Ninoslav Djelic

Premature centromere division of the X chromosome in neurons in Alzheimer's disease.

Premature centromere division (PCD) represents a loss of control over the sequential separation and segregation of chromosome centromeres. Although first described in aging women, PCD on the X chromosome (PCD,X) is markedly elevated in peripheral blood lymphocytes of individuals suffering from Alzheimer disease (AD). The present study evaluated PCD,X, using a fluorescent in situ hybridization method, in interphase nuclei of frontal cerebral cortex neurons from sporadic AD patients and age‐matched controls. The average frequency of PCD,X in AD patients (8.60 ± 1.20%) was almost three times higher (p < 0.01) than in the control group (2.96 ± 1.20). However, consistent with previous studies, no mitotic cells were found in neurons in either AD or control brain, suggesting an intrinsic inability of post‐mitotic neurons to divide. In view of the fact that it has been well‐documented that neurons in AD can re‐enter into the cell division cycle, the findings presented here of increased PCD advance the hypothesis that deregulation of the cell cycle may contribute to neuronal degeneration and subsequent cognitive deficits in AD.

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.

Analysis of premature centromere division (PCD) of the chromosome 18 in peripheral blood lymphocytes in Alzheimer disease patients.

Premature centromere division (PCD) of the chromosome 18 was analyzed by using fluorescent in situ hybridization (FISH) on interphase peripheral blood lymphocytes isolated from six sporadic Alzheimer disease (AD) patients and six healthy elderly controls. Results of FISH analysis revealed that chromosome 18 expressed PCD in 5.18% interphase nuclei of AD patients, and in 2.59% interphase nuclei of age-matched controls (p<0.05). Our study also showed that hypoploidy and hyperploidy frequency for chromosome 18 exhibited a statistically significant increase in the AD group compared to the control one. The increase in spontaneous aneuploidy of chromosome 18 in AD patients which is correlated with PCD shows that deregulation of the time of centromere separation can be considered as a manifestation of chromosome instability leading to aneuploidy.

Protective effect of dry olive leaf extract in adrenaline induced DNA damage evaluated using in vitro comet assay with human peripheral leukocytes.

Excessive release of stress hormone adrenaline is accompanied by generation of reactive oxygen species which may cause disruption of DNA integrity leading to cancer and age-related disorders. Phenolic-rich plant product dry olive leaf extract (DOLE) is known to modulate effects of various oxidants in human cells. The aim was to evaluate the effect of commercial DOLE against adrenaline induced DNA damage in human leukocytes by using comet assay. Peripheral blood leukocytes from 6 healthy subjects were treated in vitro with three final concentrations of DOLE (0.125, 0.5, and 1mg/mL) for 30 min at 37°C under two different protocols, pretreatment and post-treatment. Protective effect of DOLE was assessed from its ability to attenuate formation of DNA lesions induced by adrenaline. Compared to cells exposed only to adrenaline, DOLE displayed significant reduction (P<0.001) of DNA damage at all three concentrations and under both experimental protocols. Pearson correlation analysis revealed a significant positive association between DOLE concentration and leukocytes DNA damage (P<0.05). Antigenotoxic effect of the extract was more pronounced at smaller concentrations. Post-treatment with 0.125 mg/mL DOLE was the most effective against adrenaline genotoxicity. Results indicate genoprotective and antioxidant properties in dry olive leaf extract, strongly supporting further explorations of its underlying mechanisms of action.

Mislocalization of CDK11/PITSLRE, a regulator of the G2/M phase of the cell cycle, in Alzheimer disease

Post-mitotic neurons are typically terminally differentiated and in a quiescent status. However, in Alzheimer disease (AD), many neurons display ectopic re-expression of cell cycle-related proteins. Cyclin-dependent kinase 11 (CDK11) mRNA produces a 110-kDa protein (CDK11p110) throughout the cell cycle, a 58-kDa protein (CDK11p58) that is specifically translated from an internal ribosome entry site and expressed only in the G2/M phase of the cell cycle, and a 46-kDa protein (CDK11p46) that is considered to be apoptosis specific. CDK11 is required for sister chromatid cohesion and the completion of mitosis. In this study, we found that the expression patterns of CDK11 vary such that cytoplasmic CDK11 is increased in AD cellular processes, compared to a pronounced nuclear expression pattern in most controls. We also investigated the effect of amyloid precursor protein (APP) on CDK11 expression in vitro by using M17 cells overexpressing wild-type APP and APP Swedish mutant phenotype and found increased CDK11 expression compared to empty vector. In addition, amyloid-β25–35 resulted in increased CDK11 in M17 cells. These data suggest that 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.

Enhanced Sister-Chromatid Exchange Rate in Human Lymphocytes Exposed to 17β Estradiol in Vitro

BACKGROUND Epidemiologic data and animal experiments strongly implicate that steroid hormones are involved in the process of malignant transformation due to their capability to stimulate mitotic division and/or elevate the level of mutations in susceptible cells. METHODS The objectives of this investigation were to evaluate the effects of 17beta estradiol in sister-chromatid exchange (SCE) test on cultured human peripheral blood lymphocytes. The lowest concentration of 17beta estradiol used in this experiment (10(-10) M) was calculated as comparable with the physiologic blood level of 17beta estradiol in women. Three experimental concentrations corresponded to minimal (7 x 10(-8) M), average (3.5 x 10(-6) M), and maximal (7 x 10(-6) M) therapeutic doses in human medicine. In addition, the highest concentrations exceed maximal therapeutic dose 10-fold (7 x 10(-5) M) and 30-fold (2.1 x 10(-4) M), respectively. RESULTS The obtained results indicate that estradiol significantly elevates SCE per cell frequency at all concentrations applied except at the lowest one. However, estradiol has not influenced mitotic activity of cultured human lymphocytes significantly. CONCLUSIONS It can be concluded that 17beta estradiol expressed genotoxic effects and therefore might represent a human health risk.

Stimulating effect of sugar dusting on honey bee grooming behaviour

The aim of this research was to investigate whether or not sugar dusting can stimulate the grooming behaviour in Apis mellifera L. (Hymenoptera: Apidae), an important defensive mechanism against Varroa destructor Anderson & Trueman (Acari: Varroidae), and to assess the most effective dose and frequency of treatment. The criterion for evaluation of grooming potential was the percentage of damaged mites (PDM) among the total number collected on the bottom boards of the hives. In each sugar‐treated group PDM was significantly higher in comparison both with the negative control (no treatment) and with the values preceding the treatment. The results point to a stimulating effect of sugar on the grooming behaviour at all doses and frequencies tested. Treatment frequency influenced the stimulating effect of sugar: treatments at 3‐ and 7‐day intervals with 30 and 40 g resulted in significantly higher PDMs than the least frequent treatment (every 14 days); dusting with 20 g influenced PDM only when repeated at 3‐day intervals. Because treatments at 3‐day intervals are time‐consuming, those with 40 or 30 g repeated every 7 days may be recommended. In the positive control (hives treated with amitraz), average PDM was significantly lower than in the negative control and all sugar‐treated groups. Possible causes of the stimulating effect of sugar dusting on bee grooming behaviour are discussed.

Premature Centromere Division of Metaphase Chromosomes in Peripheral Blood Lymphocytes of Alzheimer's Disease Patients: Relation to Gender and Age

Chromosomal alterations are a feature of both aging and Alzheimers disease (AD). This study examined if premature centromere division (PCD), a chromosomal instability indicator increased in AD, is correlated with aging or, instead, represents a de novo chromosomal alteration due to accelerating aging in AD. PCD in peripheral blood lymphocytes was determined in sporadic AD patients and gender and age-matched unaffected controls. Metaphase nuclei were analyzed for chromosomes showing PCD, X chromosomes with PCD (PCD,X), and acrocentric chromosomes showing PCD. AD patients, regardless of age, demonstrated increased PCD on any chromosome and PCD on acrocentric chromosomes in both genders, whereas an increase in frequency of PCD,X was expressed only in women. This cytogenetic analysis suggests that PCD is a feature of AD, rather than an epiphenomenon of chronological aging, and may be useful as a physiological biomarker that can be used for disease diagnosis.

In vitro cytogenetic analysis of the effects of oxytocin on human peripheral blood lymphocytes.

The purpose of this study was to determine possible genotoxic and cytotoxic (or mitogenic) effects of high concentrations of oxytocin, active component of Syntocinon in cultures of human peripheral blood lymphocytes. Two test systems were used: (1) analysis of numerical and structural chromosome aberrations, and (2) the in vitro sister chromatid exchange (SCE) test. On the basis of the results obtained it can be concluded that oxytocin does not express any genotoxical properties. Furthermore, the mitotic index did not change significantly.

Dry Olive Leaf Extract Counteracts L-Thyroxine-Induced Genotoxicity in Human Peripheral Blood Leukocytes In Vitro

The thyroid hormones change the rate of basal metabolism, modulating the consumption of oxygen and causing production of reactive oxygen species, which leads to the development of oxidative stress and DNA strand breaks. Olive (Olea europaea L.) leaf contains many potentially bioactive compounds, making it one of the most potent natural antioxidants. The objective of this study was to evaluate the genotoxicity of L-thyroxine and to investigate antioxidative and antigenotoxic potential of the standardized oleuropein-rich dry olive leaf extract (DOLE) against hydrogen peroxide and L-thyroxine-induced DNA damage in human peripheral blood leukocytes by using the comet assay. Various concentrations of the extract were tested with both DNA damage inducers, under two different experimental conditions, pretreatment and posttreatment. Results indicate that L-thyroxine exhibited genotoxic effect and that DOLE displayed protective effect against thyroxine-induced genotoxicity. The number of cells with DNA damage, was significantly reduced, in both pretreated and posttreated samples (P < 0.05). Comparing the beneficial effect of all tested concentrations of DOLE, in both experimental protocols, it appears that extract was more effective in reducing DNA damage in the pretreatment, exhibiting protective role against L-thyroxine effect. This feature of DOLE can be explained by its capacity to act as potent free radical scavenger.