Journal of Sciences, Islamic Republic of Iran
Volume:14 Issue: 4, Autumn 2003

The purpose of the present study is to investigate the cytotoxicity/apoptotic effect of 2-chloro-2′-deoxyadenosine, cladribine, (2-CdA) in the human breast cancer cell line, MDA-MB468 (estrogen receptor negative, ER−). MTT [3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H tetrazolium bromide] assay, annexin V-Fluorescein/PI and Hoechst 33258 staining were used to detect cytotoxicity and cell apoptosis. The activation of caspase-3 and -9 was assayed using caspase activation assay kits. Gel electrophoresis was performed to detect DNA fragmentation. Treatment of MDA-MB468 cells with different concentrations of 2-CdA (50, 100 and 500 µM) resulted in a significant increase in the cell death. Annexin V-Fluorescein/PI and Hoechst 33258 staining revealed that the cell death was mainly of apoptotic type. DNA laddering profile was also obtained in the treated MDA-MB468 cells using DNA fragmentation analysis. A significant (p<0.05) increase in the activity of caspase-3 and -9 was observed. Pre-treatment of the cells with kinase inhibitor, 5′-amino-5′-deoxyadenosine inhibited the cytotoxicity effect of 2-CdA. This suggests that intracellular phosphorylation activation reaction plays a key role in the 2-CdA-induced apoptosis. In conclusion, this study showed that high dose of cladribine has an apoptotic effect on ER−MDA-MB468 breast cancer cells and its intracellular phosphorylation is necessary for cytotoxicity.

In order to study the periplasmic expression of human growth hormone (hGH) in Escherichia coli, the related cDNA was inserted in two expression plasmids carrying pelB signal peptide, one with lac bacterial promoter and the other with a bacteriophage T7-based promoter. The recombinant plasmids were moved to TG1 and BL21 strains of E. coli, respectively. To induce the expression systems, IPTG and its natural analog, lactose, were examined. Results show the over-expression of recombinant hGH (rhGH) in the T7-based system which is much higher than that of the lac-regulated system. In both systems, a fraction of the hGH, produced by recombinant bacteria, remains in the cytoplasm as pre-protein and the rest is transferred to the periplasmic space as mature protein. Any decline in the expression level did not lead to a complete processing and transport of mature hGH to the periplasmic space of the bacteria. It is suggested that, for an efficient expression of rhGH in the periplasmic spaceof E. coli, a combined approach including application of a suitable signal peptide, solubility of the over-expressed protein in cytoplasm in addition to the optimization of the bacterial growth and inducing conditions should be considered.

Interferon is a protein secreted by eucaryotic cells following stimulation by viruses, bacteria, and many other immunogenes. Recent medical studies indicate that interferons have effective role in the treatment of virus infections, immunodeficiency and certain types of cancer such as hairy cell leukaemia (HCL). The aim of the present study is to apply yeast strain for secreting human IFNα2b following the use of yeast mating α factor signal peptide. On the other hand, cloning of IFNα2b gene without signal peptide and with non homologous signal peptide from Aspergillus was carried out as controls for comparison. First, human IFNα2b gene was amplified using mating α factor signal peptide codons and the first 20 bases of IFNα2b gene as a forward primer. This amplicon (700 bp) was cloned in pYZ4 vector and after using suitable restriction sites, the cleaved fragment was cloned in plasmid pYES2 as expression vector and named pPMSHβ2. Since the original construct of IFNα2b contained one of the Aspergillus signal peptides and flanked with Sal І and Eco RI sites, this gene (900 bp) was isolated and cloned in plasmid pET24d as an intermediate after using suitable sites. This insert was cleaved then cloned in pYES2 vector as pPMSHαB2. Also to construct IFNα2b without signal peptide, primers were designed to exclude the Aspergillus signal peptide from the original IFNα2b gene and the amplicon (500 bp) was cloned in plasmid pET23a then in pUC18 and finally in plasmid pYES2 and named as pPMSH2. Saccharomyces cerevisiae (INVSC1) was transformed with these three mentioned constructs and for interferon gene expression, galactose was used as an inducer. Primary results of western blotting analysis showed that IFNα2b gene with α factor signal peptide was produced inside the pPMSHβ2 transformed yeast cells. The Use of α Factor signal peptide is convenient for expressing the IFNα2b gene in yeast. Studies on growth condition optimization and IFN secretion are under consideration and application.

Glucose-6-Phosphate Dehydrogenase (G6PD) is a cytosolic enzyme which its main function is to produce NADPH in the red blood cells by controlling the step from Glucose-6-Phosphate to 6-Phospho gluconate in the pentose phosphate pathway. G6PD deficiency is the most common X-chromosome linked hereditary enzymopathy in the world, that result in reduced enzyme activity and more than 125 different mutations causing it have been identified. In the present study we analyzed peripheral blood samples of 103 unrelated patients with G6PD deficiency in GilanProvince in the north of Iran. DNA samples from these subjects were analyzed for the presence of certain known G6PD mutations by the appropriate restriction enzyme digestion of fragments, amplified by PCR. In this way, it revealed that Mediterranean mutation (C563T, Ser 188 Phe) is predominant in this area and 89 samples out of 103 (86.4%) have this mutation, indicating a higher prevalence of this mutation, in Gilan in Comparison to Mazandaran (66.2%). Among other patients, 10 samples (9.71%) have Chatham mutation (G1003A, Ala 335 Thr), but none of the samples was found to have Cosenza mutation (G1367, Arg 459 Pro).

The carrier facilitated transport of lithium picrate was studied using a series of non-cyclic polyethers containing different end groups and chain lengths through Bulk Liquid Membrane (BLM) and Supported Liquid Membrane (SLM) systems. The various membrane supports used are viz. PTFE, cellulose nitrate, and dialysis membrane and onion membrane. The amount of Li+ transported depends upon the structure of the ionophores and membrane supports used. The amount of Li+ ion transported follows the trend V >> IV ≈ III ≈ II >I >VI > VII > VIII = IX in the BLMs studies. Diethylene glycol dibenzoate proved to be the most efficient carrier for the transport of the lithium ion through liquid membrane systems.

Effects of temperature and percentage of organic modifier were studied on retention and selectivity in RP-HPLC using solvation parameter model. The system constants were determined by multiple linear regression analysis from experimental values in the retention factor for a group of different solutes with known descriptors by computer using the program SPSS/PC. The experimental results showed that the variation of percentages of organic modifier (5-20% V/V CH3CN) changes cavity formation (m = +3.55 to +2.53), dipole-dipole inter actions (s = +0.62 to −0.64) and hydrogen-bond acidity (b = −2.41 to −3.27) terms. The variation of temperature (5-45°C) also changes cavity formation (m = +3.21 to +2.56) and dipole-dipole interactions (s = +0.34 to −0.22) terms.

The lower to middle Miocene Chenar granitoid stock, a part of Central Iranian volcanic belt, is intruded into Eocene volcano-sedimentary complex in northwestern Kerman in south Central Iran. The Chenar granitoid stock consists mainly of coarse to medium grain granodiorite and monzogranite with subordinate tonalite and syenogranite. Alkali granites as aplitic veins and quartz dioritic dykes frequently cut through the granitoid stock. Enclaves of various types and sizes (andesitic and microgranular mafic, few meters to several centimeters wide) also occur near the margin. The granitoid rocks represent convergent margin magmas enriched in large ionic lithophile elements (LILE) such as Rb, Ba, K, Ce and depleted in high field strength elements (HFSE) such as Y, Nb and Zr. The REE patterns have relatively smooth LREE enriched chondrite normalized REE profiles with (La/Yb)n between 4.28 to 14.66. A lack of Eu anomalies along with a continuous increase in the slope of LREE profile from tonalite to syenogranite exhibit a relatively low fractionated chondrite normalized pattern [(La/Yb)n] and also involvement of plagioclase in the melting processes. Small negative Eu anomaly in alkali granite is an indication of more differentiated rock type [(La/Yb)n=42.78] in Chenar granitoid stock. Geochemical data, various trace element discriminant diagrams, common igneous microgranitoid enclaves, and ocean ridge granite normalized multi-element diagrams indicate that the Chenar granitoid stock has characteristics of metaluminous to slightly peraluminous, calc-alkaline, I-type granite of a volcanic arc settings and is formed in an active continental margin environment. In this model, a NE-dipping subduction zone of Neotethys could have accounted for the arc volcanism of the Central Iran where felsic, Andean-type magmatic arc volcanics and plutonics were formed. As a result of subduction mafic arc magmas with significant fluids were produced following the dehydration of oceanic crust and partial melting of mantle wedge which in turn provoke partial melting of considerably metasomatised and enriched subcontinental lithosphere, leading to generation of siliceous magma. Its low-pressure crystal fractionation gave rise to generation of hydrous calc-alkaline magmas, represented in part by Chenar granitoid stock.

In many computer vision applications, segmenting and extraction of moving objects in video sequences is an essential task. Background subtraction, by which each input image is subtracted from the reference image, has often been used for this purpose. In this paper, we offer a novel background-subtraction technique for real-time dynamic background generation using color images that are taken from a static camera. The new algorithm, which is based on ‘temporal median filter with exponentially weighted moving average (EWMA) filtering’, is presented that effectively implements a temporal mode operation. The proposed method has the advantage that the parameters of the algorithm are computed automatically. In addition, the new method could start its operation for a sequence of images in which moving objects are included. The efficiency and robustness of the new algorithm is confirmed by the results obtained on a number of outdoor image sequences.

The linear adiabatic pulsation-periods of Mira variable stars have been derived. Approximately 2701[1]models were calculated for M = 0.7 MΘ to 2 MΘ stars with radii from 180 RΘ to 340 RΘ and luminosities from 2800 LΘ to 10,000 LΘ. The chemical composition of all models is (X,Z) = (0.7,0.02). From the result of this study, linear relations on Luminosity-Period-Mass relationship and luminosity-period-radius-mass relationship have been derived for the fundamental mode. The relation has been tested on a recent data set of Mira stars in the LMC. 1 Models of various computational input are available through the author.