Cell Journal (Yakhteh)
Year:2008 | Volume:10 | Issue:2 (38)|Papers Count:9

Objective: It has been suggested that low frequency stimulation (LFS) exerts its inhibitory effect on epileptogenesis through adenosine receptors activation. In the present study, effect of different LFS frequencies on A1 and A2A receptors gene expression was investigated in perforant path kindled seizures. Materials and Methods: Animals were kindled by perforant path stimulation. Afterdischarges were recorded from the dentate gyrus. LFS (0.5, 1 and 5 Hz) was applied at the end of each kindling stimulation. On the 7th day, A1 and A2A receptors gene expression were evaluated in the dentate gyrus.Results: Application of different LFS frequencies retarded the kindling acquisition. Also, it decreased the afterdischarge durations and behavioural seizure stages 4 and 5 significantly. LFS application prevented the kindling induced decrease in the A1 receptor gene expression. On the other hand, LFS attenuated the level of A2A receptor gene expression in the dentate gyrus. LFS had the most effect at the frequency of 5 Hz.Conclusion: It may be suggested that antiepileptogenic effects of LFS is mediated somehow through changes in the gene expression of adenosine A1 (which has inhibitory effects) and A2A (which has excitatory effects) receptors. These effects might be somehow frequency dependent.

Objective: Evaluation of Bax encoding plasmid for increasing efficacy of DNA vaccine plasmid encoding gB of Herpes Simplex Virus type 1.Materials and Methods: We compared three different dosages of Bax encoding plasmid (pcbax) including 10, 25 and 50 mg of plasmid DNA. They were co-injected ineradermally with glycoprotein B (gB) of herpes simplex virus (HSV)-1 encoded plasmid (pcgB) in C57BL/6 mice to elicit immune responses to protect against lethal HSV-1 challenge. Immune responses to the antigen were assessed by lymphocyte proliferative responses and cytokine (INF-g and IL-4) release assays.Results: The study demonstrates that the mice immunized with 25 mg pcbax together with pcgB have more efficient protection than the mice immunized with 10 and 50 mg of pcbax and pcgB. Analysing of cell-mediated responses show that the mice immunized with 25mg pcbax and pcgB induce stronger lymphocyte proliferative responses and higher levels of INF-g and IL-4 compared to the mice are received 10 and 50 mg of pcbax and pcgB.Conclusion: The data show that co-immunization with 25 mg of pcbax and pcgB increase immune responses compared to 10 and 50 mg of pcbax and pcgB. This can be considered a promising approach for development an efficient DNA vaccine against HSV-1 or other pathogens.

Objective: The exfoliated human deciduous tooth (SHED) contain multipotent stem cells that identified to be a population of highly proliferative and clonogenic .These cells are capable of  differentiating into a variety of cell types including neural cells, adipocytes, and odontoblasts. Material and Methods: Normal exfoliated human deciduous incisors collected from six- to nine-years-old children. The pulp was separated from the crown and digested with collagenase .Single cell solutions were cultivated in a-MEM supplemented with ES-FCS. After two to three days, the cells reached confluency and were trypsinized and cultured for further passages. The passage-4 cells were analyzed with CD34, CD45, CD105, CD166, CD31, CD90 and CD146 markers that indicated these cells had a mesenchymal stem cell (MSC) identity. We examined the cells for Alkaline Phosphatase activity to investigate the mesenchymal (stromal) nature.Finally, the cells were differentiated into the osteoblastic and adipocytic lineages in different subcultures and analysed by RT-PCR and different staining protocols.Results: Viable cells growing out of the explants showed elongated shapes in clusters. These cells showed alkaline phosphatase activity. Flow cytometry results revealed high expression of pluripotent stem cell markers .In some area of the osteoinductive cultures nodule-like structures were observed that showed red mineralizing area upon staining with Alizarin Red. In adipogenic cultures lipid vesicles appeared after five weeks of induction with Oil Red. Conclusion: This study show that pulp contains cells with high plasticity and proliferation capacity and can be easily isolated without any serious intervention.

Objective: Breast cancer is the most common cancer among women in the world. Early diagnosis of this cancer is a key element for its treatment. One of the approaches for diagnosis of breast cancer is detection of its tumour-associated markers. Hence, Her2 has been the main focus of the researches in the field.Materials and Methods: For diagnosis of Her2 overexpression, monoclonal antibodies (mAb) reacting against Her2 were produced in this study. For this purpose, two peptides from extracellular domain of Her2 were selected and the mAbs reacting against them were produced by hybrodoma technology. Reactivity of these antibodies were then evaluated in different immunological assays including ELISA, Immunoflurescence (IF), western blot (WB) and immunoprecipitation (IP(.Results: Total of 5 clones was produced from two separate fusions, and antibody is typing revealed that all clones were IgM. These mAbs showed appropriate reactivates in the following assays: ELISA, immunofluresence by staining of breast cancer cell line (SKBR3), WB and IP by detecting the 185 KD band of Her2. Conclusion: In conclusion, it seems that the mAbs are useful diagnostic tools for detection of Her2 expression in patients with breast cancer.

Objective: The aim of this study was to investigate the effect of laser assisted hatching on the development and quality of vitrified-warmed 4-cell stage mouse embryos.Materials and Methods: The vitrified-warmed 4-cell mouse embryos were divided into two groups: control group (without laser assisted hatching) and experiment group (with laser assisted hatching). All embryos in both groups were cultured in sequential media containing G1TMver3 and G2TMver3. Afterward, all expanded blastocysts were randomly selected and stained with differential (for cellularity) and TUNEL (for cell death) methods.Results: On day 1(24hrs) of culture, the difference between the control and the experimental groups was insignificant in the rate of blastocyst formation. But on day 2 (48hrs) of culture, 87.61% of embryos in the experimental group reached the blastocyst stage. This rate did not increase significantly as compared to the control group (78.14%). Finally on day 3 (72 hrs), the rate of blastocyst formation reached 94.40% and 81.75%, respectively, in both the experimental and control groups. The difference between these two groups were significant (p<0.05). The number of blastomeres and apoptotic cells were similar in the experimental and control groups.Conclusion: The laser assisted hatching has no decreasing effect on cellularity, but it has increasing effect on incidence of cell death. In addition, the assisted hatching significantly increases the blastocyst formation rate of intact vitrified-warmed 4-cell stage mouse embryos.

Objective: Hydrostatic pressure is crucial component of cell environment and fundamental physical quantity; also it is the main factor of both cell integrity and function. Pressure variation disorder, beyond physiological limits, may lead to pathological states. In this study, we examined the effect of hydrostatic pressure on apoptosis induction, viability, morphology, adhesion potency to substrate and migration of differentiated PC-12 cells. Materials and Methods: PC-12 as a neuronal cell line maintained in RPMI 1640 culture medium supplemented with 10% fetal bovine serum. Staurosporine was used for differentiating of mitotic PC-12 cells to post mitotic and differentiated neuronal cells. Exclusion Dye was used for viability assay, total neurite length of each cell as well as morphometry. TUNEL staining was also performed for apoptosis detection, adhesion potency of cells to substrate and evaluation of cell migration. Results: Hydrostatic pressure, over physiological limits, induced apoptosis in differentiated PC-12 cells. It changed cell viability gradually and reduction happened significantly after 24 hours (p<0.05). In compare to the control group, hydrostatic pressure reduced total neurite length, adhesion potency to substrate and migration of cells in the examined group (p<0.05).Conclusion: Hydrostatic pressure induced apoptosis in differentiated PC-12 cells as a result of inappropriate interaction between cells and substrate. We propose that apoptosis in differentiated PC-12 cells may be an anoikis causing to lose the attachment to the substrate.  

Objective: Ischemic preconditioning (IPC) is an endogenous phenomenon that can induce ischemic tolerance (IT) in variety of organs such as brain. In this study, we examined the intermittent and prolonged dose of normobaric hyperoxia (NBHO) in neurologic deficit scores, NF-kB activity, and TNF-a converting enzyme (TACE) expression.  Materials and Methods: The rats were divided to four main groups.  First two groups were exposed to HO divided in prolonged (24hrs; PrHO) and intermittent (4h×6 days; InHO) groups. Second two groups acted as control groups and they were exposed to 21% oxygen with the following condition: in the same chamber (room air, RA), continuously (24hrs; Pr RA) and discontinuously (4h×6days; InRA). Each group was subdivided to three subgroups. After 24hrs, first subgroup was subjected to 60mins MCAO followed by 24hrs of reperfusion. Then, IT, induced by InHO and PrHO, was measured by neurologic deficit scores and infarct volume. Second and third subgroups were respectively called sham-operated subgroup and intact subgroup designed to assess the effect of HO on NF-kB activity and TNF-α converting enzyme expression.    Results: Our findings indicate that InHO and PrHO are involved in the induction of IT. Pretreatment with InHO and PrHO reduce neurologic deficit scores and infarct volume significantly. InHO and PrHO increase NF-kB activity and TNF-a converting enzyme expression with different degrees. Also, InHO with ischemia increase NF-kB activity and TNF-a converting enzyme expression significantly.  Conclusion: Although further studies are needed to clarify the mechanisms of ischemic tolerance, InHO and PrHO seem partly to exert their effects via increasing NF-kB activity and up regulation of TNF-a converting enzyme.

Objective: To study the structure and distribution of microtubules in embryos derived from young, old and reconstructed oocytes.Materials and Methods: Embryos obtained from old (50 embryos), young (50 embryos) and reconstructed oocytes (10 embryos) were studied by immunocytochemistry. The microtubule structures of the embryos were studied by using fluroscent microscopy with FITC-PI filter and polyclonal antibody against alfa tubulin.Results: The spindle structure of MII young oocyte and the obtained embryos were normal with the suitable condensation. There was no contact between chromosome and spindle in old Oocytes as well as the obtained embryos; in addition, the spindle was extended in old group. In reconstructed embryos, thin and scattered filaments were observed.Conclusion: This study reveals that the arrangement of microtubules in reconstructed embryos was caused by repeating of injection and oocyte manipulation. Also, interactions between karyoblast, cytoplasm and microtubuls may not be suitable. This may be caused by low fertilization in these oocytes.