The long non-coding RNA MEG3 has been reported to be a tumor suppressor in a number of malignant tumors including gastric cancer. Several studies have shown that the regulation of MEG3 may attribute to the promoter hypermethylation. However, the mechanism of MEG3 regulation in gastric cancer is still not well understood. MiR-148a can suppress gastric tumorigenesis through regulating the expression of target genes such as DNA methyltransferase 1(DNMT-1). We examined the expression of MEG3 in 52 gastric cancer samples using quantitative real-time PCR and found the down-regulation of MEG3 in both gastric cancer tissues and cell lines. The positive correlation of MEG3 and miR-148a was further confirmed in SGC-7901 and BGC-823 gastric cancer cell lines. Hypermethylation of MEG3 differentially methylated regions was identified by methylation-specific PCR, and MEG3 expression was increased with the inhibition of methylation with siRNA to DNMT-1 in gastric cancer cells. In addition, transfection of MEG3 siRNA into gastric cancer cells diminished the suppression of proliferation induced by overexpression of miR-148a. Our results suggest that the suppression of miR-148a may contribute to the down-regulation of MEG3 in gastric cancer by modulation of DNMT-1.

Epithelial-mesenchymal transition (EMT), a process closely related to tumor development, is regulated by a variety of signaling pathways and growth factors, such as transforming growth factor-β1 (TGF-β1) and epidermal growth factor (EGF). Hyaluronan (HA) has been shown to induce EMT through either TGF-β1 or EGF signaling and to be a regulator of the crosstalk between these two pathways in fibroblasts. In this study, in order to clarify whether HA has the same effect in tumor cells, we utilized the lung cancer cell line, A549, and the breast cancer cell line, MCF-7, and found that the effects of stimulation with TGF-β1 were more potent than those of EGF in regulating the expression of EMT-associated proteins and in enhancing cell migration and invasion. In addition, we observed that TGF-β1 activated EGF receptor (EGFR) and its downstream AKT and extracellular signal-regulated kinase (ERK) pathways. Furthermore, we found that TGF-β1 upregulated the expression of hyaluronan synthases (HAS1, HAS2 and HAS3) and promoted the expression of CD44, a cell surface receptor for HA, which interacts with EGFR, resulting in the activation of the downstream AKT and ERK pathways. Conversely, treatment with 4-methylumbelliferone (4-MU; an inhibitor of HAS) prior to stimulation with TGF-β1, inhibited the expression of CD44 and EGFR, abolished the interaction between CD44 and EGFR. Furthermore, the use of shRNA targeting CD44 impaired the expression of EGFR, deactivated the AKT and ERK pathways, reversed EMT and decreased the migration and invasion ability of cells. In conclusion, our data demonstrate that TGF-β1 induces EMT by the transactivation of EGF signaling through HA/CD44 in lung and breast cancer cells.

Background/Aims: The rejuvenation properties of nanofat grafting have been described in recent years. However, it is not clear whether the clinical efficacy of the procedure is attributable to stem cells or linked to other components of adipose tissue. In this study we isolated nanofat-derived stem cells (NFSCs) to observe their biological characteristics and evaluate the efficacy of precise intradermal injection of nanofat combined with platelet-rich fibrin (PRF) in patients undergoing facial rejuvenation treatment. Methods: Third-passage NFSCs were isolated and cultured using a mechanical emulsification method and their surface CD markers were analyzed by flow cytometry. The adipogenic and osteogenic nature and chondrogenic differentiation capacity of NFSCs were determined using Oil Red O staining, alizarin red staining, and Alcian blue staining, respectively. Paracrine function of NFSCs was evaluated by enzyme-linked immunosorbent assay (ELISA) at 1, 3, 7, 14, and 28 days after establishing the culture. Then, the effects of PRF on NFSC proliferation were assessed in vitro. Finally, we compared the outcome in 103 patients with facial skin aging who underwent both nanofat and intradermal PRF injection (treatment group) and 128 patients who underwent hyaluronic acid (HA) injection treatment (control group). Outcomes in the two groups were compared by assessing pictures taken at the same angle before and after treatment, postoperative recovery, incidence of local absorption and cysts, and skin quality before treatment, and at 1, 12, 24 months after treatment using the VISIA Skin Image Analyzer and a SOFT5.5 skin test instrument. Results: NFSCs expressed CD29, CD44, CD49d, CD73, CD90, and CD105, but did not express CD34, CD45, and CD106. NFSCs also differentiated into adipocytes, osteoblasts, and chondrocytes under appropriate induction conditions. NFSCs released large amounts of growth factors such as VEGF, bFGF, EGF, and others, and growth factor levels increased in a time-dependent manner. At the same time, PRF enhanced proliferation of NFSCs in vitro in a dose-dependent manner, and the growth curves under different concentrations of PRF all showed plateaus 6d after seeding. Facial skin texture was improved to a greater extent after combined injection of nanofat and PRF than after control injection of HA. The nanofat-PRF group had a higher satisfaction rate. Neither treatment caused any complications such as infection, anaphylaxis, or paresthesia during long-term follow-up. Conclusion: NFSCs demonstrate excellent multipotential differentiation and paracrine function, and PRF promotes proliferation of NFSCs during the early stage after seeding. Both nanofat-PRF and HA injection improve facial skin status without serious complications, but the former was associated with greater patient satisfaction, implying that nanofat-PRF injection is a safe, highly effective, and long-lasting method for skin rejuvenation.