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Background: Cytokines play key roles in stimulating periodontal regeneration; however, their exact mechanisms of action remain unclear. Mesenchymal stem cells (MSCs) are multipotent cells that have self-renewal abilities and can differentiate into periodontal tissues such as bone, cementum, and periodontal ligaments following transplantation, like periodontal progenitor cells. Here, we used MSCs to identify the regulatory genes induced by periodontal regenerative cytokines.Methods: Human MSCs (hMSCs) were cultured under conditions of periodontal regenerative cytokine stimulation or silencing of undifferentiated hMSC transcription factors. To characterize the changes associated with periodontal regenerative cytokine-regulated microRNAs (miRNAs), miRNA, and mRNA expression was evaluated using miRNA arrays and quantitative real-time polymerase chain reaction, respectively. One of the identified miRNAs, miR-628-5p, was then overexpressed or suppressed in hMSCs during osteogenesis; the effect of these changes on osteogenesis was investigated.Results: Cytokine-stimulated MSCs showed characteristic miRNA profiles and mRNA levels of undifferentiated hMSC transcription factors ETV1, SOX11, and GATA6. Next, we silenced these transcription factors in MSCs and examined the miRNA profiles. The levels of miR-628-5p were decreased upon all cytokine treatments and were increased upon silencing of ETV1, SOX11, and GATA6. Overexpression of miR-628-5p suppressed osteogenesis; however, its inhibition enhanced OPN, ALP, OC, BMP2, and RUNX2 mRNA levels, and bone matrix mineralization, but not OSX mRNA or ALP activity.Conclusions: miR-628-5p negatively regulates MSC stemness during periodontal regeneration. Periodontal regenerative cytokines act as miR-628-5p suppressors to support periodontal regeneration. Thus, selection of effective cytokines for different MSCs, based on miRNA profiling, is important for advancing regenerative therapies. [ABSTRACT FROM AUTHOR]

Objective: Periodontitis causes periodontal tissue destruction and results in physiological tooth dysfunction. Therefore, periodontal regeneration is ideal therapy for periodontitis. Mesenchymal stem cells (MSCs) are useful for periodontal regenerative therapy as they can differentiate into periodontal cells; however, the underlying regulatory mechanism is unclear. In this study, we attempted to identify regulatory genes involved in periodontal cell differentiation and clarify the differentiation mechanism for effective periodontal regenerative therapy. Background: The cementum and periodontal ligament play important roles in physiological tooth function. Therefore, cementum and periodontal ligament regeneration are critical for periodontal regenerative therapy. Mesenchymal stem cell transplantation can be a common periodontal regenerative therapy because these cells have multipotency and self‐renewal ability, which induces new cementum or periodontal ligament formation. Moreover, MSCs can differentiate into cementoblasts. Cementoblast‐ or periodontal ligament cell–specific proteins have been reported; however, it is unclear how these proteins are regulated. MicroRNA (miRNA) can also act as a key regulator of MSC function. Therefore, in this study, we identified regulatory genes involved in cementoblast or periodontal cell differentiation and commitment. Methods: Human MSCs (hMSCs), cementoblasts (HCEM), and periodontal ligament cells (HPL cells) were cultured, and mRNA or miRNA expression was evaluated. Additionally, cementoblast‐specific genes were overexpressed or suppressed in hMSCs and their expression levels were investigated. Results: HCEM and HPL cells expressed characteristic genes, of which we focused on ets variant 1 (ETV1), miR‐628‐5p, and miR‐383 because ETV1 is a differentiation‐related transcription factor, miR‐628‐5p was the second‐highest expressed gene in HCEM and lowest expressed gene in HPL cells, and miR‐383 was the highest expressed gene in HCEM. miR‐628‐5p and miR‐383 overexpression in hMSCs regulated ETV1 mRNA expression, and miR‐383 overexpression downregulated miR‐628‐5p expression. Moreover, miR‐383 suppression decreased miR‐383 expression and enhanced ETV1 mRNA expression, but miR‐383 suppression also decreased miR‐628‐5p. Furthermore, silencing of ETV1 expression in hMSCs regulated miR‐628‐5p and miR‐383 expression. Concerning periodontal cell commitment, miR‐628‐5p, miR‐383, and ETV1 regulated the expression of HCEM‐ or HPL cell–related genes by adjusting the expression of these miRNAs. Conclusion: HCEM and HPL cells show characteristic mRNA and miRNA profiles. In particular, these cells have specific miR‐383, miR‐628‐5p, and ETV1 expression patterns, and these genes interact with each other. Therefore, miR‐383, miR‐628‐5p, and ETV1 are key genes involved in cementogenesis or HPL cell differentiation. [ABSTRACT FROM AUTHOR]

Objective: This study aimed to determine the regulatory mechanism of bone marrow-derived mesenchymal stem cell (BM-MSC) differentiation mediated by humoral factors derived from human periodontal ligament (HPL) cells and human gingival fibroblasts (HGFs). We analyzed histone deacetylase (HDAC) expression and activity involved in BM-MSC differentiation and determined their regulatory effects in co-cultures of BM-MSCs with HPL cells or HGFs., Background: BM-MSCs can differentiate into various cell types and can, thus, be used in periodontal regenerative therapy. However, the mechanism underlying their differentiation remains unclear. Transplanted BM-MSCs are affected by periodontal cells via direct contact or secretion of humoral factors. Therefore, their activity is regulated by humoral factors derived from HPL cells or HGFs., Methods: BM-MSCs were indirectly co-cultured with HPL cells or HGFs under osteogenic or growth conditions and then analyzed for osteogenesis, HDAC1 and HDAC2 expression and activity, and histone H3 acetylation. BM-MSCs were treated with trichostatin A, or their HDAC1 or HDAC2 expression was silenced or overexpressed during osteogenesis. Subsequently, they were evaluated for osteogenesis or the effects of HDAC activity., Results: BM-MSCs co-cultured with HPL cells or HGFs showed suppressed osteogenesis, HDAC1 and HDAC2 expression, and HDAC phosphorylation; however, histone H3 acetylation was enhanced. Trichostatin A treatment remarkably suppressed osteogenesis, decreasing HDAC expression and enhancing histone H3 acetylation. HDAC1 and HDAC2 silencing negatively regulated osteogenesis in BM-MSCs to the same extent as that achieved by indirect co-culture with HPL cells or HGFs. Conversely, their overexpression positively regulated osteogenesis in BM-MSCs., Conclusion: The suppressive effects of HPL cells and HGFs on BM-MSC osteogenesis were regulated by HDAC expression and histone H3 acetylation to a greater extent than that mediated by HDAC activity. Therefore, regulation of HDAC expression has prospects in clinical applications for effective periodontal regeneration, mainly, bone regeneration., (© 2022 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)