مجله سلول و بافت
سال هفتم شماره 4 (زمستان 1395)

Medicinal plants, as one of the important sources for the treatment of diseases, have been used from thousands years ago. These plants produce a major and diverse group of secondary metabolites. Secondary metabolites are the compounds derived from the primary metabolites (metabolites associated with the plant nutrition and survival) essential to sustain plant life. Manipulation of the cell culture media by the abiotic elicitors is one of the important strategies for the induction and production of valuable metabolites, because the potential of natural producing of these compounds is very limited. Elicitors induce secondary metabolites by the activation of defense mechanisms and genes through the increasing ionic flow across the plasma membrane, reactive oxygen production, structural changes in the cell wall as well as phytoalexin production. In this paper, various aspects of increasing secondary metabolites production have been reviewed in the plant cell culture systems using abiotic elicitors.

Aim: In this study, the effect of newborn rat brain extract to induce neuronal differentiation in P19 embryonal carcinoma (EC) stem cells was investigated.
Material and Newborn rat brain extract was collected in sterile condition and the concentration of its total protein was determined. Then, the amount of cell viability was determined after treatment with brain extract. In order to differentiation, the embryoid bodies resulted from cells suspension culture were exposed to culture medium containing 3% serum that supplemented by the extract, for 7-14 days. Specific staining and real-time PCR methods were applied to evaluate neural differentiation.
Cresyl violet staining confirmed neuronal morphology of the differentiated cells. The gene expression of neural specific was confirmed by real–time PCR. Developing brain extract, which contains numerous neorotrophic factors, could induce expression of synaptophysin (presynaptic membrane protein) and nestin (intermediate filament of stem cells in the neural tube) genes. In addition, the expression of transcription factor Nanog, an important factor for pluripotency and self-renewal of stem cells, decreased under the influence of newborn rat brain extract.
The results of the present study indicated that newborn rat brain extract can induce neuronal phenotype as well as neuronal specific gene expression in P19 stem cells. This study suggests the potential use of combined newborn rat brain extract and stem cell therapy to improvement of deficits in neurodegenerative diseases.

Aim: Effects of two nanoparticles were investigated on the expression of STR, DAT and D4H genes in periwinkle.
Material and For this purpose, concentrations of 0.5, 0.75 and 1 mg per liter of cobalt and zinc oxide nanoparticles were used. Sampling was carried out at 0, 8, 24 and 48 hours after treatment.
Gene expression analysis was performed by the SQ-RT-PCR method. In general, both elicitors influenced gene expression. But, the effects of zinc oxide nanoparticles on the gene expression were more pronounced than cobalt oxide nanoparticles. The highest expression of STR and D4H genes were occurred in 0.5 mg per liter of zinc oxide nanoparticles after 8 hours treatment, while in the case of DAT gene, it occurred in concentration of 1 mg per liter of this nanoparticle. Moreover, Cobalt oxide nanoparticles in most of the studied concentrations and time intervals caused decreases in gene expression.
Both nanoparticles had significant effects on gene expression of vinblastine and vincristine pathways.

Aim: This investigation was conducted to determine the amount of isolated anthocyanins from seed coats of Phaseolus vulgaris L. under magnetic field treatment and also assess their antiproliferative activity of the extracts.
Material and Dry and soaked red bean seeds were subjected to different electromagnetic field treatments in four group. The seeds of groups 1(dry) and 2 (soaked) were treated for 45 min., while in groups 3 and 4 the dry and soaked seeds treated two times with 120 min. interval with electromagnetic field. Then, they were cultured and grown, and their produced seeds were collected. Seed coats from these beans were separated and coat extract was prepared. Two groups of anthocyanins, cyanidin and pelargonidin, were identified and measured by high performance liquid chromatography (HPLC). The antiproliferative activity of the extract was determined by MTT assay against ovarian cancer cells (CP2780A).
The bean coats of the control and the treated plants contain a high content of cyanidin and pelargonidin. Among the extracts, the samples of group 1 had the highest amount of cyanidin, while seeds of group 2 had the highest amount of pelargonidin. The results of the antiproliferative activity revealed that the methanol extract of the treated red beans under 4 mT were highly antiproliferative activity of the cell (78.74 to 84.44 %). Moreover, the seeds of group 2 showed the highest antiproliferative activity (IC50: 72.63 ± 2.2).
The electromagnetic field treatment is the most effective in increasing the amounts of anthocyanins in red beans, and that the red bean coat can be used as a natural source of nutrition and drug supplement with antiproliferative properties.

Aim: Iran is considered as an arid and semi-arid area of the world and also it is one of the main countries producing cucumbers; so study the effects of drought on this plant and also its antioxidant system changes are important.
Material and In this study, cucumber cv. Isfahani plants were grown under greenhouse condition. In order to study the effect of changes in protein and antioxidant systems, cucumber plants at 3 to 5 leaf stage were treated with 4 levels of drought stress (100, 75, 50 and 25 percent) and their responses to applied stresses were investigated. The amounts of electrolyte leakage, lipid peroxidation, protein, proline, total antioxidant, catalase and guaiacol peroxidase activities, α-tocopherol and hydrogen peroxide were measured.
The results study showed that the contents of electrolyte leakage, lipid peroxidation, proline, total antioxidant, catalase, guaiacol peroxidase, α-tocopherol and hydrogen peroxide increased under drought stresses as compared to the control plants. However, protein index decreased under drought stresses in comparison with controls.
The obtained findings showed that in this cultivar of cucumber tolerance to drought was related to enzymatic and non-enzymatic antioxidant activities and also proline accumulation.

Aim: The aim of this study was to investigate the effects of different treatments of growth regulators on the amount of shooting, rooting, callus production and plant regeneration of hairless catmint (Nepeta nuda), in vitro condition.
Material and The seeds of this plant were cultured in MS/2, and then the plantlets were sub-cultured. The obtained samples were cultured in different levels of BA and Kinetin, either alone or in combination with IBA. Leaves of explants were used for callus induction with BA and NAA treatments in both medium (light and dark). Also, for regeneration, the same calluses were used with BA and NAA treatments.
The results of this study showed that, in proliferation part, the highest fresh weight (2315.9 mg) and shoot number (15.22 per each bush) of each plantlet were obtained in 1 mgL-1 BA and 0.25 mgL-1 IBA. Moreover, the BA treatment with 2 mgL-1 concentration cause to produce the longest branches (50.5 cm) and the highest number of nodes (53.6 per plantlet) in each plantlet.
The highest length, number and percentage of root were obtained at MS/2 medium and 0.5 mgL-1 IBA. In callus formation part, the highest fresh weight of callus was related to application of 1 mgL-1 BA with 0.5 mgL-1 NAA. In addition, the highest percentage of callus formation was observed in dark medium and no significant differences were observed between the growth regulator treatments. Furthermore, the highest percentage of regeneration of callus related to both growth regulators treatments included 1 mgL-1 BA with 0.2 mgL-1 NAA (83.2 %) and 1 mgL-1 BA with 0.5 mgL-1 NAA (81.66 %) with no significant differences between them.
In total, the most useful growth regulator compound for micro-propagation of hairless catmint, which is endangered extinction species, were 1 mgL-1 BA with 0.5 mgL-1 IBA and NAA.

Aim: In this century, the production of recombinant drugs such as growth hormone has increased. Different problems existed in the expressions of cytoplasmic and Periplasmic types of growth hormone. Therefore, finding a way to optimize expression is very necessary. In this study, we optimized expression of growth hormone in the form of solution state by trx-tag method. This method increase protein expression (Periplasmic problem) and also prevents formation of inclusion body (problem cytoplasmic).
Material and Gene synthesis and gene cassette was done in pET 32a expression vector. Gene cassette contains trx tag for protein solubilization, His tag for purification and enterokinase for separate rHgh from previous tags.
After cloning the gene in vector, its expression was confirmed by Western blot technique. The results showed that the expression of fusion protein was done well.
the obtained finding proved that protein can be soluble by Trx-tag and increased its expression levels. Moreover, better results can be achieved in the fermentation and downstream processing.

Aim: Papaver somniferum remains the sole commercial source for several pharmaceutical benzophenanthridine alkaloids from benzylisoquinoline branch alkaloids, which includes the narcotic analgesics codeine and the semi-synthetic drugs such as; oxycodone, buprenorphine and naltrexone. Although, most of the biosynthetic pathways genes of this alkaloid have been identified, the post-transcriptional regulation of these alkaloids pathway has not been completely determined. Virus induced gene silencing (VIGS) is a method for fast functional genomics, and in this study, this technique was used to investigate the silencing one of the most important genes in this alkaloid pathway.
In the current research VIGS technique was used for systematic reduction in the level of genes expression involves in benzylisoquinoline alkaloids pathway. For silencing of codeinone reductase gene, pTRV vector was used for cloning. The specific silencing of COR gene was evaluated in 2-3 weeks old leaves of Papaver somniferum using agro-infiltration method.
Result of cloning was confirmed by using of different molecular methods such as enzyme digestion and also PCR. PCR technique was used for confirmation of transgenic plants in some transformed plants. Using of semi-quantitative PCR and real-time PCR showed that the level of reduction in transcription of COR gene was about 89 percent.
The obtained results confirmed that by application of RNA interference method, the level of COR gene expression was significantly reduced compared with control.

Aim: The aim of the present study was to investigate the effects of royal jelly (RJ) and vitamin C (vit C) against PHZ-induced nephropathy in mice.
Material and Adult male mice were randomly divided into eight eight-membered groups. The PHZ was administered to four groups of mice at a dose of 60 mg/kg per 48 hours intraperitoneally for 35 days. Three of these groups received vit C (250 mg/kg per day) intraperitoneally, RJ (100 mg/kg per day) orally and vit C with same doses, four hours before PHZ administration, respectively. A vehicle-treated control group and vit C, RJ and vit C control groups were also included. Serum and renal tissue samples were collected 24 hours after the last treatment and subjected to biochemical and histological examinations, respectively. The data were analyzed by one-way analysis of variance followed by Duncan test for post hoc comparisons.
The PHZ treatment caused a significant (P The Vit C and RJ can minimize PHZ-induced nephropathy in mouse through oxidative reactions inhibition.

Aim: This study was performed to investigate if silymarin can prevent the adverse effects of sodium arsenite on ram sperm DNA and nuclear integrity.
Material and Epididymal sperm obtained from Farahani’s ram (Ovis aries) was swim up and divided into five groups: 1. Sperm at 0 hour, 2. sperm at 180 minutes (control), 3. sperm treated with sodium arsenite (10μM) for 180 minutes, 4. sperm treated with silymarin (20 μM) sodium arsenite (10 μM) for 180 minutes and 5. sperm treated with silymarin (20 μM) for 180 minutes. Ram´s sperm DNA integrity was assessed by SCD (Sperm Chromatine Dispersion) test to study DNA fragmentation and Acridine orange staining was used to estimate DNA denaturation (double-strand DNA versus single-strand DNA). To evaluate sperm nuclear integrity, Diff-quick staining was used and sperm nuclear diameter was measured.
DNA fragmentation percent and nuclear diameter of the spermatozoa were significantly increased and decreased, respectively, in sodium arsenite group compared to the control samples. While this toxicant had no effect on sperm DNA denaturation. In silymarin sodium arsenite group, silymarin was able to significantly ameliorate the adverse effects of sodium arsenite on these sperm parameters compared to sodium arsenite group.
Silymarin as a potent antioxidant could compensate the adverse effects of sodium arsenite on DNA fragmentation and nuclear diameter of ram sperm.