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Al-Rubai, Abdal-jabbar
Languages: English
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Generally, most of the in vitro tests used in neurotoxicology are limited to transformed cell lines which are derived from rodent or human. For an in vitro test to have high rate of predictability of neurotoxicity and teratogenicity it should undergo the important processes of embryological development, such as cell proliferation, cell migration, and differentiation. Human neural stem cells have been proposed for this purpose, which have the ability to divide, differentiate, and migrate. In this study, it was found that double coating of laminin with either poly D lysine or poly L lysine was most suitable for growing human neural stem cells rather than coating with a single extracellular molecule. Several chemicals and drugs were then chosen to assess the utility of neural stem cells as an assay for neurotoxicity: methyl mercury and lead acetate; four anti-epileptics drugs (sodium valproate, phenytoin, carbamazepine, and phenobarbitone); anti-oxidants (folic acid and melatonin). These anti-oxidants were tested alone and when added to sodium valproate and to phenytoin (which are well known in their teratogenicity), and other drugs (lithium, diazepam, and amitriptyline), which are weak teratogens. To assess the effects of these molecules on human neural stem cells cell survival, total cellular protein, neuronal process length, neurosphere sizes, migration distance, Glial Fibrillary Acidic Protein, and tubulin III protein expression were measured. \ud The study shows that methyl mercury caused significant reduction in most of the end points from the dose of 1µM and it led to significant increase in Glial Fibrillary Acidic Protein expression (which is a sign of reactive gliosis). Lead acetate led to a significant reduction in cell migration 48hours after treatment with 10µM. In the case of the anti-epileptics, sodium valproate appeared to reduce neurosphere size significantly from the dose of 500µM and decrease migration distance significantly 48hours after treatment with 1000µM. Moreover, phenytoin treatment resulted in significant reduction in neurosphere sizes from the dose of 25µM and reduced cell migration significantly from the dose of 50µM. However, the other anti-epileptics (carbamazepine and phenobarbitone) revealed their effect only at high doses which are above their therapeutic range. On the other hand, adding the anti-oxidants (Folic acid or Melatonin) to sodium valproate or phenytoin had to some extent beneficial effects, by making their toxic effect appear at doses which were higher than when used alone. Regarding the other drugs (lithium, diazepam, and amitriptyline), it seems that their toxic effect appeared only at doses which are higher than the therapeutic range.\ud Therefore, it can be concluded that human neural stem cells are a sensitive model in detecting the neurotoxicity of methyl mercury and lead acetate at low doses and can predict the neurotoxicity of sodium valproate and phenytoin at their therapeutic doses.
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    • 1 Mouse monoclonal Anti-Tubulin 50Kda III antibody
    • 2 Rabbit polyclonal Anti-GFAP 55Kda & antibody 48Kda
    • 3 Mouse monoclonal Anti-GAPDH 37Kda antibody 2. Primary and secondary antibodies in Immunocytochemistry and Immunohistochemistry: Primary Antibodies Dilution Sources
    • 1 Mouse monoclonal Anti-Tubulin III 1:500 Abcam antibody
    • 2 Rabbit polyclonal Anti-GFAP antibody 1:800 Abcam
    • 3 Rabbit polyclonal Anti-SOX2 antibody 1:2000 Abcam
    • 4 Mouse monoclonal Anti-Nestin 1:1000 BD Antibody Biosciences
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