Introduction:The mechanism of putative cytotoxicity of 4-[3-(cyclopentyloxy)-4-methoxyphenyl]-2-pyrrolidone (rolipram), a specific phosphodiesterase-4 (PDE4) inhibitor, on glioblastoma multiforme (GBM) is almost unknown. This study aimed to investigate the role of protein kinase B (Akt) pathway in the cytotoxic effect of rolipram on human GBM U87 MG cell line and Tumor-Initiating Cells (TICs) isolated from patient’s GBM specimen.Methods:TICs were characterized by using flow cytometry and quantitative real-time PCR. The cells were treated with rolipram at inhibitory concentration of 50% (IC50) in the presence or absence of SC79 (4μg/mL), a specific AKT activator, for 48 hours. The cell viability and apoptosis were measured by MTT assay and TUNEL staining, respectively. The relative expression of Phospho-Akt (Ser473), matrix metalloproteinase 2 (MMP2), and vascular endothelial growth factor A (VEGFA) were detected using Western blotting.Results:The findings showed that rolipram could suppress cell viability in both U87MG and TICs, dose-dependently. Interestingly, the rolipram-induced cytotoxicity was significantly reduced in the presence of SC79. Nevertheless, in rolipram-treated cells, the pretreatment with SC79 significantly led to increase in U87 MG cells and TICs apoptosis and decrease in viability of U87 MG cells but not TICs relative to corresponding control. In U87 MG and TICs, rolipram-induced reduction of Phospho-Akt (Ser473) and MMP2 levels were significantly suppressed by SC79.Conclusion:There is a cell type-specific mechanism of anti-proliferative action of rolipram on GBM cells. The reduction of intracellular level of MMP2 but not VEGFA by rolipram is conducted through the inhibition of Akt signal. Rolipram-induced apoptosis is mediated via Akt dependent/independent mechanisms.
Cell therapy is an important strategy for the treatment of incurable diseases including those that occur in the central nervous system (CNS). Among different strategies, the method of delivering or transplantation of cells into the brain has shown significant effects on regeneration. In this study, a new protocol has been developed for the transplantation of adipose tissue-derived stem cells into the brain through cerebrospinal fluid (CSF) in rat models. For this purpose, a wide range of ages (7-30 days old) of male neonates of Wistar rats was used. Moreover, human adipose tissue was obtained from a superficial layer of abdomen through liposuction surgery. The size of the inserted part of needle to access middle cranial fossa and subarachnoid space in animals with an average weight of 10-80 g was determined. In addition, to confirm the entrance of needle into the subarachnoid space, CSF was aspirated slowly and then injection was done within two minutes. The findings showed the presence of transplanted human adipose-derived stem cells (hADSC) in the cerebellum and basal ganglia following three days and also after two months that confirmed the entrance of transplanted cells into the cerebrospinal fluid and migration of them into the brain tissue. All the animals survived after the transplantation process, with the lowest side effects compared to the available conventional methods. It can be concluded that the cells could be efficiently transplanted into CSF through subarachnoid space by injection via superior orbital fissure with a minimally invasive technique.
Kernicterus is a neurological syndrome associated with indirect bilirubin accumulation and damages to the basal ganglia, cerebellum and brain stem nuclei particularly the cochlear nucleus. To mimic haemolysis in a rat model such that it was similar to what is observed in a preterm human, we injected phenylhydrazine in 7-day-old rats to induce haemolysis and then infused sulfisoxazole into the same rats at day 9 to block bilirubin binding sites in the albumin. We have investigated the effectiveness of human adiposity-derived stem cells as a therapeutic paradigm for perinatal neuronal repair in a kernicterus animal model. The level of total bilirubin, indirect bilirubin, brain bilirubin and brain iron was significantly increased in the modelling group. There was a significant decreased in all severity levels of the auditory brainstem response test in the two modelling group. Akinesia, bradykinesia and slip were significantly declined in the experience group. Apoptosis in basal ganglia and cerebellum were significantly decreased in the stem cell-treated group in comparison to the vehicle group. All severity levels of the auditory brainstem response tests were significantly decreased in 2-month-old rats. Transplantation results in the substantial alleviation of walking impairment, apoptosis and auditory dysfunction. This study provides important information for the development of therapeutic strategies using human adiposity-derived stem cells in prenatal brain damage to reduce potential sensori motor deficit.
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