containing electron-withdrawing nitro-groups are not readily oxidized by laccases. DHP4 with decahydroacridine moiety consisting of three condensed six-membered rings that contribute not only to the volume but also to the higher redox potential of the substrate rendered this compound not to be biotransformed with any of the mentioned enzymes. Overall, we showed that multiple analytical approaches are needed in order to assess biocatalytical reactions
Synthesis and Biological Screening of New 4-Hydroxycoumarin Derivatives and Their Palladium(II) Complexes
Get PDFTwo newly synthesized 4-hydroxycoumarin bidentate ligands (L1 and L2) and their palladium(II) complexes (C1 and C2) were screened for their biological activities, in vitro and in vivo. Structures of new compounds were established based on elemental analysis, H-1 NMR, C-13 NMR, and IR spectroscopic techniques. The obtained compounds were tested for their antioxidative and cytotoxic activities and results pointed to selective antiradical activity of palladium(II) complexes towards (OH)-O-center dot and -center dot OOH radicals and anti-ABTS (2,2 '-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) cation radical) activity comparable to that of ascorbate. Results indicated the effect of C1 and C2 on the enzymatic activity of the antioxidative defense system. In vitro cytotoxicity assay performed on different carcinoma cell lines (HCT166, A375, and MIA PaCa-2), and one healthy fibroblast cell line (MRC-5) showed a cytotoxic effect of both C1 and C2, expressed as a decrease in carcinoma cells' viability, mostly by induction of apoptosis. In vivo toxicity tests performed on zebrafish embryos indicated different effects of C1 and C2, ranging from adverse developmental effect to no toxicity, depending on tested concentration. According to docking studies, both complexes (C1 and C2) showed better inhibitory activity in comparison to other palladium(II) complexes
RNA-targeting low-molecular-weight fluorophores for nucleoli staining: synthesis, in silico modelling and cellular imaging
No full textHerein we present our work on the synthesis, investigation of the photophysical properties, interactions with nucleic acids, molecular docking, and imaging application of three carbocyanine dyes. The described low-molecular-weight compounds were found to exhibit high resistance against photobleaching and showed promising optical properties as fluorescent labeling agents for ribonucleic acids. They form strong biocomplexes (log K-s = 6.11-7.84) and revealed remarkable sensitivity towards RNA, reaching up to a 379 times increase of the emission signal when bound to AU-rich sequences. According to the score values obtained from the molecular docking studies, the compounds show strong binding affinity towards RNA macromolecules. All fluorophores exhibit significant cell tolerance since they were found to be 16 to 60 times less toxic against MRC5 cells (healthy human fibroblasts) compared to the conventional Thiazole Orange - TO. The IC50 concentrations for the compounds were calculated up to 40 mu M in human fibroblasts MRC5, A549 adenocarcinomic human alveolar basal epithelial cells, the HCT116 human colon cancer cell line, and MDA-MB-231 adenocarcinoma cells. Analyzing the dyes for preferential cytotoxicity towards cancer cell lines in comparison to the normal human fibroblasts, we found a candidate exhibiting promising anticancer potential. Based on the selectivity (Si) towards cancer cells and more specifically against difficult to treat colon HCT116 carcinoma, we can suggest these small molecules as an interesting platform for further development. We have also demonstrated the efficiency of the carbocyanines as staining agents for in vivo labeling of human cells as well as microbial and eukaryotic cells
Increased Expression of Extracellular Vesicles Is Associated With the Procoagulant State in Patients With Established Rheumatoid Arthritis
Get PDFThis study sought to identify different subpopulations of extracellular vesicles (EVs) in plasma from female patients with established rheumatoid arthritis (RA) in relation to the activation of coagulation and fibrin formation in these patients. Forty women were included in the study, 20 patients and 20 age-matched healthy controls. The mean disease duration in patients was 13.0 (5.0-25.0) years, with medium to high disease activity despite ongoing treatment with low-dose prednisolone and methotrexate. There were no differences between the investigated groups regarding the presence of traditional cardiovascular risk factors. The concentration of phosphatidylserine-positive (PS+) EVs; platelet (CD42a(+)), leucocyte (CD45(+)), monocyte (CD14(+)), and endothelial (CD144(+))-derived EVs; and EVs-expressing tissue factor (CD142(+)), P-selectin (CD62P(+)), and E-selectin (CD62E(+)) were determined by flow cytometry analysis. Overall hemostasis potential (OHP) was assessed to follow the hemostatic disturbances, including the parameters for overall coagulation potential (OCP) and overall fibrinolytic potential (OFP). Fibrin clot turbidity was measured together with clot lysis time, and scanning electron microscopy was performed. Increased concentrations of PS+, CD42a(+), CD142(+), CD45(+), CD14(+), and CD62P(+) EVs were found in plasma from patients with RA compared to healthy controls, and the concentrations of PS+, CD42a(+), CD14(+), and CD62P(+) EVs were positively correlated with the inflammatory parameters in RA patients. Positive correlations were also found between the levels of PS+ and CD42a(+) EVs and OCP as well as between the levels of PS+, CD42a(+), and CD62P(+)EVs and OHP. The levels of PS+, CD42a(+), CD14(+), CD62P(+), and CD62E(+) EVs were negatively correlated with OFP. Elevated levels of circulating EVs of different cell origins were found in patients with established RA, in relation to the inflammatory burden and coagulation activation in the disease
Electroanalysis of Candida albicans biofilms: A suitable real-time tool for antifungal testing
No full textCandida albicans is a fungal pathogen that accounts for more than a million death annually. Analysis of Candida biofilms and rapid assessment of antifungal therapy is a critical challenge in clinical practice. Current biochemical methods are time-consuming and expensive or require expert interpretation of the results. Hence, there is the need for a rapid anti-fungal and anti-biofilm test. While electroanalysis has been previously adopted to determine antibiotic susceptibility in bacterial biofilms, there are no studies on yeast and fungi. In this work, electroanalysis of C. albicans biofilm and evaluation of anti-biofilm activity of several antifungal compounds using screen-printed carbon electrodes (SPEs) are reported. Results are compared with standard biochemical and microscopic methods. The current output decreases by 47.5%, 73.4%, and 88.5% in biofilms treated with the antifungals fluconazole (Flz), amphotericin B (AmB), and complex Ag3, respectively, indicating the suitability of electrochemical testing for evaluation of anti fungal drugs. Chronoamperometry allows measuring antifungal activity on Candida biofilm as early as 10 h after inoculation, which show promises for the development of bioelectrochemical sensors in clinical settings
Case Report: Successful Therapy of Spontaneously Occurring Canine Degenerative Lumbosacral Stenosis Using Autologous Adipose Tissue-Derived Mesenchymal Stem Cells
Get PDFThe management of degenerative lumbosacral stenosis (DLSS) in dogs usually requires aggressive, costly surgical treatments that may themselves present complications, while do not fully resolve the symptoms of the disease. In this study, the dog diagnosed with severe DLSS, with hind limb paresis, was treated using a new and least invasive treatment. Cultured autologous adipose tissue-derived mesenchymal stem cells (AT-MSCs) were injected bilaterally at the level of L7-S1, in the vicinity of the external aperture of the intervertebral foramen of DLSS patient. In the previously described treatments of spontaneous intervertebral disc degeneration in dogs, intradiscal injections of MSCs did not lead to positive effects. Here, we report a marked improvement in clinical outcome measures related to the ability of a dog to walk and trot, which were expressed by a numeric rating scale based on a veterinary assessment questionnaire. The improved status persisted throughout the observed time course of 4.5 years after the AT-MSC transplantation. To the best of our knowledge, this is the first case of successful therapy, with long-term positive effect, of spontaneously occurring canine DLSS using presented treatment that, we believe, represents a contribution to current knowledge in this field and may shape both animal and human DLSS treatment options