A comprehensive evaluation of the genotoxic potential of chemicals requires the assessment of the ability to induce gene mutations and structural chromosome (clastogenic activity) and numerical chromosome (aneugenic activity) aberrations. Aneuploidy is a major cause of human reproductive failure and an important contributor to cancer and it is therefore important that any increase in its frequency due to chemical exposures should be recognized and controlled. The in vitro binucleate cell micronucleus assay provides a powerful tool to determine the ability of a chemical to induce chromosome damage. The application of an anti-kinetochore antibody to micronuclei allows their classification into kinetochore-positive and kinetochorenegative, indicating their origin by aneugenic or clastogenic mechanisms, respectively. The availability of chromosomespecific centromere probes allows the analysis of the segregation of chromosomes into the daughter nuclei of binucleate cells to evaluate chromosome non-disjunction. Quantitative relationships between the two major causes of aneuploidy, chromosome loss and non-disjunction, can be determined. The mechanisms leading to chromosome loss and non-disjunction can be investigated by the analysis of morphological and structural changes in the cell division apparatus by the application of specific stains and antibodies for various cell division components. We illustrate such analyses by the demonstration of the interaction of the monomer bisphenol-A with the centrosome of the mitotic spindle and the folic acid antagonist pyrimethamine with the centromeres of chromosomes. Both types of modifications lead to the induction of aneuploidy in exposed cells. Our studies also implicate the products of the p53 and XPD genes in the regulation of the fidelity of chromosome segregation at mitosis.

This study compares the lipid peroxidation marker urinary thiobarbituric acid reactive substances (TBARS) and antioxidants including plasma alpha-tocopherol (vitamin E), plasma (P-GSH-Px) and erythrocyte glutathione peroxidase (E-GSH-Px) activities, and plasma selenium levels in two groups of type 2 diabetic subjects (both n=20) with a disease duration of < or =2 (GP1) and 4-6 years (GP2), and non-diabetic age and gender-matched control subjects (CG, n=20). The mean (standard deviation [SD]) age of the groups was similar at 41(10) years. Fasting blood and midstream urine samples were obtained from diabetic and non-diabetic subjects attending the diabetic clinic and HbAlc, fructosamine, urine TBARS, total antioxidant (TAS) levels, P-GSH-Px, E-GSHPx and plasma selenium and vitamin E concentrations were measured. HbA1c (%) and fructosamine levels in the GP1 and GP2 diabetic subjects and the controls were 5.75 (0.67), 11.43 (2.01) and 4.33 (0.47), and 3.09 (0.57), 6.09 (1.15) and 1.67 (0.31), respectively (GP1 vs. GP2, GP1 vs. GC and GP2 vs. CG, all P < 0.001). Elevated urinary TBARS (micromol/mmol urinary creatinine) in the GP1, GP2 and GC groups were 2.47 (0.37), 3.73 (0.93) and 1.18 (0.24), respectively (GP1 vs. GP2, GP1 vs. GC and GP2 vs. CG, all P < 0.001). A significant correlation between HbA1c and TBARS was also noted (r2 = 0.894, P < 0.001) but only in the GP2 subjects. TAS levels were only decreased in the GP2 group compared to control values (0.59 [0.18] vs. 1.74 [0.21], P < 0.001). Plasma vitamin E concentrations (micromol/L) of 34.11 (3.31), 9.57 (2.20) and 39.01 (2.91) were observed in the GP1, GP2 and GC groups, respectively (GP1 vs. CG, P < 0.05 and GP1 vs. GP2 and GP vs. CG, both P < 0.001). E-GSH-Px (U/g Hb) and P-GSH-Px (U/L) activities in GP1, GP2 and CG groups were also decreased at 57.04 (4.31), 24.0 (8.94) and 67.6 (4.29), and 6.16 (1.56), 2.67 (0.47) and 8.72 (0.31), respectively (E-GSH-Px: CG vs. GP1, P < 0.01, CG vs. GP2 and GP1 vs. GP2, both P < 0.001; P-GSH-Px: CG vs. GP1, CG vs. GP2 and GP1 vs. GP2, all P < 0.001). Plasma selenium levels (miromol/L) were only significantly decreased in GP2 compared to both GP1 and CG values (0.49 [0.29] vs. 1.67 [0.80] vs. 1.79 [0.26], both P < 0.001). These observations support the suggestion that chronic hyperglycaemia can influence the generation of free radicals, which may lead ultimately to increased lipid peroxidation and depletion of antioxidants, and thereby enhanced oxidative stress in subjects with type 2 diabetes mellitus.

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The neural retina is subjected to various degenerative conditions. Regenerative stem-cell-based therapy holds great promise for treating severe retinal degeneration diseases, although many drawbacks remain to be overcome. One important problem is to gain authentically differentiated cells for replacement. Paired box 6 protein (5a) (PAX6 (5a)) is a highly conserved master control gene that has an essential role in the development of the vertebrate visual system. Human adipose-tissue-derived stem cell (hADSC) isolation was performed by using fat tissues and was confirmed by the differentiation potential of the cells into adipocytes and osteocytes and by their surface marker profile. The coding region of the human PAX6 (5a) gene isoform was cloned and lentiviral particles were propagated in HEK293T. The differentiation of hADSCs into retinal cells was characterized by morphological characteristics, quantitative real-time reverse transcription plus the polymerase chain reaction (qPCR) and immunocytochemistry (ICC) for some retinal cell-specific and retinal pigmented epithelial (RPE) cell-specific markers. hADSCs were successfully isolated. Flow cytometric analysis of surface markers indicated the high purity (~97 %) of isolated hADSCs. After 30 h of post-transduction, cells gradually showed the characteristic morphology of neuronal cells and small axon-like processes emerged. qPCR and ICC confirmed the differentiation of some neural retinal cells and RPE cells. Thus, PAX6 (5a) transcription factor expression, together with medium supplemented with fibronectin, is able to induce the differentiation of hADSCs into retinal progenitors, RPE cells and photoreceptors.

Colorectal cancer is a growing health concern with increasing mortality rates, and resistance to anticancer drugs and radiotherapy is a serious drawback in its treatment. Auraptene is a natural prenyloxycoumarin with valuable anticancer effects. The aim of current study was to determine the synergy between auraptene, ionizing radiation, and chemotherapeutic drugs in colon adenocarcinoma cells for the first time. To do so, HT29 cells were treated with combination of auraptene + cisplatin, + doxorubicin, or + vincristine. Furthermore, cells were pretreated with nontoxic auraptene and then exposed to various doses of X-radiation. Assessment of cell viability not only indicated significant (p < 0.05) synergic effects of auraptene and anticancer agents, also revealed more significant (p < 0.01) increase in the toxicity of applied radiations in auraptene pretreated cells. Interesting synergy between auraptene and radiotherapy was then confirmed by morphological alterations, DAPI staining, and flow cytometric analysis of the cell cycle. Moreover, real-time reverse transcription polymerase chain reaction analysis indicated significant (p < 0.01) overexpression of p21, but not GATA6, in auraptene pretreated cells after radiotherapy, and also significant (p < 0.01) down regulation of CD44 and ALDH1 by auraptene. According to present results, auraptene could be considered as an effective natural coumarin to improve the outcome of current chemoradiotherapy options. Copyright © 2017 John Wiley & Sons, Ltd.

Environmental pollution caused by heavy metals such as mercury is one of the most important human problems. It might have severe teratogenic effects on embryonic development. Some pharmacological and physiological aspects of fruit flies (Drosophila melanogaster) are similar to humans. So the stages of egg to adult fruit fly, as a developmental model, were employed in the study. Wild adult insects were maintained in glass dishes containing standard medium at 25 °C in complete darkness. Five pairs of 3-day old flies were then transferred to standard culture dishes containing different concentrations of mercury ion. They were removed after 8 hours. We considered the following: The rate of larvae becoming pupae and pupae to adults; the time required for the development; the hatching rate in the second generation without mercury in the culture; the morphometric changes during development in both length and width of the eggs through two generations; larvae, pupae and adult thorax length and width. The results showed that mercury in culture (20–100 mg/l) increase the duration of larvae (p<0.01) and pupae (p<0.01) development, the rate of larvae becoming pupae (p<0.001); pupae maturation (p<0.05), the hatching rate (p<0.01), the length (p<0.05) and width of larvae (p<0.01) and pupae (p<0.001) and the length in the adult thorax (p<0.01) decreased significantly. There was no effect upon the size of eggs. There were also no larvae hatching in concentrations of 200 mg/l of mercury. Negative effects of mercury as a heavy metal are possibly due to the interference of this metal in cellular signaling pathways, such as: Notch signaling and protein synthesis during the period of development. Since it bonds chemically with the sulfur hydride groups of proteins, it causes damage to the cell membrane and decreases the amount of RNA. This is the cause of failure of many enzyme mechanisms.

The association between urine microalbumin, alpha1-microglobulin concentration (alpha1MG) and the urinary enzyme activities of alanine aminopeptidase (AAP), N-acetyl-beta-D-glucosaminidase (NAG), alpha-glutathione-S-transferase (alphaGST) and pi-glutathione-S-transferase (piGST) is investigated in 36 type 2 diabetic and 15 age- and sex-matched non-diabetic subjects. Diabetic subjects were grouped into those with microalbuminuria <3 mg/L (group A: 7M/5F), 3-30 mg/L (group B: 5M/7F) and 30-300 mg/L (group C: 6M/6F). While serum creatinine concentration remained within the laboratory reference range (<115 mmol/L) in all experimental groups, alpha1MG excretion increased with the severity of microalbuminuria (control group and groups A, B and C mean [SD] values were 1.3 [0.21], 1.6 [0.11], 2.18 [0.42] and 2.8 [0.51] mg/mmol urinary creatinine, respectively). Activities of NAG (U/mmol creatinine) were significantly elevated in groups A, B and C at 98.7 (8.6), 112.8 (12.9) and 147.4(16.2), respectively, compared with the reference range <35 U/mmol creatinine (group C vs. groups A and B: P < 0.01). Activity of AAP (U/mmol creatinine) was significantly elevated in groups B and C at 7.6 (0.5) and 7.9 (0.6), respectively (both P < 0.001), compared to the control and group A values (2.5 [0.2]). Activity of piGST (U/mmol creatinine) was elevated in groups B and C at 2.6 (0.4) and 2.8 (0.5), respectively (both P < 0.001), compared to the control and group A values (1.1 [0.1]). Similarly, urine piGST activity was also elevated in groups B and C at 2.9 (0.6) and 3.1 (0.5), respectively (both P < 0.001), compared to control and group A values (1.3 [0.1] and 1.4 [0.2]). These results suggests that site-specific urinary biochemical markers provide valuable information about early renal proximal and distal tubular insult that ultimately may precede enhanced glomerular permeability in subjects with type 2 diabetes.

Acute physiological stress induces remarkable effects on the nervous, endocrine, and immune systems and also on cellular metabolism and cell division processes. Stress-induced instability of cellular mechanisms might play an important role in increasing cell division disorders. In this study, a relationship between stress and micronucleus (MN) induction in mouse (balb/c) bone marrow cells following vinblastine treatment, or stress or stress and vinblastine treatment in comparison to a non-stressed control group was investigated. In order to test the effects of treatments on MN induction, an in vivo MN assay was performed on bone marrow cells. The results revealed a significantly greater increase in MNs in bone marrow cells (polychromatic erythrocytes) from the stressed/vinblastine treated mice. The data indicate the ability of exposure to an emotional stressor to enhance the damaging actions on bone marrow cells of an aneugenic agent.