Deregulation of the imprinted DLK1-DIO3 locus ncRNAs is associated with replicative senescence of human adipose-derived stem cells

Background Human adult adipose-derived stem cells (hADSCs) have become the most promising cell source for regenerative medicine. However the prolonged ex vivo expansion periods required to obtain the necessary therapeutic dose promotes progressive senescence, with the concomitant reduction of their therapeutic potential. Aim and scope A better understanding of the determinants of hADSC senescence is needed to improve biosafety while preserving therapeutic efficiency. Here, we investigated the association between deregulation of the imprinted DLK1-DIO3 region and replicative senescence in hADSC cultures. Methods We compared hADSC cultures at short (P S ) and prolonged (P L ) passages, both in standard and low [O 2 ] (21 and 3%, respectively), in relation to replicative senescence. hADSCs were evaluated for expression alterations in the DLK1-DIO3 region on chromosome 14q32, and particularly in its main miRNA cluster. Results Comparison of hADSCs cultured at P L or P S surprisingly showed a quite significant fraction (69%) of upregulated miRNAs in P L cultures mapping to the imprinted 14q32 locus, the largest miRNA cluster described in the genome. In agreement, expression of the lncRNA MEG3 (Maternally Expressed 3; Meg3/Gtl2), cultured at 21 and 3% [O 2 ], was also significantly higher in P L than in P S passages. During hADSC replicative senescence the AcK16H4 activating mark was found to be significantly associated with the deregulation of the entire DLK1-DIO3 locus, with a secondary regulatory role for the methylation of DMR regions. Conclusion A direct relationship between DLK1-DIO3 deregulation and replicative senescence of hADSCs is reported, involving upregulation of a very significant fraction of its largest miRNA cluster (14q32.31), paralleled by the progressive overexpression of the lncRNA MEG3, which plays a central role in the regulation of Dlk1/Dio3 activation status in mice.This work was supported by grants to AB from the Spanish Ministry of Economy, Industry (SAF2015-70882-R; AEI/FEDER, UE), Comunidad Autónoma de Madrid (S2010/BMD-2420), Instituto Salud Carlos III (RETICS TerCel, RD12/0019/0018) and the European Commission (FP7-HEALTH- 2009/CARE-MI). AMS was supported by grants from the MINECO (SAF2010–17167) and Instituto Salud Carlos III (RETICS TerCel, RD12/0019/0013), and MFF and RGU by grants from the Plan Nacional de I+D+I 2013-2016/FEDER (PI15/ 00892), the Asturias Regional Government (GRUPIN14-052), the IUOPA (Obra Social Cajastur) and the Fundación Científica de la AECC. SGL held a predoctoral fellowship from the Spanish Programa de Formación del Profesorado Universitari

Argonaute 2 drives miR-145-5p-dependent gene expression program in breast cancer cells

To perform their regulatory functions, microRNAs (miRNAs) must assemble with any of the four mammalian Argonaute (Ago) family of proteins, Ago1–4, into an effector complex known as the RNA-induced silencing complex (RISC). While the mature miRNA guides the RISC complex to its target mRNA, the Ago protein represses mRNA translation. The specific roles of the various Ago members in mediating miRNAs activity, however, haven’t been clearly established. In this study, we investigated the contribution of Ago2, the only human Ago protein endowed with nuclease activity, to the function of tumor-suppressor miR-145-5p in breast cancer (BC). We show that miR-145-5p and Ago2 protein are concomitantly downregulated in BC tissues and that restoration of miR-145-5p expression in BC cells leads to Ago2 protein induction through the loosening of Ago2 mRNA translational repression. Functionally, miR-145-5p exerts its inhibitory activity on cell migration only in presence of Ago2, while, upon Ago2 depletion, we observed increased miR-145/Ago1 complex and enhanced cell motility. Profiling by microarray of miR-145-5p target mRNAs, in BC cells depleted or not of Ago2, revealed that miR-145-5p drives Ago2-dependent and -independent activities. Our results highlight that the Ago2 protein in cancer cells strictly dictates miR-145-5p tumor suppressor activity

Post-transcriptional regulation of microRNA biogenesis and localization in mammalian neurons

The remarkable cognitive capabilities of our brain require a complex and dynamic network of neurons that is able to quickly and precisely react to changes. Adapting to an ever-changing environment and the storage of information requires that sensory information is transformed into long-lasting structural changes. At the molecular level, highly sophisticated and tightly regulated gene expression programs are necessary to alter synaptic connections in the brain without disrupting the existing network. MicroRNAs are important regulators in this neuronal network as they are able to precisely regulate local gene expression. These small non-coding RNAs bind complementary sequences in target mRNAs, thereby repressing their translation into protein. This plays an important role in activity-dependent synapse development, where local protein synthesis in dendrites is required to implement long-lasting changes in synaptic strength. The latter serves as the molecular basis for learning and memory processes. This cumulative dissertation presents two studies that investigate how neuronal micro- RNAs are regulated at the level of biogenesis and localization. The first publication "The DEAH-box helicase DHX36 mediates dendritic localization of the neuronal precursor-microRNA-134" investigates the transport of miRNA-134 to its final destination in the synapto-dendritic compartment. The study describes that already the precursor (pre-miRNA) is located at the synapse and identifies DHX36 as a protein that specifically binds pre-miR-134 and is important for its transport. Knockdown of DHX36 further shows that the localization of pre-miR-134 to the dendrite is of functional importance. The absence of DHX36 leads to elevated expression of known miR-134 targets, accompanied by an increase in dendritic spine volume. The second study presented in this thesis entitled "The nuclear matrix protein Matr3 regulates processing of the synaptic microRNA-138-5p" investigates the expression of microRNA-138. Two distinct precursor forms are known for miR-138, pre-miR-138-1 and pre-miR-138-2. In our study, we demonstrate that pre-miR-138-2 is the primary source for mature miR-138 in neurons. Using pulldown assays we identify the nuclear matrix protein Matrin-3 (Matr3) as a specific interactor of the hairpin structure of both the primary and precursor form of miR-138-2 (pri-/pre-miR-138-2). Knockdown of its expression demonstrates an inhibitory function of Matr3 in the nuclear processing of pri-miR-138-2, resulting in decreased mature miR-138 levels. In summary, this thesis describes novel post-transcriptional regulatory mechanisms that control the expression and sub-cellular localization of two neuronal microRNAs, miR- 134 and miR-138. Both microRNAs have important roles in synaptic plasticity and a precise regulation of their expression is crucial for maintaining a stable and functional neuronal network. A further understanding of the regulation of these microRNAs and their downstream processes is an important step to gain insight into the complex regulatory processes involved in learning and memory, as well as into malfunctions of these systems that occur in neurological diseases

Effect of oxidative stress and 3-hydroxytyrosol on DNA methylation levels of miR-9 promoters

No abstract availabl

MicroRNAs and Neutrophil Activation Markers Predict Venous Thrombosis in Pancreatic Ductal Adenocarcinoma and Distal Extrahepatic Cholangiocarcinoma

Cancer-associated venous thrombosis (VTE) increases mortality and morbidity. However, limited tools are available to identify high risk patients. Upon activation, neutrophils release their content through different mechanisms, thereby prompting thrombosis. We explored plasma microRNAs (miRNAs) and neutrophil activation markers to predict VTE in pancreatic ductal adenocarcinoma (PDAC) and distal extrahepatic cholangiocarcinoma (DECC). Twenty-six PDAC and 6 DECC patients recruited at cancer diagnosis, were examined for deep vein thrombosis and pulmonary embolisms, and were then followed-up with clinical examinations, blood collections, and biCUS. Ten patients developed VTE and were compared with 22 age- and sex-matched controls. miRNA expression levels were measured at diagnosis and right before VTE, and neutrophil activation markers (cell-free DNA, nucleosomes, calprotectin, and myeloperoxidase) were measured in every sample obtained during follow-up. We obtained a profile of 7 miRNAs able to estimate the risk of future VTE at diagnosis (AUC = 0.95; 95% Confidence Interval (CI) (0.987, 1)) with targets involved in the pancreatic cancer and complement and coagulation cascades pathways. Seven miRNAs were up- or down-regulated before VTE compared with diagnosis. We obtained a predictive model of VTE with calprotectin as predictor (AUC = 0.77; 95% CI (0.57, 0.95)). This is the first study that addresses the ability of plasma miRNAs and neutrophil activation markers to predict VTE in PDAC and DECC

The Beacon, April 28, 2008

Vol. 20, Issue 68, 16 pageshttps://digitalcommons.fiu.edu/student_newspaper/1222/thumbnail.jp

Evaluation of subcutaneous proleukin (Interleukin-2) in a randomized international trial (ESPRIT): Geographical and gender differences in the baseline characteristics of participants

Background: ESPRIT, is a phase III, open-label, randomized, international clinical trial evaluating the effects of subcutaneous recombinant interleukin-2 (rIL-2) plus antiretroviral therapy (ART) versus ART alone on HIV-disease progression and death in HIV-1-infected individuals with CD4+ T-cells ≥300 cells/μL. Objectives: To describe the baseline characteristics of participants randomized to ESPRIT overall and by geographic location. Method: Baseline characteristics of randomized participants were summarized by region. Results: 4,150 patients were enrolled in ESPRIT from 254 sites in 25 countries. 41%, 27%, 16%, 11%, and 5% were enrolled in Europe, North America, South America, Asia, and Australia, respectively. The median age was 40 years, 81% were men, and 76%, 11%, and 9% were Caucasian, Asian, and African American or African, respectively. 44% of women enrolled (n = 769) were enrolled in Thailand and Argentina. Overall, 55% and 38% of the cohort acquired HIV through male homosexual and heterosexual contact, respectively. 25% had a prior history of AIDS-defining illness; Pneumocystis jirovecii pneumonia, M. tuberculosis, and esophageal candida were most commonly reported. Median nadir and baseline CD4+ T-cell counts were 199 and 458 cells/μL, respectively. 6% and 13% were hepatitis B or C virus coinfected, respectively. Median duration of antiretroviral therapy (ART) was 4.2 years; the longest median duration was in Australia (5.2 years) and the shortest was in Asia (2.3 years). 17%, 13%, and 69% of participants began ART before 1995, between 1996 and 1997, and from 1998 onward, respectively. 86% used ART from two or more ART classes, with 49% using a protease inhibitor-based regimen and 46% using a nonnucleoside reverse transcriptase inhibitor-based regimen. 78% had plasma HIV RNA below detection (<500 cp/mL). Conclusion: ESPRIT has enrolled a diverse population of HIV-infected individuals including large populations of women and patients of African-American/African and Asian ethnicity often underrepresented in HIV research. As a consequence, the results of the study may have wide global applicability

Circulating microRNA profiles predict the severity of COVID-19 in hospitalized patients

We aimed to examine the circulating microRNA (miRNA) profile of hospitalized COVID-19 patients and evaluate its potential as a source of biomarkers for the management of the disease. This was an observational and multicenter study that included 84 patients with a positive nasopharyngeal swab Polymerase chain reaction (PCR) test for SARS-CoV-2 recruited during the first pandemic wave in Spain (March-June 2020). Patients were stratified according to disease severity: hospitalized patients admitted to the clinical wards without requiring critical care and patients admitted to the intensive care unit (ICU). An additional study was completed including ICU nonsurvivors and survivors. Plasma miRNA profiling was performed using reverse transcription polymerase quantitative chain reaction (RT-qPCR). Predictive models were constructed using least absolute shrinkage and selection operator (LASSO) regression. Ten circulating miRNAs were dysregulated in ICU patients compared to ward patients. LASSO analysis identified a signature of three miRNAs (miR-148a-3p, miR-451a and miR-486-5p) that distinguishes between ICU and ward patients [AUC (95% CI) = 0.89 (0.81-0.97)]. Among critically ill patients, six miRNAs were downregulated between nonsurvivors and survivors. A signature based on two miRNAs (miR-192-5p and miR-323a-3p) differentiated ICU nonsurvivors from survivors [AUC (95% CI) = 0.80 (0.64–0.96)]. The discriminatory potential of the signature was higher than that observed for laboratory parameters such as leukocyte counts, C-reactive protein (CRP) or D-dimer [maximum AUC (95% CI) for these variables = 0.73 (0.55–0.92)]. miRNA levels were correlated with the duration of ICU stay. Specific circulating miRNA profiles are associated with the severity of COVID-19. Plasma miRNA signatures emerge as a novel tool to assist in the early prediction of vital status deterioration among ICU patients.Supported by ISCIII (CIBERESUCICOVID, COV20/00110), co-funded by ERDF, “Una manera de hacer Europa”. DdGC acknowledges receiving financial support from Instituto de Salud Carlos III (ISCIII), Miguel Servet 2020 (CP20/00041), co-funded by the European Social Fund (ESF), “Investing in your future”. LP is the recipient of a predoctoral fellowship from the Ministry of Universities of Spain (FPU19/03526). This work partially supported by IRBLleida Biobank (B.0000682) and “Plataforma Biobancos PT17/0015/0027/”.Peer Reviewed"Article signat per 33 autors/es: David de Gonzalo-Calvo, Iván D. Benítez, Lucía Pinilla, Amara Carratalá, Anna Moncusí-Moix, Clara Gort-Paniello, Marta Molinero, Jessica González, Gerard Torres, María Bernal, Silvia Pico, Raquel Almansa, Noelia Jorge, Alicia Ortega, Elena Bustamante-Munguira, José Manuel Gómez, Milagros González-Rivera, Dariela Micheloud, Pablo Ryan, Amalia Martinez, Luis Tamayo, César Aldecoa, Ricard Ferrer, Adrián Ceccato, Laia Fernández-Barat, Ana Motos, Jordi Riera, Rosario Menéndez, Dario Garcia-Gasulla, Oscar Peñuelas, Antoni Torres, Jesús F. Bermejo-Martin, Ferran Barbé on behalf of the Ciberesucicovid Project (Cov20/00110, Isciii)"Postprint (author's final draft