Antisense oligonucleotide treatment enhances the recovery of acute lung injury through IL-10-secreting M2-like macrophage-induced expansion of CD4+ regulatory T cells.

MicroRNAs (miRNAs) have been shown as an important regulator in the pathologies of acute lung injury (ALI). However, the potential effect of miRNA-based therapeutic studies in ALI remains poorly understood. We assessed the effect of antisense oligonucleotides (ASOs) against miR-155 on the development of ALI using a murine ALI model. We found that miR-155 ASO treatment could enhance the recovery of ALI as evidenced by accelerated body weight back, reduced level of bronchoalveolar lavage (BAL) protein and proinflammatory cytokines, and reduced number of BAL cells. Adoptive cell transfer assay in RAG1(-/-) mice showed that CD4(+)CD25(+) regulatory T cells (Tregs) mediated the enhanced recovery of ALI. Mechanistic evidence showed that enhanced expansion of Tregs in vivo, dominantly induced by IL-10-secreting M2-like macrophages, was critical for their elevated proportion in miR-155 ASO-treated ALI mice. Finally, we report that C/EBPß, a target molecule of miR-155, was upregulated and associated with IL-10 secretion and M2-like phenotype of macrophages. These data provided a previously unknown mechanism for miRNA-based therapy against ALI, which could ultimately aid the understanding of recovery of ALI and the development of new therapeutic strategies against clinical inflammatory lung disease.

MicroRNA-126 regulates the induction and function of CD4(+) Foxp3(+) regulatory T cells through PI3K/AKT pathway.

Recent evidence showed that limited activation of PI3K/Akt pathway was critical for induction and function sustainment of CD4(+) Foxp3(+) regulatory T cells (Tregs). However, the underlying mechanism remains largely unknown. In this study, we reported that miR-126 was expressed in mouse and human Tregs. Further study showed that silencing of miR-126 using miR-126 antisense oligonucleotides (ASO) could significantly reduce the induction of Tregs in vitro. Furthermore, miR-126 silencing could obviously reduce the expression of Foxp3 on Tregs, which was accompanied by decreased expression of CTLA-4 and GITR, as well as IL-10 and TGF-ß, and impair its suppressive function. Mechanistic evidence showed that silencing of miR-126 enhanced the expression of its target p85ß and subsequently altered the activation of PI3K/Akt pathway, which was ultimately responsible for reduced induction and suppressive function of Tregs. Finally, we further revealed that miR-126 silencing could impair the suppressive function of Tregs in vivo and endow effectively antitumour effect of CD8(+) T cells in adoptive cell transfer assay using a murine breast cancer model. Therefore, our study showed that miR-126 could act as fine-tuner in regulation of PI3K-Akt pathway transduction in the induction and sustained suppressive function of Tregs and provided a novel insight into the development of therapeutic strategies for promoting T-cell immunity by regulating Tregs through targeting specific miRNAs.

Myocardial Mitochondrial DNA Drives Macrophage Inflammatory Response through STING Signaling in Coxsackievirus B3-Induced Viral Myocarditis.

Coxsackievirus B3 (CVB3), a single-stranded positive RNA virus, primarily infects cardiac myocytes and is a major causative pathogen for viral myocarditis (VMC), driving cardiac inflammation and organ dysfunction. However, whether and how myocardial damage is involved in CVB3-induced VMC remains unclear. Herein, we demonstrate that the CVB3 infection of cardiac myocytes results in the release of mitochondrial DNA (mtDNA), which functions as an important driver of cardiac macrophage inflammation through the stimulator of interferon genes (STING) dependent mechanism. More specifically, the CVB3 infection of cardiac myocytes promotes the accumulation of extracellular mtDNA. Such myocardial mtDNA is indispensable for CVB3-infected myocytes in that it induces a macrophage inflammatory response. Mechanistically, a CVB3 infection upregulates the expression of the classical DNA sensor STING, which is predominantly localized within cardiac macrophages in VMC murine models. Myocardial mtDNA efficiently triggers STING signaling in those macrophages, resulting in strong NF-kB activation when inducing the inflammatory response. Accordingly, STING-deficient mice are able to resist CVB3-induced cardiac inflammation, exhibiting minimal inflammation with regard to their functional cardiac capacities, and they exhibit higher survival rates. Moreover, our findings pinpoint myocardial mtDNA as a central element driving the cardiac inflammation of CVB3-induced VMC, and we consider the DNA sensor, STING, to be a promising therapeutic target for protecting against RNA viral infections.

Down-regulation of GAS5 has diagnostic value for tuberculosis and regulates the inflammatory response in mycobacterium tuberculosis infected THP-1 cells.

OBJECTIVE: This study aimed to investigate the expression of long non-coding RNA (lncRNA) growth arrest-special transcript 5 (GAS5) in the serum of tuberculosis (TB) patients and discuss the mechanism of GAS5 in TB by establishing an in-vitro TB cell model. METHODS: Serum expressions of GAS5 and miR-18a-5p were determined by quantitative real-time PCR (qRT-PCR). The effects of GAS5 on macrophage cell viability and the inflammatory response after MTB infection were assessed by CCK-8 and ELISA. Luciferase reporter gene assay was applied to delve into the potential target gene of GAS5. RESULTS: The expression of GAS5 in TB patients was down-regulated, while miR-18a-5p was up-regulated, and the serum inflammatory factors were negatively correlated with the expression level of GAS5. MTB infection induced significant upregulation on the cell viability and inflammatory response but the acceleration effect could be rescued by GAS5-overexpression. Meanwhile, miR-18a-5p was recognized as the target gene of GAS5. CONCLUSION: This study indicated that the expression level of GAS5 in the serum of TB patients was decreased, while in the cells infected with MTB, the down-regulated GAS5 might develop a role in facilitating the cell vitality and the inflammatory response by adsorbing miR-18a-5p in the form of molecular sponge.

MicroRNA-1271 inhibits cellular proliferation of hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the most common causes of cancer-associated mortality worldwide, particularly in China. MicroRNAs (miRs) serve important roles in the pathogenesis of HCC. The present study investigated the function of miR-1271 in HCC. The miR-1271 levels were analyzed by quantitative reverse transcription polymerase chain reaction. Cells growth was examined by MTT assay. Bioinformatics algorithms from TargetScanHuman were used to predict the target genes of miR-1271. The protein level was assayed by western blotting. miR-1271 demonstrated a lower expression level in HCC tissues. Upregulation of miR-1271 suppressed the growth of HepG-2 and Huh-7 cells and induced apoptosis of cells. Forkhead box Q1 (FOXQ1) was targeted by miR-1271. In conclusion, miR-1271 is a novel tumor suppressor that inhibits HCC proliferation and induces cellular apoptosis by targeting FOXQ1 in HCC. The results of the present study may provide a novel therapeutic target of HCC.

[Effects of microRNA-155 on the growth of human lung cancer cell line 95D 
in vitro].

BACKGROUND AND OBJECTIVE: Recent studies suggest that miR-155 is involved in lung tumorgenesis, whereas the precise mechanism has not yet been characterized. The aim of this study is to investigate the effects of over-expression of miR-155 on the growth of human lung cancer 95D cells in vitro and its possible mechanism, and thus to provide experimental evidence for further researching on the role of miR-155 in the pathogenesis and development of lung cancer. METHODS: miR-155 mimics control and miR-155 mimics were tranfected into human lung cancer 95D cells by FuGENE®HD Transfection Reagent respectively in vitro. The relative expression level of miR-155 in 95D cells was determined using specific probe of real-time PCR after transfection. The proliferation of 95D cells was detected by MTT assay. The cell cycle was analyzed by flow cytometry. The expression of SOS1 protein was measured by Western blot. RESULTS: Compared with control groups, the expression level of miR-155 was significantly increased in miR-155 mimics transfected group (P<0.05). The proliferation of miR-155-transfected 95D cells was significantly inhibited (P<0.05). The percentage of G0/G1 phase cells was increased significantly in miR-155-transfected 95D cells, while the percentage of S phase was remarkably reduced (P<0.05). Furthermore, the expression of SOS1 in miR-155-transfected 95D cells was significantly down-regulated (P<0.05). CONCLUSIONS: miR-155 could significantly inhibit the growth of human lung cancer 95D cells in vitro, which might be closely related to miR-155 induced G0/G1 phase arrest.