Knockdown of lncRNA HAGLR promotes Treg cell differentiation through increasing the RUNX3 level in dermatomyositis.

Dermatomyositis (DM) is a systemic autoimmune disease. It's known that the number of regulatory T (Treg) cells was decreased in DM. Besides, Treg cells were increased after treatment in DM patients. Forkhead box P3 (Foxp3) is specifically expressed in Treg cells and Runt-related transcription factor (RUNX3) could regulate Foxp3 transcription. And our previous experiment showed that Homeobox D gene cluster antisense growth-associated long noncoding RNA (HAGLR) was up-regulated in DM patients. Here, we aimed to explore whether HAGLR regulated the differentiation of Treg cells through RUNX3-mediated transcription of Foxp3, thus affecting the progression of DM. The levels of HAGLR, Foxp3, and RUNX3 were examined by quantitative real-time PCR. The protein levels of Foxp3 and RUNX3 were examined by western blot. The proportions of Treg cells in CD4+ T cells were detected by flow cytometry. Hematoxylin and eosin staining was conducted to observe the histopathological changes of the muscle. RNA pull-down assay was performed to detect the interaction between HAGLR and RUNX3. A dual-luciferase reporter gene assay was conducted to examine the effect of HAGLR on the transcriptional regulation of Foxp3 by RUNX3. HAGLR was up-regulated and Foxp3 was down-regulated in DM patients. Besides, RUNX3 protein levels were decreased in DM patients, while its mRNA levels did not change significantly. The proportion of Treg cells was down-regulated in DM patients. In addition, interference with HAGLR could increase the levels of Foxp3, RUNX3 protein level, and the proportion of Treg cells. Besides, there was an interaction between HAGLR and RUNX3. We also found that knockdown of HAGLR and RUNX3 restored the increased Treg cells induced by HAGLR knockdown alone. In vivo experiments indicated that injection with adv-HAGLR increased Treg cell proportion and attenuated DM development. Interference with HAGLR could increase the protein levels of RUNX3, high levels of RUNX3 further promoted the expression of the Foxp3, thus restoring the number of Treg cells and easing the development of DM.

Antitumor activity of cell-permeable RUNX3 protein in gastric cancer cells.

PURPOSE: Gastric cancer is a leading cause of cancer death worldwide. Limited therapeutic options highlight the need to understand the molecular changes responsible for the disease and to develop therapies based on this understanding. The goal of this study was to develop cell-permeable (CP-) forms of the RUNT-related transcription factor 3, RUNX3-a candidate tumor suppressor implicated in gastric and other epithelial cancers-to study the therapeutic potential of RUNX3 in the treatment of gastric cancer. EXPERIMENTAL DESIGN: We developed novel macromolecule transduction domains (MTD) which were tested for the ability to promote protein uptake by mammalian cells and tissues and used to deliver of biologically active RUNX3 into human gastric cancer cells. The therapeutic potential CP-RUNX3 was tested in the NCI-N87 human tumor xenograft animal model. RESULTS: RUNX3 fusion proteins, HM(57)R and HM(85)R, containing hydrophobic MTDs enter gastric cancer cells and suppress cell phenotypes (e.g., cell-cycle progression, wounded monolayer healing, and survival) and induce changes in biomarker expression (e.g., p21(Waf1) and VEGF) consistent with previously described effects of RUNX3 on TGF-ß signaling. CP-RUNX3 also suppressed the growth of subcutaneous human gastric tumor xenografts. The therapeutic response was comparable with studies augmenting RUNX3 gene expression in tumor cell lines; however, the protein was most active when administered locally, rather than systemically (i.e., intravenously). CONCLUSIONS: These results provide further evidence that RUNX3 can function as a tumor suppressor and suggest that practical methods to augment RUNX3 function could be useful in treating of some types of gastric cancer.