Human bone marrow-derived mesenchymal stem cells (hBMSCs) are important for tissue regeneration but their epigenetic regulation is not well understood. Here we investigate the ability of a non-nucleoside DNA methylation inhibitor, RG108 to induce epigenetic changes at both global and gene-specific levels in order to enhance mesenchymal cell markers, in hBMSCs. hBMSCs were treated with complete culture medium, 50 μM RG108 and DMSO for three days and subjected to viability and apoptosis assays, global and gene-specific methylation/hydroxymethylation, transcript levels’ analysis of epigenetic machinery enzymes and multipotency markers, protein activities of DNMTs and TETs, immunofluorescence staining and western blot analysis for NANOG and OCT4 and flow cytometry for CD105. The RG108, when used at 50 μM, did not affect the viability, apoptosis and proliferation rates of hBMSCs or hydroxymethylation global levels while leading to 75% decrease in DNMTs activity and 42% loss of global DNA methylation levels. In addition, DNMT1 was significantly downregulated while TET1 was upregulated, potentially contributing to the substantial loss of methylation observed. Most importantly, the mesenchymal cell markers CD105, NANOG and OCT4 were upregulated being NANOG and OCT4 epigenetically modulated by RG108, at their gene promoters. We propose that RG108 could be used for epigenetic modulation, promoting epigenetic activation of NANOG and OCT4, without affecting the viability of hBMSCs. DMSO can be considered a modulator of epigenetic machinery enzymes, although with milder effect compared to RG108.

Chromatin conformation, DNA methylation pattern, transcriptional profile, and non-coding RNAs (ncRNAs) interactions constitute an epigenetic pattern that influences the cellular phenotypic commitment and impacts the clinical outcomes in regenerative therapies. Here, we investigated the epigenetic landscape of the SP7 transcriptor factor (SP7) and Distal-Less Homeobox 4 (DLX4) osteoblastic transcription factors (TFs), in human periodontal ligament mesenchymal cells (PDLCs) with low (l-PDLCs) and high (h-PDLCs) osteogenic potential. Chromatin accessibility (ATAC-seq), genome DNA methylation (Methylome), and RNA sequencing (RNA-seq) assays were performed in l- and h-PDLCs, cultured at 10 days in non-induced (DMEM) and osteogenic (OM) medium in vitro. Data were processed in HOMER, Genome Studio, and edgeR programs, and metadata was analyzed by online bioinformatics tools and in R and Python environments. ATAC-seq analyses showed the TFs genomic regions are more accessible in l-PDLCs than in h-PDLCs. In Methylome analyses, the TFs presented similar average methylation intensities (AMIs), without differently methylated probes (DMPs) between l- and h-PDLCs; in addition, there were no differences in the expression profiles of TFs signaling pathways. Interestingly, we identified the long non-coding RNAs (lncRNAs), MIR31HG and LINC00939, as upregulated in l-PDLCs, in both DMEM and OM. In the following analysis, the web-based prediction tool LncRRIsearch predicted RNA:RNA base-pairing interactions between SP7, DLX4, MIR31HG, and LINC00939 transcripts. The machine learning program TriplexFPP predicted DNA:RNA triplex-forming potential for the SP7 DNA site and for one of the LINC00939 transcripts (ENST00000502479). PCR data confirmed the upregulation of MIR31HG and LINC00939 transcripts in l-PDLCs (× h-PDLCs) in both DMEM and OM (p < 0.05); conversely, SP7 and DLX4 were downregulated, confirming those results observed in the RNA-Seq analysis. Together, these results indicate the lncRNAs MIR31HG and LINC00939 as possible epigenetic inhibitors of the osteogenic differentiation in PDLCs by (post)transcriptional and translational repression of the SP7 and DLX4 TFs.

A iniciação científica é uma oportunidade aos acadêmicos de desenvolver senso crítico, ético e profissional, com o intuito de aumentar a evidenciação científica e conduzir o que de melhor a prática odontológica pode oferecer. A universidade é um cenário importante de estímulo e incentivo à pesquisa, tendo um papel fundamental no desenvolvimento desta atividade, sendo necessário o apoio institucional e de professores interessados. O objetivo é conhecer o interesse dos graduandos do curso de Odontologia da Universidade Federal do Espírito Santo em relação à pesquisa científica. O estudo se caracteriza como descritivo transversal com abordagem quanti/qualitativa. A coleta de dados foi realizada com 101 acadêmicos do curso de Odontologia da Universidade Federal do Espírito Santo (UFES) através de um questionário semiestruturado, pré-validado, contendo perguntas sobre o interesse em pesquisa, participação em jornadas científicas e sobre o Trabalho de Conclusão de Curso (TCC). Os resultados encontrados foram: 79,2% relatam gostar de pesquisa científica e 81,19% têm interesse em pesquisar. É principalmente no 3o e 4o períodos que os alunos começam a frequentar eventos científicos, e na sua maioria, no 5o e 6o períodos começam a elaborar e apresentar trabalhos científicos. Conclui-se que um número considerável de acadêmicos de Odontologia mostra interesse e participa de pesquisas científicas, mesmo que a falta de incentivo financeiro, apoio dos professores e da infraestrutura não colabore para a realização de estudos com maior evidência científica.