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Pulmonary hypertension (PH) is a deadly vascular disease with enigmatic molecular origins. We found that vascular extracellular matrix (ECM) remodeling and stiffening are early and pervasive processes that promote PH. In multiple pulmonary vascular cell types, such ECM stiffening induced the microRNA-130/301 family via activation of the co-transcription factors YAP and TAZ. MicroRNA-130/301 controlled a PPAR?-APOE-LRP8 axis, promoting collagen deposition and LOX-dependent remodeling and further upregulating YAP/TAZ via a mechanoactive feedback loop. In turn, ECM remodeling controlled pulmonary vascular cell crosstalk via such mechanotransduction, modulation of secreted vasoactive effectors, and regulation of associated microRNA pathways. In vivo, pharmacologic inhibition of microRNA-130/301, APOE, or LOX activity ameliorated ECM remodeling and PH. Thus, ECM remodeling, as controlled by the YAP/TAZ-miR-130/301 feedback circuit, is an early PH trigger and offers combinatorial therapeutic targets for this devastating disease.

Analysis of extracellular vesicles (EVs) derived from plasma or cerebrospinal fluid (CSF) has emerged as a promising biomarker platform for therapeutic monitoring in glioblastoma patients. However, the contents of the various subpopulations of EVs in these clinical specimens remain poorly defined. Here we characterize the relative abundance of miRNA species in EVs derived from the serum and cerebrospinal fluid of glioblastoma patients. EVs were isolated from glioblastoma cell lines as well as the plasma and CSF of glioblastoma patients. The microvesicle subpopulation was isolated by pelleting at 10,000×g for 30 min after cellular debris was cleared by a 2000×g (20 min) spin. The exosome subpopulation was isolated by pelleting the microvesicle supernatant at 120,000×g (120 min). qRT-PCR was performed to examine the distribution of miR-21, miR-103, miR-24, and miR-125. Global miRNA profiling was performed in select glioblastoma CSF samples. In plasma and cell line derived EVs, the relative abundance of miRNAs in exosome and microvesicles were highly variable. In some specimens, the majority of the miRNA species were found in exosomes while in other, they were found in microvesicles. In contrast, CSF exosomes were enriched for miRNAs relative to CSF microvesicles. In CSF, there is an average of one molecule of miRNA per 150-25,000 EVs. Most EVs derived from clinical biofluids are devoid of miRNA content. The relative distribution of miRNA species in plasma exosomes or microvesicles is unpredictable. In contrast, CSF exosomes are the major EV compartment that harbor miRNAs.

Introduction MicroRNAs (miRNAs) represent distinct small noncoding RNAs that function as important posttranscriptional regulators of gene expression (1). A single miRNA can silence the expression of a number of functionally [...], MicroRNA-21 (miR-21) contributes to the pathogenesis of fibrogenic diseases in multiple organs, including the kidneys, potentially by silencing metabolic pathways that are critical for cellular ATP generation, ROS production, and inflammatory signaling. Here, we developed highly specific oligonucleotides that distribute to the kidney and inhibit miR-21 function when administered subcutaneously and evaluated the therapeutic potential of these anti-miR-21 oligonucleotides in chronic kidney disease. In a murine model of Alport nephropathy, miR-21 silencing did not produce any adverse effects and resulted in substantially milder kidney disease, with minimal albuminuria and dysfunction, compared with vehicle-treated mice. miR- 21 silencing dramatically improved survival of Alport mice and reduced histological end points, including glomerulosclerosis, interstitial fibrosis, tubular injury, and inflammation. Anti-miR-21 enhanced PPARα/retinoid X receptor (PPARα/RXR) activity and downstream signaling pathways in glomerular, tubular, and interstitial cells. Moreover, miR-21 silencing enhanced mitochondrial function, which reduced mitochondrial ROS production and thus preserved tubular functions. Inhibition of miR-21 was protective against TGF-β-induced fibrogenesis and inflammation in glomerular and interstitial cells, likely as the result of enhanced PPARα/RXR activity and improved mitochondrial function. Together, these results demonstrate that inhibition of miR-21 represents a potential therapeutic strategy for chronic kidney diseases including Alport nephropathy.

Recent advances in the understanding of lipid metabolism have revealed that the genetic loci encoding the transcription factors SREBP1 and SREBP2 (known as SREBF1 and SREBF2) also encode the miRNAs [...], Cardiovascular disease remains the leading cause of mortality in westernized countries, despite optimum medical therapy to reduce the levels of low-density lipoprotein (LDL)-associated cholesterol. The pursuit of novel therapies to target the residual risk has focused on raising the levels of high-density lipoprotein (HDL)-associated cholesterol in order to exploit its atheroprotective effects (1). MicroRNAs (miRNAs) have emerged as important post-transcriptional regulators of lipid metabolism and are thus a new class of target for therapeutic intervention (2). MicroRNA-33a and microRNA-33b (miR-33a/b) are intronic miRNAs whose encoding regions are embedded in the sterol-response-element-binding protein genes SREBF2 and SREBF1 (refs 3-5), respectively. These miRNAs repress expression of the cholesterol transporter ABCA1, which is a key regulator of HDL biogenesis. Recent studies in mice suggest that antagonizing miR-33a may be an effective strategy for raising plasma HDL levels (3-5) and providing protection against atherosclerosis (6); however, extrapolating these findings to humans is complicated by the fact that mice lack miR-33b, which is present only in the SREBF1 gene of medium and large mammals. Here we show in African green monkeys that systemic delivery of an anti-miRNA oligonucleotide that targets both miR-33a and miR-33b increased hepatic expression of ABCA1 and induced a sustained increase in plasma HDL levels over 12 weeks. Notably, miR-33 antagonism in this non-human primate model also increased the expression of miR-33 target genes involved in fatty acid oxidation (CROT, CPT1A, HADHB and PRKAA1) and reduced the expression of genes involved in fatty acid synthesis (SREBF1, FASN, ACLY and ACACA), resulting in a marked suppression of the plasma levels of very-low-density lipoprotein (VLDL)-associated triglycerides, a finding that has not previously been observed in mice. These data establish, in a model that is highly relevant to humans, that pharmacological inhibition of miR-33a and miR33b is a promising therapeutic strategy to raise plasma HDL and lower VLDL triglyceride levels for the treatment of dyslipidaemias that increase cardiovascular disease risk.

Abstract Background The master regulator p53 tumor-suppressor protein through coordination of several downstream target genes and upstream transcription factors controls many pathways important for tumor suppression. While it has been reported that some of the p53's functions are microRNA-mediated, it is not known as to how many other microRNAs might contribute to the p53-mediated tumorigenesis. Results Here, we use bioinformatics-based integrative approach to identify and prioritize putative p53-regulated miRNAs, and unravel the miRNA-based microregulation of the p53 master regulatory network. Specifically, we identify putative microRNA regulators of a) transcription factors that are upstream or downstream to p53 and b) p53 interactants. The putative p53-miRs and their targets are prioritized using current knowledge of cancer biology and literature-reported cancer-miRNAs. Conclusion Our predicted p53-miRNA-gene networks strongly suggest that coordinated transcriptional and p53-miR mediated networks could be integral to tumorigenesis and the underlying processes and pathways.

Although a number of computational approaches have been developed to integrate data from multiple sources for the purpose of predicting or prioritizing candidate disease genes, relatively few of them focus on identifying or ranking drug targets. To address this deficit, we have developed an approach to specifically identify and prioritize disease and drug candidate genes. In this chapter, we demonstrate the applicability of integrative systems-biology-based approaches to identify potential drug targets and candidate genes by employing information extracted from public databases. We illustrate the method in detail using examples of two neurodegenerative diseases (Alzheimer's and Parkinson's) and one neuropsychiatric disease (Schizophrenia).