Methods: Embryonic day 18 (E-18) rat hippocampus neurons were cultured with poly-L-lysine coated glass coverslips. Following optimisation, KA (0.5 μM), a chemoconvulsant agent, was administered at three different time-points (30, 60 and 90 min) to induce seizure in rat hippocampal neuronal cell culture. We examined cell viability, neurite outgrowth density and immunoreactivity of the hippocampus neuron culture by measuring brain derived neurotrophic factor (BDNF), γ-amino butyric acid A (GABAA) subunit α-1 (GABRA1), tyrosine receptor kinase B (TrkB), and inositol trisphosphate receptor (IP3R/IP3) levels.
Results: The results revealed significantly decreased and increased immunoreactivity changes in TrkB (a BDNF receptor) and IP3R, respectively, at 60 min time point.
Conclusion: The current findings suggest that TrkB and IP3 could have a neuroprotective role which could be a potential pharmacological target for anti-epilepsy drugs.
MATERIALS AND METHODS: MCF-7 cells were plated at a density of 15105 cells/well in 6-well plates. After 24h, cells were treated with a series of concentrations of rapamycin while only adding DMEM medium with PEG for the control regiment and grown at 37oC, 5% CO2 and 95% air for 72h. Trypan blue was used to determine the cell viability and proliferation. Untreated and rapamycin-treated MCF-7 cells were also examined for morphological changes with an inverted-phase contrast microscope. Alteration in cell morphology was ascertained, along with a stage in the cell cycle and proliferation. In addition, cytotoxicity testing was performed using normal mouse breast mammary pads.
RESULTS: Our results clearly showed that rapamycin exhibited inhibitory activity on MCF-7 cell lines. The IC50 value of rapamycin on the MCF-7 cells was determined as 0.4μg/ml (p<0.05). Direct observation by inverted microscopy demonstrated that the MCF-7 cells treated with rapamycin showed characteristic features of apoptosis including cell shrinkage, vascularization and autophagy. Cells underwent early apoptosis up to 24% after 72h. Analysis of the cell cycle showed an increase in the G0G1 phase cell population and a corresponding decrease in the S and G2M phase populations, from 81.5% to 91.3% and 17.3% to 7.9%, respectively.
CONCLUSIONS: This study demonstrated that rapamycin may potentially act as an anti-cancer agent via the inhibition of growth with some morphological changes of the MCF-7 cancer cells, arrest cell cycle progression at G0/G1 phase and induction of apoptosis in late stage of apoptosis. Further studies are needed to further characterize the mode of action of rapamycin as an anti-cancer agent.
(: MVD) is the quantification method of various aspects of tumor vasculature that indicates angiogenic activity. This study aims to analyze the correlation between MVD to the expression of VEGFRs on breast cancer tissue.
Materials and Method: A total of 60 N-methyl-N-nitrosourea (MNU)-induced breast carcinomas in rats were suppressed by using antiangiogenic drugs. The rats were then sacrificed, and the tumor was fixed in 10% formalin, paraffin embedded, and immunohistochemistry stained using VEGFRs and CD34.
Result: One-way ANOVA test showed a significant difference in all markers that have been used (P < 0.05) on MNU-breast tumor treated with rapamycin (M= 90.1664, SD= 7.4487), PF4 (M= 93.7946, SD= 7.1303) and rapamycin + PF4 (M= 93.6990, SD= 1.8432). We obtained a significant reduction of MVD count on breast carcinoma for rapamycin group (M= 25.6786, SD= 9.7075) and rapamycin + PF4 group (M= 30.5250, SD= 13.6928) while PF4 group (M=47.7985, SD=4.8892) showed slightly increase compared to control (M= 45.1875, SD= 4.4786). There was a moderately strong, positive correlation between angiogenic markers; Flt-1 (r= 0.544, n=60, P < 0.005) and Flt-4 (r= 0.555, n= 60, P < 0.005) while Flk-1 (r= 0.797, n= 60, P < 0.005) showed a strong, positive correlation with MVD.
Conclusion: MVD was strongly correlated to the VEGFRs expression on breast carcinoma.
METHODS: Thirty-two female Sprague Dawley rats at age 21-days old were administered intraperitoneally with N-Methyl-N-Nitroso Urea (NMU), dosed at 70mg/kg body weight. The rats were divided into 4 groups; Group 1 (Control, n=8), Group 2 (Sirolimus, n=8), Group 3 (Sunitinib, n=8) and Group 4 (Sirolimus+Sunitinib, n=8), being treated twice when the tumor reached the size of 14.5±0.5 mm and subsequently sacrificed after 5 days. The protein expressions of ER, PgR and HER2/neu of the tumor tissues were evaluated by using immunohistochemistry analysis.
RESULTS: Treatment with sirolimus alone lowered expressions of ER and PgR of breast cancer and reduced tumor size. There was no significant difference of ER and PgR expressions between control and sunitinib treated tumor. Sunitinib treated tumors reduce in diameter after the first treatment, however the diameter increases after the second treatment. Histologically, sunitinib treated tumor did not show any aggressive invasive carcinoma of no special type (NST) histological subtypes. In addition, all NMU-induced tumors are HER2/neu-negative scoring.
CONCLUSION: Sirolimus is neither synergistic nor additive with sunitinib for breast cancer treatment.
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