Fig. 4

Identification and validation of GACAT2 as a key lncRNA associated with the cementoblastic differentiation of PDLSCs. a–h Identification of differentially expressed lncRNAs between the PDLSCs incubated in noninflammatory environments and the PDLSCs incubated in inflammatory environments. The cells were incubated in medium with the cementoblastic inducer EMD alone (EMD) or EMD plus inflammatory cytokines (Infla-EMD). a Volcano plots of the differentially expressed (fold change >1.5 and adjusted P < 0.05) lncRNAs in the PDLSCs between the EMD and Infla-EMD groups. b Heatmap of the top 8 lncRNAs with upregulated expression and the top 8 lncRNAs with downregulated expression in the PDLSCs of the EMD group compared with those in the Infla-EMD group. Six lncRNAs, AL390957.1 (c), LINC01638 (d), AC010247.2 (e), AC096773.1 (f), LINC01133 (g), and GACAT2 (h), were selected for qRT-PCR verification according to their RNA length (<2 000 nt), intensity (>100), relation (without overlapping with coding transcripts) and database source (in the GENCODE or RefSeq public databases), and 10 other lncRNAs were excluded. c and d No significant changes in AL390957.1 and LINC01638 expression were observed in the cells incubated in the noninflammatory or inflammatory environments. e and f AC010247.2 and AC096773.1 expression was significantly increased in an inflammatory environment. g and h LINC01133 and GACAT2 expression was significantly decreased in an inflammatory environment. i The inhibition of AC096773.1 (si-AC096773.1), but not AC010247.2 (si-AC010247.2), increased the relative cementoblastic differentiation-related gene expression levels of BSP, CAP and CEMP-1 in the PDLSCs incubated in an inflammatory environment (qRT-PCR). j The inhibition of LINC01133 (si-LINC01133) or GACAT2 (si-GACAT2) decreased the relative cementoblastic differentiation-related gene expression levels of BSP, CAP and CEMP-1 in the PDLSCs incubated in a noninflammatory environment (qRT-PCR); the most significant change was observed in response to GACAT2 inhibition. The data are shown as the mean ± SD for n from 3 to 6; *P < 0.05, **P < 0.01 and ***P < 0.001 indicate significant differences between the indicated columns