The in vitro effects of pure and street methamphetamine on the proliferation and cell cycle of mouse brain endothelial (bend5) cells
Mafunda, Patrick Siyambulela
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The blood-brain barrier (BBB) is an interface between the brain parenchyma and the circulating system. This barrier plays a vital role in protecting the CNS by restricting free paracellular diffusion of molecules from the systemic circulation. Methamphetamine (MA) is a highly addictive psychostimulant and has demonstrated neurotoxic properties as well as the ability to compromise the BBB. MA exposure is strongly linked with increased oxidative stress which can result in a decrease in the integrity of the BBB.The aim of this study was to investigate in vitro effects of pure and street MA "tik" on DNA proliferation and cell cycles in mouse brain endothelial (bEnd5) cells. Trypan blue was used to determine effects of MA (0.0001M-1mM) on cell viability and % cell growth. The Cell Titer Glo® luminescent assay and nonradioactive analogue, 5-bromo-2'-deoxyuridine (BrdU) was used to detect ATP and DNA levels, respectively. Cell cycles (propidium iodide incorporation) were analysed using flow cytometry. Statistical analysis was performed using Wilcoxin Rank Sum Test in which P<0.05 was denoted as significant. Results of this study showed that: 1. Viability of bEnd5 cells exposed to all selected concentrations of MA were unaffected when compared to controls (P>0.05). 2. % Cell growth was suppressed by MA exposure at 96hrs in comparison to that of controls (P≤0.03). 3. Cells exposed to MA had significant higher ATP concentrations than control cells at 96hrs (P ≤.0.03) 4. DNA synthesis was markedly suppressed in cells exposed to pure MA and street MA sample 4 (P≤0.03), while was similar and higher in cells exposed to street MA sample 1 (P=0.39), and street MA sample 2 and 3 (P≤0.04), respectively at 96hrs. 5. bEnd5 cell were arrested between 72 and 96hrs at the G1-S phase. In conclusion, this study demonstrated pure and illicit samples of MA obtained from forensic police did not affect the viability of bEnd5 cells, however resulted in the significant suppression of their cell numbers. This growth inhibition may be due to MA-induced cell cycle arrest at the G1-S phase. The study also showed that compounds found in the samples of street MA produced results significantly different to that of pure MA.