Long noncoding RNA expression profile of mouse cementoblasts under compressive force

Abstract

Objectives: To investigate the long noncoding RNA (lncRNA) expression profile of cementoblasts under compressive force.

Materials and methods: Mouse cementoblasts were exposed to compression (1.5 g/cm²) for 8 hours. RNA sequencing (RNA-seq) was performed to compare the transcriptomes of the compressed and control cells. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to validate five of the differentially expressed lncRNAs of interest. Gene Ontology (GO) functional annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were also performed.

Results: A total of 70 lncRNAs and 521 mRNAs were differentially regulated in cementoblasts subjected to compressive loading. Among the differentially expressed lncRNAs, 57 were upregulated and 13 downregulated. The expression levels of the five selected lncRNAs (Prkcz2, Hklos, Trp53cor1, Gdap10, and Ak312-ps) were validated by qRT-PCR and consistent with the RNA-seq results. GO functional annotation demonstrated upregulation of genes associated with cellular response to hypoxia and apoptotic processes during compressive loading. KEGG analysis identified the crucial pathways involving the hypoxia-inducing factor-1α, forkhead box O, and mammalian target of rapamycin signaling pathways.

Conclusions: Mechanical compression changes the lncRNA expression profile of cementoblasts, providing important references for further investigation into the role and regulation of lncRNAs in compressed cementoblasts and root resorption during orthodontic treatment.

Keywords: Cementoblasts; Compressive stress; Long noncoding RNA; RNA-seq; Root resorption.

Figure 1

Cell viability and cementoblast differentiation of OCCM-30 under mechanical pressure. (A) Diagram showing the application of compressive force. (B) Relative cell growth with or without compression. (C) DAPI staining (left) and relative capase-3/7 activity (right) of OCCM-30 with or without compression. Scale bar, 50 μm. (D) ALP staining images (left) and ALP activity (right) of these cells. (E) Relative mRNA expression of Runx2, Ocn, Ptpla, and Bsp in these cells. Results are presented as mean ± SD (**P < .01).

Figure 2

The differentially expressed lncRNAs and mRNAs under compressive stress. (A) Chromosomal read distributions of control cells (left) and compression loaded cells (right). (B) Volcano plot of differentially expressed transcripts (lncRNAs and mRNAs) in the control and force group. Red points: upregulated mRNAs or lncRNAs; blue points: downregulated mRNAs or lncRNAs.

Figure 3

Differentially expressed lncRNAs (Prkcz2, Hklos, Trp53cor1, Gdap10, and Ak312-ps) validated by qRT-PCR. Results are presented as mean ± SD (*P < .05; **P < .01).

Figure 4

The top 20 enrichment in the GO terms (A) and the KEGG pathway analysis (B) of differentially expressed genes.

Figure 5

KEGG maps of differentially expressed genes in HIF-1, FoxO, and mTOR signaling pathways. Red boxes: upregulated genes; green boxes: downregulated genes.