Abstract
Purpose
The long non-coding RNA GAS8 antisense RNA 1 (lncRNA GAS8-AS1) is a tumor suppressor in papillary thyroid cancer (PTC), but the mechanisms underlying how GAS8-AS1 regulates PTC biology remain unclear. Here, we evaluated the molecular function of GAS8-AS1 in regulating autophagy in PTC cell lines.
Methods
GAS8-AS1 was overexpressed and knocked down in PTC cell lines by transfecting with expression plasmids or short interfering RNAs (siRNAs). Cell proliferation was evaluated using the Cell Counting Kit-8 (CCK-8). qRT-PCR and western blot were used to determine changes in expression of autophagy-related genes. Autophagy was evaluated by immunofluorescence and transmission electron microscopy.
Results
Relative GAS8-AS1 expression was lower in the PTC cell lines, TPC1 and BCPAP, compared to a normal thyroid cell line. Overexpression of GAS8-AS1 inhibited proliferation, significantly increased the ratio of LC3-II/LC3-I, and reduced p62 expression, whereas GAS8-AS1 knockdown demonstrated opposite effects. In GAS8-AS1 overexpressing cell lines, LC3 immunofluorescence staining demonstrated increased punctate aggregates of LC3 staining, and transmission electron microscopy revealed increased numbers of autophagosomes. Autophagy-related gene 5 (ATG5) was markedly upregulated by GAS8-AS1 overexpression and downregulated by GAS8-AS1 knockdown. Finally, silencing of ATG5 attenuated autophagy activation and rescued the inhibition of cell proliferation caused by GAS8-AS1.
Conclusions
In PTC cell lines, GAS8-AS1 inhibited proliferation, activated autophagy, and increased ATG5 expression. Downregulation of ATG5 reversed GAS8-AS1-mediated activation of autophagy leading to cell death, revealing a novel mechanism of the GAS8-AS1-ATG5 axis in PTC cell lines. This provided a new experimental basis to explore the effects of lncRNA on autophagy in the treatment of thyroid cancer.
from #ORL-AlexandrosSfakianakis via ola Kala on Inoreader http://ift.tt/2AStupW
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