Lada Zivkovic

Analysis of premature centromere division (PCD) of the X chromosome in Alzheimer patients through the cell cycle

Cytogenetic analysis of the X chromosome in phytohaemagglutinin stimulated peripheral blood lymphocytes was evaluated in 12 sporadic Alzheimer disease (AD) patients and in 11 healthy subjects. For chromosome analysis two methods were used: (1) standard analysis of G-banded metaphase chromosomes and; (2) fluorescent in situ hybridization (FISH) for the detection of the X chromosome centromeric region in interphase nuclei. Cytogenetic analysis revealed that the X chromosome expresses premature centromere division (PCD) in AD females in 10.53% of metaphase cells and in 15.22% of interphase nuclei. In AD men the percentages were 3.98 and 6.06%, respectively. X chromosome PCD in the female control group showed a percentage of 7.46% in metaphase cells and 9.35% in interphase nuclei and in male controls the percentages were 2.84% in metaphases and 5.54% in interphase nuclei. The results of FISH analysis showed that PCD could occur much earlier than metaphase of mitosis, i.e. in interphase of the cell cycle, immediately after replication. The FISH method can be used for PCD verification in all phases of the cell cycle in various disorders including AD.

Enhanced Viability of Endothelial Colony Forming Cells in Fibrin Microbeads for Sensor Vascularization.

Enhanced vascularization at sensor interfaces can improve long-term function. Fibrin, a natural polymer, has shown promise as a biomaterial for sensor coating due to its ability to sustain endothelial cell growth and promote local vascularization. However, the culture of cells, particularly endothelial cells (EC), within 3D scaffolds for more than a few days is challenging due to rapid loss of EC viability. In this manuscript, a robust method for developing fibrin microbead scaffolds for long-term culture of encapsulated ECs is described. Fibrin microbeads are formed using sodium alginate as a structural template. The size, swelling and structural properties of the microbeads were varied with needle gauge and composition and concentration of the pre-gel solution. Endothelial colony-forming cells (ECFCs) were suspended in the fibrin beads and cultured within a perfusion bioreactor system. The perfusion bioreactor enhanced ECFCs viability and genome stability in fibrin beads relative to static culture. Perfusion bioreactors enable 3D culture of ECs within fibrin beads for potential application as a sensor coating.

Cyclin Dependent Kinase 11, Neuroinflammation and Alzheimer's Disease: A Review

Aging is the main risk factor for Alzheimer’s disease (AD). With aging, inflammation has been recognized as potential trigger for starting the neurodegenerative cascade leading to neuronal death. Before Aβ and tau accumulation, evidence has put alterations of the cell cycle at the core of these processes. Still, a number of features of the cell cycle re-entry phenotype have remained elusive to the role of ectopic protein expression in the process of neuroinflammation and consequently neuronal cell death. Recently, a novel cyclin dependent kinase CDK11 has been found to be involved in astrocyte mediated inflammatory response and Alzheimer’s disease. In this review, we aim to establish the missing part of the puzzle between neuroinflammation and APP / Aβ deregulation in AD by evaluating the role of a cyclin, CDK11. CDK11 may play a vital role in cell cycle re-entry in AD neurons in an APP-dependent manner, thus presenting an intriguing novel function of the APP signaling pathway in AD.

Evaluation of the effects of ephedrine on human lymphocytes in the comet assay.

Ephedrine, a natural alkaloid from plants of the genus Ephedra, has a chemical structure similar to catecholamines. It is well established that catecholamines (adraneline, noradrenaline and dopamine) cause genotoxic and mutagenic effects. Therefore, the objectives of this investigation were to examine weather ephedrine can exhibit genotoxic effects on isolated human lymphocytes in the Comet assay. Dose-response of human lymphocytes was determined at the concentration range of ephedrine from 0.0005 μM to 500 μM. Three concentrations of ephedrine (1, 50 and 300 μM) which had acceptable cell viability (over 90%) were used for further experiments with inhibitors of DNA reparation (cytosine arabinoside and hydroxyurea). The obtained results showed that ephedrine did not induce DNA damage in isolated human lymphocytes. However, co-treatment of the negative control with DNA repair inhibitors caused a slight but significant increase of DNA damage, due to an endogenous DNA damage. Interestingly, cells treated with ephedrine and DNA repair inhibitors did not express increased DNA damage. On the basis of the obtained results it can be concluded that ephedrine did not exhibit genotoxic effects on isolated human lymphocytes. This result is in accordance with previous investigations showing negative genotoxicological results for ephedrine using bacterial gene mutation test-systems and in vitro cytogenetic analysis.

Genoprotective Capacity of Alternatively Cultivated Lingzhi or Reishi Medicinal Mushroom, Ganoderma lucidum (Agaricomycetes), Basidiocarps.

Ganoderma lucidum is traditionally used in Eastern medicine to preserve vitality, promote longevity, and treat disease. It possesses immunomodulatory, antitumor, antimicrobial, and antiaging activities, among others, but one of the most important is its antioxidant property, which is the basis for other effects, because free radicals trigger many diseases. The substrate commonly used for commercial cultivation of G. lucidum is not environmentally friendly nor economically justified, so there is a need to find new alternative substrates. The aim of this study was to analyze the effect of substrate composition on the bioactivity of G. lucidum basidiocarps. G. lucidum was cultivated on 2 different substrates: (1) a mixture of wheat straw, grapevine branches, and wheat bran, and (2) wheat straw. Commercial fruiting bodies, cultivated on oak sawdust, were used as the control. 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging ability, total phenols, and flavonoid content were determined spectrophotometrically to define the antioxidative potential of basidiocarp extracts. The comet test was performed to detect the degree of DNA damage in the cells that were exposed to G. lucidum extracts before and after the effect of oxidants. Higher antioxidative potential was observed for the extract of G. lucidum basidiocarps cultivated on wheat straw compared with that from the mixed substrate and especially with commercial ones. The alternatively cultivated basidiocarps also showed stronger antigenotoxic potential compared with commercial ones. The study showed that fruiting bodies produced on wheat straw, one of the most accessible and cheapest crop residues, are more potent antioxidant and antigenotoxic agents than commercially cultivated ones.

Lack of clastogenic effects of L-thyroxine in whole-blood cultured human lymphocytes

Abstract Thyroidhormonesstimulateaerobicmetabolismwhichmayleadtooxidativestressaccompaniedbydamagetovari-ous cellular macromolecules, including DNA. Previous comet assay studies have shown that thyroid hormonescause DNA damage due to the creation of reactive oxygen species (ROS). However, cytogenetic studies have beenequivocal because although an increase in the sister-chromatid exchange frequency per cell has been reported in-creasedmicronucleifrequencyhasnot.Weusedcytogeneticexaminationofchromosomebreakageandaberrationsinwhole-bloodculturesofhumanperipheralbloodlymphocytestoinvestigatepossibleclastogeniceffectswhenlym-phocytes were exposed to 0.002 μMto50μM of L-thyroxine for 24 h and 48 h, these concentrations being chosenbecausetheyhadbeenusedinpreviousstudiesofsister-chromatidexchangeandmicronucleifrequency.Underourexperimental conditions thyroxine did not induced any statistically significant increase in chromosome breakage oraberrations. This lack of clastogenic effects is in contrast to the reported comet assay results obtained using purifiedlymphocytes, possibly because whole-blood cultures contain catalase and glutathione peroxidase capable of reduc-ing the effects of reactive oxygen species.

Curcumin-loaded low-energy nanoemulsions as a prototype of multifunctional vehicles for different administration routes: Physicochemical and in vitro peculiarities important for dermal application

Graphical abstract Figure. No Caption available. Abstract The objective of this work was to investigate and profoundly characterize low‐energy nanoemulsions as multifunctional carriers, with slight reference to dermal administration. An evidence‐based approach was offered for deepening the knowledge on their formation via spontaneous emulsification. Curcumin, a compound of natural origin, potentially powerful therapeutic, was chosen as a model API. Due to curcumins demanding properties (instability, poor solubility, low permeability), its potentials remain unreached. Low‐energy nanoemulsions were considered carriers capable of overcoming imposed obstacles. Formulation consisting of Polysorbate 80 and soybean lecithin as stabilizers (9:1, 10%), medium‐chain triglycerides as the oil phase (10%) and ultrapure water was selected for curcumin incorporation in 3 different concentrations (1, 2 and 3 mg/mL). Physicochemical stability was demonstrated during 3 months of monitoring (mean droplet size: 111.3–146.8 nm; PDI < 0.2; pH: 4.73–5.73). Curcumin’s release from developed vehicles followed Higuchis kinetics. DPPH (IC50 = 0.1187 mg/mL) and FRAP (1.19 ± 0.02 mmol/g) assays confirmed that curcumin acts as a potent antioxidant through different mechanisms, with no alterations after incorporation in the formulation. High biocompatibility in line with antigenotoxic activity of curcumin‐loaded formulations (protective and reparative) was estimated through Comet assay. A multidisciplinary approach is needed to fully characterize developed systems, directing them to more concrete application possibilities.

The influence of the induction of farrowing on live birth, body mass, appearance of dystocia, mortality and surviving of neonatal pigs in litter during the first ten days

* Rad primljen za štampu 11.03.2015. ** Dr sc. vet. med. Jović Slavoljub, van. profesor, dr sc. vet. med. Ćupić Vitomir, red. profesor, Fakultet veterinarske medicine, Univerzitet u Beogradu; Ristić Gordana, dr vet. med. spec., Delta Agrar, Beograd, dr sc. vet. med. Vakanjac Slobodanka, vanr. profesor, dr sc. vet. med. Dimitrijević Blagoje, docent, Fakultet veterinarske medicine, Univerzitet u Beogradu, Miladinović Dejana, dr vet. med., doktorant, Fakultet veterinarske medicine, Univerzitet u Beogradu, dr sc. biol. Živković Lada, docent, Farmaceutski fakultet, Univerzitet u Beogradu ORIGINALNI RAD / ORIGINAL PAPER

Alterations of the x chromosome in lymphocytes of Alzheimer disease patients

Chromosomal alterations as a sign of genetic instability are a feature of Alzheimers disease (AD). Assessment of the genetic instability of non-neuronal cells of AD patients may provide a method to diagnose or monitor prognosis of the disease. Considering the importance of X chromosome alterations in the possible etiology of AD females, we used fluorescent in situ hybridization (FISH) for the centromere region of the X chromosome to determine aneuploidy, for a possible correlation with premature centromere division (PCD, X) in lymphocytes of AD females and age-matched controls. In AD patients, our results showed a marked and significant increase in the frequency of the X chromosome aneuploidy comparing with age matched controls (p<0.001). Also, a significant difference was detected in the PCD, X frequency between AD females when compared with age matched controls (p<0.001). In addition, a strong (R2=0.97, n=20) and significant (p<0.001) correlation was found between the frequency of aneuploidy and PCD, X in the AD group. Our results support the view that AD is a generalized systematic disease where PCD is to be considered as a stable sign of disease leading to aneuploidy.

X chromosome premature centromere division in neurons of Alzheimer patients

interaction of Grb2-SH2 with AICD and the consequent movement of the complex mediated by the SH3 domains. En route this complex is localized in vesicular structures inside the cell. Conclusions: Grb2 mediated rerouting of APP/AICD traffic might have long term implications in AD pathogenesis. Especially since SH3 domain of Grb2 is supposed to regulate the movement by interacting with cytoskeletal proteins, it would be interesting to know the fate of the traffic in a degenerating neuron. Further, the differential role of N-terminal and C-terminal SH3 domain in this trafficking is also to be deciphered.