Mechanism of Long Noncoding RNA HOTAIR in Nucleus Pulposus Cell Autophagy and Apoptosis in Intervertebral Disc Degeneration

Figure 3

HOTAIR acting as a ceRNA competitively binds to miR-148a to upregulate PTEN mRNA expression. (a) The lncRNA HOTAIR location was predicted through the database (https://www.csbio.sjtu.edu.cn/bioinf/lncLocator/). (b) The fluorescence localization of HOTAIR in NP cells was detected using RNA-FISH. (c) The expression of HOTAIR was detected using RT-qPCR after nuclear and cytoplasmic fractionation assay. (d) The binding sites of HOTAIR and miR-148a were predicted through the bioinformatics website (https://starbase.sysu.edu.cn/agoClipRNA.php?source=lncRNA). (e) Dual-luciferase reporter gene assay was used to verify the binding relationship between HOTAIR and miR-148a. (f) RNA pull-down was used to verify the binding relationship between HOTAIR and miR-148a in NP cells. (g) RT-qPCR was used to detect miR-148a expression after HOTAIR knockdown. (h) Bioinformatics website (http://starbase.sysu.edu.cn/agoClipRNA.php?source=lncRNA) was used to predict the binding sites of miR-148a and PTEN. (i) Dual-luciferase reporter gene assay was used to verify the binding relationship between miR-148a and PTEN. (j) RT-qPCR was used to detect the expression of miR-148a. (k) RT-qPCR was used to detect PTEN mRNA expression. The experiment was repeated three times, and the data were expressed as mean ± standard deviation. Data in panels E/G/I were analyzed using t-test; data in panels C/F/J/K were analyzed using one-way ANOVA, followed by Tukey’s multiple comparisons test. ; .

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