مجله سلول و بافت
سال ششم شماره 4 (زمستان 1394)

Aim: The purpose of the present research project was evaluation of the effect of silicon on the severity of peroxidase genes expression and some morphological traits in both resistant and susceptible barley lines under drought stress.
Material and Total soluble protein content, photosynthetic pigments, and total RNA were extracted from the leaves affected by various treatments. Target gene was evaluated by Semi-quantitative RT-PCR analysis using synthesized cDNA. Both the peroxidase enzyme activity by Chance method and proline with Bates method were also evaluated under drought stress treatment. Accordingly, the experiment was analyzed in a factorial test based on the completely randomized design with three treatments of control, drought and silicon-drought (sodium silicate 2 mg / 1 Kg soil) with three replications in a greenhouse.
Silicon application caused to increase the amount of total soluble protein and photosynthetic pigments in both lines under drought stress. Semi-quantitative RT-PCR analysis of treatments observed significant differences. Maximum of peroxidase gene expression was observed in the silicon-drought treatment. Antioxidant enzyme activities were at the highest level as shown by gene expression pattern of polyacrylamide gel analysis for treated silicon samples. A further increase was exhibited for proline accumulation caused by silicon application rather than such accumulation in stress treatment compared to control.
Silicon reduces oxidative damage induced by reactive oxygen species and with the increase in antioxidant enzyme activities caused to protect plants against environmental stresses.

Aim: Chalcone synthase (CHS) is a key enzyme in the flavonoid biosynthesis pathway. This enzyme catalyses the first step of flavonoid biosynthesis pathway. The purpose of this study was to identify chalcone synthase gene in Calotropis procera.
Material and The genomic DNA was extracted from fresh leaves and used as template in PCR. Primers were designed according to the conserved regions of this gene in other plants. The PCR product was purified and sequenced. Multiple sequence alignment and sequence homology analyses were carried out with DNAMAN software using CHS sequences retrieved from Genbank. A neighbor-joining phylogenetic tree based on CpCHS and other CHS sequences were constructed using MEGA6 software.
PCR results indicated the existence of this gene and amplification of 570 nucleotides fragment that belong to exon2 of CHS gene. This gene was denominated as CpCHS and deposited at the GenBank accession KM878672. Comparison analysis based on nucleotide sequence alignment revealed that CpCHS shared a high degree of similarity to CHS from Arabidopsis thaliana (59%), Nicotiana tabacum (63%), Olea europaea (63%) and Petunia X hybrida (64%). Maximum similarity was observed between CpCHS and CHS from Catharanthus roseus (67%). Phylogenetic analysis showed that CpCHS was grouped into a branch with Catharanthus roseus.
These results suggest that CpCHS may play a similar role as other CHS in regulation of flavonoids biosynthesis pathway. Identification of this gene is an effective step which may lead to increase in the accumulation of useful medicinal extract in this plant.

Aim: In this study, we explored the effect of epigenetic modifications on planarian regeneration process by treating with small molecule epigenetic modifiers
Material and Planaria worms were cut into 3 sections including head, trunk, and tail regions and treated with 6 small molecules with epigenetic modification effects. All samples were evaluated morphologically for 20 days to monitor the effects on regeneration process.
Morphological analysis of samples treated with selected six small molecules with different epigenetic modification functions revealed that treating with two of them including Sodium butyrate and Valporic acid will result in delayed head, trunk, and tail fragments regeneration, and delayed eye formation in trunk and tail fragments, respectively. In addition, samples treated with other four small molecules including BIX01294, RG108, SAHA, and Tranylcypromine with other epigenetic modification functions, regenerated normally.
These results indicating that different epigenetic levels and activity of histone deacetylases play a key role in planarian regeneration and inhibition of histone deacetylases activity will result in abnormal regeneration. However, further work is needed to determine the exact mechanism of histone deacetylases activity effect on planarian regeneration process. Finally, we hope to understand more about the underlying mechanism of planarian and vertebrate’s regeneration inhibition and activation using these findings.

Aim: The aim of the current study was to isolate microglial cells from neonatal rat brain, activate the cells using lipopolysaccharide (LPS) and examine the effect of the inflammatory factors that they express on dopaminergic (DAergic) neurons.
mixed glial cells were isolated from 1-3 day rat neonatal brains and then microglial cells were extracted from them. Upon treatment of the cells with LPS, their conditioned media (CM) were collected. DAergic SH-SY5Y cell line was seeded in 96-well plates and fed with the collected CM. The rate of viability and apoptosis of the neuronal cells was examined using MTT and apoptosis tests and the data were analyzed statistically.
Ten to thirteen days after isolation, mixed glial cells were composed of three types: astroglia, oligodendrocytes and microglia. Microglia were floating while semi-attached to the surface. Twenty four hours post-isolation, these cells were transferred to and cultured in separate dishes where they formed spindled and ramified phenotypes. At this stage, the cells were treated with LPS to be activated. This activation was demonstrated by morphological changes from spindle-like form to amoeboid form, detection of increase in NO expression using Griess test and induction of inflammatory iNOS and TNF-α gene expression. Also teatment of SH-SY5Y cell line with conditioned medium of activated microglia led to both apoptotic and necrotic cell death.
LPS-mediated activation of microglia results in overexpression of neuroinflammatory markers. The overproduction of these markers results in both apoptotic and necrotic cell death amongst DAergic neurons via neuroinflammation.

Aim: In this study, the effect of salinity stress on quantitative gene expression of sucrose synthase1, relative water content and the uptake ratio of sodium to potassium in the leaves of wheat were evaluated.
Material and Wheat seedlings treated with 100 and 200 mM sodium chloride and at times zero, 6, 12, 24 and 36 hours after treatment relative water content, the uptake ratio of sodium to potassium in the leaves and quantitative gene expression of Sus1 by qRT-PCR method were studied.
Results showed a significant decrease in relative water content, increase in the uptake ratio of sodium to potassium by leaves and increase in the relative expression of sucrose synthase1 gene at treated plants compared to control group.
Generally, increase in the expression of sucrose synthase 1 gene could be one of the possible mechanisms in the process of tolerance to salinity stress.

Aim: The goal of the present research was evaluation of the effects of industrial area air pollution of Arak Aluminum Company on Robinia pseudoacacia (acacia) and tree-of-heaven anatomical changes.
Material and Information from the Environmental Protection Agency of Markazi province, Aluminum park as the infected area and Haftad-Ghole in 35 kilometers of Arak as the area clean were used. Leaf samples of Robinia and Alianthus were harvested in August 2013 simultaneously. Density and length of trichomes, stomatal density and opening, length of the palisade cells and diameter of the spongy cells of leaf were measured. Data analysis were performed by SPSS 16 and Excel software and means comparison by T-Test.
Under air pollution, trichome density in abaxial epidermis and trichome length in both surfaces of epidermis of acacia leaf increased significantly. Also in tree-of-heaven leaf, trichome density decreased and trichome length increased on abaxial epidermis significantly. Stomatal density on abaxial epidermis of acasia and tree-of-heaven leaves exposed to air pollution decreased and increased, respectively. Stomatal opening of Robinia and Ailanthus leaves had no significant effect. Length of the palisade cells of leaf in infected area decreased in acacia and increased in tree-of-heaven as compared with clean area, diameter of the spongy cells had no significant change in acacia leaf but decreased in tree-of-heaven leaf.
Results showed the anatomical changes in both plants for further compromise with regional air pollution.

Aim: This study aimed to investigate the effect of Bisphenol­ A on the cell viability, morphologic changes and induction of apoptosis in bone marrow mesenchymal stem cells of an adult rat.
Material and The bone marrow mesenchymal stem cells were extracted using flashing-out method. At the end of the third passage, cells were divided into 11 groups of control and experimental. Experimental cells were treated with the different doses of Bisphenol­ A (1, 5, 10, 50, 100, 250, 500, 1000, 2000 and 4000 nM) for four periods of 5, 10, 15 and 21 days in the osteogenic media containing 10% of fetal bovine serum. The cell viability, DNA damage, expression of genes and the morphologic changes of the cells were then investigated during the procedure of osteogenesis. The study data was analyzed using one-way ANOVA and T-Test setting the significant P value at P Within Bisphenol­ A treated cells, the mean viability, the mean of nuclei diameter and the expression of anti-apoptotic Bcl-2 gene of the mesenchymal stem cells treated with Bisphenol­ A significantly decreased (P Our data suggest that BPA decreases cell viability and induces apoptosis in mesenchymal stem cells, in a dose dependant manner.

Aim: The aim of this study is to compare sperm parameters (concentration, motility and morphology), DNA damage, percentage and intensity of ROS (Reactive Oxygen Species) and relationship between these parameters in the fertile and infertile men with varicocele.
Material and In this study, the semen samples from 83 individuals with varicocele and 32 fertile men were studied. Sperm parameters according WHO (World Health Organization) guidelines, protamine deficiency (chromomycin A3 staining), sperm DNA damage (TUNEL assay), percentage and intensity of ROS (DCFH-DA staining) were assessed in fertile and infertile men with varicocele.
The results of this study showed that sperm concentration, percentage of motility and normal morphology were significantly lower in individuals with varicocele compared to fertile individuals. In addition, percentage of sperm DNA damage, protamine deficiency and ROS positive were significantly higher in infertile men with varicocele.
The increase of stress oxidative and testicular temperature in infertile men with varicocele, lead to decrease the quality of sperm parameters as well as negative effects on sperm chromatin integrity. Therefore the process of spermatogenesis and the rate of fertility were affected in these individuals.

Aim: The aim of this study was to assess side effects of clinical dose of busulfan on testis and epididymal sperm of adult male mouse.
Material and Male adult NMRI mice (25-35 g) were divided into two groups of sixteen each. Control and busulfan treated group were administered 100 μL DMSO (Dimethyl Sulfoxide) and 3.2 mg/kg busulfan for 4 days, respectively. The animals were sacrificed 35 days after starting treatment. The histological change on germinal epithelium, sperm quality parameters (count and normal morphology), viability and DNA fragmentation on sperm were analyzed by light microscopy, CASA (Computer–aided Sperm Analyzer), MTT (3-[4, 5-Dimethylthiazol-2-yl]-2, 5 Diphenyltetrazolium Bromide) and TUNEL assays, respectively.
Busulfan administration significantly decreased parameters of sperm (count and normal morphology) and increased germinal epithelium destruction. The head, mid piece and tail abnormalities of treated group were increased significantly versus control. The lower levels of MTT in treated group were significant compared with control group. Higher levels of TUNEL positive cells that were found in treated groups demonstrated the increasing of DNA fragmentation in sperms following busulfan treatment.
The results showed that clinical dose of busufan induces genotoxic, pre-apoptotic and cytotoxic effects on spermatogenesis. Also, busulfan can increase cytotoxicity on testis tissue of adult male mouse.

Aim: The aim of the present study was to investigate the effect of lead concentrations on the growth, antocyanin, chlorophyll, carotenoid, total soluble sugars, reducing sugars and free amino acids of harmal (Peganum harmala L.) populations.
Material and Plants of two different Peganum harmala populations (metallicolous and non metallicolous) were grown in hydroponic condition. Then plants were exposed to 0, 5, 10, 25 and 50 ppm Pb for 14 days. Finally, parameters were measured and calculated by spectrophotometer.
Results showed that the increase of Pb concentrations in the nutrient solution reduced root length and there were significant difference between two populations. Chlorophylls and carotenoid pigments were generally decreased by Pb concentrations, especially under the highest Pb treatment (50 ppm) in both populations. Anthocyanin, total soluble sugars and free amino acids were increased in both populations.
The higher amounts of anthocyanin, total soluble sugars and free amino acids in metallicolous population reflecting the higher degree of tolerance to Pb in this population.