Activation of the YAP/KLF5 transcriptional cascade in renal tubular cells aggravates kidney injury.

The Hippo/YAP pathway plays a critical role in tissue homeostasis. Our previous work demonstrated that renal tubular YAP activation induced by double knockout (dKO) of the upstream Hippo kinases Mst1 and Mst2 promotes tubular injury and renal inflammation under basal conditions. However, the importance of tubular YAP activation remains to be established in injured kidneys in which many other injurious pathways are simultaneously activated. Here, we show that tubular YAP was already activated 6 h after unilateral ureteral obstruction (UUO). Tubular YAP deficiency greatly attenuated tubular cell overproliferation, tubular injury, and renal inflammation induced by UUO or cisplatin. YAP promoted the transcription of the transcription factor KLF5. Consistent with this, the elevated expression of KLF5 and its target genes in Mst1/2 dKO or UUO kidneys was blocked by ablation of Yap in tubular cells. Inhibition of KLF5 prevented tubular cell overproliferation, tubular injury, and renal inflammation in Mst1/2 dKO kidneys. Therefore, our results demonstrate that tubular YAP is a key player in kidney injury. YAP and KLF5 form a transcriptional cascade, where tubular YAP activation induced by kidney injury promotes KLF5 transcription. Activation of this cascade induces tubular cell overproliferation, tubular injury, and renal inflammation.

12-inch growth of uniform MoS2 monolayer for integrated circuit manufacture.

Two-dimensional (2D) semiconductors, such as transition metal dichalcogenides, provide an opportunity for beyond-silicon exploration. However, the lab to fab transition of 2D semiconductors is still in its preliminary stages, and it has been challenging to meet manufacturing standards of stability and repeatability. Thus, there is a natural eagerness to grow wafer-level, high-quality films with industrially acceptable scale-cost-performance metrics. Here we report an improved chemical vapour deposition synthesis method in which the controlled release of precursors and substrates predeposited with amorphous Al2O3 ensure the uniform synthesis of monolayer MoS2 as large as 12 inches while also enabling fast and non-toxic growth to reduce manufacturing costs. Transistor arrays were fabricated to further confirm the high quality of the film and its integrated circuit application potential. This work achieves the co-optimization of scale-cost-performance metrics and lays the foundation for advancing the integration of 2D semiconductors in industry-standard pilot lines.

Pharmacokinetics of IMM-H012 in rats using ultra-performance liquid chromatography-tandem mass spectrometry.

The present study examined the pharmacokinetics of IMM-H012 in rat plasma, utilizing ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Internal standard cilostazol was employed, and plasma samples were processed using acetonitrile precipitation. A mobile phase (acetonitrile-0.1% formic acid in water) with gradient elution was used to achieve chromatographic separation using a UPLC BEH C18 column. In multiple reaction monitoring mode, electrospray ionization MS/MS was utilized in positive ionization mode. Based on findings, the lower limit of quantification was 2 ng/mL, and the linearity of IMM-H012 in rat plasma was found to be acceptable within the range of 2-2000 ng/mL (R2 > 0.995). The intra-day and inter-day precision relative standard deviation was less than 14% of IMM-H012 in rat plasma. The matrix effect was within the range of 102%-107%, and the accuracy ranged from 92% to 113%. Pharmacokinetics of IMM-H012 in rats after oral administration were successfully studied using UPLC-MS/MS.

The MuvB complex safeguards embryonic stem cell identity through regulation of the cell cycle machinery.

Increasing evidences indicate that unlimited capacity for self-renewal and pluripotency, two unique properties of embryonic stem cells (ESCs), are intrinsically linked to cell cycle control. However, the precise mechanisms coordinating cell fate decisions and cell cycle regulation remain to be fully explored. Here, using CRISPR/Cas9-mediated genome editing, we show that in ESCs, deficiency of components of the cell cycle regulatory MuvB complex Lin54 or Lin52, but not Lin9 or Lin37, triggers G2/M arrest, loss of pluripotency, and spontaneous differentiation. Further dissection of these phenotypes demonstrated that this cell cycle arrest is accompanied by the gradual activation of mesoendodermal lineage-specifying genes. Strikingly, the abnormalities observed in Lin54-null ESCs were partially but significantly rescued by ectopic coexpression of genes encoding G2/M proteins Cyclin B1 and Cdk1. Thus, our study provides new insights into the mechanisms by which the MuvB complex determines cell fate through regulation of the cell cycle machinery.

Follistatin-like 1 (FSTL1) interacts with Wnt ligands and Frizzled receptors to enhance Wnt/ß-catenin signaling in obstructed kidneys in vivo.

Follistatin (FS)-like 1 (FSTL1) is a member of the FS-SPARC (secreted protein, acidic and rich in cysteine) family of secreted and extracellular matrix proteins. The functions of FSTL1 have been studied in heart and lung injury as well as in wound healing; however, the role of FSTL1 in the kidney is largely unknown. Here, we show using single-cell RNA-Seq that Fstl1 was enriched in stromal cells in obstructed mouse kidneys. In addition, immunofluorescence demonstrated that FSTL1 expression was induced in fibroblasts during kidney fibrogenesis in mice and human patients. We demonstrate that FSTL1 overexpression increased renal fibrosis and activated the Wnt/ß-catenin signaling pathway, known to promote kidney fibrosis, but not the transforming growth factor ß (TGF-ß), Notch, Hedgehog, or Yes-associated protein (YAP) signaling pathways in obstructed mouse kidneys, whereas inhibition of FSTL1 lowered Wnt/ß-catenin signaling. Importantly, we show that FSTL1 interacted with Wnt ligands and the Frizzled (FZD) receptors but not the coreceptor lipoprotein receptor-related protein 6 (LRP6). Specifically, we found FSTL1 interacted with Wnt3a through its extracellular calcium-binding (EC) domain and von Willebrand factor type C-like (VWC) domain, and with FZD4 through its EC domain. Furthermore, we show that FSTL1 increased the association of Wnt3a with FZD4 and promoted Wnt/ß-catenin signaling and fibrogenesis. The EC domain interacting with both Wnt3a and FZD4 also enhanced Wnt3a signaling. Therefore, we conclude that FSTL1 is a novel extracellular enhancer of the Wnt/ß-catenin pathway.

YAP promotes AP-1 expression in tubular epithelial cells in the kidney.

Chronic kidney disease (CKD) is a major health problem. Kidney fibrosis is a hallmark and final common pathway of CKD. The Hippo/yes-associated protein (YAP) pathway regulates organ size, inflammation, and tumorigenesis. Our previous study demonstrated tubular YAP activation by tubule-specific double knockout of mammalian STE20-like protein kinase 1/2 (Mst1/2) induced CKD in mice, but the underlying mechanisms remain to be fully elucidated. Activator protein (AP)-1 activation was found to promote tubular atrophy and tubulointerstitial fibrosis. Therefore, we studied whether YAP regulates AP-1 expression in the kidney. We found that expression of various AP-1 components was induced in kidneys subjected to unilateral ureteric obstruction and in Mst1/2 double knockout kidneys, and these inductions were blocked by deletion of Yap in tubular cells, with Fosl1 being most affected compared with other AP-1 genes. Inhibition of Yap also most highly suppressed Fosl1 expression among AP-1 genes in HK-2 and IMCD3 renal tubular cells. YAP bound to the Fosl1 promoter and promoted Fosl1 promoter-luciferase activity. Our results suggest that YAP controls AP-1 expression and that Fosl1 is the primary target of YAP in renal tubular cells.NEW & NOTEWORTHY Yes-associated protein (YAP) activation leads to tubular injury, renal inflammation, and fibrosis, but the underlying mechanisms are not fully understood. We now provide genetic evidence that YAP promotes activator protein-1 expression and that Fosl1 is the primary target of YAP in renal tubular cells.

Kidney tubular transcription co-activator, Yes-associated protein 1 (YAP), controls the expression of collecting duct aquaporins and water homeostasis.

Final urine volume and concentration are defined by water reabsorption through the water channel proteins aquaporin (AQP)-2, -3 and -4 in the collecting duct. However, the transcriptional regulation of these AQPs is not well understood. The Hippo/Yes-associated protein 1 (YAP) pathway plays an important role in organ size control and tissue homeostasis. When the Hippo pathway including the Mst1/Mst2 kinases is inhibited, YAP is activated and functions as a transcription co-activator. Our previous work revealed a pathological role of tubular YAP activation in chronic kidney disease, but the physiological role of YAP in the kidney remains to be established. Here, we found that tubule-specific Yap knockout mice showed increased urine output and decreased urinary osmolality. Decreases in Aqp2, -3 and -4 mRNA and protein abundance in the kidney were evident in Yap knockout mice. Analysis of Mst1/Mst2 double knockout and Mst1/Mst2/Yap triple knockout mice showed that expression of Aqp2 and Aqp4 but not Aqp3 was dependent on YAP. Furthermore, YAP was recruited to the promoters of the Aqp2 and Aqp4 genes and stimulated their transcription. Interestingly, YAP was found to interact with transcription factors GATA2, GATA3 and NFATc1. These three factors promoted Aqp2 transcription in a YAP dependent manner in collecting duct cells. These three factors also promoted Aqp4 transcription whereas only GATA2 and GATA3 enhanced Aqp3 transcription. Thus, our results suggest that YAP promotes Aqp2 and Aqp4 transcription, interacts with GATA2, GATA3 and NFATc1 to control Aqp2 expression, while Aqp-2, -3 and -4 exploit overlapping mechanisms for their baseline transcriptional regulation.

KLF2 Mediates the Suppressive Effect of Laminar Flow on Vascular Calcification by Inhibiting Endothelial BMP/SMAD1/5 Signaling.

[Figure: see text].

Thioridazine protects against disturbed flow-induced atherosclerosis by inhibiting RhoA/YAP-mediated endothelial inflammation.

Atherosclerotic diseases remain the leading cause of adult mortality and impose heavy burdens on health systems globally. Our previous study found that disturbed flow enhanced YAP activity to provoke endothelial activation and atherosclerosis, and targeting YAP alleviated endothelial inflammation and atherogenesis. Therefore, we established a luciferase reporter assay-based drug screening platform to seek out new YAP inhibitors for anti-atherosclerotic treatment. By screening the FDA-approved drug library, we identified that an anti-psychotic drug thioridazine markedly suppressed YAP activity in human endothelial cells. Thioridazine inhibited disturbed flow-induced endothelial inflammatory response in vivo and in vitro. We verified that the anti-inflammatory effects of thioridazine were mediated by inhibition of YAP. Thioridazine regulated YAP activity via restraining RhoA. Moreover, administration of thioridazine attenuated partial carotid ligation- and western diet-induced atherosclerosis in two mouse models. Overall, this work opens up the possibility of repurposing thioridazine for intervention of atherosclerotic diseases. This study also shed light on the underlying mechanisms that thioridazine inhibited endothelial activation and atherogenesis via repression of RhoA-YAP axis. As a new YAP inhibitor, thioridazine might need further investigation and development for the treatment of atherosclerotic diseases in clinical practice.

Effects of Surgical Timing of Facial Nerve Decompression for Patients With Severe Bell's Palsy.

Context: Bell's palsy is a form of idiopathic, facial nerve palsy. Initial treatment includes the use of oral corticosteroids and/or antiviral agents, but facial paralysis may persist. Some surgeons suggest that surgical decompression of the facial nerve can be a beneficial, but the optimal surgical approach, extent of nerve decompression, and timing of surgery remain unclear. Objective: This study intended to evaluate the efficacy of delayed, facial nerve decompression for severe Bell's palsy (BP) and to explore the relationship of opportunity timing for operations, with postoperative recovery for facial nerve function. Design: The research team performed a retrospective study. Setting: The study took place at Beijing Tiantan Hospital of Capital Medical University in Beijing, China. Participants: Participants were 45 patients who had been diagnosed with BP between 2015 and 2021 and who had undergone facial nerve decompression using the transmastoid approach, between 30 and 180 days after the onset of BP. According to the operation's timing, the research team divided the participants into three groups, consisting of participants who underwent surgery: (1) at 30-60-days after BP onset-19 participants, (2) at 61-90 days after BP onset-18 participants, and (3) at more than 90 days after BP onset-8 participants. Outcome Measures: The research team: (1) analyzed participants' demographic and preoperative and postoperative clinical characteristics, (2) compared the surgical outcomes with participants' House-Brackmann (HB) scales, and (3) analyzed the factors affecting the recovery of facial nerve function using logistic regression. Results: Decompression surgery was effective for 29 participants (64.4%), with similar rates for participants who underwent surgery after 30-60 days (73.7%) and 61-90 days (77.8%), but the surgery' success was significantly higher for those groups than for participants who underwent surgery after >90 days (12.5%), with P = .008 and P = .003, respectively. Multivariate logistic regression analysis showed that disease duration was the only factor significantly associated with the effectiveness of surgery (odds ratio = 120.337; 95% confidence interval 2.997-4832.267, P = .011). Conclusions: For patients with severe Bell's palsy with ineffective conservative treatment, surgery performed 30 to 90 days after the onset of paralysis can have therapeutic benefits, whereas surgery performed after 3 months is relatively ineffective.

Long-term Outcome of Sound Localization with Baha ® Attract System.

BACKGROUND: Spatial hearing is a critical feature in daily life. However, there is quite a range in hearing loss patients regarding the effect of bone conduction device on localization performance. OBJECTIVES: To analyze the localization performance in patients with bilateral conductive or mixed hearing loss fitted with one Baha® Attract system. MATERIALS AND METHODS: This prospective study included 12 patients who had followed up for more than one year. The parameters analyzed included (1) audiological results: sound field threshold, speech discrimination scores (SDSs), and sound localization test, and (2) functional results: scores for the Speech, Spatial and Qualities of Hearing Scale (SSQ) and the Chinese translation of the Spatial Hearing Questionnaire (C-SHQ). RESULTS: The audiological assessments showed a reduction of 28.5 dB in the mean sound field thresholds and improvements of 61.7% in the SDSs for disyllabic words. The root mean square error improved slightly with the Baha® Attract system. Patients showed promising results in the functional questionnaire assessments, with significant improvements in the SSQ and C-SHQ scores. CONCLUSIONS: Although most patients were not able to localize sound accurately after surgery, the change in the scores of the SSQ and C-SHQ indicated that the Baha® Attract system could improve spatial hearing.

Value of the Hyomental Distance Measured With Ultrasound in Forecasting Difficult Laryngoscopy in Newborns.

PURPOSE: Preoperative evaluations of difficult airways are imperative, especially in newborns. The hyomental distance is a reliable index for predicting difficult airways in adults. However, few studies have evaluated the value of the hyomental distance for predicting difficult airways in newborns. It is unclear whether the hyomental distance forecasts a restricted or difficult view when using direct laryngoscopy. We intended to develop an effective system for predicting difficult tracheal intubation in newborns. DESIGN: A prospective observational clinical study. METHODS: Newborns aged 0 to 28 days undergoing oral endotracheal intubation with direct laryngoscopy for elective surgery under general anesthesia were enrolled. The hyomental distance and hyoid level tissue thickness were assessed by ultrasound. Other parameters, such as the mandibular length and sternomental distance, were also evaluated before anesthesia. The glottic structure view under laryngoscopy was graded according to the Cormack-Lehane classification. The patients with Grade 1 and 2 laryngeal views were assigned to Group E. Those with Grade 3 and 4 views were assigned to Group D. FINDINGS: A total of 123 newborns were recruited for our study. The incidence of poor visualization of the larynx during laryngoscopy in our study was 10.6%. The multifactor logistic regression results showed that the hyomental distance was a powerful predictor of difficult laryngoscopy (OR = 0.16, 95% CI 0.03-0.74, P = .019). The curve with the highest sensitivity and specificity and the maximum area under the curve (AUC) was the hyomental distance. The receiver operating characteristic (ROC) curve for the hyomental distance suggested that the best cut-off value was less than equal to 2.74 cm, with an AUC of 0.80 (95% CI 0.64-0.95). CONCLUSIONS: It is noninvasive and feasible to accurately measure the hyomental distance with ultrasound in newborns, and the results are reliable. We believe that the hyomental distance measured with ultrasound could be used as a marker for predicting difficult laryngoscopy in newborns.

Supramolecular Polymer Ion Conductor with Weakened Li Ion Solvation Enables Room Temperature All-Solid-State Lithium Metal Batteries.

Improved durability, enhanced interfacial stability, and room temperature applicability are desirable properties for all-solid-state lithium metal batteries (ASSLMBs), yet these desired properties are rarely achieved simultaneously. Here, in this work, it is noticed that the huge resistance at Li metal/electrolyte interface dominantly impeded the normal cycling of ASSLMBs especially at around room temperature (<30 °C). Accordingly, a supramolecular polymer ion conductor (SPC) with "weak solvation" of Li+ was prepared. Benefiting from the halogen-bonding interaction between the electron-deficient iodine atom (on 1,4-diiodotetrafluorobenzene) and electron-rich oxygen atoms (on ethylene oxide), the O-Li+ coordination was significantly weakened. Therefore, the SPC achieves rapid Li+ transport with high Li+ transference number, and importantly, derives a unique Li2 O-rich SEI with low interfacial resistance on lithium metal surface, therefore enabling stable cycling of ASSLMBs even down to 10 °C. This work is a new exploration of halogen-bonding chemistry in solid polymer electrolyte and highlights the importance of "weak solvation" of Li+ in the solid-state electrolyte for room temperature ASSLMBs.

Paired test of matrix graphs and brain connectivity analysis.

Inferring brain connectivity network and quantifying the significance of interactions between brain regions are of paramount importance in neuroscience. Although there have recently emerged some tests for graph inference based on independent samples, there is no readily available solution to test the change of brain network for paired and correlated samples. In this article, we develop a paired test of matrix graphs to infer brain connectivity network when the groups of samples are correlated. The proposed test statistic is both bias corrected and variance corrected, and achieves a small estimation error rate. The subsequent multiple testing procedure built on this test statistic is guaranteed to asymptotically control the false discovery rate at the pre-specified level. Both the methodology and theory of the new test are considerably different from the two independent samples framework, owing to the strong correlations of measurements on the same subjects before and after the stimulus activity. We illustrate the efficacy of our proposal through simulations and an analysis of an Alzheimer's Disease Neuroimaging Initiative dataset.

Wafer-Scale Demonstration of MBC-FET and C-FET Arrays Based on Two-Dimensional Semiconductors.

Two-dimentional semiconductors have shown potential applications in multi-bridge channel field-effect transistors (MBC-FETs) and complementary field-effect transistors (C-FETs) due to their atomic thickness, stackability, and excellent electrical properties. However, the exploration of MBC-FET and C-FET based on large-scale 2D semiconductors is still lacking. Here, based on a reliable vertical stacking of wafer-scale 2D semiconductors, large-scale MBC-FETs and C-FETs using n-type MoS2 and p-type MoTe2 are successfully fabricated. The drive current of an MBC-FET with two layers of MoS2 channel (20 µm/10 µm) is up to 60 µA under 1 V bias. Compared with the single-gate MoS2 FET, the carrier mobility of MBC-FET is 2.3 times higher and the sub-threshold swing is 70% smaller. Furthermore, NAND and NOR logic circuits are also constructed based on two vertically stacked MoS2 channels. Then, C-FET arrays are fabricated by 3D integrating n-type MoS2 FET and p-type MoTe2 FET, which exhibit a voltage gain of 7 V/V when VDD  = 4 V. In addition, this C-FET device can directly convert light signals to an electrical digital signal within a single device. The demonstration of MBC-FET and C-FET based on large-scale 2D semiconductors will promote the application of 2D semiconductors in next-generation circuits.

The effects of gestational diabetes mellitus with maternal age between 35 and 40 years on the metabolite profiles of plasma and urine.

BACKGROUND: Gestational diabetes mellitus (GDM) is defined as impaired glucose tolerance in pregnancy and without a history of diabetes mellitus. While there are limited metabolomic studies involving advanced maternal age in China, we aim to investigate the metabolomic profiling of plasma and urine in pregnancies complicated with GDM aged at 35-40 years at early and late gestation. METHODS: Twenty normal and 20 GDM pregnant participants (≥ 35 years old) were enlisted from the Complex Lipids in Mothers and Babies (CLIMB) study. Maternal plasma and urine collected at the first and third trimester were detected using gas chromatography-mass spectrometry (GC-MS). RESULTS: One hundred sixty-five metabolites and 192 metabolites were found in plasma and urine respectively. Urine metabolomic profiles were incapable to distinguish GDM from controls, in comparison, there were 14 and 39 significantly different plasma metabolites between the two groups in first and third trimester respectively. Especially, by integrating seven metabolites including cysteine, malonic acid, alanine, 11,14-eicosadienoic acid, stearic acid, arachidic acid, and 2-methyloctadecanoic acid using multivariant receiver operating characteristic models, we were capable of discriminating GDM from normal pregnancies with an area under curve of 0.928 at first trimester. CONCLUSION: This study explores metabolomic profiles between GDM and normal pregnancies at the age of 35-40 years longitudinally. Several compounds have the potential to be biomarkers to predict GDM with advanced maternal age. Moreover, the discordant metabolome profiles between the two groups could be useful to understand the etiology of GDM with advanced maternal age.

Hypothesis Testing for Network Data with Power Enhancement.

Comparing two population means of network data is of paramount importance in a wide range of scientific applications. Numerous existing network inference solutions focus on global testing of entire networks, without comparing individual network links. The observed data often take the form of vectors or matrices, and the problem is formulated as comparing two covariance or precision matrices under a normal or matrix normal distribution. Moreover, many tests suffer from a limited power under a small sample size. In this article, we tackle the problem of network comparison, both global and simultaneous inferences, when the data come in a different format, i.e., in the form of a collection of symmetric matrices, each of which encodes the network structure of an individual subject. Such data format commonly arises in applications such as brain connectivity analysis and clinical genomics. We no longer require the underlying data to follow a normal distribution, but instead impose some moment conditions that are easily satisfied for numerous types of network data. Furthermore, we propose a power enhancement procedure, and show that it can control the false discovery, while it has the potential to substantially enhance the power of the test. We investigate the efficacy of our testing procedure through both an asymptotic analysis and a simulation study under a finite sample size. We further illustrate our method with examples of brain connectivity analysis.

The polycomb group protein PCGF6 mediates germline gene silencing by recruiting histone-modifying proteins to target gene promoters.

Polycomb group (PcG) proteins are essential for maintenance of lineage fidelity by coordinating developmental gene expression programs. Polycomb group ring finger 6 (PCGF6) has been previously reported to repress expression of lineage-specific genes, especially germ cell-related genes in mouse embryonic stem cells (ESCs) via the noncanonical polycomb repressive complex PRC1.6. However, the molecular mechanism of this repression remains largely unknown. Here, using RNA-Seq, real-time RT-PCR, immunohistochemistry, immunoprecipitation, and ChIP analyses, we demonstrate that PCGF6 plays an essential role in embryonic development, indicated by the partially penetrant embryonic lethality in homozygous PCGF6 (Pcgf6-/-)-deficient mice. We also found that surviving Pcgf6-deficient mice exhibit reduced fertility. Using the Pcgf6-deficient mice, we observed that ablation of Pcgf6 in somatic tissues robustly derepresses germ cell-related genes. We further provide evidence that these genes are direct targets of PCGF6 in ESCs and that endogenous PCGF6 co-localizes with the histone-modifying proteins G9A histone methyltransferase (G9A)/G9a-like protein (GLP) and histone deacetylase 1/2 (HDAC1/2) on the promoters of the germ cell-related genes. Moreover, the binding of these proteins to their target genes correlated with methylation of Lys-9 of histone 3 and with the status of histone acetylation at these genes. Moreover, the recruitment of G9A/GLP and HDAC1/2 to target promoters depended on the binding of PCGF6. Our findings indicate that PCGF6 has a critical role in safeguarding lineage decisions and in preventing aberrant expression of germ cell-related genes.

Long noncoding RNA PVT1 facilitates high glucose-induced cardiomyocyte death through the miR-23a-3p/CASP10 axis.

Dilated cardiomyopathy (DCM) is the leading cause of morbidity and mortality in diabetic patients. Long noncoding RNA plasmacytoma variant translocation 1 (PVT1) has been shown to be related to the pathogenesis of DCM. However, the mechanism by which PVT1 regulates DCM pathogenesis is unclear. High glucose level was employed to construct a DCM cell model in vitro. Cell viability was determined via cell counting kit-8 assay. The level of lactate dehydrogenase (LDH) was measured with the corresponding kit. Expression levels of PVT1, miR-23a-3p, and caspase-10 (CASP10) messenger RNA were evaluated with a quantitative real-time polymerase chain reaction. Cell apoptosis was assessed by flow cytometry assay. Protein levels of B-cell lymphoma 2-associated X (Bax), cleaved-caspase-3 (cleaved-casp-3), and CASP10 were examined via western blot analysis. The relationship between PVT1 or CASP10 and miR-23a-3p was verified with dual-luciferase reporter assay. We observed that PVT1 and CASP10 were upregulated while miR-23a-3p was downregulated in high glucose-induced cardiomyocytes. High glucose levels repressed cardiomyocyte activity and induced cardiomyocyte apoptosis, but this influence was antagonized by PVT1 knockdown or miR-23a-3p overexpression. Furthermore, PVT1 acted as a sponge for miR-23a-3p, and miR-23a-3p inhibition counterbalanced the influence of PVT1 silencing on viability and apoptosis of cardiomyocytes under high glucose level treatment. PVT1 could increase CASP10 expression via sponging miR-23a-3p. In conclusion, PVT1 acted as a deleterious lncRNA in DCM. PVT1 facilitated cardiomyocyte death by regulating the miR-23a-3p/CASP10, which offered a new mechanism to comprehend the pathogenesis of DCM.

Development and application of SYBR Green â real-time quantitative reverse transcription PCR assay for detection of swine Getah virus.

Getah virus (GETV), a mosquito-borne virus belonging to the Alphavirus genus of family Togaviridae, has become increasingly problematic, which poses a huge threat to the safety of animals and public health. In order to detect GETV quickly and accurately, we have developed a SYBR Green I real-time quantitative reverse transcription PCR (RT-qPCR) assay for GETV with the detection limit of 66 copies/µL, excellent correlation coefficient (R2) of 0.9975, and amplification efficiency (E) of 98.90%, the target selected was the non-structural protein 3 of GETV. The sensitivity of it was higher than that of ordinary RT-PCR by 1000 folds, and the inter-assay and intra-assay CV values were all less than 0.99%. The newly developed RT-qPCR assay exhibited good sensitivity and reproducibility, which will provide technical support for the reliable and specific rapid diagnosis, and quantitative analysis of GETV infection.