Study on induction of apoptosis on HeLa and Vero cells by recombinant shiga toxin and its subunits

Verotoxin (VT) or shiga toxin (Stx) produced by enterohemorrhagic Escherichia coli (EHEC) and Shigella dysenteriae is AB5 holotoxin with potent protein synthesis inhibitor. VT can induce both apoptosis and necrosis depending on the cell type, it has been shown that VT-induced apoptosis and cytotoxicity are distinct processes, and the A subunit can be necessary for apoptosis. In other words, the precise role of each subunit in apoptosis signaling has yet to be established. In this study, induction of apoptosis has been examined by using both recombinant A and B subunits, and recombinant Stx (rStx) with different doses in HeLa and Vero cells. For this purpose, the polymyxin B extract of constructs expressing A, B and AB5 recombinant proteins was used. Therefore, amounts greater than normally reported were used to induce desire effects on cell lines. The apoptotic effect of A and B subunits appear at higher doses than that of rStx. The highest apoptotic effect was observed for rStx at low concentration, compared to A and B subunits. A or B subunits separately cannot induce the signaling pathway stimulated by holotoxin though A subunit, does induce laddering pattern similar to holotoxin. We concluded that both subunits are important in complete death signaling pathway. Since different concentration of A and B subunits and rStx was required in different assay, therefore, it could be emphasized that cell death or even apoptosis caused by either of the subunits or holotoxin depends on sensitivity or specificity of the assay and cell types used.     

Spatiotemporal Correlations between Blood-Brain Barrier Permeability and Apparent Diffusion Coefficient in a Rat Model of Ischemic Stroke

Variations in apparent diffusion coefficient of water (ADC) and blood-brain barrier (BBB) permeability after ischemia have been suggested, though the correlation between ADC alterations and BBB opening remains to be studied. We hypothesized that there are correlations between the alteration of ADC and BBB permeability. Rats were subjected to 2 h of transient middle cerebral artery occlusion and studied at 3 and 48 h of reperfusion, which are crucial times of BBB opening. BBB permeability and ADC values were measured by dynamic contrast-enhanced MRI and diffusion-weighted imaging, respectively. Temporal and spatial analyses of the evolution of BBB permeability and ADC alteration in cortical and subcortical regions were conducted along with the correlation between ADC and BBB permeability data. We found significant increases in BBB leakage and reduction in ADC values between 3 and 48 h of reperfusion. We identified three MR tissue signature models: high K_i and low ADC, high K_i and normal ADC, and normal K_i and low ADC. Over time, areas with normal K_i and low ADC transformed into areas with high K_i. We observed a pattern of lesion evolution where the extent of initial ischemic injury reflected by ADC abnormalities determines vascular integrity. Our results suggest that regions with vasogenic edema alone are not likely to develop low ADC by 48 h and may undergo recovery.     

Study of defense-related gene expression in grapevine infested by <Emphasis Type="Italic">Colomerus vitis</Emphasis> (Acari: Eriophyidae)

As the main source of lipids and proteins in honey bees, pollen is a major nutrient provider involved in development and health and has been studied for tolerance stimulation against pathogens and parasites. In the case of Varroa destructor Anderson & Trueman (Acari, Mesostigmata: Varroidae) parasitization, the lack of a complete laboratory system to rear both the bee larva and the acarian parasite limited the studies concerning larval nutrition effects on the bee tolerance and resistance against varroatosis. Due to the development of this complete rearing protocol, we managed to feed young honey bee larvae with pollen supplemented solutions and to study the effect on their later development under parasitism conditions. In our experimental conditions, pollen influences neither the deformity rate, nor the survival of bees both parasitized and unparasitized. However, pollen extract supplementation seems to significantly impact the weight of the spinning bee larvae without having an effect on the physiological weight loss during pupation, so the differences found at the larval stage remain the same as at emergence. Varroa has a deleterious effect on bee pupae and led to a steady increase of the physiological weight loss experienced during metamorphosis. Interestingly, this ponderal loss associated with Varroa parasitization seems to be reduced in the polyfloral pollen supplementation condition. Altogether, this work is to our knowledge the first to study in laboratory conditions the impact of larval nutrition on the tolerance to parasitism. A diverse pollen diet may be beneficial to the bees’ tolerance against V. destructor parasitism.     

Widespread environmental contamination in agricultural soils and human health risk at the vicinity of an oil refinery site

In this research, carcinogenic and non-carcinogenic human health risks due to polycyclic aromatic hydrocarbons (PAHs) were investigated via three exposure pathways: accidental ingestion of soil, dermal contact of soils, and contaminated vegetable ingestion. To determine the contaminant concentrations in soil, samples were collected from areas adjacent to the Tehran oil refinery, located in Shahr-e-Ray city, Iran. Analyses of the samples indicated that the average of PAHs concentration in the soil samples were greater than clean-up level guidelines. Cancer risk of contaminants due to ingestion of cultivated vegetables that are sold in Tehran markets was significant in comparison with other exposure pathways. Moreover, the total cancer risk for 5th percentile, 95 upper confidence limit, and 95th percentile concentration of contaminants were 5.69E-04, 8.78E-02, and 2.13E-01, classifying the site as having a significant cancer risk potential. Furthermore, non-carcinogenic health risk analyses for the contaminants demonstrated hazard index of less than 1. Remediation of the soils is highly recommended to eliminate the potential cancer risks and prevent the contamination of the food chain for approximately 10 million Tehran residents.     

Effect of cotton cultivar on performance of Aphis gossypii (Homoptera: Aphididae) in Iran

Cotton aphids, Aphis gossypii Glover (Homoptera: Aphididae), obtained from cotton, Gossypium hirsutum L., fields in the Gorgan region of northern Iran, were colonized on 'Varamin' cotton plants in a growth chamber. The development, survivorship, and life table parameters of the cotton aphid were evaluated at 27.5 +/- 1 degrees C, 65 +/- 10% RH, and aphotoperiod of 14:10 (L:D) h of artificial light on five commonly growing cotton cultivars: Varamin, 'Sealand' (relatively resistant cultivar), 'Bakhtegan', 'Sahel' (both relatively susceptible cultivars), and 'Siokra' [resistant to Bemisia tabaci (Gennadius)]. The developmental times of immature stages ranged from 4.6 d on Bakhtegan and Varamin to 6.3 d on Sealand, whereas the immature survival was 97.5 to 65% on Sahel and Siokra, respectively. On average, there were 28.7, 28.3, 23.5, 20.1, and 16.8 nymphs produced per female on Sahel, Bakhtegan, Varamin, Sealand, and Siokra, respectively. The intrinsic rate of natural increase (r(m)) for cotton aphids on Sahel was the highest, whereas the values for r(m) varied from 0.284 (nymphs per female per d) on Siokra to 0.368 on Sahel. Jackknife estimates of generation times (T), net reproductive rate (R(0)), doubling time (DT), and finite rate of increase (lambda) on these cultivars were as follows: 9.79-10.84 d for T, 9.23-23.81 nymphs per female for R(0), 2.17-3.19 d for DT, and 1.28-1.38 nymphs per female per d for lambda. Cotton aphid performance was at its highest on Sahel and lowest on Siokra.     

Effect of periplasmic expression of recombinant mouse interleukin-4 on hydrogen peroxide concentration and catalase activity in Escherichia coli

Oxidative stress occurs as a result of imbalance between generation and detoxification of reactive oxygen species (ROS). This kind of stress was rarely discussed in connection with foreign protein production in Escherichia coli. Relation between cytoplasmic recombinant protein expression with H₂O₂ concentration and catalase activity variation was already reported. The periplasmic space of E. coli has different oxidative environment in relative to cytoplasm and there are some benefits in periplasmic expression of recombinant proteins. In this study, hydrogen peroxide concentration and catalase activity following periplasmic expression of mouse IL-4 were measured in E. coli. After construction of pET2mIL4 plasmid, the expression of recombinant mouse interleukin-4 (mIL-4) was confirmed. Then, the H₂O₂ concentration and catalase activity variation in the cells were studied in exponential and stationary phases at various ODs and were compared to those of wild type cells and empty vector transformed cells. It was revealed that empty vector introduction and periplasmic recombinant protein expression increased significantly the H₂O₂ concentration of the cells. However, the H₂O₂ concentration in mIL-4 expressing cells was significantly higher than its concentration in empty vector transformed cells, demonstrating more effects of recombinant mIL-4 expression on H₂O₂ elevation. Likewise, although catalase activity was reduced in foreign DNA introduced cells, it was more lowered following expression of recombinant proteins. Correlation between H₂O₂ concentration elevation and catalase activity reduction with cell growth depletion is also demonstrated. It was also found that recombinant protein expression results in cell size increase.     

Effect of exposure to extremely low electro-magnetic field during prenatal period on mice spleen

Total body weight of newborns, the volume of spleen, and the number of megakaryocytes decreased following the exposure to ELF-MF (6 x 10(-3) T and 50 Hz) at 1-5, 6-10, 11-15, and 16-20 days of pregnancy of mice. The complete period of gestation was sensitive to ELF-MF exposure; the initial days were more prone to exposure. The results suggest that the use of ELF-MF producing instruments should be limited during gestation.