RESUMEN
microRNA is necessary for osteoclast differentiation, function and survival. It has been reported that miR-199/214 cluster plays important roles in vertebrate skeletal development and miR-214 inhibits osteoblast function by targeting ATF4. Here, we show that miR-214 is up-regulated during osteoclastogenesis from bone marrow monocytes (BMMs) with macrophage colony stimulating factor (M-CSF) and receptor activator of nuclear factor-κB ligand (RANKL) induction, which indicates that miR-214 plays a critical role in osteoclast differentiation. Overexpression of miR-214 in BMMs promotes osteoclastogenesis, whereas inhibition of miR-214 attenuates it. We further find that miR-214 functions through PI3K/Akt pathway by targeting phosphatase and tensin homolog (Pten). In vivo, osteoclast specific miR-214 transgenic mice (OC-TG214) exhibit down-regulated Pten levels, increased osteoclast activity, and reduced bone mineral density. These results reveal a crucial role of miR-214 in the differentiation of osteoclasts, which will provide a potential therapeutic target for osteoporosis.
Asunto(s)
MicroARNs/genética , Osteoclastos/metabolismo , Osteoporosis/genética , Fosfohidrolasa PTEN/genética , Fosfatidilinositol 3-Quinasas/genética , Proteínas Proto-Oncogénicas c-akt/genética , Animales , Secuencia de Bases , Densidad Ósea , Células de la Médula Ósea/citología , Células de la Médula Ósea/efectos de los fármacos , Células de la Médula Ósea/metabolismo , Bovinos , Diferenciación Celular , Regulación de la Expresión Génica , Factor Estimulante de Colonias de Macrófagos/farmacología , Ratones , Ratones Transgénicos , MicroARNs/agonistas , MicroARNs/antagonistas & inhibidores , MicroARNs/metabolismo , Datos de Secuencia Molecular , Monocitos/citología , Monocitos/efectos de los fármacos , Monocitos/metabolismo , Oligonucleótidos/genética , Oligonucleótidos/metabolismo , Osteoclastos/citología , Osteoclastos/efectos de los fármacos , Osteoporosis/metabolismo , Osteoporosis/patología , Fosfohidrolasa PTEN/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Ligando RANK/farmacología , Transducción de SeñalRESUMEN
Long noncoding RNAs (lncRNAs) are a class of RNA with 200 nucleotides or longer that are not translated into protein. lncRNAs are highly abundant; a study estimates that at least four times more lncRNAs are typically present than coding RNAs in humans. However, function of more than 95% of human lncRNAs are still unknown. Synthetic antisense oligonucleotides called gapmers are powerful tools for lncRNA loss-of-function studies. Gapmers contain a central DNA part, which activates RNase H-mediated RNA degradation, flanked by modified oligonucleotides, such as 2'-O-methyl RNA (2'OMe), 2'-O-methoxyethyl RNA (2'MOE), constrained ethyl nucleosides (cEt), and locked nucleic acids (LNAs). In contrast to siRNA or RNAi-based methods, antisense oligonucleotide gapmer-based knockdown is often more effective against nuclear-localized lncRNA targets, since RNase H is mainly localized in nuclei. As such, gapmers are also potentially a powerful tool for therapeutics targeting lncRNAs in various diseases, including cancer, cardiovascular diseases, lung fibrosis, and neurological/neuromuscular diseases. This chapter will discuss the development and applications of gapmers for lncRNA loss-of-function studies and tips to design effective antisense oligonucleotides.