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To investigate the effects of ginsenosides on the memory impairment in Sprague–Dawley rats (SD rats) after anesthesia through the administration of propofol SPF, SD rats were randomly divided into four groups: control group (Group I), propofol-treated group (Group II), low dose of ginsenosides-treated group (Group III) and high dose of ginsenosides-treated group (Group IV). These rats were subjected to fear memory test in shuttle box, Y-maze test and Morris water maze test. Immediately after the test, the expression levels of nerve growth factor (NGF) and brain derived neurotrophic factor (BDNF) were further detected by ELISA method. Ginsenosides could ameliorate the impairment on the functions of fear memory, working memory and spatial memory in rats caused by anesthesia via the injection of Propofol. Furthermore, the expression levels of NGF and BDNF on rat hippocampus were significant increased by the treatment of ginsenosides at both two doses compared with the control group (both P

Merazin hydrate (MH), an essential ingredient of Fructus aurantii, has been identified to have an antidepressant-like effect. However, the molecular mechanisms of MH modulate depressive behavior are largely uncharacterized. Here, in lipopolysaccharide-induced mice, we identified that a single administration of MH recovered depressive behaviors and down-regulated the expressions of neuronal nitric oxide synthase (nNOS) in the hippocampus after 1 day. Activation of nNOS by l-arginine led to depressive behaviors, and inhibition of nNOS contributed to antidepressive behaviors. Notably, MH only reversed the expression of nNOS's downstream NF-κB and not the CREB/BDNF pathway in the hippocampus, and MH's antidepressant-like effects were prevented by Asatone (an agonist of NF-κB) and not H89 (an antagonist of CREB). MH also normalized the expressions of GFAP and IB-1 in dentate gyrus in the hippocampus and inflammatory factors such as IL-1β, IL-10, and TNF-α in serum. Overall, our studies reveal the molecular mechanisms of MH's antidepressant-like effect.

This study aimed to investigate the electrophysiological characteristics of idiopathic ventricular arrhythmias (VAs) originating from the left ventricular posterior papillary muscles (LPPM) and explore the efficiency of catheter ablation using three-dimensional intracardiac ultrasound technology.Twenty-seven cases of premature ventricular contraction/ventricular tachycardia (PVC/VT) originating from the left ventricular posterior papillary muscles were recorded from July 2015 to June 2019 in the Central Hospital of Shengli Oil Field and the First Affiliated Hospital of Zhengzhou University. Electrophysiological mapping and radiofrequency catheter ablation (RFCA) were performed using three-dimensional intracardiac ultrasound technology. The characteristics of the body surface and intracavity electrocardiogram were analyzed. All cases were followed up for 24 months after the operation.The VAs of all 27 cases were successfully eliminated by catheter ablation. QRS complexes were observed with a right bundle branch block (RBBB) pattern and a steep slope in the initial segment. Lead I appeared with an Rs pattern, and inferior leads (lead II, III, and aVF) were usually with an S wave. The lead aVR appeared with a qR pattern, while the R wave was commonly found in aVL. The main wave in leads VVentricular arrhythmias originating from the left ventricular posterior papillary muscles have similar electrophysiological characteristics. The origin site was accurately located using three-dimensional intracardiac ultrasound technology. Catheter ablation effectively eliminated VAs.

Major depressive disorder has become an increasingly serious disease in the world. However, convenient antidepressants have low efficacy and slow onset defects, which is dangerous for suicidal tendency patients. Nowadays, rapid antidepressant research has become the focus. Merazin hydrate (MH), a component of the natural herb Fructus Aurantii, has been shown to produce rapid antidepressant-like effects in animal models. However, the mechanism of its rapid antidepressant-like effects was still elusive like that of ketamine. The study aimed to reveal the relationship between the rapid antidepressant-like effects of MH and mTOR signaling, which is closely related to rapid antidepressants. The results showed that a single administration of MH was capable of reversing the behavioral defects at 2 h in two classic depressive models including learned helplessness (LH) and chronic mild stress (CMS). Moreover, the phosphorylated expression of mTOR, reduced by LH or CMS, was upregulated after a single administration of MH, and the expressions of BDNF and synaptic proteins in the hippocampus were also reversed 2 h later, similar to ketamine. Moreover, LH increased the expressions of eNOS, IL-10, and TNF-α in serum, which were all reversed by a single dose of MH at 2 h, similar to ketamine. Furthermore, we used rapamycin, an antagonist of mTOR, to confirm whether the rapid antidepressant-like effects of MH depend on mTOR or not. We found that inhibiting the activation of mTOR blocked the rapid antidepressant-like effects of MH, which also inhibited the upregulation of expressions of BDNF and PSD95. To sum up, the rapid antidepressant effect of MH depended on the activation of mTOR to regulate downstream BNDF and synaptic protein expressions.

INTRODUCTION Linear ablation in addition to pulmonary vein antrum isolation (PVAI) has failed to improve the success rate for persistent atrial fibrillation (PeAF), due to incomplete block of ablation lines, especially in the mitral isthmus (MI). METHODS AND RESULTS The study enrolled 191 patients (66 in group 1 and 125 in group 2). In group 1, ethanol infusion into the vein of Marshall was first performed, followed by radiofrequency (RF) applications targeting bilateral PVAI and bidirectional block in the roofline, cavotricuspid isthmus, and MI. In group 2, PVAI and the three linear ablations were completed using only RF energy. MI block was achieved in 63 (95.5%) and 101 (80.8%) patients in groups 1 and 2, respectively (p = .006). Patients in group 1 had shorter ablation time for left pulmonary vein antrum (8.15 vs. 12.59 min, p

Aesthetic quality assessment and semantic recognition are the two fundamental aspects of image perception and understanding tasks. Though these two tasks are related, most of the current research generally treats them as independent problems without any interaction. In this paper, we explore the relationships between aesthetic quality assessment and semantic recognition task, and employ a multi-task convolutional neural network with feature enhancement mechanism to effectively integrate these two tasks. A novel Enhanced Aggregation of Features Network (EAFNet) for joint learning of the two tasks is proposed to enhance the valid features and suppress the invalid features of each task in both channel and spatial dimensions. Experiments conducted on two benchmark datasets well verify the superior performance of EAFNet in handling aesthetic quality assessment and semantic recognition tasks.

The study of genetic syndromes characterized by sensitivity to DNA damaging agents has provided important insights into the mechanisms that maintain genome stability and identified novel targets for cancer therapies. Here, we used exome sequencing to study 51 unrelated individuals with previously reported hypersensitivity to ionizing radiation as well as a range of neurologic, immunologic, and developmental features, but who did not clearly fit any previously defined genetic syndrome. Based on the combination of variant identification, computational evidence of deleteriousness, and functional screening, we identified three groups of subjects. Two subjects carried the bi-allelic loss of function variants in causative genes for known DNA damage response syndromes. Eight subjects carried the single loss of function variants in causative genes for DNA damage response syndromes, six of whom also carried predicted deleterious variants in other genes with DNA damage-related functions. Three subjects carried deleterious mutations in genes without obvious roles in DNA damage responses. However, treatment of U2OS cells with small interfering RNA targeting these genes resulted in significantly increased radiation sensitivity. Our results suggest that gene-gene interaction may contribute to ionizing radiation sensitivity as well as highlighting possible roles for several genes not obviously involved in the response to DNA damage.

In the traditional surgical intervention procedure, residual tumor cells may potentially cause tumor recurrence. In addition, large bone defects caused by surgery are difficult to self-repair. Thus, it is necessary to design a bioactive scaffold that can not only kill residual tumor cells but also promote bone defect regeneration simultaneously. Here, we successfully developed Cu-containing mesoporous silica nanosphere-modified β-tricalcium phosphate (Cu-MSN-TCP) scaffolds, with uniform and dense nanolayers with spherical morphology via 3D printing and spin coating. The scaffolds exhibited coating time- and laser power density-dependent photothermal performance, which favored the effective killing of tumor cells under near-infrared laser irradiation. Furthermore, the prepared scaffolds favored the proliferation and attachment of rabbit bone marrow-derived mesenchymal stem cells and stimulated the gene expression of osteogenic markers. Overall, Cu-MSN-TCP scaffolds can be considered for complete eradication of residual bone tumor cells and simultaneous healing of large bone defects, which may provide a novel and effective strategy for bone tumor therapy. In the future, such Cu-MSN-TCP scaffolds may function as carriers of anti-cancer drugs or immune checkpoint inhibitors in chemo-/photothermal or immune-/photothermal therapy of bone tumors, favoring for effective treatment.

The emission of NOx in thermal power plants would cause serious pollution to the environment, and effective prediction of NOx has important significance. With the development of deep learning, many methods can effectively solve the prediction problem. In this paper, a long short-term memory (LSTM) neural network based on the generalized correntropy (GC) loss function, which is called the GC-LSTM neural network here, is proposed to predict the NOx emission. In order to make the characteristics of high-dimensional data have the same measurement scale, normalization is used to preprocess data firstly. Then, the processed historical data is used as training data to train the GC-LSTM neural network. After that, the emission of the NOx at each time instant can be predicted according to the online measured relevant data based on the established GC-LSTM neural network. Finally, prediction results under the LSTM neural network with GC, mean square error (MSE) and the mean absolute error (MAE) loss functions are given. And according to the simulation results, it can be seen that the loss function curve of the LSTM network based on GC is more stable and has better convergence. It can be seen that the proposed GC-LSTM prediction is more accurate, by comparing the probability density function (PDF) and root mean square error (RMSE) of the error between the actual values and the prediction values.

Background Depression and functional dyspepsia (FD) are characterized by comorbidity, overlapping depression, and nausea. The pathogenesis of depression and FD is mediated by α2-adrenoreceptor and/or ghrelin. Antidepressant (A) or prokinetic (P) agents are numerous, but few have been investigated in this context. Ancient Gan-zhu-shu-xie (GZSX), whose representative traditional Chinese medicine(TCM) is Chaihu-Shugan-San (CSS), may exert antidepressant effects with prokinetic meranzin hydr-ate (MH) via α2-adrenoreceptors in the acute forced swimming (FS) test in rats.Therefore, the main aim of the study is to investigate the acute antidepressant and prokinet-ic effects of CSS and MH after acutely FS on rats, and its possible mechanism. Methods FS rats were treated with CSS, MH, fluoxetine, ghrelin antagonist [D-Lys3]-GHRP-6, and take a series of behavior tests and gastrointestina motility tests, and via 7.0 T fMRI-BOLD signal, compared with well-known mechanism of positive control. Results MH has similar effects to CSS-stimulated deactivation when comparedto those of fluoxetine (4.02-fold for hippocampus and 1.45-fold for thalamus). The ghrelin antagonist [D-Lys3]-GHRP-6 synchronously inhibited A&P and BOLD HTB foci. Prokinetic mosapride had effects on the thalamus and basal ganglia but not the hippocampus. Within the HTB, the hippocampus is implicated in depression and FD. Conclusion These data show that on acute FS-stimulated DB&H, MH-induced rapid A&P, and ghrelin-related regulation coupled to BOLD signals in brain areas before, providing insight into a unified theory of depression pathogenesis and pharmacotherapy.

The demand response (DR) strategy enables household users to actively optimize and dispatch the household energy management system (HEMS), which may significantly reduce the cost of user energy consumptions. However, the uncertainty of home users behaviors, the diversity of electrical equipment types, as well as the complexity of the working status of various devices have brought severe challenges to the home energy management system. This article proposes a new forecasting based optimization method to deliver the real-time scheduling considering the future environment trend. A recent proposed dueling based deep reinforcement learning approach is adopted to optimally dispatch the HEMS. In addition, due to the delay of heating, ventilation, and air Conditioning (HVAC) indoor and outdoor temperature data non-Gaussian, a new generalized corr-entropy assisted long short-term memory (GC-LSTM) neural network is proposed where the generalized correntropy (GC) loss function is adopted to predict the outdoor temperature. The proposed method is verified in a featured HEMS benchmark problem and the experimental results show that the user costs are effectively reduced while the user satisfaction is maintained utilizing the proposed method.

Nanostructure-enhanced detection is promising for a number of applications such as early cancer diagnosis, environmental monitoring and mine safety, among which nanostructures integrated microfluidic chips offers unique advantage of ultra-low quantitative analyses. Here, dense ZnO nanowires of varied diameter and length were obtained by changing the content of polyethyleneimine (PEI) and growth time via simple hydrothermal growth in microfluidic channels for protein detection. We showed that this approach was superiorly efficient compared to the conventional hydrothermal method due to the flow-induced replenishment of nutrient and the effect of shear stress. When immobilizing FITC conjugated anti-bovine immunoglobulin G (IgG) on ZnO nanowires, the fluorescence emission was significantly amplified compared to glass substrate and ZnO seed layer. Under the different growth conditions, the most remarkable fluorescence enhancement was observed on the ZnO nanowire substrate grown for 3h with 5mM PEI in solution. It is ascribed not only to the increase of the binding surface area of proteins but also the intrinsic fluorescence enhancement of ZnO nanowires as waveguides. We further used the optimized ZnO nanowires to demonstrate multiple detection of cancer biomarkers, achieving a superior limit of detection (LOD) as low as 1pg/mL in human α-fetoprotein (AFP) assay and 100 fg/mL in carcinoembryonic antigen (CEA) assay with large dynamic range of 6-7 orders, which suggests that ZnO nanowire integrated microfluidic chips are promising for high-throughput fluorescence-based diagnostic assays.

Depression medication is still facing challenges, where the key issue is to explore a desirable delivery platform to assist drugs in penetrating blood-brain barrier (BBB). Herein, we prepared beard nanoscale particles (BNPs) from the beard of young Asian males, which were mainly composed by keratin and melanin. BNPs were able to load antidepressants (Fluoxetine) and exhibited therapeutic effect on depression. Moreover, in vitro and in vivo studies showed that BNPs held favorable photothermal conversion capability, prominent BBB permeability under near-infrared (NIR) irradiation and reliable biocompatibility. Deserved to be mentioned, BNPs also performed a pivotal role in redox equilibrium, which was thus capable of scavenging ROS induced by depression. Subsequently, behavioral and biochemical experiments also demonstrated significant depression-ameliorating effects in vivo. In summary, our data for the first time suggested that biological organism derived beard could serve as ideal low-cost nanovectors for depression therapy.

The burden of depression is enormous, and numerous studies have found that major depressive disorder (MDD) induces cardiovascular disorders (CVD) and functional dyspepsia (FD). Excitingly, meranzin hydrate (MH), an absorbed bioactive compound of Aurantii Fructus Immaturus, reverses psychosocial stress-induced mood disorders, gastrointestinal dysfunction and cardiac disease. Pharmacological methods have repeatedly failed in antidepressant development over the past few decades, but repairing aberrant neural circuits might be a reasonable strategy. This article aimed to explore antidepressant-like effects and potential mechanisms of MH in a rat model of unpredictable chronic mild stress (UCMS). Utilizing blood oxygen level-dependent (BOLD) functional magnetic resonance imaging (fMRI), we sought to find reliable neurocircuits or a dominant brain region revealing the multiple effects of MH. The results show that compared with UCMS rats, MH (10 mg/kg/day for 1 week i.g.)-treated rats exhibited decreased depression-like behaviour; increased expression of brain-derived neurotrophic factor (BDNF) in the hippocampal dentate gyrus; and normalized levels of adrenocorticotropic hormone (ACTH), corticosterone (CORT), and acylated ghrelin (AG). Additionally, the UCMS-induced rise in BOLD activation in the reward system was attenuated after MH treatment. A literature search shown that nucleus accumbens (NAc) and hypothalamus of the reward system might reveal multiple effects of MH on MDD-FD-CVD comorbidity. Further research will focus on the role of these two brain regions in treating depression associated with comorbidities.

Purpose Mutations in the gene encoding 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase/IMP cyclohydrolase (ATIC), a bifunctional enzyme that catalyzes the final 2 steps of the purine de novo biosynthetic pathway, were identified in a subject referred for radiation sensitivity testing. Functional studies were performed to determine whether ATIC inhibition was radiosensitizing and, if so, to elucidate the mechanism of this effect and determine whether small molecule inhibitors of ATIC could act as effective radiosensitizing agents. Methods and Materials Both small interfering RNA knockdown and small molecule inhibitors were used to inactivate ATIC in cell culture. Clonogenic survival assays, the neutral comet assay, and γH2AX staining were used to assess the effects of ATIC inhibition or depletion on cellular DNA damage responses. Results Depletion of ATIC or inhibition of its transformylase activity significantly reduced the surviving fraction of cells in clonogenic survival assays in multiple cancer cell lines. In the absence of ionizing radiation exposure, ATIC knockdown or chemical inhibition activated cell cycle checkpoints, shifting cells to the more radiosensitive G2/M phase of the cell cycle, and depleted cellular adenosine triphosphate but did not result in detectable DNA damage. Cells in which ATIC was knocked down or inhibited and then treated with ionizing radiation displayed increased numbers of DNA double-strand breaks and a delay in the repair of those breaks relative to irradiated, but otherwise untreated, controls. Supplementation of culture media with exogenous adenosine triphosphate ameliorated the DNA repair phenotypes. Conclusions These findings implicate ATIC as an effective, and previously unrecognized, target for chemoradiosensitization and, more broadly, suggest that purine levels in cells might have an underappreciated role in modulating the efficiency of DNA damage responses that could be exploited in radiosensitizing strategies.

Myelodysplastic syndrome (MDS) is a primary bone marrow disorder whose hallmark is the development of peripheral cytopenias and a predilection toward the development of acute myeloid leukemia (AML). Patients often have hypercellular bone marrows with dysplastic features that may involve multiple lineages. An increased awareness of MDS has led to the reporting of a number of associated autoimmune and paraneoplastic conditions in the medical literature. We present the case of an elderly man who was transferred to our institution with persistent, refractory bleeding several weeks after the resection of a sebaceous cyst. Despite reoperation, treatment with topical and intravenous hemostatic agents, and transfusion of blood products, the patient's bleeding persisted. A comprehensive evaluation for the cause of his coagulopathy was undertaken. Bone marrow evaluation was consistent with MDS. A paraneoplastic consumptive coagulopathy or fibrinolytic process in conjunction with MDS-related platelet dysfunction was felt to be the most likely etiology of the patient's bleeding.

Protein kinase R (PKR) is an interferon (IFN)-inducible, double-stranded RNA-activated kinase that initiates apoptosis in response to cellular stress. To determine the role of PKR in hematopoiesis, we developed transgenic mouse models that express either human PKR (TgPKR) or a dominant-negative PKR (TgDNPKR) mutant specifically in hematopoietic tissues. Significantly, peripheral blood counts from TgPKR mice decrease with age in association with dysplastic marrow changes. TgPKR mice have reduced colony-forming capacity and the colonies also are more sensitive to hematopoietic stresses. Furthermore, TgPKR mice have fewer hematopoietic stem/progenitor cells (HSPCs), and the percentage of quiescent (G0) HSPCs is increased. Importantly, treatment of TgPKR bone marrow (BM) with a PKR inhibitor specifically rescues sensitivity to growth factor deprivation. In contrast, marrow from PKR knockout (PKRKO) mice has increased potential for colony formation and HSPCs are more actively proliferating and resistant to stress. Significantly, TgPKR HSPCs have increased expression of p21 and IFN regulatory factor, whereas cells from PKRKO mice display mechanisms indicative of proliferation such as reduced eukaryotic initiation factor 2α phosphorylation, increased extracellular signal-regulated protein kinases 1 and 2 phosphorylation, and increased CDK2 expression. Collectively, data reveal that PKR is an unrecognized but important regulator of HSPC cell fate and may play a role in the pathogenesis of BM failure.

The effect of Cr doping on magnetic and electrical properties in the bilayer manganites La1.4Sr1.6(Mn1−yCry)2O7 (y=0–0.1) has been investigated. When y≤0.025, Cr doping enhances the three-dimensional magnetic transition temperature TC and the insulator–metal transition temperature TIM as well as decreases the peak resistivity at TIM, and the saturated magnetization decreases slightly. When y≥0.035, TIM decreases gradually accompanied by the increase of peak resistivity, but TC remains nearly constant, and the saturated magnetization decreases heavily. In the whole doping region, the two-dimensional magnetic transition temperature T⁎ monotonously decreases with an increasing of Cr doping level. These results can be explained by considering different magnetic (including ferromagnetic and antiferromagnetic) interactions between Mn ions and Cr ions.

Total phenolic content (TPC) and antioxidant capacity of lettuce were evaluated using the Folin–Ciocalteu method and DPPH assay, respectively, on 25 cultivars of lettuce, including leaf, romaine, crisphead, and butterhead types, cultivated over two harvest periods. Leaf lettuce possessed the highest TPC and highest DPPH scavenging ability, followed by romaine, butterhead and Batavia (crisphead subtype). Within a lettuce type, red pigmented lettuce cultivars had higher TPC and antioxidant capacity than did green cultivars grown under the same conditions. In addition, lettuce harvested in July possessed higher TPC and antioxidant capacity than did lettuce harvested in September, suggesting that environmental conditions could influence the phenolic content and antioxidant activity of lettuce. These results suggest that Colorado grown lettuce may serve as potential dietary sources of natural phenolic antioxidants.

To investigate microvessel density (MVD), vascular endothelial growth factor (VEGF), endothelial nitric oxide synthase (eNOS), and interleukin-6 (IL-6) mRNA expression in peritoneal tissues, and their relationships with baseline peritoneal transport in uremia. Thirty uremic patients with a peritoneal dialysis catheter were selected in the Department of Nephrology in Dalian Central Hospital, Liaoning, China between 2010 and 2012. Peritoneal specimens were harvested for assessment of MVD, VEGF, eNOS, and IL-6 mRNA expression. One month after continuous ambulatory peritoneal dialysis, a peritoneal equilibration test was conducted. According to the 4-h peritoneal dialysate and plasma creatinine ratio (D/P Cr), patients were divided into high (n = 16) and low (n = 14) transport groups. General clinical data of high and low transport groups were similar (P > 0.05). The MVD in peritoneal tissues was significantly higher in the high than in the low transport group (P

Reduced expression and activity of the proapoptotic, double-stranded RNA-dependent protein kinase, PKR (protein kinase R) is observed in breast, lung and various leukemias, suggesting that loss of PKR potentiates transformation. Now we report that decreased PKR activity inhibits chemotherapy-induced apoptosis of leukemia cells both in vitro and in vivo. Inhibition of PKR expression or activity reduces protein phosphatase 2A (PP2A) activity, a B-cell lymphoma 2 (Bcl-2) phosphatase, resulting in enhanced Bcl-2 phosphorylation. Thus, inhibition of PKR activity leads to hyperphosphorylation of Bcl-2, stabilization of Bcl-2/Bax interaction and decreased Bax insertion into the outer mitochondrial membrane. Treatment with the PP2A activator, FTY720, restores Bcl-2 dephosphorylation and apoptosis in cells with reduced PKR expression following stress. Significantly, xenografts of REH leukemic cells with reduced PKR display significantly increased tumor volume, increased resistance to doxorubicin treatment and shorter survival. Importantly, FTY720 treatment restores sensitivity to chemotherapy and prolongs overall survival of these mice. Collectively, these findings suggest that PP2A activation is a downstream target of PKR and the PKR/PP2A signaling axis is required for rapid and potent stress-induced apoptosis. Importantly, loss of PKR promotes leukemia progression and may serve as a biomarker for predicting chemosensitivity.

The interferon-inducible, dsRNA-dependent protein kinase, PKR, is activated by diverse cellular stresses. Loss of PKR expression or activity has been found in tumors including various types of leukemia, suggesting that loss of PKR may contribute to malignancy. However, no convincing conclusion has been reached regarding the relationship between PKR and leukemia progression. We analyzed 511 AML primary bone marrow samples by reverse phase protein array (RPPA) and discovered that PKR protein expression is > 50% reduced in CD34+ blasts from AML patients compared to CD34+ cells from healthy donors. This finding prompted us to further study how PKR affected progression of leukemia. PKR expression was either knocked down (by 82%) by specific SiRNA or directly inhibited by specific PKR inhibitor in leukemia cell lines. Significantly, inhibition of PKR decreased the rate of apoptosis following treatment with hydrogen peroxide (H2O2) or doxorubicin for 48 hours. In addition, bone marrow mononuclear cells form wild type, PKR null (-/-) or PKR transgenic mice were harvested and subjected to growth factor withdrawal for up to 120 hours. Cells isolated from PKR null mice had decreased apoptosis while cells from PKR transgenic mice had increased apoptosis compared to cells from wild type littermate mice. Further investigation showed that PKR inhibition led to reduction of PP2A activity, and a 2.4 fold increase in Bcl-2 phosphorylation. In addition, okadaic acid (an inhibitor of PP2A) had a similar effect to PKR inhibition on both Bcl-2 phosphorylation (1.74 fold increase) and stress-induced apoptosis. By contrast, treatment of cells with a PP2A activator, FTY 720, resulted in decreased Bcl-2 phosphorylation (by 2.24 fold) and enhanced apoptosis in response to H2O2. Furthermore, our study showed that following PKR inhibition, phosphorylation of Bcl-2 stabilized Bcl-2/Bax complex following H2O2 treatment, preventing Bax from being released, anchoring and perforating the mitochondrial outer membrane, which would initiate apoptotic process. To confirm our in vitro findings, a leukemia xenograft model was established in NSG mice using REH cells expressing luciferase. Mice receiving REH SiPKR cells displayed significantly faster tumor progression compared to mice receiving Sicontrol cells. Furthermore, tumors from REH SiPKR cells were more resistant to doxorubicin treatment compared to tumors from REH sicontrol cells. Taken together our results demonstrate that loss of PKR function inhibits PP2A-dependent Bcl-2 dephosphorylation resulting in increased leukemia progression and survival. In addition, since our studies indicate that PKR activation enhances the effectiveness of chemotherapy, PKR level/function status may serve as a clinical predictor of prognosis, disease relapse and resistance to chemotherapy. Citation Format: Xiaodong Cheng, Richard L. Bennett, Xiangfei Liu, Stratford May. Loss of PKR promotes leukemia progression by activating Bcl-2 to inhibit apoptosis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4606. doi:10.1158/1538-7445.AM2013-4606

It has been reported that the expression and activity of the interferon-inducible, dsRNA-dependent protein kinase, PKR, is increased in mammary carcinoma cell lines and primary tumor samples. To extend these findings and determine how PKR signaling may affect breast cancer cell sensitivity to chemotherapy, we measured PKR expression by immunohistochemical staining of 538 cases of primary breast cancer and normal tissues. Significantly, PKR expression was elevated in ductal, lobular and squamous cell carcinomas or lymph node metastases but not in either benign tumor specimens or cases of inflammation compared to normal tissues. Furthermore, PKR expression was increased in precancerous stages of mammary cell hyperplasia and dysplasia compared to normal tissues, indicating that PKR expression may be upregulated by the process of tumorigenesis. To test the function of PKR in breast cancer, we generated MCF7, T-47D and MDA-MB-231 breast cancer cell lines with significantly reduced PKR expression by siRNA knockdown. Importantly, while knockdown of PKR expression had no effect on cell proliferation under normal growth conditions, MCF7, T-47D or MDA-MB-231 cells with reduced PKR expression or treated with a small molecule PKR inhibitor were significantly less sensitive to doxorubicin or H(2)O(2)-induced toxicity compared to control cells. In addition, the rate of eIF2α phosphorylation following treatment with doxorubicin was delayed in breast cancer cell lines with decreased PKR expression. Significantly, treatment of breast cancer lines with reduced PKR expression with either interferon-α, which increases PKR expression, or salubrinal, which increases eIF2α phosphorylation, restored doxorubicin sensitivity to normal levels. Taken together these results indicate that increased PKR expression in primary breast cancer tissues may serve as a biomarker for response to doxorubicin-containing chemotherapy and that future therapeutic approaches to promote PKR expression/activation and eIF2α phosphorylation may be beneficial for the treatment of breast cancer.

The Interferon-induced double-stranded RNA-activated protein kinase (PKR) was originally found to be activated by viral infection leading to inhibition of new protein synthesis and apoptosis. Recently multiple studies have reported decreased expression and function of PKR in leukemia cells, although the exact role of PKR in the process of oncogenesis remains unclear. Since leukemias are clonal disorders originating in a primitive pluripotent leukemic stem cell (LSC) which is regulated by multi-potential hematopoietic growth factors including interleukin-3 (IL-3). Thus, studying how PKR regulates apoptosis in IL-3 dependent (CD123+) pluripotent stem/progenitor cells is expected to reveal PKR's role in leukemia. PKR expression was either knocked down (by 78%) using a specific ShRNA or directly inhibited (57%) by treating cells with a specific PKR inhibitor. Results revealed that inhibition of PKR was associated with a reduction of PP2A activity, a 2.3 fold increase in Bcl-2 phosphorylation and inhibition of apoptosis following treatment with 100uM hydrogen peroxide for 48 hours. In addition, okadaic acid (an inhibitor of PP2A) had a similar effect to PKR inhibition on both Bcl-2 phosphorylation (1.74 fold increase) and stress-induced apoptosis. By contrast, treatment of cells with the PP2A activator, FTY 720, resulted in decreased Bcl-2 phosphorylation (by 2.24 fold) and enhanced apoptosis in response to treatment of cells with chemotherapy (etoposide or doxorubicin) or hydrogen peroxide. To test the role of PKR in growth factor- withdrawal mediated apoptosis in bone marrow stem/progenitor cells, CD123+(IL-3R[[Unsupported Character - Symbol Font ]]+) bone marrow cells from wild type, PKR transgenic (over-expressed) and PKR null(-/-) mice were exposed to different stresses including IL-3 deprivation or hydrogen peroxide. CD123+ cells isolated from PKR null mice had a significantly lower percentage of apoptosis while CD123+ cells from PKR transgenic mice had increased apoptosis compared to cells from wild type littermate mice. These results demonstrate that PKR negatively affects hematopoietic cell survival by enhancing apoptosis following stress in a mechanism that involves PP2A activation and Bcl2 dephosphorylation. Taken together these results may indicate that inhibition of PKR's pro-apoptotic function in bone marrow pluripotent stem/progenitor cells may contribute to tumorigenesis in hematopoietic malignancies. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2009. doi:1538-7445.AM2012-2009

Double stranded (ds) RNA activated protein kinase PKR was discovered over 30 years ago as a downstream mediator of the interferon signal pathway. Initial studies focused on PKR's anti-virus and pro-inflammatory effects. More recently, PKR has been found to be involved in signal transduction pathways critical for the negative regulation of cell growth and initiation of apoptosis. Significantly, loss of PKR expression/activity has been associated with increased growth of human breast carcinoma, B-cell CLL, and T-cell ALL using patient samples. Furthermore, PKR activity is increased in bone marrow progenitor cells isolated from patients with myelodysplastic syndrome (MDS) which undergo significant apoptosis compared to normal marrow progenitors not observed when evolved to AML. Thus, in the process of MDS evolution to leukemia, PKR expression/activity may be downregulated by a process that is currently unclear. To explore the role of PKR in MDS/AML we have constructed a transgenic mouse model expressing human PKR (TgPKR) under control of vav regulatory elements to drive expression specifically in hematopoietic-tissue. We hypothesized transgenic expression of human PKR would exert an inhibitory effect on mouse hematopoiesis, cause ineffective blood cell production, and thus simulate a disease process similar to human MDS. In our preliminary data, quantitative PCR and immunoblotting results confirm that human PKR is specifically expressed in spleen, thymus and bone marrow (BM) but not in liver, kidney, intestine, muscle and other tissues outside hematopoiesis in TgPKR mice. Aberrant activation of transcription factor STAT1 and MAPKinases ERK1/2 occurs in hematopoietic cells from the TgPKR mice. In addition, inhibitory cytokines including interferon gamma, CXCL9 and 10 are overexpressed. Initial characterization of BM cells from TgPKR mice indicates a higher level of uninduced/basal apoptosis in growth medium and increased apoptosis upon withdrawal of growth factors compared to cells from wild type (WT) littermates. Furthermore, BM cells from TgPKR mice display >20% reduced colonies (CFU-GM and CFU-GEMM) which contain 40% less cells than those from WT littermates. Significantly, histopathological analysis of BM and spleen from TgPKR mice reveals a ∼20% increase in bone marrow cellularity with a striking increase in dysplastic megakaryocytes that are either small without nuclear lobation or of normal size but with hypolobation or separated nuclear lobes. Taken together our results demonstrate that transgenic expression of hPKR induces an MDS-like condition in mice characterized by reduced BM cell survival, increased apoptosis and reduced proliferation and differentiation of BM progenitor/stem cells. Thus, increased PKR expression in patients may, at least in part, lead to a BM failure state similar to MDS. Thus, future anti-MDS therapies that inhibit PKR may be clinically useful. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1350. doi:1538-7445.AM2012-1350