Elevated miR-210 Research Articles

From animal models to patients: the role of placental microRNAs, miR-210, miR-126, and miR-148a/152 in preeclampsia.

Placental microRNAs (miRNAs) regulate the placental transcriptome and play a pathological role in preeclampsia (PE), a hypertensive disorder of pregnancy. Three PE rodent model studies explored the role of placental miRNAs, miR-210, miR-126, and miR-148/152 respectively, by examining expression of the miRNAs, their inducers, and potential gene targets. This review evaluates the role of miR-210, miR-126, and miR-148/152 in PE by comparing findings from the three rodent model studies with in vitro studies, other animal models, and preeclamptic patients to provide comprehensive insight into genetic components and pathological processes in the placenta contributing to PE. The majority of studies demonstrate miR-210 is upregulated in PE in part driven by HIF-1α and NF-κBp50, stimulated by hypoxia and/or immune-mediated processes. Elevated miR-210 may contribute to PE via inhibiting anti-inflammatory Th2-cytokines. Studies report an up- and downregulation of miR-126, arguably reflecting differences in expression between cell types and its multifunctional capacity. MiR-126 may play a pro-angiogenic role by mediating the PI3K-Akt pathway. Most studies report miR-148/152 family members are upregulated in PE. Evidence suggests they may inhibit DNA methylation of genes involved in metabolic and inflammatory pathways. Given the genetic heterogeneity of PE, it is unlikely that a single placental miRNA is a suitable therapeutic target for all patients. Investigating miRNAs in PE subtypes in patients and animal models may represent a more appropriate approach going forward. Developing methods for targeting placental miRNAs and specific placental cell types remains crucial for research seeking to target placental miRNAs as a novel treatment for PE.

Exosomes derived from cancer stem cells of gemcitabine-resistant pancreatic cancer cells enhance drug resistance by delivering miR-210

PurposeGemcitabine (GEM)-based chemotherapy is the first-line treatment for locally advanced pancreatic cancer. GEM resistance, however, remains a significant clinical challenge. Here, we investigated whether exosomes derived from GEM-resistant pancreatic cancer stem cells (CSCs) mediate cell-cell communication between cells that are sensitive or resistant to GEM and, by doing so, regulate drug resistance.MethodsGEM-sensitive BxPC-3-derived BxS and PANC-1 pancreatic cancer cells were cultured with exosomes extracted from CSCs isolated from GEM-resistant BxPC-3-derived BxR cells (BxR-CSC). The effect of exosomes on drug resistance, cell cycle progression, apoptosis and miRNA expression was evaluated in BxS and PANC-1 cells. Relevant miRNAs associated with GEM resistance were identified and the role of miR-210 in conferring drug resistance was examined in vitro and in vivo.ResultsBxR-CSC-derived exosomes induced GEM resistance, inhibited GEM-induced cell cycle arrest, antagonized GEM-induced apoptosis, and promoted tube formation and cell migration in BxS and PANC-1 cells. Elevated miR-210 expression levels were detected in BxR-CSCs and BxR-CSC-derived exosomes compared to those in BxS-CSCs and BxS-CSC-derived exosomes. In addition, increased expression levels of miR-210 were observed in BxS and PANC-1 cells cultured with BxR-CSC-derived exosomes upon exposure to GEM in a dose-dependent manner. Also, a series of biological changes was observed in BxS cells after transfection with miR-210 mimics, including activation of the mammalian target of rapamycin (mTOR) signaling pathway, and these changes were similar to those triggered by BxR-CSC-derived exosomes.ConclusionsOur findings suggest that exosomes derived from GEM-resistant pancreatic cancer stem cells mediate the horizontal transfer of drug-resistant traits to GEM-sensitive pancreatic cancer cells by delivering miR-210.

HIF-1α Stabilization Increases miR-210 Eliciting First Trimester Extravillous Trophoblast Mitochondrial Dysfunction.

Preeclampsia is associated with first trimester placental dysfunction. miR-210, a small non-coding RNA, is increased in the preeclamptic placenta. The effects of elevated miR-210 on placental function remain unclear. The objectives of this study were to identify targets of miR-210 in first trimester primary extravillous trophoblasts (EVTs) and to investigate functional pathways altered by elevated placental miR-210 during early pregnancy. EVTs isolated from first trimester placentas were exposed to cobalt chloride (CoCl2), a HIF-1α stabilizer and hypoxia mimetic, and miR-210 expression by qPCR, HIF1α protein levels by western blot and cell invasion were assessed. A custom TruSeq RNA array, including all known/predicted miR-210 targets, was run using miR-210 and miR-negative control transfected EVTs. Mitochondrial function was assessed by high resolution respirometry in transfected EVTs. EVTs exposed to CoCl2 showed a dose and time-dependent increase in miR-210 and HIF1α and reductions in cell invasion. The TruSeq array identified 49 altered genes in miR-210 transfected EVTs with 27 genes repressed and 22 enhanced. Three of the top six significantly repressed genes, NDUFA4, SDHD, and ISCU, are associated with mitochondrial function. miR-210 transfected EVTs had decreased maximal, complex II and complex I+II mitochondrial respiration. This study suggests that miR-210 alters first trimester trophoblast function. miR-210 overexpression alters EVT mitochondrial function in early pregnancy. Mitochondrial dysfunction may lead to increased reactive oxygen species, trophoblast cell damage and likely contributes to the pathogenesis of preeclampsia.

Salidroside protects hypoxia-induced injury by up-regulation of miR-210 in rat neural stem cells

Neonatal brain hypoxia is a disease that affects the nervous system in children. Salidroside is a compound that has an anti-hypoxic effect, but the mechanism of salidroside in neonatal cerebral hypoxia is unclear. Hence, we investigated the regulatory effect and mechanism of salidroside on hypoxic-induced injury of neural stem cells (NSCs). NSCs derived from embryo 14 Sprague-Dawley rats were treated by hypoxia, followed by the treatment of 0.8 mM salidroside. The expression levels of miR-210 and BTG3 in NSCs were altered by transfection. Cell viability and apoptosis were examined by CCK-8 and flow cytometry analysis. qRT-PCR and Western blot were performed to assess the expression changes of miR-210, BTG3, apoptosis-related factors and core factors in PI3K/AKT/mTOR pathway. We found that hypoxia induced an apoptosis-dependent death in NSCs. Salidroside exerted bFGF-like effect, as it alleviated hypoxia-induced viability impairment and apoptosis in NSCs. Further studies showed that hypoxia plus salidroside elevated miR-210 expression, and the protective actions of salidroside on hypoxia-modulated death in NSCs were attenuated by miR-210 suppression, while were enhanced by miR-210 overexpression. Besides, BTG3 was negatively regulated by miR-210. Overexpression of BTG3 inhibited the activation of PI3K/AKT/mTOR signaling pathway; of contrast, suppression of BTG3 promoted it. To conclude, this study provide in vitro evidence that salidroside protected NSCs against hypoxia-induced injury by up-regulation of miR-210, which in turn inhibited the expression of BTG3 and activated PI3K/AKT/mTOR signaling pathway.

MiR-210 and GPD1L regulate EDN2 in primary and immortalized human granulosa-lutein cells.

Endothelin-2 (EDN2), expressed at a narrow window during the periovulatory period, critically affects ovulation and corpus luteum (CL) formation. LH (acting mainly via cAMP) and hypoxia are implicated in CL formation; therefore, we aimed to elucidate how these signals regulate EDN2 using human primary (hGLCs) and immortalized (SVOG) granulosa-lutein cells. The hypoxiamiR, microRNA-210 (miR-210) was identified as a new essential player in EDN2 expression. Hypoxia (either mimetic compound-CoCl2, or low O2) elevated hypoxia-inducible factor 1A (HIF1A), miR-210 and EDN2 Hypoxia-induced miR-210 was suppressed in HIF1A-silenced SVOG cells, suggesting that miR-210 is HIF1A dependent. Elevated miR-210 levels in hypoxia or by miR-210 overexpression, increased EDN2 Conversely, miR-210 inhibition reduced EDN2 levels, even in the presence of CoCl2, indicating the importance of miR-210 in the hypoxic induction of EDN2 A molecule that destabilizes HIF1A protein, glycerol-3-phosphate dehydrogenase 1-like gene-GPD1L, was established as a miR-210 target in both cell types. It was decreased by miR-210-mimic and was increased by miR-inhibitor. Furthermore, reducing GPD1L by endogenously elevated miR-210 (in hypoxia), miR-210-mimic or by GPD1L siRNA resulted in elevated HIF1A protein and EDN2 levels, implying a vital role for GPD1L in the hypoxic induction of EDN2 Under normoxic conditions, forskolin (adenylyl cyclase activator) triggered changes typical of hypoxia. It elevated HIF1A, EDN2 and miR-210 while inhibiting GPD1L Furthermore, HIF1A silencing greatly reduced forskolin's ability to elevate EDN2 and miR-210. This study highlights the novel regulatory roles of miR-210 and its gene target, GPD1L, in hypoxia and cAMP-induced EDN2 by human granulosa-lutein cells.

Loading MiR-210 in Endothelial Progenitor Cells Derived Exosomes Boosts Their Beneficial Effects on Hypoxia/Reoxygeneation-Injured Human Endothelial Cells via Protecting Mitochondrial Function

Background/Aims: Stem cell-derived exosomes (EXs) offer protective effects on various cells via their carried microRNAs (miRs). Meanwhile, miR-210 has been shown to reduce mitochondrial reactive oxygen species (ROS) overproduction. In this study, we determined the potential effects of endothelial progenitor cell-derived EXs (EPC-EXs) on hypoxia/ reoxygenation (H/R) injured endothelial cells (ECs) and investigated whether these effects could be boosted by miR-210 loading. Methods: Human EPCs were used to generate EPC-EXs, or transfected with scrambler control or miR-210 mimics to generate EPC-EXs^sc and EPC-EXs^miR-210. H/R-injured human ECs were used as a model for functional analysis of EXs on apoptosis, viability, ROS production and angiogenic ability (migration and tube formation) by flow cytometry, MTT, dihydroethidium and angiogenesis assay kits, respectively. For mechanism analysis, the mitochondrion morphology, membrane potential (MMP), ATP level and the expression of fission/fusion proteins (dynamin-related protein 1: drp1 and mitofusin-2: mfn2) were assessed by using JC-1 staining, ELISA and western blot, respectively. Results: 1) Transfection of miR-210 mimics into EPCs induced increase of miR-210 in EPC-EXs^miR-210 without change of average size; 2) EPC-EXs^miR-210, but not EPC-EXs or EPC-EXs^sc, significantly elevated miR-210 level in ECs; 3) EPC-EXs^miR-210 were more effective than EPC-EXs and EPC-EXs^sc in reducing H/R-induced EC apoptosis, ROS overproduction and angiogenic dysfunction; 4) EPC-EXs decreased mitochondrial fragmentation, elevated MMP and ATP level, as well as improved mitochondrial mfn2 and drp1 dysregulation, which were more effective in EPC-EXs^miR-210. Conclusion: Our results suggest that EPC-EXs protect ECs against H/R injury via improving mitochondrial function and miR-210 enrichment could boost their effects.

MicroRNA-210 contributes to peripheral nerve regeneration through promoting the proliferation and migration of Schwann cells.

Peripheral nerve injury impacts the daily life of affected individuals. MicroRNA (miR)-210 is a multifunctional miR and has effects on the proliferation, migration and differentiation of cells. However, whether miR-210 has effects on peripheral nerve regeneration has remained elusive. In the present study, the miR-210 levels in a rat model of sciatic nerve injury were evaluated by reverse-transcription quantitative PCR and the effects of miR-210 on the proliferation and migration of Schwann cells were explored. Elevated miR-210 levels were discovered in the sciatic nerve injury rat model. miR-210 mimics were found to promote the proliferation and migration of Schwann cells, while miR-210 inhibitor was found to inhibit the proliferation and migration of Schwann cells. Further study showed that miR-210 had effects on the expression of growth-associated protein-43, myelin-associated glycoprotein and myelin basic protein. These results showed that miR-210 had effects on the proliferation and migration of Schwann cells and may be involved in the peripheral nerve regeneration.

Identification of genes dysregulated by elevation of microRNA-210 levels in human trophoblasts cell line, Swan 71.

Preeclampsia is a serious pregnancy disorder characterized by gestational hypertension and proteinuria. miR-210 is significantly overexpressed in the placentas of preeclampsia patients. Swan 71 cells, first-trimester human trophoblastic cell line, were transfected with hsa-miR-210-3p oligonucleotides by electroporation. Altered transcriptome was analyzed using microarray technique. Differentially expressed genes (DEGs) were clustered into Gene Ontology annotation biological processes. The extent of physical interaction between miR-210 and IGFBP3 mRNA was assessed via ribonucleoprotein immunoprecipitation. Microarray analysis showed 408 DEGs by elevated levels of miR-210 in Swan 71 cells. These genes were enriched in several biological processes involved in the pathogenesis of preeclampsia. IGFBP3, a gene associated with preeclampsia pathophysiology, was validated as a target gene of miR-210. We have demonstrated that elevated miR-210 levels in human trophoblast alter the expression profile of known preeclampsia-associated genes, and of gene targets involved in various biological processes essential to preeclampsia progression.

AntihypoxamiR functionalized gramicidin lipid nanoparticles rescue against ischemic memory improving cutaneous wound healing

Peripheral vasculopathies cause severe wound hypoxia inducing the hypoxamiR miR-210. High level of miR-210, persisting in wound-edge tissue as ischemic memory, suppresses oxidative metabolism and inhibits cell proliferation necessary for healing. In wound-edge tissue of chronic wound patients, elevated miR-210 was tightly associated with inhibition of epidermal cell proliferation as evident by lowered Ki67 immunoreactivity. To inhibit miR-210 in murine ischemic wound-edge tissue, we report the formulation of antihypoxamiR functionalized gramicidin lipid nanoparticles (AFGLN). A single intradermal delivery of AFGLN encapsulating LNA-conjugated anti-hypoximiR-210 (AFGLNmiR-210) lowered miR-210 level in the ischemic wound-edge tissue. In repTOP™mitoIRE mice, AFGLNmiR-210 rescued keratinocyte proliferation as visualized by in vivo imaging system (IVIS). 31P NMR studies showed elevated ATP content at the ischemic wound-edge tissue following AFGLNmiR-210 treatment indicating recovering bioenergetics necessary for healing. Consistently, AFGLNmiR-210 improved ischemic wound closure. The nanoparticle based approach reported herein is effective for miR-directed wound therapeutics warranting further translational development.

Overexpression of miR-210 is associated with SDH-related pheochromocytomas, paragangliomas, and gastrointestinal stromal tumours

miR-210 is a key regulator of response to hypoxia. Pheochromocytomas (PCs) and paragangliomas (PGLs) with germline SDHx or VHL mutations have pseudohypoxic gene expression signatures. We hypothesised that PC/PGLs containing SDHx or VHL mutations, and succinate dehydrogenase (SDH)-deficient gastrointestinal stromal tumours (GISTs), would overexpress miR-210 relative to non-SDH or -VHL-mutated counterparts. miR-210 was analysed by quantitative PCR in i) 39 PC/PGLs, according to genotype (one SDHA, five SDHB, seven VHL, three NF1, seven RET, 15 sporadic, one unknown) and pathology (18 benign, eight atypical, 11 malignant, two unknown); ii) 18 GISTs, according to SDHB immunoreactivity (nine SDH-deficient and nine SDH-proficient) and iii) two novel SDHB-mutant neurosphere cell lines. miR-210 was higher in SDHx- or VHL-mutated PC/PGLs (7.6-fold) compared with tumours without SDHx or VHL mutations (P=0.0016). miR-210 was higher in malignant than in unequivocally benign PC/PGLs (P=0.05), but significance was lost when benign and atypical tumours were combined (P=0.08). In multivariate analysis, elevated miR-210 was significantly associated with SDHx or VHL mutation, but not with malignancy. In GISTs, miR-210 was higher in SDH-deficient (median 2.58) compared with SDH-proficient tumours (median 0.60; P=0.0078). miR-210 was higher in patient-derived neurosphere cell lines containing SDHB mutations (6.5-fold increase) compared with normal controls, in normoxic conditions (P<0.01). Furthermore, siRNA-knockdown of SDHB in HEK293 cells increased miR-210 by 2.7-fold (P=0.001) under normoxia. Overall, our results suggest that SDH deficiency in PC, PGL and GISTs induces miR-210 expression and substantiates the role of aberrant hypoxic-type cellular responses in the development of these tumours.

Survival of hypoxic human mesenchymal stem cells is enhanced by a positive feedback loop involving miR-210 and hypoxia-inducible factor 1

The use of mesenchymal stem cells (MSCs) has emerged as a potential new treatment for myocardial infarction. However, the poor viability of MSCs after transplantation critically limits the efficacy of this new strategy. The expression of microRNA-210 (miR-210) is induced by hypoxia and is important for cell survival under hypoxic conditions. Hypoxia increases the levels of hypoxia inducible factor-1 (HIF-1) protein and miR-210 in human MSCs (hMSCs). miR-210 positively regulates HIF-1α activity. Furthermore, miR-210 expression is also induced by hypoxia through the regulation of HIF-1α. To investigate the effect of miR-210 on hMSC survival under hypoxic conditions, survival rates along with signaling related to cell survival were evaluated in hMSCs over-expressing miR-210 or ones that lacked HIF-1α expression. Elevated miR-210 expression increased survival rates along with Akt and ERK activity in hMSCs with hypoxia. These data demonstrated that a positive feedback loop involving miR-210 and HIF-1α was important for MSC survival under hypoxic conditions.

Ischemic Preconditioning Augments Survival of Stem Cells via miR-210 Expression by Targeting Caspase-8-associated Protein 2

MicroRNAs (miRs) participate in most cellular functions by posttranscriptional regulation of gene expression albeit with little information regarding their role in ischemic preconditioning (IP) of stem cells. We report that IP of bone marrow-derived mesenchymal stem cells (MSCs) with two cycles of 30-min ischemia/reoxygenation (I/R) supported their survival under subsequent longer exposure to anoxia and following engraftment in the infarcted heart. IP significantly reduced apoptosis in MSCs through activation of Akt (Ser(473)) and ERK1/2 (Thr(202)/Tyr(204)) and nuclear translocation of hypoxia-inducible factor-1alpha (HIF-1alpha). We observed concomitant induction of miR-210 in the preconditioned MSCs ((PC)MSCs). Inhibition of HIF-1alpha or of miR-210 abrogated the cytoprotective effects of preconditioning. Extrapolation of these data to in vivo studies in a rat model of acute myocardial infarction predominantly improved stem cell survival after engraftment with a role for miR-210. Notably, multiple I/R cycles more effectively regulated the miR-210 and hence promoted MSC survival compared with single-cycle hypoxia of an equal duration. Real time PCR array for rat apoptotic genes, computational target gene analyses, and luciferase reporter assay identified FLICE-associated huge protein (FLASH)/caspase-8-associated protein-2 (Casp8ap2) in (PC)MSCs as the target gene of miR-210. Induction of FLASH/CASP8AP2 in miR-210 knocked-down (PC)MSCs resulted in increased cell apoptosis. Taken together, these data demonstrated that cytoprotection afforded by IP was regulated by miR-210 induction via FLASH/Casp8ap2 suppression. These results highlighted that IP by multiple short episodes of I/R is a novel strategy to promote stem cell survival.