Transcriptomes of human primary skin fibroblasts of healthy individuals reveal age-associated mRNAs and long noncoding RNAs.

Changes in the transcriptomes of human tissues with advancing age are poorly cataloged. Here, we sought to identify the coding and long noncoding RNAs present in cultured primary skin fibroblasts collected from 82 healthy individuals across a wide age spectrum (22-89 years old) who participated in the GESTALT (Genetic and Epigenetic Signatures of Translational Aging Laboratory Testing) study of the National Institute on Aging, NIH. Using high-throughput RNA sequencing and a linear regression model, we identified 1437 coding RNAs (mRNAs) and 1177 linear and circular long noncoding (lncRNAs) that were differentially abundant as a function of age. Gene set enrichment analysis (GSEA) revealed select transcription factors implicated in coordinating the transcription of subsets of differentially abundant mRNAs, while long noncoding RNA enrichment analysis (LncSEA) identified RNA-binding proteins predicted to participate in the age-associated lncRNA profiles. In summary, we report age-associated changes in the global transcriptome, coding and noncoding, from healthy human skin fibroblasts and propose that these transcripts may serve as biomarkers and therapeutic targets in aging skin.

Mechanistic Investigation of Curcuma Protection against Oral Submucous Fibrosis.

Objective: Oral submucous fibrosis (OSMF) is a chronic, fibrotic disease that affects the oral cavity, showing a high rate of malignant transformation. Curcuma exerts therapeutic potentials in many diseases including OSMF. However, the potential targets and pathways to explain the therapeutic effects of curcuma on OSMF are outside the scope of present knowledge. Herein we intend to reveal the predictive targets and potential pathways of curcuma against OSMF by a network pharmacology-based approach followed by molecular docking technology. Methods: We searched the SymMap, GeneCards, and OMIM database to obtain curcuma and OSMF common targets. The protein-protein interaction (PPI) of curcuma and OSMF common targets were then analyzed, followed by functional enrichment analysis. The best binding mode of curcuma and target proteins was analyzed by molecular docking technology. Results: We collected 290 putative targets of curcuma molecules and 600 known therapeutic targets of OSMF, with 64 curcuma and OSMF common targets sorted out. In the PPI network, there were 63 nodes with 922 edges. The node indicates protein and the line indicates PPI relation. The most enriched GO term in the BP level is "gland development", followed by "cellular response to chemical stress", and then "response to oxygen levels", while the most enriched GO term in CC and MF is "membrane raft" and "cytokine receptor binding", respectively. We also found 131 KEGG pathways significantly enriched by curcuma and OSMF common targets. The binding energy of curcuma to ALB, TNF, TP53, IL6, and VEGFA was -9.5 kcal/mol, -3.9 kcal/mol, -3.5 kcal/mol, -3.6 kcal/mol, and -8.9 kcal/mol, respectively, which suggested ALB and VEGFA were regarded as main targets involving in the potential mechanism of curcuma against OSMF. Conclusion: The present study illustrated that the therapeutic effects of curcuma on OSMF were achieved by targeting ALB and VEGFA, which giving reference to further drug design and development for OSMF.

Kaempferol promotes osteogenic differentiation of mouse bone marrow mesenchymal cells under tension stress via the mTORC1 signaling pathway / 口腔疾病防治

Objective @#To investigate the activation of the mammalian target of rapamycin complex 1 (mTORC1) signaling pathway molecules during the process by which kaempferol (Kae) promotes osteogenic differentiation of mouse bone marrow mesenchymal cells (BMMCs) under cyclic and uniaxial tension.@*Methods @#BMMCs isolated and cultured in vitro were subjected to uniaxial dynamic tension with a 10% shape variable. The appropriate concentration of Kae was selected by cytotoxicity testing. The endogenous mTOR signal was inhibited by pp242. Four hours after traction, alkaline phosphatase (ALP) and osteocalcin (OCN) were detected by chemical colorimetry and ELISA, and the relative concentration of intracellular calcium was detected by flow cytometry. Phosphorylation of mTOR, 4E/BP1, and ribosomal protein S6 kinases (S6K), which are the main molecules of the endogenous mTORC1 signaling pathway, and expression of osteogenic transcription factors (Runx2 and Osterix) were detected by western blotting (WB), and mRNA expression levels of the above factors were detected by qRT-PCR.@*Results @# The cytotoxicity test showed that 10 μmol/L Kae had little inhibitory effect on cell proliferation but had the strongest osteogenic ability. Four hours after stretching, Kae effectively promoted the osteogenic differentiation of BMMCs. The expression of ALP was （153.04 ± 18.72） U/mg, the expression of OCN was （1.64 ± 0.25） U. The mRNA and protein levels of Runx2 and Osterix were upregulated, and the intracellular calcium content was decreased. The mRNA and protein phosphorylation of mTOR and S6K was upregulated, and the opposite effect was observed with 4E/BP1. After pp242 was added to inhibit mTOR signaling, mTOR and S6K mRNA and protein phosphorylation were downregulated, but 4E/BP1 mRNA and protein phosphorylation was upregulated. The osteogenic differentiation of BMMCs was also significantly inhibited, mRNA and protein expression of Runx2 and Osterix were significantly downregulated, ALP and OCN expression were downregulated, and intracellular calcium content was increased. @* Conclusion@#Kae promotes osteogenic differentiation of mouse BMMCs under uniaxial dynamic tension through the mTORC1 signaling pathway.