Hypoxia stimulates pulmonary artery smooth muscle cell (PASMC) proliferation. Recent studies have implicated an important role for microRNAs (miRNAs) in hypoxia-mediated responses in various cellular processes, including cell proliferation. In this study, we investigated the role of microRNA-21 (miR-21) in hypoxia-induced PASMC proliferation and migration. We first demonstrated that miR-21 expression increased by ∼3-fold in human PASMC after 6 h of hypoxia (3% O₂) and remained high (∼2-fold) after 24 h of hypoxia. Knockdown of miR-21 with anti-miR-21 inhibitors significantly reduced hypoxia-induced cell proliferation, whereas miR-21 overexpression in normoxia enhanced cell proliferation. We also found that miR-21 is essential for hypoxia-induced cell migration. Protein expression of miR-21 target genes, specifically programmed cell death protein 4 (PDCD4), Sprouty 2 (SPRY2), and peroxisome proliferator-activated receptor-α (PPARα), was decreased in hypoxia and in PASMC overexpressing miR-21 in normoxia and increased in hypoxic cells in which miR-21 was knocked down. In addition, PPARα 3'-untranslated region (UTR) luciferase-based reporter gene assays demonstrated that PPARα is a direct target of miR-21. Taken together, our findings indicate that miR-21 plays a significant role in hypoxia-induced pulmonary vascular smooth muscle cell proliferation and migration by regulating multiple gene targets.

MicroRNAs (miRNAs) are small endogenous RNAs and are widely regarded as one of the most important regulators of gene expression in both plants and animals. To define the roles of miRNAs in fetal lung development, we profiled the miRNA expression pattern during lung development with a miRNA microarray. We identified 21 miRNAs that showed significant changes in expression during lung development. These miRNAs were grouped into four distinct clusters based on their expression pattern. Cluster 1 contained miRNAs whose expression increased as development progressed, while clusters 2 and 3 showed the opposite trend of expression. miRNAs in cluster 4 including miRNA-127 (miR-127) had the highest expression at the late stage of fetal lung development. Quantitative real-time PCR validated the microarray results of six selected miRNAs. In situ hybridization demonstrated that miR-127 expression gradually shifted from mesenchymal cells to epithelial cells as development progressed. Overexpression of miR-127 in fetal lung organ culture significantly decreased the terminal bud count, increased terminal and internal bud sizes, and caused unevenness in bud sizes, indicating improper development. These findings suggest that miR-127 may have an important role in fetal lung development.

Rationale: Recent studies suggest that microRNAs (miRNAs) play important roles in regulation of pulmonary artery smooth muscle cell (PASMC) phenotype and are implicated in pulmonary arterial hypertension (PAH). However, the underlying molecular mechanisms remain elusive.Objectives: This study aims to understand the mechanisms regulating PASMC proliferation and differentiation by microRNA-17z92 (miR-17z92) and to elucidate its implication in PAH. Methods:We generated smooth muscle cell (SMC)-specific miR17z92 and PDZ and LIM domain 5 (PDLIM5) knockout mice and overexpressed miR-17z92 and PDLIM5 by injection of miR-17z92 mimics or PDLIM5-V5-His plasmids and measured their responses to hypoxia. We used miR-17z92 mimics, inhibitors, overexpression vectors, small interfering RNAs against PDLIM5, Smad, and transforming growth factor (TGF)-b to determine the role of miR-17z92 and its downstream targets in PASMC proliferation and differentiation.Measurements and Main Results: We found that human PASMC (HPASMC) from patients with PAH expressed decreased levels of the miR-17z92 cluster, TGF-b, and SMC markers. Overexpression of miR-17z92 increased and restored the expression of TGF-b 3 , Smad3, and SMC markers in HPASMC of normal subjects and patients with idiopathic PAH, respectively. Knockdown of Smad3 but not Smad2 prevented miR-17z92-induced expression of SMC markers. SMC-specific knockout of miR-17z92 attenuated hypoxiainduced pulmonary hypertension (PH) in mice, whereas reconstitution of miR-17z92 restored hypoxia-induced PH in these mice. We also found that PDLIM5 is a direct target of miR-17/20a, and hypertensive HPASMC and mouse PASMC expressed elevated PDLIM5 levels. Suppression of PDLIM5 increased expression of SMC markers and enhanced TGF-b/Smad2/3 activity in vitro and enhanced hypoxia-induced PH in vivo, whereas overexpression of PDLIM5 attenuated hypoxia-induced PH. Conclusions:We provided the first evidence that miR-17z92 inhibits PDLIM5 to induce the TGF-b 3 /SMAD3 pathway, contributing to the pathogenesis of PAH.

Edible berries have a broad spectrum of biomedical functions, including improving immune responses and reducing risk for chronic diseases. In this study, the anti-inflammatory activities of crude extracts (CEs), anthocyanin-rich fractions (ARFs), and des-anthocyanin fractions (DAFs) from seven berries were evaluated based on their inhibitory effects on nitric oxide (NO) production in lipopolysaccharide (LPS)/IFN-γ-activated RAW264.7 macrophages. ARFs from red raspberries (RR-ARFs) exhibited the highest efficiency in suppressing NO synthesis. The anti-inflammatory properties were also demonstrated by reducing the expression levels of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), interleukin-1 beta (IL-1β) and IL-6 in RAW264.7 cells. The luciferase reporter assay demonstrated that the activities of NF-κB and AP-1 signaling pathways were significantly suppressed by RR-ARFs. Further studies showed that RR-ARFs decreased the phosphorylation of IKK, IκBα, p65 and JNK and the nuclear translocation of p65 in LPS/IFN-γ-stimulated RAW264.7 cells. In a mouse colitis model, dextran sulfate sodium (DSS)-induced weight loss and histological damage were significantly ameliorated by RR-ARFs treatment. Taken together, our results indicate that RR-ARFs attenuate inflammation both in vitro and in vivo primarily by inhibiting the activation of NF-κB and MAPKs. The anti-inflammatory of RR-ARFs could be harnessed and applied in animal agriculture, drug and food industries.