Vascular smooth muscle cell-specific miRNA-214 deficiency alleviates simulated microgravity-induced vascular remodeling.

The human cardiovascular system has evolved to accommodate the gravity of Earth. Microgravity during spaceflight has been shown to induce vascular remodeling, leading to a decline in vascular function. The underlying mechanisms are not yet fully understood. Our previous study demonstrated that miR-214 plays a critical role in angiotensin II-induced vascular remodeling by reducing the levels of Smad7 and increasing the phosphorylation of Smad3. However, its role in vascular remodeling evoked by microgravity is not yet known. This study aimed to determine the contribution of miR-214 to the regulation of microgravity-induced vascular remodeling. The results of our study revealed that miR-214 expression was increased in the forebody arteries of both mice and monkeys after simulated microgravity treatment. In vitro, rotation-simulated microgravity-induced VSMC migration, hypertrophy, fibrosis, and inflammation were repressed by miR-214 knockout (KO) in VSMCs. Additionally, miR-214 KO increased the level of Smad7 and decreased the phosphorylation of Smad3, leading to a decrease in downstream gene expression. Furthermore, miR-214 cKO protected against simulated microgravity induced the decline in aorta function and the increase in stiffness. Histological analysis showed that miR-214 cKO inhibited the increases in vascular medial thickness that occurred after simulated microgravity treatment. Altogether, these results demonstrate that miR-214 has potential as a therapeutic target for the treatment of vascular remodeling caused by simulated microgravity.

Deep annotation of long noncoding RNAs by assembling RNA-seq and small RNA-seq data.

Long noncoding RNAs (lncRNAs) are increasingly being recognized as modulators in various biological processes. However, due to their low expression, their systematic characterization is difficult to determine. Here, we performed transcript annotation by a newly developed computational pipeline, termed RNA-seq and small RNA-seq combined strategy (RSCS), in a wide variety of cellular contexts. Thousands of high-confidence potential novel transcripts were identified by the RSCS, and the reliability of the transcriptome was verified by analysis of transcript structure, base composition, and sequence complexity. Evidenced by the length comparison, the frequency of the core promoter and the polyadenylation signal motifs, and the locations of transcription start and end sites, the transcripts appear to be full length. Furthermore, taking advantage of our strategy, we identified a large number of endogenous retrovirus-associated lncRNAs, and a novel endogenous retrovirus-lncRNA that was functionally involved in control of Yap1 expression and essential for early embryogenesis was identified. In summary, the RSCS can generate a more complete and precise transcriptome, and our findings greatly expanded the transcriptome annotation for the mammalian community.

Targeting E3 Ubiquitin Ligase WWP1 Prevents Cardiac Hypertrophy Through Destabilizing DVL2 via Inhibition of K27-Linked Ubiquitination.

BACKGROUND: Without adequate treatment, pathological cardiac hypertrophy induced by sustained pressure overload eventually leads to heart failure. WWP1 (WW domain-containing E3 ubiquitin protein ligase 1) is an important regulator of aging-related pathologies, including cancer and cardiovascular diseases. However, the role of WWP1 in pressure overload-induced cardiac remodeling and heart failure is yet to be determined. METHODS: To examine the correlation of WWP1 with hypertrophy, we analyzed WWP1 expression in patients with heart failure and mice subjected to transverse aortic constriction (TAC) by Western blotting and immunohistochemical staining. TAC surgery was performed on WWP1 knockout mice to assess the role of WWP1 in cardiac hypertrophy, heart function was examined by echocardiography, and related cellular and molecular markers were examined. Mass spectrometry and coimmunoprecipitation assays were conducted to identify the proteins that interacted with WWP1. Pulse-chase assay, ubiquitination assay, reporter gene assay, and an in vivo mouse model via AAV9 (adeno-associated virus serotype 9) were used to explore the mechanisms by which WWP1 regulates cardiac remodeling. AAV9 carrying cardiac troponin T (cTnT) promoter-driven small hairpin RNA targeting WWP1 (AAV9-cTnT-shWWP1) was administered to investigate its rescue role in TAC-induced cardiac dysfunction. RESULTS: The WWP1 level was significantly increased in the hypertrophic hearts from patients with heart failure and mice subjected to TAC. The results of echocardiography and histology demonstrated that WWP1 knockout protected the heart from TAC-induced hypertrophy. There was a direct interaction between WWP1 and DVL2 (disheveled segment polarity protein 2). DVL2 was stabilized by WWP1-mediated K27-linked polyubiquitination. The role of WWP1 in pressure overload-induced cardiac hypertrophy was mediated by the DVL2/CaMKII/HDAC4/MEF2C signaling pathway. Therapeutic targeting WWP1 almost abolished TAC induced heart dysfunction, suggesting WWP1 as a potential target for treating cardiac hypertrophy and failure. CONCLUSIONS: We identified WWP1 as a key therapeutic target for pressure overload induced cardiac remodeling. We also found a novel mechanism regulated by WWP1. WWP1 promotes atypical K27-linked ubiquitin multichain assembly on DVL2 and exacerbates cardiac hypertrophy by the DVL2/CaMKII/HDAC4/MEF2C pathway.

Vascular smooth muscle cell-specific miRNA-214 knockout inhibits angiotensin II-induced hypertension through upregulation of Smad7.

Vascular remodeling is a prominent trait during the development of hypertension, attributable to the phenotypic transition of vascular smooth muscle cells (VSMCs). Increasing studies demonstrate that microRNA plays an important role in this process. Here, we surprisingly found that smooth muscle cell-specific miR-214 knockout (miR-214 cKO) significantly alleviates angiotensin II (Ang II)-induced hypertension, which has the same effect as that of miR-214 global knockout mice in response to Ang II stimulation. Under the treatment of Ang II, miR-214 cKO mice exhibit substantially reduced systolic blood pressure. The vascular medial thickness and area in miR-214 cKO blood vessels were obviously reduced, the expression of collagen I and proinflammatory factors were also inhibited. VSMC-specific deletion of miR-214 blunts the response of blood vessels to the stimulation of endothelium-dependent and -independent vasorelaxation and phenylephrine and 5-HT induced vasocontraction. In vitro, Ang II-induced VSMC proliferation, migration, contraction, hypertrophy, and stiffness were all repressed with miR-214 KO in VSMC. To further explore the mechanism of miR-214 in the regulation of the VSMC function, it is very interesting to find that the TGF-ß signaling pathway is mostly enriched in miR-214 KO VSMC. Smad7, the potent negative regulator of the TGF-ß/Smad pathway, is identified to be the target of miR-214 in VSMC. By which, miR-214 KO sharply enhances Smad7 levels and decreases the phosphorylation of Smad3, and accordingly alleviates the downstream gene expression. Further, Ang II-induced hypertension and vascular dysfunction were reversed by antagomir-214. These results indicate that miR-214 in VSMC established a crosstalk between Ang II-induced AT1R signaling and TGF-ß induced TßRI /Smad signaling, by which it exerts a pivotal role in vascular remodeling and hypertension and imply that miR-214 has the potential as a therapeutic target for the treatment of hypertension.

3' untranslated region of Ckip-1 inhibits cardiac hypertrophy independently of its cognate protein.

AIMS: 3' untranslated region (3' UTR) of mRNA is more conserved than other non-coding sequences in vertebrate genomes, and its sequence space has substantially expanded during the evolution of higher organisms, which substantiates their significance in biological regulation. However, the independent role of 3' UTR in cardiovascular disease was largely unknown. METHODS AND RESULTS: Using bioinformatics, RNA fluorescent in situ hybridization and quantitative real-time polymerase chain reaction, we found that 3' UTR and coding sequence regions of Ckip-1 mRNA exhibited diverse expression and localization in cardiomyocytes. We generated cardiac-specific Ckip-1 3' UTR overexpression mice under wild type and casein kinase 2 interacting protein-1 (CKIP-1) knockout background. Cardiac remodelling was assessed by histological, echocardiography, and molecular analyses at 4 weeks after transverse aortic constriction (TAC) surgery. The results showed that cardiac Ckip-1 3' UTR significantly inhibited TAC-induced cardiac hypertrophy independent of CKIP-1 protein. To determine the mechanism of Ckip-1 3' UTR in cardiac hypertrophy, we performed transcriptome and metabolomics analyses, RNA immunoprecipitation, biotin-based RNA pull-down, and reporter gene assays. We found that Ckip-1 3' UTR promoted fatty acid metabolism through AMPK-PPARα-CPT1b axis, leading to its protection against pathological cardiac hypertrophy. Moreover, Ckip-1 3' UTR RNA therapy using adeno-associated virus obviously alleviates cardiac hypertrophy and improves heart function. CONCLUSIONS: These findings disclose that Ckip-1 3' UTR inhibits cardiac hypertrophy independently of its cognate protein. Ckip-1 3' UTR is an effective RNA-based therapy tool for treating cardiac hypertrophy and heart failure.

miR-214 stimulated by IL-17A regulates bone loss in patients with ankylosing spondylitis.

OBJECTIVE: Bone loss is common in AS, and miR-214 plays an important role in regulating bone formation. The aim of this study was to investigate the effect of miR-214, the production of which is stimulated by IL-17A, on bone loss in AS. METHODS: Peripheral blood was obtained from 32 patients with AS and 24 healthy controls. Levels of IL-17A, soluble RANK ligand (RANKL) and osteoprotegerin in serum were evaluated by ELISA, and the relative level of miR-214 in serum was detected by real-time quantitative PCR. In addition, we assessed the relationship between levels of miR-214, IL-17A and bone loss in primary murine osteoblasts and mouse bone marrow cells. RESULTS: The expression of RANKL and miR-214 in osteoblasts was increased following stimulation by IL-17A, and osteoblasts stimulated by IL-17A promoted the expression of miR-214 in osteoclasts and the activity of osteoclasts. We showed that osteoblast-derived miR-214 could be transferred to osteoclasts and could then regulate their activity. The levels of IL-17A and miR-214 were much higher in the serum of patients with AS than in that of healthy controls, and the relative level of miR-214 was positively correlated with the level of IL-17A in the serum and synovial fluid of the patients with AS, not healthy controls. The level of miR-214 in the serum of AS patients has potential diagnostic value. CONCLUSION: The production of miR-214 in osteoblasts is stimulated by IL-17A. It is an important inhibitor of bone formation in AS, and the serum level of miR-214 might be of potential diagnostic value for AS.

Hematopoietic stem cells and lineage cells undergo dynamic alterations under microgravity and recovery conditions.

Spaceflight leads to health risks including bone demineralization, skeletal muscle atrophy, cardiovascular dysfunction, and disorders of almost all physiologic systems. However, the impacts of microgravity on blood lineage cells and hematopoietic stem cells (HSCs) in vivo are largely unknown. In this study, we analyzed peripheral blood samples from 6 astronauts who had participated in spaceflight missions and found significant changes in several cell populations at different time points. These dynamic alterations of lineage cells and the role of HSCs were further studied in a mouse model, using hindlimb unloading (HU) to simulate microgravity. Large reductions in the frequency of NK cells, B cells, and erythrocyte precursors in the bone marrow of the HU mice were observed, together with an increased frequency of T cells, neutrophils, and HSCs. T cell levels recovered faster than those of B cells and erythrocyte precursors, whereas the recovery rates of NK cells and granulocytes were slow. In addition, competitive reconstitution experiments demonstrated the impaired function of HSCs, although these changes were reversible. Deep sequencing showed changes in the expression of regulatory molecules important for the differentiation of HSCs. This study provides the first determination of altered HSC function under simulated microgravity in vivo. The impairment of HSC function and differentiation provides an explanation for the immune disorders that occur under simulated microgravity. Thus, our findings demonstrated that spaceflight and simulated microgravity disrupt the homeostasis of immune system and cause dynamic alterations on both HSCs and lineage cells.-Cao, D., Song, J., Ling, S., Niu, S., Lu, L., Cui, Z., Li, Y., Hao, S., Zhong, G., Qi, Z., Sun, W., Yuan, X., Li, H., Zhao, D., Jin, X., Liu, C., Wu, X., Kan, G., Cao, H., Kang, Y., Yu, S., Li, Y. Hematopoietic stem cells and lineage cells undergo dynamic alterations under microgravity and recovery conditions.

[Mechanism of cardiac atrophy under weightlessness/simulated weightlessness].

Cardiac remodeling is the heart's response to external or internal stimuli. Weightlessness/simulated weightlessness leads to cardiac atrophy and heart function declining. Understanding the mechanism of cardiac atrophy under weightlessness is important to help astronaut recover from unloading-induced cardiovascular changes after spaceflight. Unloading-induced changes of hemodynamics, metabolic demands and neurohumoral regulation contribute to cardiac atrophy and function declining. During this process, Ca(2+)-related signaling, NF-κB signaling, ERK signaling, ubiquitin-proteasome pathway and autophagy are involved in weightlessness-induced cardiac atrophy. This article reviews the underlying mechanism of cardiac atrophy under weightlessness/simulated weightlessness.

Chronic treatment with ticagrelor limits myocardial infarct size: an adenosine and cyclooxygenase-2-dependent effect.

OBJECTIVE: In a phase III clinical trial (PLATelet inhibition and patient Outcomes, PLATO), ticagrelor provided better clinical outcomes than clopidogrel in patients with acute coronary syndromes. In addition to P2Y12-receptor antagonism, ticagrelor prevents cell uptake of adenosine and has proven able to augment adenosine effects. Adenosine protects the heart against ischemia-reperfusion injury. We compared the effects of clopidogrel and ticagrelor on myocardial infarct size (IS). APPROACH AND RESULTS: Rats received oral ticagrelor (0, 75, 150, or 300 mg/kg/d) or clopidogrel (30 or 90 mg/kg/d) for 7 days and underwent 30-minute coronary artery ligation and 24-hour reperfusion. Area at risk was assessed by blue dye and IS by 2,3,5-triphenyl-tetrazolium-chloride. Cyclooxygenase-2 (COX2) enzyme activity was assessed by ELISA and expression by real-time polymerase chain reaction. Mechanism responsible was explored using adenosine-receptor antagonist (CGS15943, an A2A/A1 antagonist) or cyclooxygenase inhibition by either aspirin (5, 10, or 25 mg/kg) or specific cyclooxygenase-1 (SC560) or COX2 (SC5815) inhibitors. Ticagrelor, dose-dependently, reduced IS, whereas clopidogrel had no effect. Adenosine-receptor antagonism blocked the ticagrelor effect and COX2 inhibition by SC5815, or high-dose aspirin attenuated the IS-limiting effect of ticagrelor, whereas cyclooxygenase-1 inhibition or low-dose aspirin had no effect. Ticagrelor, but not clopidogrel, upregulated COX2 expression and activity. Also this effect was blocked by adenosine-receptor antagonism. Ticagrelor, but not clopidogrel, increased Akt and endothelial nitric oxide synthase phosphorylation. CONCLUSIONS: Ticagrelor, but not clopidogrel, reduces myocardial IS. The protective effect of ticagrelor was dependent on adenosine-receptor activation with downstream upregulation of endothelial nitric oxide synthase and COX2 activity.

miR-214 promotes osteoclastogenesis by targeting Pten/PI3k/Akt pathway.

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.

PTEN upregulation may explain the development of insulin resistance and type 2 diabetes with high dose statins.

PURPOSE: Statins increase the incidence of new onset diabetes. Prolonged statin therapy upregulates PTEN expression. PTEN levels are also elevated in diabetic animals. Activation of protein kinase A by cAMP decreases PTEN expression. We assessed whether prolonged treatment with rosuvastatin (ROS) induces glucose intolerance by upregulating Phosphatase and Tensin Homologue on Chromosome 10 (PTEN) in mice receiving normal (ND) or Western Diet (WD) and whether concomitant treatment with cilostazol (CIL, a phosphodiesterase-3 inhibitor) attenuates the effects. METHODS: PTEN(loxp/cre) or PTEN(+/-) mice received ND or WD without or with ROS (10 mg/kg/day). Wild-type mice received ND or WD without or with ROS, CIL (10 mg/kg/day), or ROS+CIL for 30 days. Fasting insulin and glucose tolerance test were measured as well as PTEN and P-AKT levels in skeletal muscle. RESULTS: Serum glucose after intraperitoneal injection of glucose was higher in PTEN(loxp/cre) mice receiving WD or ROS and especially WD+ROS. Levels were lower in PTEN(+/-) mice compared to PTEN(loxp/cre) in each treatment group. CIL decreased glucose levels in mice receiving WD, ROS and their combination. Insulin levels were higher in the WD+ROS group. CIL decreased insulin in mice receiving WD+ROS. WD, ROS and especially their combination increased PTEN and decreased P-AKT levels. CIL attenuated the effect of WD, ROS and their combination. CONCLUSIONS: Long-term ROS can induce diabetes by upregulating PTEN. CIL attenuates these changes. Partial knockdown of PTEN also ameliorates ROS-induced insulin resistance. Further studies are needed to assess the effects of increasing cAMP levels to prevent the induction of diabetes by statins.

Ckip-1 3'UTR alleviates prolonged sleep deprivation induced cardiac dysfunction by activating CaMKK2/AMPK/cTNI pathway.

Sleep deprivation (SD) has emerged as a critical concern impacting human health, leading to significant damage to the cardiovascular system. However, the underlying mechanisms are still unclear, and the development of targeted drugs is lagging. Here, we used mice to explore the effects of prolonged SD on cardiac structure and function. Echocardiography analysis revealed that cardiac function was significantly decreased in mice after five weeks of SD. Real-time quantitative PCR (RT-q-PCR) and Masson staining analysis showed that cardiac remodeling marker gene Anp (atrial natriuretic peptide) and fibrosis were increased, Elisa assay of serum showed that the levels of creatine kinase (CK), creatine kinase-MB (CK-MB), ANP, brain natriuretic peptide (BNP) and cardiac troponin T (cTn-T) were increased after SD, suggesting that cardiac remodeling and injury occurred. Transcript sequencing analysis indicated that genes involved in the regulation of calcium signaling pathway, dilated cardiomyopathy, and cardiac muscle contraction were changed after SD. Accordingly, Western blotting analysis demonstrated that the cardiac-contraction associated CaMKK2/AMPK/cTNI pathway was inhibited. Since our preliminary research has confirmed the vital role of Casein Kinase-2 -Interacting Protein-1 (CKIP-1, also known as PLEKHO1) in cardiac remodeling regulation. Here, we found the levels of the 3' untranslated region of Ckip-1 (Ckip-1 3'UTR) decreased, while the coding sequence of Ckip-1 (Ckip-1 CDS) remained unchanged after SD. Significantly, adenovirus-mediated overexpression of Ckip-1 3'UTR alleviated SD-induced cardiac dysfunction and remodeling by activating CaMKK2/AMPK/cTNI pathway, which proposed the therapeutic potential of Ckip-1 3'UTR in treating SD-induced heart disease.

Modulation of microRNAs in hypertension-induced arterial remodeling through the ß1 and ß3-adrenoreceptor pathways.

BACKGROUND: Dysregulation of microRNAs (miRNAs) in arterial dysfunction and hypertension has not been extensively investigated yet. This project determined the effects of two anti-hypertensive ß1 adrenergic selective blockers on miRNA expression in the Dahl Salt Sensitive (DSS) hypertensive rat model. METHODS AND RESULTS: Microarray analysis showed that a set of miRNAs is differently expressed in the aorta of high salt (HS) treated rats with miR-320 increased and miR-26b and -21 decreased. All of these changes were reverted to normal by nebivolol (NEB, a ß1 selective-blocker and ß3 activator). The selective ß3-adrenoceptor antagonist S-(-)-cyanopindolol (Syc) counteracted the effect of NEB on these miRNAs. Atenolol (ATN, a pure ß1-blocker) combined with specific ß3 agonist BRL37344 restored the expression of all three miRNAs, similar to NEB, while ATN alone had only a partial effect on miR-320 expression. Computational analysis found Insulin Growth Factor-1 Receptor (IGF1R) as a putative target of miR-320, and Phosphatase and tensin homolog on chromosome ten (PTEN) as a putative target of miR-26b and -21. The targets were verified by luciferase reporter assays. Inhibition of miR-320 by an antisense inhibitor or NEB increased IGF1R expression, while miR-320 overexpression reversed the effect of NEB. Overexpression of miR-26b or -21 or NEB decreased PTEN levels, while inhibition of miR-26b or -21 attenuated the effect of NEB. HS diet induced downregulation of IGF1R and upregulation of PTEN in the aorta. NEB normalized the aberrant expression of IGF1R and PTEN and also improved the impairment of vascular AKT/eNOS signaling. Moreover, both NEB and ATN showed to have protective effects on salt-induced hypertension, oxidative stress, and vascular remodeling. NEB had a greater effect than ATN. CONCLUSIONS: Our data supports a differential miRNA expression profile in salt-induced hypertension. Manipulation of dysregulated miRNAs by ß-blockers may substantially induce alterations of gene expression and prevent arterial dysfunction and remodeling.

CKIP-1 inhibits cardiac hypertrophy by regulating class II histone deacetylase phosphorylation through recruiting PP2A.

BACKGROUND: Sustained cardiac pressure overload-induced hypertrophy and pathological remodeling frequently leads to heart failure. Casein kinase-2 interacting protein-1 (CKIP-1) has been identified to be an important regulator of cell proliferation, differentiation, and apoptosis. However, the physiological role of CKIP-1 in the heart is unknown. METHODS AND RESULTS: The results of echocardiography and histology demonstrate that CKIP-1-deficient mice exhibit spontaneous cardiac hypertrophy with aging and hypersensitivity to pressure overload-induced pathological cardiac hypertrophy, as well. Transgenic mice with cardiac-specific overexpression of CKIP-1 showed resistance to cardiac hypertrophy in response to pressure overload. The results of GST pull-down and coimmunoprecipitation assays showed the interaction between CKIP-1 and histone deacetylase 4 (HDAC4), through which they synergistically inhibited transcriptional activity of myocyte-specific enhancer factor 2C. By directly interacting with the catalytic subunit of phosphatase 2A, CKIP-1 overexpression enhanced the binding of catalytic subunit of phosphatase-2A to HDAC4 and promoted HDAC4 dephosphorylation. CONCLUSIONS: CKIP-1 was found to be an inhibitor of cardiac hypertrophy by upregulating the dephosphorylation of HDAC4 through the recruitment of protein phosphatase 2A. These results demonstrated a unique function of CKIP-1, by which it suppresses cardiac hypertrophy through its capacity to regulate HDAC4 dephosphorylation and fetal cardiac genes expression.

Phosphodiesterase-3 inhibition augments the myocardial infarct size-limiting effects of exenatide in mice with type 2 diabetes.

Glucagon-like peptide (GLP)-1 receptor activation increases intracellular cAMP with downstream activation of PKA. Cilostazol (CIL), a phosphodiesterase-3 inhibitor, prevents cAMP degradation. We assessed whether CIL amplifies the exenatide (EX)-induced increase in myocardial cAMP levels and PKA activity and augments the infarct size (IS)-limiting effects of EX in db/db mice. Mice fed a Western diet received oral CIL (10 mg/kg) or vehicle by oral gavage 24 h before surgery. One hour before surgery, mice received EX (1 µg/kg sc) or vehicle. Additional mice received H-89, a PKA inhibitor, alone or with CIL + EX. Mice underwent 30 min of coronary artery occlusion and 24 h of reperfusion. Both EX and CIL increased myocardial cAMP levels and PKA activity. Levels were significantly higher in the EX + CIL group. Both EX and CIL reduced IS. IS was the smallest in the CIL + EX group. H-89 completely blocked the IS-limiting effects of EX + CIL. EX + CIL decreased phosphatase and tensin homolog on chromosome 10 upregulation and increased Akt and ERK1/2 phosphorylation after ischemia-reperfusion. These effects were blocked by H-89. In conclusion, EX and CIL have additive effects on IS limitation in diabetic mice. The additive effects are related to cAMP-induced PKA activation, as H-89 blocked the protective effect of CIL + EX.

Dickkopf-1 (DKK1) phosphatase and tensin homolog on chromosome 10 (PTEN) crosstalk via microRNA interference in the diabetic heart.

Competitive endogenous RNAs (ceRNAs) regulate mRNA transcripts containing common microRNA (miRNA) recognition elements (MREs) through sequestration of shared miRNAs. Interactions of ceRNA have been demonstrated in cancerous cells. However, a paucity of information is available relative to the interactions of ceRNAs interaction in diabetes mellitus and the myocardium. The purpose of this study is to assess the potential role of DKK1 and PTEN in ceRNA regulation utilizing their common miRNAs in diabetic cardiomyocytes. The interactions' regulation between PTEN and DKK1 were determined in two diabetic models in vivo (streptozotocin-induced type-1 DM mice and db/db mice) and in vitro (human cardiomyocytes cells exposed to hyperglycemia). The levels of DKK1 and PTEN (mRNA and protein) were upregulated in parallel in all three diabetic models. DKK1 modulates PTEN protein levels in a miRNA and 3'UTR-dependent manner. RNAi-mediated DKK1 gene silencing resulted in a decreased PTEN expression and vice versa. The effect was blocked when Dicer was inhibited. Silencing either PTEN or DKK1 resulted in an increase of the availabilities of shared miRNAs. The silencing of either PTEN or DKKI resulted in a suppression end of the luciferase-PTEN 3'UTR activity. However, the over expression of DKK1 3'UTR or PTEN 3'UTR resulted in an increase in the activity. The attenuation of DKK1 increased AKT phosphorylation, improved glucose uptake and decreased apoptosis in HCMs exposed to hyperglycemia. The effects were blocked by PI3K inhibition. DKK1 and PTEN transcripts are co-upregulated in DM and hyperglycemia. DKK1 and PTEN serve as ceRNA, affecting the expression of each other via competition for miRNAs binding.

HuR-mediated nucleocytoplasmic translocation of HOTAIR relieves its inhibition of osteogenic differentiation and promotes bone formation.

Bone marrow mesenchymal stem cell (BMSC) osteogenic differentiation and osteoblast function play critical roles in bone formation, which is a highly regulated process. Long noncoding RNAs (lncRNAs) perform diverse functions in a variety of biological processes, including BMSC osteogenic differentiation. Although several studies have reported that HOX transcript antisense RNA (HOTAIR) is involved in BMSC osteogenic differentiation, its effect on bone formation in vivo remains unclear. Here, by constructing transgenic mice with BMSC (Prx1-HOTAIR)- and osteoblast (Bglap-HOTAIR)-specific overexpression of HOTAIR, we found that Prx1-HOTAIR and Bglap-HOTAIR transgenic mice show different bone phenotypes in vivo. Specifically, Prx1-HOTAIR mice showed delayed bone formation, while Bglap-HOTAIR mice showed increased bone formation. HOTAIR inhibits BMSC osteogenic differentiation but promotes osteoblast function in vitro. Furthermore, we identified that HOTAIR is mainly located in the nucleus of BMSCs and in the cytoplasm of osteoblasts. HOTAIR displays a nucleocytoplasmic translocation pattern during BMSC osteogenic differentiation. We first identified that the RNA-binding protein human antigen R (HuR) is responsible for HOTAIR nucleocytoplasmic translocation. HOTAIR is essential for osteoblast function, and cytoplasmic HOTAIR binds to miR-214 and acts as a ceRNA to increase Atf4 protein levels and osteoblast function. Bglap-HOTAIR mice, but not Prx1-HOTAIR mice, showed alleviation of bone loss induced by unloading. This study reveals the importance of temporal and spatial regulation of HOTAIR in BMSC osteogenic differentiation and bone formation, which provides new insights into precise regulation as a target for bone loss.

Effects of 60 days of 6° head-down bed rest on the composition and function of the human gut microbiota.

Spaceflight is rigorous and dangerous environment which can negatively affect astronauts' health and the entire mission. The 60 days of 6° head-down bed rest (HDBR) experiment provided us with an opportunity to trace the change of gut microbiota under simulated microgravity. The gut microbiota of volunteers was analyzed and characterized by 16S rRNA gene sequencing and metagenomic sequencing. Our results showed that the composition and function of the volunteers' gut microbiota were markedly was affected by 60 days of 6° HDBR. We further confirmed the species and diversity fluctuations. Resistance and virulence genes in the gut microbiota were also affected by 60 days of 6° HDBR, but the species attributions remained stable. The human gut microbiota affected by 60 days of 6° HDBR which was partially consistent with the effect of spaceflight, this implied that HDBR was a simulation of how spaceflight affects the human gut microbiota.

Simulated spaceflight-induced cardiac remodeling is modulated by gut microbial-derived trimethylamine N-oxide.

Spaceflight is physically demanding and can negatively affect astronauts' health. It has been shown that the human gut microbiota and cardiac function are affected by spaceflight and simulated spaceflight. This study investigated the effects of the gut microbiota on simulated spaceflight-induced cardiac remodeling using 10° of head-down bed rest (HDBR) in rhesus macaques and 30° of hindlimb unloading (HU) in mice. The gut microbiota, fecal metabolites, and cardiac remodeling were markedly affected by HDBR in macaques and HU in mice, cardiac remodeling in control mice was affected by the gut microbiota of HU mice and that of HU mice was protected by the gut microbiota of control mice, and there was a correlation between cardiac remodeling and the gut microbial-derived metabolite trimethylamine N-oxide. These findings suggest that spaceflight can affect cardiac remodeling by modulating the gut microbiota and fecal metabolites.

Mechanical stimulation controls osteoclast function through the regulation of Ca2+-activated Cl- channel Anoctamin 1.

Mechanical force loading is essential for maintaining bone homeostasis, and unloading exposure can lead to bone loss. Osteoclasts are the only bone resorbing cells and play a crucial role in bone remodeling. The molecular mechanisms underlying mechanical stimulation-induced changes in osteoclast function remain to be fully elucidated. Our previous research found Ca2+-activated Cl- channel Anoctamin 1 (Ano1) was an essential regulator for osteoclast function. Here, we report that Ano1 mediates osteoclast responses to mechanical stimulation. In vitro, osteoclast activities are obviously affected by mechanical stress, which is accompanied by the changes of Ano1 levels, intracellular Cl- concentration and Ca2+ downstream signaling. Ano1 knockout or calcium binding mutants blunts the response of osteoclast to mechanical stimulation. In vivo, Ano1 knockout in osteoclast blunts loading induced osteoclast inhibition and unloading induced bone loss and. These results demonstrate that Ano1 plays an important role in mechanical stimulation induced osteoclast activity changes.