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Titanium with nanotopography attenuates the osteoclast-induced disruption of osteoblast differentiation by regulating histone methylation

ABSTRACT: We report the application of RNA sequencing to evaluate the effect of titanium (Ti) with nanotopography, compared to Ti control, on the whole transcriptome of osteoblastic cells in isolated cultures grown on these surfaces and to evaluate the effect of Ti with nanotopography, compared to Ti control, on the whole transcriptome of osteoblastic cells, grown on these surfaces, in co-culture with osteoclastic cells. MC3T3-E1 osteoblastic cells were plated at the density of 1x10^4 cells/well on the Ti discs with nanotopography and Ti control, in 24-well plates in osteogenic medium and kept isolated for 5 days to allow cell adhesion. Osteoclast cells from the RAW 264.7 lineage were cultured on osteoclastogenic medium on ThinCert™ cell culture inserts (Greiner Bio-One), with 0.4 μm pore, at the density of 1x104 cells/membrane, and was also be kept isolated for 5 days to allow cell adhesion. At the end of this period theThinCert™ cell culture inserts (Greiner Bio-One), containing the osteoclast cells were positioned on the osteoblastic cell cultures, thereby establishing indirect co-culture. The controls were osteoblastic cells of the MC3T3-E1 lineage grown on the Ti discs in the absence of osteoclast cells of the RAW 264.7 lineage, i.e., not maintained in co-culture.

ORGANISM(S): Mus musculus

PROVIDER: GSE189164 | GEO | 2021/11/23

REPOSITORIES: GEO

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Project description:The biological effects of 1α,25-dihydroxyvitamin D3 (1,25(OH)2D3) on osteoblast differentiation and function differ significantly depending upon the cellular state of maturation. To explore this phenomenon mechanistically, we examined the impact of 1,25(OH)2D3 on the transcriptomes of both pre-osteoblastic (POBs) and differentiated osteoblastic (OBs) MC3T3-E1 cells, and assessed localization of the vitamin D receptor (VDR) at sites of action on a genome-scale using ChIP-seq analysis. We observed that the 1,25(OH)2D3-induced transcriptomes of POBs and OBs were quantitatively and qualitatively different, supporting not only the altered biology observed but the potential for a change in VDR interaction at the genome as well. This idea was confirmed through discovery that VDR cistromes in POBs and OBs were also strikingly different. Depletion of VDR binding sites in OBs, due in part to reduced VDR expression, was the likely cause of the loss of VDR-target gene interaction. Continued novel regulation by 1,25(OH)2D3, however, suggested that factors in addition to the VDR might also be involved. Accordingly, we show that transcriptomic modifications are also accompanied by changes in genome binding of the master osteoblast regulator RUNX2 and the chromatin remodeler C/EBPβ. Importantly, genome occupancy was also highlighted by the presence of epigenetic enhancer signatures which were selectively changed in response to both differentiation and 1,25(OH)2D3. The impact of VDR, RUNX2, and C/EBPβ on osteoblast differentiation is exemplified by their actions at the Runx2 and Sp7 gene loci. We conclude that each of these mechanisms may contribute to the diverse actions of 1,25(OH)2D3 on differentiating osteoblasts. RNA was isolated and applied to gene expression microarrays in undifferentiated MC3T3-E1 cells as well as post 15 day osteogenic differentiation MC3T3-E1 cells, which were treated for 24 hours with 10-7M 1,25(OH)2D3. For the vehicle matched samples, please refer to study GSE41955. The samples were completed in biological triplicate.

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Titanium with nanotopography attenuates the osteoclast-induced disruption of osteoblast differentiation by regulating histone methylation

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