STING Agonist cGAMP Attenuates Sleep Deprivation-Induced Neuroinflammation and Cognitive Deficits via TREM2 Up-Regulation.

Sleep deprivation (SD) has been associated with several adverse effects, including cognitive deficit. Emerging evidence suggests microglia-associated neuroinflammation is a potential trigger of cognitive deficit after SD. Stimulator of interferon genes (STING) constitutes an important factor in host immune response to pathogenic organisms and is found in multiple cells, including microglia. STING is involved in neuroinflammation during neuronal degeneration, although how STING signaling affects SD-induced neuroinflammation remains unexplored. In the present study, the chronic sleep restriction (CSR) model was applied to examine the effects of STING signaling on cognition. The results revealed that cGAMP, a high-affinity and selective STING agonist, significantly improved cognitive deficit, alleviated neural injury, and relieved neuroinflammation in CSR mice by activating the STING-TBK1-IRF3 pathway. Moreover, triggering receptor expressed on myeloid cells 2 (TREM2) was upregulated in CSR mice treated with cGAMP, and this effect was abolished by STING knockout. TREM2 upregulation induced by cGAMP regulated the microglia from pro-inflammatory state to anti-inflammatory state, thereby relieving neuroinflammation in CSR mice. These findings indicate cGAMP-induced STING signaling activation alleviates SD-associated neuroinflammation and cognitive deficit by upregulating TREM2, providing a novel approach for the treatment of SD-related nerve injury.

Single-embryo transcriptomic atlas of oxygen response reveals the critical role of HIF-1α in prompting embryonic zygotic genome activation.

Adaptive response to physiological oxygen levels (physO2; 5% O2) enables embryonic survival in a low-oxygen developmental environment. However, the mechanism underlying the role of physO2 in supporting preimplantation development, remains elusive. Here, we systematically studied oxygen responses of hallmark events in preimplantation development. Focusing on impeded transcriptional upregulation under atmospheric oxygen levels (atmosO2; 20% O2) during the 2-cell stage, we functionally identified a novel role of HIF-1α in promoting major zygotic genome activation by serving as an oxygen-sensitive transcription factor. Moreover, during blastocyst formation, atmosO2 impeded H3K4me3 and H3K27me3 deposition by deregulating histone-lysine methyltransferases, thus impairing X-chromosome inactivation in blastocysts. In addition, we found atmosO2 impedes metabolic shift to glycolysis before blastocyst formation, thus resulting a low-level histone lactylation deposition. Notably, we also reported an increased sex-dimorphic oxygen response of embryos upon preimplantation development. Together, focusing on genetic and epigenetic events that are essential for embryonic survival and development, the present study advances current knowledge of embryonic adaptive responses to physO2, and provides novel insight into mechanism underlying irreversibly impaired developmental potential due to a short-term atmosO2 exposure.

METTL3 Modulates TXNIP Expression to Affect the Activation of NLRP3 Inflammasome in Hepatic Cells Under Oxygen-Glucose Deprivation/Reperfusion Injury.

Hepatic ischemia-reperfusion (I/R) injury is still a major risk factor and unsolved problem in hepatic surgery. Methyltransferase-like 3 (METTL3), an important m6A-modified methylase, regulates inflammation and cellular stress response. In this study, we demonstrated the special role of METTL3 and its underlying mechanism in hepatic I/R injury. In the mouse model of hepatic I/R and in the oxygen-glucose deprivation and reoxygenation (OGD/R)-induced AML12 and NCTC 1469 cells, the expression of METTL3 was significantly upregulated. Inhibition of METTL3 in OGD/R-induced AML12 and NCTC 1469 cells both increased the cell viability, declined the cell apoptosis, and decreased the reactive oxygen species (ROS) and the release levels of interleukin-1ß (IL-1ß) and interleukin-18 (IL-18), diminishing NLRP3 and Caspase1-p20 expressions. Moreover, METTL3 positively modulated TXNIP expression in an m6A manner. TXNIP overexpression reversed the effects of METTL3 knockdown on OGD/R-induced injury in AML12 cells. Furthermore, inhibition of NLRP3 inflammasome activity contributed to the protective effects of TXNIP knockdown in OGD/R-induced AML12 cells. In conclusion, METTL3 knockdown alleviated OGD/R-induced hepatocyte injury, and the specific mechanism was associated with the inhibition of NLRP3 inflammasome activation, which was attributed to the reduction of TXNIP in an m6A-dependent manner.

Progesterone Activates the Histone Lactylation-Hif1α-glycolysis Feedback Loop to Promote Decidualization.

Decidualization is a progesterone-dependent cellular differentiation process that is essential for establishing pregnancy. Robust activation of glycolysis and lactate synthesis during decidualization is remarkable, but their developmental functions remain largely unknown. Herein, we identify that endometrial lactate production plays a critical role in establishing local histone lactylation, a newly identified histone modification, and is important for ensuring normal decidualization. Enhanced endometrial glycolysis is the hallmark metabolic change and is tightly coupled with H4K12la during decidualization. Inhibition of histone lactylation impaired decidualization, in either physiological conception or in vivo and in vitro induced decidualization models. Mechanistic study based on CUT&Tag and ATAC-seq revealed that a transcriptional factor hypoxia-inducible factor 1 α (Hif1α) is the critical regulatory target of H4K12la, and in turn forms an H4K12la-Hif1α-glycolysis feedback loop to drive decidualization. Moreover, we demonstrate that the loop is directly activated by progesterone during decidualization. Our study not only advances the current knowledge of the role of lactate in regulating uterine function, but also establishes a novel functional link among the major endocrine factors, endometrial metabolic change, and epigenetic modification during endometrial remodeling. These findings present valuable clues to develop clinical intervention strategies to improve pregnancy outcomes following both natural conception and assisted reproduction.

WNT Co-Receptor LRP6 Is Critical for Zygotic Genome Activation and Embryonic Developmental Potential by Interacting with Oviductal Paracrine Ligand WNT2.

Mammalian preimplantation development depends on the interaction between embryonic autocrine and maternal paracrine signaling. Despite the robust independence of preimplantation embryos, oviductal factors are thought to be critical to pregnancy success. However, how oviductal factors regulate embryonic development and the underlying mechanism remain unknown. In the present study, focusing on WNT signaling, which has been reported to be essential for developmental reprogramming after fertilization, we analyzed the receptor-ligand repertoire of preimplantation embryonic WNT signaling, and identified that the WNT co-receptor LRP6 is necessary for early cleavage and has a prolonged effect on preimplantation development. LRP6 inhibition significantly impeded zygotic genome activation and disrupted relevant epigenetic reprogramming. Focusing on the potential oviductal WNT ligands, we found WNT2 as the candidate interacting with embryonic LRP6. More importantly, we found that WNT2 supplementation in culture medium significantly promoted zygotic genome activation (ZGA) and improved blastocyst formation and quality following in vitro fertilization (IVF). In addition, WNT2 supplementation significantly improved implantation rate and pregnancy outcomes following embryo transfer. Collectively, our findings not only provide novel insight into how maternal factors regulate preimplantation development through maternal-embryonic communication, but they also propose a promising strategy for improving current IVF systems.

Bending Resistance Covalent Organic Framework Superlattice: "Nano-Hourglass"-Induced Charge Accumulation for Flexible In-Plane Micro-Supercapacitors.

Covalent organic framework (COF) film with highly exposed active sites is considered as the promising flexible self-supported electrode for in-plane micro-supercapacitor (MSC). Superlattice configuration assembled alternately by different nanofilms based on van der Waals force can integrate the advantages of each isolated layer to exhibit unexpected performances as MSC film electrodes, which may be a novel option to ensure energy output. Herein, a mesoporous free-standing A-COF nanofilm (pore size is 3.9 nm, averaged thickness is 4.1 nm) with imine bond linkage and a microporous B-COF nanofilm (pore size is 1.5 nm, averaged thickness is 9.3 nm) with ß-keto-enamine-linkages are prepared, and for the first time, we assembly the two lattice matching films into sandwich-type superlattices via layer-by-layer transfer, in which ABA-COF superlattice stacking into a "nano-hourglass" steric configuration that can accelerate the dynamic charge transportation/accumulation and promote the sufficient redox reactions to energy storage. The fabricated flexible MSC-ABA-COF exhibits the highest intrinsic CV of 927.9 F cm-3 at 10 mV s-1 than reported two-dimensional alloy, graphite-like carbon and undoped COF-based MSC devices so far, and shows a bending-resistant energy density of 63.2 mWh cm-3 even after high-angle and repeat arbitrary bending from 0 to 180°. This work provides a feasible way to meet the demand for future miniaturization and wearable electronics.

Modulating the Band Structure of Metal Coordinated Salen COFs and an In Situ Constructed Charge Transfer Heterostructure for Electrocatalysis Hydrogen Evolution.

A series of crystalline, stable Metal (Metal = Zn, Cu, Ni, Co, Fe, and Mn)-Salen covalent organic framework (COF)EDA complex are prepared to continuously tune the band structure of Metal-Salen COFEDA , with the purpose of optimizing the free energy intermediate species during the hydrogen evolution reaction (HER) process. The conductive macromolecular poly(3,4-ethylenedioxythiophene) (PEDOT) is subsequently integrated into the one-dimensional (1D) channel arrays of Metal-Salen COFEDA to form heterostructure PEDOT@Metal-Salen COFEDA via the in situ solid-state polymerization method. Among the Metal-Salen COFEDA and PEDOT@Metal-Salen COFEDA complexes, the optimized PEDOT@Mn-Salen COFEDA displays prominent electrochemical activity with an overpotential of 150 mV and a Tafel slope of 43 mV dec-1 . The experimental results and density of states data show that the continuous energy band structure modulation in Metal-Salen COFEDA has the ability to make the metal d-orbital interact better with the s-orbital of H, which is conducive to electron transport in the HER process. Moreover, the calculated charge density difference indicates that the heterostructures composed of PEDOT and Metal-Salen COFEDA induce an intramolecular charge transfer and construct highly active interfacial sites.

Vitamin C Rescues in vitro Embryonic Development by Correcting Impaired Active DNA Demethylation.

During preimplantation development, a wave of genome-wide DNA demethylation occurs to acquire a hypomethylated genome of the blastocyst. As an essential epigenomic event, postfertilization DNA demethylation is critical to establish full developmental potential. Despite its importance, this process is prone to be disrupted due to environmental perturbations such as manipulation and culture of embryos during in vitro fertilization (IVF), and thus leading to epigenetic errors. However, since the first case of aberrant DNA demethylation reported in IVF embryos, its underlying mechanism remains unclear and the strategy for correcting this error remains unavailable in the past decade. Thus, understanding the mechanism responsible for DNA demethylation defects, may provide a potential approach for preventing or correcting IVF-associated complications. Herein, using mouse and bovine IVF embryos as the model, we reported that ten-eleven translocation (TET)-mediated active DNA demethylation, an important contributor to the postfertilization epigenome reprogramming, was impaired throughout preimplantation development. Focusing on modulation of TET dioxygenases, we found vitamin C and α-ketoglutarate, the well-established important co-factors for stimulating TET enzymatic activity, were synthesized in both embryos and the oviduct during preimplantation development. Accordingly, impaired active DNA demethylation can be corrected by incubation of IVF embryos with vitamin C, and thus improving their lineage differentiation and developmental potential. Together, our data not only provides a promising approach for preventing or correcting IVF-associated epigenetic errors, but also highlights the critical role of small molecules or metabolites from maternal paracrine in finetuning embryonic epigenomic reprogramming during early development.

MiR-513b-5p represses autophagy during the malignant progression of hepatocellular carcinoma by targeting PIK3R3.

Hepatocellular carcinoma (HCC) serves as a prevailing global malignancy with severe mortality and extremely unsatisfactory prognosis, in which autophagy is a fundamental process in liver cancer pathogenesis, but the mechanisms are poorly understood. MicroRNAs (miRNAs) serve as a type of well-recognized non-coding regulators and contribute to the modulation of liver cancer development, from the aspects of diagnosis, progression, and therapy. Here, we aimed to investigate the function of hsa_microRNA-513b-5p (miR-513b-5p) in regulating autophagy during HCC progression. Specifically, our data showed that miR-513b-5p mimic reduced the LC3-II and beclin1 expression but enhanced p62 expression in HCC cells. MiR-513b-5p repressed liver cancer cell proliferation, migration/invasion, and induced apoptosis in vitro. Crucially, miR-513b-5p attenuated tumor growth of liver cancer cells in vivo. In the mechanical investigation, we identified that PIK3R3 mRNA 3'UTR was targeted by miR-513b-5p and miR-513b-5p suppressed PIK3R3 expression. PIK3R3 overexpression partly reversed miR-513b-5p-mediated autophagy, proliferation, and apoptosis of liver cancer cells. Consequently, we concluded that miR-513b-5p repressed autophagy during the malignant progression of HCC by targeting PIK3R3. MiR-513b-5p may be applied as a therapeutic target for HCC.

The mRNA-destabilizing protein Tristetraprolin targets "meiosis arrester" Nppc mRNA in mammalian preovulatory follicles.

C-natriuretic peptide (CNP) and its receptor guanylyl cyclase, natriuretic peptide receptor 2 (NPR2), are key regulators of cyclic guanosine monophosphate (cGMP) homeostasis. The CNP-NPR2-cGMP signaling cascade plays an important role in the progression of oocyte meiosis, which is essential for fertility in female mammals. In preovulatory ovarian follicles, the luteinizing hormone (LH)-induced decrease in CNP and its encoding messenger RNA (mRNA) natriuretic peptide precursor C (Nppc) are a prerequisite for oocyte meiotic resumption. However, it has never been determined how LH decreases CNP/Nppc In the present study, we identified that tristetraprolin (TTP), also known as zinc finger protein 36 (ZFP36), a ubiquitously expressed mRNA-destabilizing protein, is the critical mechanism that underlies the LH-induced decrease in Nppc mRNA. Zfp36 mRNA was transiently up-regulated in mural granulosa cells (MGCs) in response to the LH surge. Loss- and gain-of-function analyses indicated that TTP is required for Nppc mRNA degradation in preovulatory MGCs by targeting the rare noncanonical AU-rich element harbored in the Nppc 3' UTR. Moreover, MGC-specific knockout of Zfp36, as well as lentivirus-mediated knockdown in vivo, impaired the LH/hCG-induced Nppc mRNA decline and oocyte meiotic resumption. Furthermore, we found that LH/hCG activates Zfp36/TTP expression through the EGFR-ERK1/2-dependent pathway. Our findings reveal a functional role of TTP-induced mRNA degradation, a global posttranscriptional regulation mechanism, in orchestrating the progression of oocyte meiosis. We also provided a mechanism for understanding CNP-dependent cGMP homeostasis in diverse cellular processes.

Mitochondrial transfer from induced pluripotent stem cells rescues developmental potential of in vitro fertilized embryos from aging females†.

Conventional heterologous mitochondrial replacement therapy is clinically complicated by "tri-parental" ethical concerns and limited source of healthy donor oocytes or zygotes. Autologous mitochondrial transfer is a promising alternative in rescuing poor oocyte quality and impaired embryo developmental potential associated with mitochondrial disorders, including aging. However, the efficacy and safety of mitochondrial transfer from somatic cells remains largely controversial, and unsatisfying outcomes may be due to distinct mitochondrial state in somatic cells from that in oocytes. Here, we propose a potential strategy for improving in vitro fertilization (IVF) outcomes of aging female patients via mitochondrial transfer from induced pluripotent stem (iPS) cells. Using naturally aging mice and well-established cell lines as models, we found iPS cells and oocytes share similar mitochondrial morphology and functions, whereas the mitochondrial state in differentiated somatic cells is substantially different. By microinjection of isolated mitochondria into fertilized oocytes following IVF, our results indicate that mitochondrial transfer from iPS, but not MEF cells, can rescue the impaired developmental potential of embryos from aging female mice and obtain an enhanced implantation rate following embryo transfer. The beneficial effect may be explained by the fact that mitochondrial transfer from iPS cells not only compensates for aging-associated loss of mtDNA, but also rescues mitochondrial metabolism of subsequent preimplantation embryos. Using mitochondria from iPS cells as the donor, our study not only proposes a promising strategy for improving IVF outcomes of aging females, but also highlights the importance of synchronous mitochondrial state in supporting embryo developmental potential.

Downregulation of microRNA-302b-3p relieves oxygen-glucose deprivation/re-oxygenation induced injury in murine hippocampal neurons through up-regulating Nrf2 signaling by targeting fibroblast growth factor 15/19.

MicroRNAs have emerged as critical mediators of cerebral ischaemia/reperfusion injury. Recent studies have demonstrated that microRNA-302b-3p (miR-302b-3p) plays an important role in regulating apoptosis and oxidative stress in various cells. However, whether miR-302b-3p is involved in regulating cerebral ischaemia/reperfusion injury-induced neuronal apoptosis and oxidative stress remains unknown. In the present study, we explored the potential function and molecular mechanism of miR-302b-3p in oxygen-glucose deprivation/re-oxygenation (OGD/R)-induced neuronal injury, using an in vitro model of cerebral ischaemia/reperfusion injury. We found that miR-302b-3p expression was up-regulated by OGD/R treatment in neurons. The inhibition of miR-302b-3p improved cell viability, and reduced apoptosis and the production of reactive oxygen species, showing a protective effect against OGD/R-induced injury. Interestingly, miR-302b-3p was shown to target and modulate murine fibroblast growth factor 15 (FGF15). Moreover, our results showed that miR-302b-3p down-regulation contributed to the promotion of nuclear factor erythroid 2-related factor 2 (Nrf2)/antioxidant response element (ARE)-mediated antioxidant signaling associated with the inactivation of glycogen synthase kinase-3ß. However, the knockdown of FGF15 significantly reversed the miR-302b-3p inhibition-mediated protective effect in OGD/R-treated neurons. Overall, these results demonstrated that miR-302b-3p inhibition confers a neuroprotective effect in OGD/R-treated neurons by up-regulating Nrf2/ARE antioxidant signaling via targeting FGF15, providing a novel target for neuroprotection in cerebral ischaemia/reperfusion injury.

LncRNA MALAT1 cessation antagonizes hypoxia/reoxygenation injury in hepatocytes by inhibiting apoptosis and inflammation via the HMGB1-TLR4 axis.

Hepatic ischemia-reperfusion (I/R) injury frequently occurs after liver transplantation, stroke, and trauma, resulting in organ dysfunction and failure. Hepatocyte apoptosis and inflammation are identified as the hallmarks of liver I/R injury. Long non-coding RNA (lncRNA) metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) is induced following hypoxia or ischemic stimulation, and exerts the contradictory roles in various injury progression. However, its role and mechanism lying beneath hepatic I/R remains ill defined. In this study, elevation of MALAT1 expression was corroborated in human hepatocytes under hypoxia/reoxygenation (H/R)H/R condition. Of interest, depression of MALAT1 blunted H/R-inhibited cell viability, and counteracted lactate dehydrogenase (LDH) and malondialdehyde release. Additionally, MALAT1 cessation antagonized H/R-evoked cell apoptosis and caspase-3 activity. Simultaneously, the increased inflammatory reaction triggered by H/R stimulation was also abrogated following MALAT1 suppression by reducing pro-inflammatory cytokine transcripts and productions including IL-1ß and TNF-α. Mechanistically, H/R exposure activated the pathway of high-mobility group box1 (HMGB1)-TLR4, which was muted after MALAT1 inhibition. More importantly, elevation of HMGB1 reversed MALAT1 down-regulation-mediated inhibition in cell injury and inflammation. Moreover, blocking the TLR4 signaling also ameliorated H/R-evoked hepatocyte apoptosis and inflammatory response. Consequently, these data suggest that MALAT1 may aggravate hepatic I/R injury by regulating the HMGB1-TLR4-triggered cell apoptosis and inflammation, implying a promising therapeutic strategy to fight liver I/R injury.

MicroRNA-449b-5p targets HMGB1 to attenuate hepatocyte injury in liver ischemia and reperfusion.

MicroRNAs (miRNAs) participate in the pathological process of liver ischemia/reperfusion (I/R) injury. MiR-449b-5p is the target miRNA of high mobility group box 1 (HMGB1). Its role and molecular mechanism in liver I/R injury remain unidentified. In this study, we found a protective effect of miR-449b-5p against hepatic I/R injury. HMGB1 expression significantly increased, whereas miR-449b-5p dramatically decreased in patients after liver transplant and in L02 cells exposed to hypoxia/reoxygenation (H/R). A dual-luciferase reporter assay confirmed the direct interaction between miR-449b-5p and the 3' untranslated region of HMGB1 messenger RNA. We also found that overexpression of miR-449b-5p significantly promoted cell viability and inhibited cell apoptosis of L02 cells exposed to H/R. Moreover, miR-449b-5p repressed HMGB1 protein expression and nuclear factor-κB (NF-κB) pathway activation in these L02 cells. In an in vivo rat model of hepatic I/R injury, overexpression of miR-449b-5p significantly decreased alanine aminotransferase and aspartate aminotransferase and inhibited the HMGB1/NF-κB pathway. Our study thus suggests that miR-449b-5p alleviated hepatic I/R injury by targeting HMGB1 and deactivating the NF-κB pathway, which may provide a novel and promising therapeutic target for hepatic I/R injury.

Melatonin protects in vitro matured porcine oocytes from toxicity of Aflatoxin B1.

Aflatoxin B1 (AFB1) is a major food and feed contaminant that threaten public health. Previous studies indicate that AFB1 exposure disrupted oocyte maturation. However, an effective and feasible method is unavailable for protecting oocytes against toxicity of AFB1. In the present study, using in vitro matured porcine oocytes and parthenogenetic embryos as model, we confirmed that AFB1 exposure during in vitro oocyte maturation (IVM) significantly impaired both nuclear and cytoplasmic maturation in a dose- and time-dependent manner. The different concentrations of melatonin were also tested for their protective effects on oocytes against the AFB1-induced toxicity. Our results showed that supplementation of a relative high concentration of melatonin (10-3 mol/L) during IVM efficiently reversed the impaired development rate and blastocyst quality, to the levels comparable to those of the control group. Further analysis indicated that melatonin application efficiently alleviated reactive oxygen species accumulation and initiation of apoptosis induced by AFB1 exposure. In addition, disrupted GSH/GPX system, as well as inhibited mitochondrial DNA (mtDNA) replication and mitochondrial biogenesis in AFB1-treated oocytes, can be notably reversed by melatonin application. Furthermore, cumulus cells may be important in mediating the toxicity of AFB1 to oocytes, and the metabolism of AFB1 in cumulus cells can be depressed by melatonin. To the best of our knowledge, this is the first report to confirm that melatonin application can efficiently protect oocytes from AFB1-induced toxicity. Our study provides a promising and practical strategy for alleviating or reversing AFB1-induced female reproductive toxicity in both clinical treatment and domestic reproductive management.

Chlorogenic acid against palmitic acid in endoplasmic reticulum stress-mediated apoptosis resulting in protective effect of primary rat hepatocytes.

BACKGROUND: We demonstrated growing evidence supports a protective role of chlorogenic acid of rat hepatocytes elicited by two compounds, i.e. thapsigargin and palmitic acid. Nevertheless, little is known about the mechanisms of palmitic acid induced endoplasmic reticulum (ER) stress and cell death. METHODS: The proliferation of primary rat hepatocytes was detected by MTT assay. The expression of GRP78, CHOP and GRP94 was detected by Western blot analyses. Caspase-3 activity was detected by a Caspase-3 substrate kit. Cell apoptosis was detected by Hoechst 33342 staining. RESULTS: We demonstrated that incubation of hepatocytes for 16 h with palmitic acid elevated cell death. Moreover, Western blot analyses demonstrated increased levels of the endoplasmic reticulum stress markers - glucose regulated protein 78 (GRP78), C/EBP homologous protein (CHOP), and glucose regulated protein 94 (GRP94). Chlorogenic acid could inhibit ER stress induced cell death and levels of indicators of ER stress caused by palmitic acid. The effect of thapsigargin, which evokes ER stress were reversed by chlorogenic acid. CONCLUSIONS: Altogether, our data indicate that in primary rat hepatocytes, chlorogenic acid prevents ER stress-mediated apoptosis of palmitic acid.

Fentanyl inhibits proliferation and invasion via enhancing miR-302b expression in esophageal squamous cell carcinoma.

Fentanyl is one of the most commonly used intravenous anesthetic agents during cancer resection surgery, but the effect of fentanyl on esophageal squamous cell carcinoma (ESCC) remains unclear. The aim of the present study was to investigate the involvement of microRNA 302b (miR-302b) in the anti-proliferation and anti-invasion effects of fentanyl in ESCC. In the present study, the effects of fentanyl on cell proliferation, apoptosis and invasion were detected using MTT assays, flow cytometry and Transwell assays in ESCC Eca109 and TE1 cell lines. Subsequently, expression of miR-302b was determined using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). RT-qPCR and western blot analysis were performed in order to evaluate the expression of ErbB4, a target of miR-302b. Furthermore, anti-miR were used to inhibit miR-302b in fentanyl-treated ESCC cells in order to evaluate the role of miR-302b in the effect of fentanyl on malignant behaviors. Fentanyl inhibited the proliferation of Eca109 and TE1 cells in a dose- and time-dependent manner. Following exposure to fentanyl for 48 h, Eca109 and TE1 cells exhibited increased apoptosis and decreased invasion. Furthermore, fentanyl upregulated miR-302b expression, but downregulated ErbB4 expression. Finally, loss of miR-302b using the anti-miR technique reversed the effect of fentanyl on cell proliferation, apoptosis and invasion in the two ESCC cell lines. Taken together, the results of the present study indicated that fentanyl inhibits the proliferation and invasion of ESCC cells through upregulation of miR-302b.

TXNIP knockdown alleviates hepatocyte ischemia reperfusion injury through preventing p38/JNK pathway activation.

Hepatic ischemia and reperfusion (I/R) injury is a major cause of liver damage during liver transplantation, resection surgery, shock, and trauma. It has been reported that TXNIP expression was upregulated in a rat model of hepatic I/R injury. However, the role of TXNIP in the hepatic I/R injury is little known. In our study, we investigated the biological role of TXNIP and its potential molecular mechanism in the human hepatic cell line (HL7702 cells). Using oxygen-glucose deprivation and reoxygenation (OGD/R) to create a cell model of hepatic I/R injury, we found that the mRNA and protein expression levels of TXNIP were upregulated in HL7702 cells exposed to OGD/R. TXNIP overexpression remarkably promoted OGD/R-induced cell apoptosis and lactate dehydrogenase (LDH) release, both of which were significantly decreased by TXNIP knockdown. The production of malondialdehyde (MDA) was also increased by TXNIP overexpression, but was reduced by TXNIP knockdown. Moreover, TXNIP overexpression significantly upregulated the phosphorylation of p38 and JNK, which was remarkably inhibited by TXNIP knockdown. Additionally, p38-specific inhibitor SB203580 abrogated the effect of TXNIP overexpression on OGD/R-induced cell injury. Taken together, these results indicated that TXNIP knockdown alleviated hepatocyte I/R injury through preventing p38/JNK pathway activation. Thus, TXNIP might offer a novel potential therapeutic target for the treatment of hepatic I/R injury.

Down-regulation of microRNA-142-5p attenuates oxygen-glucose deprivation and reoxygenation-induced neuron injury through up-regulating Nrf2/ARE signaling pathway.

MicroRNAs (miRNAs) play vital roles in regulating neuron survival during cerebral ischemia/reperfusion injury. miR-142-5p is reported to be an important regulator of cellular survival. However, little is known about the role of miR-142-5p in regulating neuron survival during cerebral ischemia/reperfusion injury. In this study, we aimed to investigate the precise function and mechanism of miR-142-5p in the regulation of neuron ischemia/reperfusion injury using a cellular model of oxygen-glucose deprivation and reoxygenation (OGD/R)-induced injury in hippocampal neurons in vitro. We found that miR-142-5p was induced in hippocampal neurons with OGD/R treatment. The inhibition of miR-142-5p attenuated OGD/R-induced cell injury and oxidative stress, whereas the overexpression of miR-142-5p aggravated them. Nuclear factor erythroid 2-related factor 2 (Nrf2) was identified as a target gene of miR-142-5p. Moreover, miR-142-5p regulated Nrf2 expression and downstream signaling. Knockdown of Nrf2 abolished the protective effects of miR-142-5p suppression. In addition, we showed an inverse correlation relationship between miR-142-5p and Nrf2 in an in vivo model of middle cerebral artery occlusion in rats. Taken together, these results suggest that miR-142-5p contributes to OGD/R-induced cell injury and the down-regulation of miR-142-5p attenuates OGD/R-induced neuron injury through promoting Nrf2 expression. Our study provides a novel insight into understanding the molecular pathogenesis of cerebral ischemia/reperfusion injury and indicates a potential therapeutic target for the treatment of cerebral ischemia/reperfusion injury.

Combined over-expression of the hypoxia-inducible factor 2α gene and its long non-coding RNA predicts unfavorable prognosis of patients with osteosarcoma.

BACKGROUND: LncRNA hypoxia-inducible factor-2α (HIF-2α) promoter upstream transcript (HIF2PUT) functions as a novel regulatory factor of osteosarcoma stem cells partly by controlling HIF-2α expression. The aim of this study was to investigate the clinical significance of HIF-2α and HIF2PUT in human osteosarcoma. MATERIALS AND METHODS: Quantitative real-time PCR was performed to detect the expression levels of HIF-2α mRNA and HIF2PUT in 82 surgical specimens of primary osteosarcoma and matched non-cancerous bone tissues. Then, the associations of HIF-2α and/or HIF2PUT expression with various clinicopathological features of osteosarcoma patients were statistically analyzed. Moreover, their prognostic value was further evaluated. RESULTS: Compared with non-cancerous bone tissues, HIF-2α mRNA and HIF2PUT expression were both significantly upregulated in osteosarcoma tissues (all P<0.001). Interestingly, the expression levels of HIF-2α mRNA in osteosarcoma tissues were positively correlated with those of HIF2PUT (r=0.28, P=0.009). Additionally, osteosarcoma patients with HIF-2α mRNA and/or HIF2PUT over-expression more frequently had large tumor size (all P<0.05), advanced clinical stage (all P<0.01) and positive distant metastasis (all P<0.01). Moreover, osteosarcoma patients with HIF-2α mRNA and/or HIF2PUT over-expression had a significantly shorter overall and disease-free survival (all P<0.05). Furthermore, Cox multivariate analysis identified that HIF-2α mRNA and/or HIF2PUT expression, clinical stage and distant metastasis were all independent and significant prognostic factors for both overall and disease-free survival (all P<0.05). CONCLUSIONS: HIF-2α and HIF2PUT upregulation may be a common feature in human osteosarcomas with aggressive potency. The over-expression of the two molecules, alone or combined, may predict poor prognosis in osteosarcoma patients.