N-n-butyl haloperidol iodide mitigates myocardial ischemia/reperfusion injury through activation of SIRT1-Nrf2 signaling loop.

ABSTRACT: N-n-butyl haloperidol iodide (F2), a derivative of haloperidol developed by our group, exhibits potent antioxidative properties and confers protection against cardiac ischemia/reperfusion (I/R) injury. The protective mechanisms by which F2 ameliorates I/R injury remain obscure. The activation of nuclear factor erythroid 2-related factor 2 (Nrf2), a key transcription factor transactivating many antioxidative genes, also attenuates I/R-induced myocardial damage. The present study investigated whether the cardioprotective effect of F2 depends on Nrf2 using a mouse heart I/R model. F2 (0.1, 0.2 or 0.4 mg/kg) or vehicle was intravenously injected to mice 5 min before reperfusion. Systemic administration of 0.4 mg/kg F2 led to a significant reduction in I/R injury, which was accompanied by enhanced activation of Nrf2 signaling. The cardioprotection conferred by F2 was largely abrogated in Nrf2-deficient mice. Importantly, we found F2-induced activation of Nrf2 is SIRT1-dependent, as pharmacologically inhibiting SIRT1 by the specific inhibitor EX527 blocked Nrf2 activation. Moreover, F2-upregulated expression of SIRT1 was also Nrf2-dependent, as Nrf2 deficiency inhibited SIRT1 upregulation. These results indicate that SIRT1-Nrf2 signaling loop activation is indispensable for the protective effect of F2 against myocardial I/R injury, and may provide new insights for the treatment of ischemic heart disease.

N-n-butyl haloperidol iodide mediates cardioprotection via regulating AMPK/FoxO1 signalling.

Derangement of redox condition largely contributes to cardiac ischemia/reperfusion (I/R) injury. FoxO1 is a transcription factor which transcripts a series of antioxidants to antagonize I/R-induced oxidative myocardial damage. N-n-butyl haloperidol iodide (F2 ) is a derivative derived from haloperidol structural modification with potent capacity of inhibiting oxidative stress. This investigation intends to validate whether cardio-protection of F2 is dependent on FoxO1 using an in vivo mouse I/R model and if so, to further elucidate the molecular regulating mechanism. This study initially revealed that F2 preconditioning led to a profound reduction in I/R injury, which was accompanied by attenuated oxidative stress and upregulation of antioxidants (SOD2 and catalase), nuclear FoxO1 and phosphorylation of AMPK. Furthermore, inactivation of FoxO1 with AS1842856 abolished the cardio-protective effect of F2 . Importantly, we identified F2 -mediated nuclear accumulation of FoxO1 is dependent on AMPK, as blockage of AMPK with compound C induced nuclear exit of FoxO1. Collectively, our data uncover that F2 pretreatment exerts significant protection against post ischemic myocardial injury by its regulation of AMPK/FoxO1 pathway, which may provide a new avenue for treating ischemic disease.

RankProd Combined with Genetic Algorithm Optimized Artificial Neural Network Establishes a Diagnostic and Prognostic Prediction Model that Revealed C1QTNF3 as a Biomarker for Prostate Cancer.

Prostate cancer (PCa) is the most commonly diagnosed cancer in males in the Western world. Although prostate-specific antigen (PSA) has been widely used as a biomarker for PCa diagnosis, its results can be controversial. Therefore, new biomarkers are needed to enhance the clinical management of PCa. From publicly available microarray data, differentially expressed genes (DEGs) were identified by meta-analysis with RankProd. Genetic algorithm optimized artificial neural network (GA-ANN) was introduced to establish a diagnostic prediction model and to filter candidate genes. The diagnostic and prognostic capability of the prediction model and candidate genes were investigated in both GEO and TCGA datasets. Candidate genes were further validated by qPCR, Western Blot and Tissue microarray. By RankProd meta-analyses, 2306 significantly up- and 1311 down-regulated probes were found in 133 cases and 30 controls microarray data. The overall accuracy rate of the PCa diagnostic prediction model, consisting of a 15-gene signature, reached up to 100% in both the training and test dataset. The prediction model also showed good results for the diagnosis (AUC = 0.953) and prognosis (AUC of 5 years overall survival time = 0.808) of PCa in the TCGA database. The expression levels of three genes, FABP5, C1QTNF3 and LPHN3, were validated by qPCR. C1QTNF3 high expression was further validated in PCa tissue by Western Blot and Tissue microarray. In the GEO datasets, C1QTNF3 was a good predictor for the diagnosis of PCa (GSE6956: AUC = 0.791; GSE8218: AUC = 0.868; GSE26910: AUC = 0.972). In the TCGA database, C1QTNF3 was significantly associated with PCa patient recurrence free survival (P < .001, AUC = 0.57). In this study, we have developed a diagnostic and prognostic prediction model for PCa. C1QTNF3 was revealed as a promising biomarker for PCa. This approach can be applied to other high-throughput data from different platforms for the discovery of oncogenes or biomarkers in different kinds of diseases.

Hypoxia Suppresses TGF-B1-Induced Cardiac Myocyte Myofibroblast Transformation by Inhibiting Smad2/3 and Rhoa Signaling Pathways.

BACKGROUND/AIMS: Hypoxia modulation of transforming growth factor (TGF)- ß-induced signaling during myofibroblast transformation is dependent on the specific cell type. The purpose of this study was to explore the effects of hypoxia on myofibroblast transformation of TGF-ß1-induced cardiomyocyte H9c2 cells. METHODS: H9c2 cells were cultured for intermittent hypoxia treatment and TGF-ß1 treatment. α-Smooth muscle actin (α-SMA) expression was examined by western blotting and immunofluorescence after treatment. To further explore the possible mechanism for this effect, the effects of hypoxia on three early TGF-ß-dependent signaling pathways, i.e. the Smad2/3, RhoA and mitogen-activated protein kinase (MAPK) pathways, were screened by western blotting. RESULTS: Intermittent hypoxia induced TGF-ß1 expression, but had no effect on α-SMA expression. Exogenous TGF-ß1 alone upregulated α-SMA expression in H9c2 cells in a concentration- and time-dependent manner. α-SMA expression declined with the duration of hypoxia after intermittent hypoxia and exogenous TGF-ß1 co-treatment. Phospho-JNK and phospho-p38 levels were not significantly altered after TGF-ß1 and hypoxia treatment. However, levels of phospho-ERK increased after TGF-ß1 treatment and continued to increase after hypoxia co-treatment. The activation of phospho-Smad2/3 and phospho-RhoA induced by TGFß1 was significantly reduced after hypoxia co-treatment. CONCLUSION: Hypoxia can inhibit TGF-ß1-induced H9c2 myofibroblast transformation, based on inhibition of α-SMA expression by suppressing signaling downstream of TGF-ß1, Smad2/3 and RhoA. It suggested that TGF-ß-mediated cardiomyocyte transformation is not involved in hypoxia-mediated fibrosis.