ABSTRACT
SUMMARY OBJECTIVE: The phosphoinositide 3-kinase/protein kinase AKT/mammalian target of rapamycin signaling pathway is essential for proper cellular metabolism and cell growth. However, aberrant activation of this pathway has been linked to the progression and metastasis of breast cancer. Recently, the role of long non-coding RNAs in interfering with the cell signaling pathways involved in cell growth and metabolism has been identified. HOX antisense intergenic RNA is an long non-coding RNA whose abnormal expression has been associated with development, therapy resistance, and metastasis of breast cancer. The purpose of this study was to investigate whether the long non-coding RNA HOX antisense intergenic RNA is linked to the phosphoinositide 3-kinase/protein kinase AKT/mammalian target of rapamycin signaling pathway in breast cancer cells. METHODS: HOX antisense intergenic RNA was silenced in the breast cancer cell line MCF-7 using siRNAs. Subsequently, the gene expression level of HOX antisense intergenic RNA, PI3K, AKT, and mTOR was assessed using real-time RT-PCR. Also, the 3-(4,5-dimethylthiazol- 2-yl)-2,5-diphenyl-tetrazolium bromide) assay was used to analyze cell proliferation. RESULTS: The results revealed that HOX antisense intergenic RNA knockdown can downregulate the expression of PI3K, AKT, and mTOR RNAs compared to negative control in MCF-7 cells. In addition, the proliferation of breast cancer cells was significantly reduced following the HOX antisense intergenic RNA silencing. CONCLUSION: This study may introduce HOX antisense intergenic RNA as a molecule involved in the upregulation of the phosphoinositide 3-kinase/protein kinase AKT/mammalian target of rapamycin signaling pathway in breast cancer cells that may contribute to breast cancer cell proliferation.
ABSTRACT
Objective:To observe the effect of LncRNA HOTAIR on the expression of HIF-1α in HaCat cell under low oxygen condition, and to explore the role of LncRNA HOTAIR in the pathogenesis and development of keloid.Methods:From Jan. 2018 to Dec. 2018 in Chinese Academy of Medical Science, recombinant plasmids were designed and constructed by specific shRNA-HOTAIR. After transfected HaCat cells with sh-LncRNA HOTAIR, RT-PCR was used to detect the expression level of HOTAIR. HaCat cells were cultured in different conditions and divided into four groups: group A cultured under normoxia condition, the other three groups cultured under hypoxia condition. Group C was transfected with sh-control before hypoxic culture, while group D were transfected with sh-LncRNA HOTAIR. After 24 hours′ culture, dual luciferase reporter gene assay was used to verify the relationship of LncRNA HOTAIR and HIF-1α. Expression of HIF-1α in four groups of HaCat cells were determined by Western blot analysis.Results:Recombinant plasmids of shRNA-HOTAIR were successfully constructed, and the HOTAIR expression was significantly decreased. The relative luciferase activity was 0.94±0.30 in group A, 20.39±1.15 in group B, 18.09±0.80 in group C and 3.04±1.15 in group D. The relative luciferase activity of group B was higher than those of group A ( t=29.03, P<0.05), and the difference was statistically significant ( P<0.05). According to the results of Western blotting, group B (1.19±0.07) and group D (1.15±0.06) had higher expression of HIF-1α than group A (0.56±0.29) and group C (0.37±0.38). Down-regulation of HOTAIR significantly inhibited the protein level of HIF-1α. Conclusions:LncRNA HOTAIR plays a positive role in upregulation of HIF-1α in HaCat cells under hypoxia condition. Thus LncRNA HOTAIR may take part in the pathogenesis and development of keloid through HIF-1α pathway.