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Hypoxia leads to significant cellular stress that has diverse pathological consequences such as cardiovascular diseases and cancers. MicroRNAs (miRNAs) are one of regulators of the adaptive pathway in hypoxia. We identified a hypoxia-induced miRNA, miR-34c, that was significantly upregulated in hypoxic human umbilical cord vein endothelial cells (HUVECs) and in murine blood vessels on day 3 of hindlimb ischemia (HLI). miR-34c directly inhibited BCL2 expression, acting as a toggle switch between apoptosis and autophagy

This file contains Supplemental figures, tables, and legends that accompany the main figures, as indicated in each legend. There are 3 supplemental tables and 6 supplemental figures. Table S1. List of potential target of the GNS compounds that were profiled in Figure 1 of the main text. Table S2. Half-maximal inhibitor concentration of the EGFR inhibitors tested. Table S3. Blood-brain-barrier penetration of the GNS compounds as assessed by the indicated pharmacokinetic measurements. Figure S1. Structural modeling studies show the binding characteristics of the indicated EGFR inhibitors. Figure S2. The effects of each EGFR inhibitor in the cell lines expressing mutant or wild type EGFR. Figure S3. In vivo effects of the GNS compounds. Figure S4. In vivo anti-tumor effects of the GNS compounds. Figure S5. In vivo anti-tumor effects of the GNS compounds in intracranial EGFR-mutant lung cancer. Figure S6. In vivo anti-tumor effects of the indicated EGFR inhibitors in intracranial EGFR-mutant lung cancer.

The clinical utility of approved EGFR small-molecule kinase inhibitors is plagued both by toxicity against wild-type EGFR and by metastatic progression in the central nervous system, a disease sanctuary site. Here, we report the discovery and preclinical efficacy of GNS-1486 and GNS-1481, two novel small-molecule EGFR kinase inhibitors that are selective for T790M-mutant isoforms of EGFR. Both agents were effective in multiple mouse xenograft models of human lung adenocarcinoma (T790M-positive or -negative), exhibiting less activity against wild-type EGFR than existing approved EGFR kinase inhibitors (including osimertinib). In addition, GNS-1486 showed superior potency against intracranial metastasis of EGFR-mutant lung adenocarcinoma. Our results offer a preclinical proof of concept for new EGFR kinase inhibitors with the potential to improve therapeutic index and efficacy against brain metastases in patients. Cancer Res; 77(5); 1200–11. ©2017 AACR.

C1q/tumor necrosis factor–related protein 9 (CTRP9) is an adipokine and has high potential as a therapeutic target. However, the role of CTRP9 in cardiovascular disease pathogenesis remains unclear. We found CTRP9 to induce HDAC7 and p38 MAPK phosphorylation via tight regulation of AMPK in vascular endothelial cells, leading to angiogenesis through increased MEF2 activity. The expression of CTRP9 and atheroprotective MEF2 was decreased in plaque tissue of atherosclerotic patients and the ventricle of post-infarction mice. CTRP9 treatment inhibited the formation of atherosclerotic plaques in ApoE KO and CTRP9 KO mice. In addition, CTRP9 induced significant ischemic injury prevention in the post-MI mice. Clinically, serum CTRP9 levels were reduced in patients with MI compared with healthy controls. In summary, CTRP9 induces a vasoprotective response via the AMPK/HDAC7/p38 MAPK pathway in vascular endothelial cells, whereas its absence can contribute to atherosclerosis and MI. Hence, CTRP9 may represent a valuable therapeutic target and biomarker in cardiovascular diseases.

Purpose MicroRNA could be biomarker and therapeutic target for rejection. The aim of this study was to investigate the role of miR-142-5p in allogeneic immune responses using in vitro and in vivo models. Materials and Methods Primary and immortalized human umbilical vein endothelial cells (HUVECs) were cultured with unrelated blood mononuclear cells to induce allogeneic immune responses. Syngeneic and allogeneic skin graft was performed in mice. Flow cytometry, quantitative reverse transcription–polymerase chain reaction, and Western blotting was performed to understand the underlying mechanisms. Results miR-142-5p was up-regulated in primary HUVEC and a HUVEC line when allogeneic immune responses were elicited. miR-142-5p was also up-regulated in the murine allogeneic skin graft. Overexpression of miR-142-5p in HUVEC increased the expression of HLA-ABC and HLA-DR additively to allogeneic immune responses, suggesting a possible increase in alloantigen presentation. Inhibition of miR-142-5p reduced the expression of HLA-DR. ZEB1, a putative target gene of miR-142-5p, was down-regulated in HUVEC on allogeneic immune response as well as in murine allogeneic skin graft. Conclusion These results suggest that the up-regulation of miR-142-5p on allogeneic immune response might facilitate endothelial activation to exacerbate rejection.

The role of yeast-derived β-glucan in angiogenesis has not been elucidated because there have been few specific studies on its clinical and physiological significance. Therefore, this study investigated the correlation between β-glucan and histone deacetylase 5 (HDAC5) in human umbilical vein endothelial cells (HUVECs), revealing the role of β-glucan in angiogenesis. We confirmed that HDAC5 was phosphorylated by β-glucan stimulation and released from the nucleus to the cytoplasm. Furthermore, we found that β-glucan-stimulated HDAC5 translocation mediates the transcriptional activation of MEF2. As a result, the expression of KLF2, EGR2, and NR4A2, whose expression is MEF2-dependent and involved in angiogenesis, increased. Thus, we showed the activity of β-glucan in angiogenesis through in vitro and ex vivo assays including cell migration, tube formation, and aortic ring analyses. Specifically, application of an HDAC5 inhibitor repressed MEF2 transcriptional activation in both in vitro and ex vivo angiogenesis. HDAC5 inhibitor LMK235 inhibited the proangiogenic activity of beta-glucan, suggesting that β-glucan induces angiogenesis through HDAC5. These findings suggest that HDAC5 is essential for angiogenesis, and that β-glucan induces angiogenesis. In conclusion, this study demonstrates that β-glucan induces angiogenesis through HDAC5. It also suggests that β-glucan has potential value as a novel therapeutic agent for modulating angiogenesis.

Background Epigenetic regulations frequently appear in Glioblastoma (GBM) and are highly associated with metabolic alterations. Especially, Histone deacetylases (HDACs) correlates with the regulation of tumorigenesis and cell metabolism in GBM progression, and HDAC inhibitors report to have therapeutic efficacy in GBM and other neurological diseases; however, GBM prevention and therapy by HDAC inhibition lacks a mechanism in the focus of metabolic reprogramming. Methods HDAC2 highly express in GBM and is analyzed in TCGA/GEPIA databases. Therefore, HDAC2 knockdown affects GBM cell death. Analysis of RNA sequencing and qRT-PCR reveals that miR-3189 increases and GLUT3 decreases by HDAC2 knockdown. GBM tumorigenesis also examines by using in vivo orthotopic xenograft tumor models. The metabolism change in HDAC2 knockdown GBM cells measures by glucose uptake, lactate production, and OCR/ECAR analysis, indicating that HDAC2 knockdown induces GBM cell death by inhibiting GLUT3. Results Notably, GLUT3 was suppressed by increasing miR-3189, demonstrating that miR-3189-mediated GLUT3 inhibition shows an anti-tumorigenic effect and cell death by regulating glucose metabolism in HDAC2 knockdown GBM. Conclusions Our findings will demonstrate the central role of HDAC2 in GBM tumorigenesis through the reprogramming of glucose metabolism by controlling miR-3189-inhibited GLUT3 expression, providing a potential new therapeutic strategy for GBM treatment.

Background: Tumor-derived exosomal microRNAs are key elements of the cell-cell communications response to lots of stimuli. However, various functions of the exosome in tumor suppression by radiotherapy (RT) are not clearly understood. Our study showed a previously unknown interaction of p53 and histone deacetylase 5 (HDAC5) by radiation exposure in hepatocellular carcinoma (HCC). Methods: Using serial ultracentrifugation methods, radiation and non-radiation exosomes were purified to investigate the radioresistance of miRNA151a-3p. Radiation doses were treated in 2gy and 4gy using radiation equipment X-RAD 320 to observe the expression of HDAC5 and p53 in hepatic cancer cells. Exosomal miRNA bioinformatics analysis was conducted to find a variation in the miRNA configuration inside Exosome after radiation exposure.Results: HDAC5 and p53 interacted by exposure to radiation, which increased exosome release and altered microRNAs' composition within exosomes. Also, we have described the intercommunication occurring between irradiated and untreated cells via exosomal microRNAs that affect tumor proliferation. In particular, the expression of exosomal microR151a-3p was markedly reduced by radiation treatment. We confirmed that inhibition of exosomal microR151a-3p promotes suppression of non-irradiated cancer cells, thereby increasing RT sensitivity. Conclusion: our present findings demonstrated HDAC5 is a key component of the p53-mediated release of exosomes resulting in tumor suppression through exosomal microRNA-151a-3p in response to radiation. Finally, we highlight the important role of exosomal microRNA 151a-3p as a biomarker in enhancing RT sensitivity.

Despite the strong influence of the gut microbiota on atherosclerosis, a causal relationship between atherosclerosis pathophysiology and gut microbiota is still unverified. This study was performed to determine the impact of the gut microbiota on the pathogenesis of atherosclerosis caused by genetic deficiency. To elucidate the influence of the gut microbiota on atherosclerosis pathogenesis, an atherosclerosis-prone mouse model (C1q/TNF-related protein 9-knockout (CTRP9-KO) mice) was generated. The gut microbial compositions of CTRP9-KO and WT control mice were compared. Fecal microbiota transplantation (FMT) was performed to confirm the association between gut microbial composition and the progression of atherosclerosis. FMT largely affected the gut microbiota in both CTRP9-KO and WT mice, and all transplanted mice acquired the gut microbiotas of the donor mice. Atherosclerotic lesions in the carotid arteries were decreased in transplanted CTRP9-KO mice compared to CTRP9-KO mice prior to transplantation. Conversely, WT mice transplanted with the gut microbiotas of CTRP9-KO mice showed the opposite effect as that of CTRP9-KO mice transplanted with the gut microbiotas of WT mice. Here, we show that CTRP9 gene deficiency is related to the distribution of the gut microbiota in subjects with atherosclerosis. Transplantation of WT microbiotas into CTRP9-KO mice protected against the progression of atherosclerosis. Conversely, the transplantation of CTRP9-KO microbiotas into WT mice promoted the progression of atherosclerosis. Treating atherosclerosis by restoring gut microbial homeostasis may be an effective therapeutic strategy.

Mannoproteins (MPs) are a major component of yeast cell walls and consist of high levels of mannose in covalent complexes with proteins. MPs complexly enhance the immune system. We previously isolated a mutant yeast, K48L3, with a higher yield of MP from its cell wall than wild-type Saccharomyces cerevisiae, YPH499. We determined that K48L3 induces the release of nitric oxide in macrophage cells. The present study reports nitric-oxide-mediated angiogenesis by MP from K48L3 and the induction of the Akt/eNOS signal pathway. Western blotting and RT-PCR were used to demonstrate that MP treatment resulted in the upregulation of p-Akt, p-eNOS, and angiogenesis-mediated gene expression. Moreover, the angiogenesis activity of the MPs was demonstrated using three angiogenesis assays, namely, a cell migration assay, a tube-forming assay, and an ex vivo aorta ring assay. Thus, this study demonstrates for the first time that MPs from S. cerevisiae K48L3 induce angiogenesis in HUVECs via the Akt-eNOS-dependent signaling pathway.

// Je-won Ryu 1, * , Sung Sik Choe 2, * , Seung-Hee Ryu 1 , Eun-Young Park 1 , Byoung Wook Lee 3 , Tae Keun Kim 4 , Chang Hoon Ha 3 , Sang-wook Lee 1 1 Department of Radiation Oncology, Asan Medical Center and University of Ulsan College of Medicine, Seoul, Republic of Korea 2 Department of Biological Sciences, Institute of Molecular Biology and Genetics, National Creative Research Institutive Center for Adipose Tissue Remodeling, Seoul National University, Seoul, Republic of Korea 3 Asan Institute for Life Science, Asan Medical Center and University of Ulsan College of Medicine, Seoul, Republic of Korea 4 Department of Life Science, College of Natural Science, Hallym University, Kyeongki Province, Republic of Korea * These authors contributed equally to this work Correspondence to: Chang Hoon Ha, email: chhoonha@gmail.com Sang-Wook Lee, email: lsw@amc.seoul.kr Keywords: Epidermal Growth Factor (EGF), Purinergic Receptor 2(P2Y), Redox Factor-1(Ref-1), Zinc Finger-containing Transcriptional Regulator 1(EGR1), Phosphatase and Tensin Homolog (PTEN) Received: September 30, 2016 Accepted: November 24, 2016 Published: December 07, 2016 ABSTRACT Epidermal growth factor (EGF) signaling promotes cell proliferation and survival in several types of cancer. Here, however, we showed that EGF inhibits proliferation and promotes programmed cell death in non-small cell lung cancer (NSCLC) cells. In A549 cells, EGF increased redox factor-1 (Ref-1) expression and the association of Ref-1 with zinc finger-containing transcriptional regulator (EGR1) via activation of p22 phox , RAC1, and an NADPH oxidase subunit. EGF increased p22 phox and RAC1 expression through activation of purinergic receptors (P2Y). Elevated Ref-1/EGR1 levels increased phosphatase and tensin homolog (PTEN) levels, leading to inhibition of the Akt pathway. EGF-induced PTEN upregulation increased apoptosis and autophagy-induced damage in A549 cells, whereas Ref-1 knockdown blocked EGF-induced PTEN upregulation in an NADPH oxidase p22 phox subunit-independent manner. In addition, p22 phox knockdown restored EGF-induced effects, implying that changes in P2Y activity caused by EGF, which activates NADPH oxidase via RAC1, influenced Ref-1-mediated redox regulation. Finally, EGF similarly attenuated cell proliferation and promoted autophagy and apoptosis in vivo in a xenograft model using A549 cells. These findings reveal that EGF-induced redox signaling is linked to Ref-1-induced death in NSCLC cells.

Tube integrity is an important aspect for safe and reliable operation of nuclear power plant steam generators. As a U.S. industry and licensing requirement, all in-service steam generator tubes shall retain structural integrity over the full range of normal operating conditions and design basis accidents by meeting the structural integrity performance criterion (SIPC) as given in NEI 97-06. The SIPC margin shall be maintained during plant operation between tube examinations. The burst strength of tubes subjected to wall thinning will depend on the extent and mode of degradation, and the magnitude of design loads to include pressure differential across the tube wall during normal operation and postulated accident conditions. In addition, non-pressure loads that can occur during postulated accident events shall be evaluated and included in the assessment of tube integrity if determined to significantly reduce the tube burst strength. The EPRI Flaw Handbook provides burst pressure relationships for flaws which include a reduction factor that accounts for the effect of applied bending stress on circumferential degradation. However, this previous industry work was only for planar crack-like flaws and did not directly address uniform volumetric wall loss which can have both axial and circumferential extent. This paper describes a test program to determine the effect of bending loads on the burst pressure of a tube with uniform thinning over a given axial length. The uniform thinning geometry was selected since it represented a bounding case of general wall loss and is conservative for calculating a tube repair limit for volumetric degradation for a given steam generator design. Tube repair limits are required for defining an upper limit on in-service degradation for which a tube is to be removed from service. Tube repair limits are cited in the Plant Technical Specifications, which is an important part of the licensing basis.

In the event of a mass casualty radiation scenario, biodosimetry has the potential to quantify individual exposures for triaging and providing dose-appropriate medical intervention. Structural maintenance of chromosomes 1 (SMC1) is phosphorylated in response to ionizing radiation. The goal of this study was to develop a new biodosimetry method using SMC1 phosphorylation as a measure of exposure to radiation. In the initial experiments, two normal human cell lines (WI-38VA-13 and HaCaT) and four lymphoblastoid cell lines were irradiated, and the levels of SMC1 phosphorylation at Ser-360 and Ser-957 were assessed using Western blotting. Subsequently, similar experiments were performed using peripheral blood mononuclear cells (PBMCs) obtained from 20 healthy adults. Phosphorylation of SMC1 at Ser-957 and Ser-360 was increased by exposure in a dose-dependent manner, peaked at 1-3 h postirradiation and then decreased gradually. Ser-360 was identified as a new phosphorylation site and was more sensitive to radiation than Ser-957, especially at doses below 1 Gy. Our results demonstrate a robust ex vivo response of phospho-SMC1-(Ser-360) to ionizing radiation in human PBMCs. Detection of phosphorylation at Ser-360 in SMC1 could be used as a marker of radiation exposure. Our findings suggest that it is feasible to measure blood cell-based changes in the phosphorylation level of a protein as an ex vivo radiation exposure detection method, even after low-dose exposure.

The rat is a time-honored traditional experimental model animal, but its use is limited due to the difficulty of genetic modification. Although engineered endonucleases enable us to manipulate the rat genome, it is not known whether the newly identified endonuclease Cpf1 system is applicable to rats. Here we report the first application of CRISPR-Cpf1 in rats and investigate whether Apoe knockout rat can be used as an atherosclerosis model. We generated Apoe- and/or Ldlr-deficient rats via CRISPR-Cpf1 system, characterized by high efficiency, successful germline transmission, multiple gene targeting capacity, and minimal off-target effect. The resulting Apoe knockout rats displayed hyperlipidemia and aortic lesions. In partially ligated carotid arteries of rats and mice fed with high-fat diet, in contrast to Apoe knockout mice showing atherosclerotic lesions, Apoe knockout rats showed only adventitial immune infiltrates comprising T lymphocytes and mainly macrophages with no plaque. In addition, adventitial macrophage progenitor cells (AMPCs) were more abundant in Apoe knockout rats than in mice. Our data suggest that the Cpf1 system can target single or multiple genes efficiently and specifically in rats with genetic heritability and that Apoe knockout rats may help understand initial-stage atherosclerosis.

Shikonin, which derives from Lithospermum erythrorhizon, has been traditionally used against a variety of diseases, including cancer, in Eastern Asia. Here we determined that shikonin inhibits proliferation of gastric cancer cells by inducing apoptosis. Shikonin's biological activity was validated by observing cell viability, caspase 3 activity, reactive oxygen species (ROS) generation, and apoptotic marker expressions in AGS stomach cancer cells. The concentration range of shikonin was 35-250 nM with the incubation time of 6 h. Protein levels of Nrf2 and p53 were evaluated by western blotting and confirmed by real-time PCR. Our results revealed that shikonin induced the generation of ROS as well as caspase 3-dependent apoptosis. c-Jun-N-terminal kinases (JNK) activity was significantly elevated in shikonin-treated cells, thereby linking JNK to apoptosis. Furthermore, our results revealed that shikonin induced p53 expression but repressed Nrf2 expression. Moreover, our results suggested that there may be a co-regulation between p53 and Nrf2, in which transfection with siNrf2 induced the p53 expression. We demonstrated for the first time that shikonin activated cell apoptosis in AGS cells via caspase 3- and JNK-dependent pathways, as well as through the p53-Nrf2 mediated signal pathway. Our study validates in partly the contribution of shikonin as a new therapeutic approaches/ agent for cancer chemotherapy.

Programmed cell death 5 (PDCD5) has been associated with human cancers as a regulator of cell death; however, the role of PDCD5 in the endothelium has not been revealed. Thus, we investigated whether PDCD5 regulates protein kinase B (PKB/AKT)-endothelial nitric oxide synthase (eNOS)–dependent signal transduction in the endothelium and affects atherosclerosis. Endothelial-specific PDCD5 knockout mice showed significantly reduced vascular remodeling compared with wild-type (WT) mice after partial carotid ligation. WT PDCD5 competitively inhibited interaction between histone deacetylase 3 (HDAC3) and AKT, but PDCD5L6R, an HDAC3-binding–deficient mutant, did not. Knockdown of PDCD5 accelerated HDAC3–AKT interaction, AKT and eNOS phosphorylation, and nitric oxide (NO) production in human umbilical vein endothelial cells. Moreover, we found that serum PDCD5 levels reflect endothelial NO production and are correlated with diabetes mellitus, high-density lipoprotein cholesterol, and coronary calcium in human samples obtained from the cardiovascular high-risk cohort. Therefore, we conclude that PDCD5 is associated with endothelial dysfunction and may be a novel therapeutic target in atherosclerosis.

Natural compounds are becoming important candidates in cancer therapy due to their cytotoxic effects on cancer cells by inducing various types of programmed cell deaths. In this study, we investigated whether genipin induces programmed cell deaths and mediates in Egr1/p21 signaling pathways in gastric cancer cells. Effects of genipin in AGS cancer cell lines were observed via evaluation of cell viability, ROS generation, cell cycle arrest, and protein and RNA levels of p21, Egr1, as well as apoptotic marker genes. The cell viability of AGS cells reduced by genipin treatment via induction of the caspase 3-dependent apoptosis. Cell cycle arrest was observed at the G2/M phase along with induction of p21 and p21-dependent cyclins. As an upstream mediator of p21, the transcription factor early growth response-1 (Egr1) upregulated p21 through nuclear translocation and binding to the p21 promoter site. Silencing Egr1 expression inhibited the expression of p21 and downstream molecules involved in apoptosis. We demonstrated that genipin treatment in AGS human gastric cancer cell line induces apoptosis via p53-independent Egr1/p21 signaling pathway in a dose-dependent manner.

Roles of dietary phytochemicals in cancer chemoprevention via induction of nuclear factor-erythroid-2-related factor 2 (Nrf2)-mediated antioxidant enzymes have been well established in a number of studies. In this study, FACS analysis was used to reveal that the intracellular reactive oxygen species level decreased at 0-25 μM of genipin treatment. Furthermore, immunofluorescence analysis and Western blotting were used to demonstrate that genipin treatment resulted in the upregulation and nuclear translocation of Nrf2, as well as upregulation of gastrointestinal glutathione peroxidase. Finally, we found that C-Jun-NH2-kinase (JNK) was also dose-dependently activated, where depleting JNK by using a biochemical inhibitor indicated that JNK was upstream of Nrf2. Interestingly, the antioxidant effects were limited to the treatment in the lower dosage of genipin, where higher dosage of genipin treatment resulted in the increased reactive oxygen species level and cytotoxicity. Thus, this study demonstrates for the first time that lower dosage of genipin results in the induction of JNK/Nrf2/ARE signaling pathway and protection from cell death.

The flow induced vibration occurs frequently in a steam generator in the nuclear power plant. The large-scale steam generator has a large number of tube supports whose cell has rhombus-type shape, and there is a tiny clearance between tube and its support grid. The damping is very complex because of non-linearity and randomness. The experiment for damping was performed to investigate it with a number of 13 support spans both in air and water environment. The lower part of multi-span fixture was excited by root-mean-square random force with the range of 1∼10 newton to get the frequency response function. The half-power bandwidth method was applied to obtain the damping ratio. The sensitivity of a number of spans was investigated in the range of 9 ∼ 13. In addition, the damping was reviewed from a comparison with Pettigrew [1∼4] and ASME B&PV Code [5].

The clinical utility of approved EGFR small-molecule kinase inhibitors is plagued both by toxicity against wild-type EGFR and by metastatic progression in the central nervous system, a disease sanctuary site. Here, we report the discovery and preclinical efficacy of GNS-1486 and GNS-1481, two novel small-molecule EGFR kinase inhibitors that are selective for T790M-mutant isoforms of EGFR. Both agents were effective in multiple mouse xenograft models of human lung adenocarcinoma (T790M-positive or -negative), exhibiting less activity against wild-type EGFR than existing approved EGFR kinase inhibitors (including osimertinib). In addition, GNS-1486 showed superior potency against intracranial metastasis of EGFR-mutant lung adenocarcinoma. Our results offer a preclinical proof of concept for new EGFR kinase inhibitors with the potential to improve therapeutic index and efficacy against brain metastases in patients. Cancer Res; 77(5); 1200–11. ©2017 AACR.

Background Extracorporeal shock wave has been used in the noninvasive treatment of various diseases including musculoskeletal disorders. In particular, shock wave with low energy level showed anti-inflammatory effect and increased angiogenesis in ischemic tissues. However, the detailed cellular pathway in endothelial signaling is not fully understood. We investigate the role of shock wave with low energy level in angiogenic gene expression and underlying molecular mechanism by comparing the laminar and oscillatory fluid shear stresses in endothelial cells. Methods and results We show that shock wave with low energy level (0.012–0.045mJ/mm 2 ) stimulated phosphorylation of Akt, eNOS and Erk 1/2 in a time-dependent manner which is similar to the effect of laminar fluid shear stress. The transfection of endothelial cells with siRNA encoding VEGFR2, VE-cadherin and PECAM-1 inhibited shock wave-induced phosphorylation of Akt, eNOS and Erk 1/2 and angiogenic gene expressions, including Akt, eNOS, KLF2/4, and Nur77. Moreover, mechanical stimulation through extracorporeal shock wave induced endothelial cell migration and tube formation. Conclusions Our results demonstrate that shock wave-induced Akt/eNOS phosphorylation and angiogenic gene expression were mediated through the mechanosensory complex formation involving VEGFR-2, VE-cadherin and PECAM-1 which was similar to the effect of laminar shear stress.

In this study, we present an analysis and experiment on the fluid damping of a Steam Generator (SG) tube with one and several plate-type supports that are submerged under water. For the damping measurement, a short tube of 2.2 meters in length with one gap support as well as a long tube of up to 10 meters with 11 and 13 supports are used. To see a fluid effect through an analytical approach, Mulcahy’s theory on a tube with finite-length gap support and Pettigrew’s empirical correlation for SG design were reviewed. The theoretical model is good at predicting the fluid damping at the first mode in a low frequency range which is an inactive mode at the support, while the practical model is much better at the higher modes, which are active modes, rocking modes, at the support(s). Experimental results from the short length tube with single support shows good agreement with a previous measurement by other researchers. A theoretical model and empirical correlation are in good agreement. Our measurements from a longer tube with several supports are reasonable compared with the others. There is, however, a wide discrepancy between predictions by the two models. As compared with the whole measurements, the theoretical model seems to be a good guideline in a low frequency range, while an empirical model is relatively good guideline in a high frequency range. Based on the whole collected data, a new correlation modified from Pettigrew’s correlation is suggested.

EFC-1 integrase is a site-specific recombinase that belongs to the large family of serine recombinase. In previously study, we isolated the temperate phage EFC-1, and characterized its genomic sequence. Within its genome, Orf28 was predicted encode a 464 amino acid of a putative integrase gene. In this study, EFC-1 integrase was characterized in vitro and in vivo. In vitro assay was performed using purified His-tag fusion integrase. Also, to identify which serine is involved in the catalytic domain, we used site-directed mutagenesis and analyzed by a recombination assay in vitro. In vivo assay involved PCR and confocal microscopy in HEK293 cells, and determined the minimal lengths of attP and attB sites. According to our results, the EFC-1 integrase-mediated recombination was site-specific and unidirectional system in vitro and in vivo. Serine 21 of EFC-1 integrase plays a major role in the catalytic domain, and minimal sizes of attB and attP was defined 48 and 54 bp. Our finding may help develop a useful tool for gene therapy and gene delivery system.

Endothelial dysfunction, characterized by impaired activation of endothelial nitric oxide (NO) synthase (eNOS) and ensued decrease of NO production, is a common mechanism of various cardiovascular pathologies, including hypertension and atherosclerosis. Laminar blood flow-mediated specific signaling cascades modulate vascular endothelial cells (ECs) structure and functions. We have previously shown that flow-stimulated Gab1 (Grb2-associated binder-1) tyrosine phosphorylation mediates eNOS activation in ECs, which in part confers laminar flow atheroprotective action. However, the molecular mechanisms whereby flow regulates Gab1 tyrosine phosphorylation and its downstream signaling events remain unclear. Here we show that platelet endothelial cell adhesion molecule-1 (PECAM1), a key molecule in an endothelial mechanosensing complex, specifically mediates Gab1 tyrosine phosphorylation and its downstream Akt and eNOS activation in ECs upon flow rather than hepatocyte growth factor (HGF) stimulation. Small interfering RNA (siRNA) targeting PECAM1 abolished flow- but not HGF-induced Gab1 tyrosine phosphorylation and Akt, eNOS activation as well as Gab1 membrane translocation. Protein-tyrosine phosphatase SHP2, which has been shown to interact with Gab1, was involved in flow signaling and HGF signaling, as SHP2 siRNA diminished the flow- and HGF-induced Gab1 tyrosine phosphorylation, membrane localization and downstream signaling. Pharmacological inhibition of PI3K decreased flow-, but not HGF-mediated Gab1 phosphorylation and membrane localization as well as eNOS activation. Finally, we observed that flow-mediated Gab1 and eNOS phosphorylation in vivo induced by voluntary wheel running was reduced in PECAM1 knockout mice. These results demonstrate a specific role of PECAM1 in flow-mediated Gab1 tyrosine phosphorylation and eNOS signaling in ECs.

Vascular endothelial growth factor (VEGF) is essential for many angiogenic processes both in normal and pathological condition. However, the signaling pathways involved in VEGF-induced angiogenesis are incompletely understood. The protein kinase D1 (PKD1), a newly described the calcium/calmodulin-dependent serine/threonine kinase, has been implicated in cell migration, proliferation and membrane trafficking. Increasing evidence suggest critical roles for PKD1-mediated signaling pathways in the endothelial cells, particularly in the regulation of VEGF-induced angiogenesis. Recent studies show that class IIa histone deacetylases (HDACs) are PKD1 substrates and VEGF signal-responsive repressors of myocyte enhancer factor-2 (MEF2) transcriptional activation in endothelial cells. This review provides a guide on PKD1 signaling pathway and the direct downstream targets of PKD1 in VEGF signaling, and suggests important functions of PKD1 in angiogenesis.

Metabolic syndrome accelerates the atherosclerotic process, and the earliest event of which is endothelial dysfunction. Ghrelin, a newly discovered gastric peptide, improves endothelial function and inhibits proatherogenic changes. In particular, low ghrelin concentration has been associated with several features of metabolic syndrome, including obesity, insulin resistance, and high blood pressure. However, the molecular mechanisms underlying ghrelin vascular actions remain largely unclear. Here, we showed that ghrelin activated endothelial nitric oxide (NO) synthase (eNOS) in cultured endothelial cells (ECs) and in intact vessels. Specifically, ghrelin rapidly induced phosphorylation of eNOS on an activation site and production of NO in human umbilical vein ECs and bovine aortic ECs. The eNOS phosphorylation was also observed in mouse aortas ex vivo perfused with ghrelin and in aortic tissues isolated from mice injected with ghrelin. Mechanistically, ghrelin stimulated AMP-activated protein kinase (AMPK) and Akt activation in cultured ECs and intact vessels. Inhibiting AMPK and Akt with their pharmacological inhibitors, small interference RNA and adenoviruses carried dominant-negative mutants, markedly attenuated ghrelin-induced eNOS activation, and NO production. Furthermore, ghrelin receptor/Gq protein/calcium-dependent pathway mediates activation of AMPK, Akt, and eNOS, and calmodulin-dependent kinase kinase is a potential convergent point to regulate Akt and AMPK activation in ghrelin signaling. Importantly, eNOS activation is critical for ghrelin inhibition of vascular inflammation. Together, both in vitro and in vivo data demonstrate a new role of ghrelin signaling for eNOS activation, and highlight the therapeutic potential for ghrelin to correct endothelial dysfunction associated with atherosclerotic vascular diseases and metabolic syndrome.

Vascular endothelial growth factor (VEGF) is essential for normal and pathological angiogenesis. However, the signaling pathways linked to gene regulation in VEGF-induced angiogenesis are not fully understood. Here we demonstrate a critical role of protein kinase D (PKD) and histone deacetylase 5 (HDAC5) in VEGF-induced gene expression and angiogenesis. We found that VEGF stimulated HDAC5 phosphorylation and nuclear export in endothelial cells through a VEGF receptor 2-phospholipase Cgamma-protein kinase C-PKD-dependent pathway. We further showed that the PKD-HDAC5 pathway mediated myocyte enhancer factor-2 transcriptional activation and a specific subset of gene expression in response to VEGF, including NR4A1, an orphan nuclear receptor involved in angiogenesis. Specifically, inhibition of PKD by overexpression of the PKD kinase-negative mutant prevents VEGF-induced HDAC5 phosphorylation and nuclear export as well as NR4A1 induction. Moreover, a mutant of HDAC5 specifically deficient in PKD-dependent phosphorylation inhibited VEGF-mediated NR4A1 expression, endothelial cell migration, and in vitro angiogenesis. These findings suggest that the PKD-HDAC5 pathway plays an important role in VEGF regulation of gene transcription and angiogenesis.

Extracorporeal shock wave (SW) therapy has been studied in the transfection of naked nucleic acids into various cell lines through the process of sonoporation, a process that affects the permeation of cell membranes, which can be an effect of cavitation. In this study, siRNAs were efficiently transfected into primary cultured cells and mouse tumor tissue via SW treatment. Furthermore SW-induced siRNA transfection was not mediated by SW-induced sonoporation, but by microparticles (MPs) secreted from the cells. Interestingly, the transfection effect of the siRNAs was transferable through the secreted MPs from human umbilical vein endothelial cell (HUVEC) culture medium after treatment with SW, into HUVECs in another culture plate without SW treatment. In this study, we suggest for the first time a mechanism of gene transfection induced by low-energy SW through secreted MPs and show that it is an efficient physical gene transfection method in vitro and represents a safe therapeutic strategy for site-specific gene delivery in vivo.

Roles of dietary phytochemicals in cancer chemoprevention via induction of nuclear factor-erythroid-2-related factor 2 (Nrf2)-mediated antioxidant enzymes have been well established in a number of studies. In this study, FACS analysis was used to reveal that the intracellular reactive oxygen species level decreased at 0-25 μM of genipin treatment. Furthermore, immunofluorescence analysis and Western blotting were used to demonstrate that genipin treatment resulted in the upregulation and nuclear translocation of Nrf2, as well as upregulation of gastrointestinal glutathione peroxidase. Finally, we found that C-Jun-NH2-kinase (JNK) was also dose-dependently activated, where depleting JNK by using a biochemical inhibitor indicated that JNK was upstream of Nrf2. Interestingly, the antioxidant effects were limited to the treatment in the lower dosage of genipin, where higher dosage of genipin treatment resulted in the increased reactive oxygen species level and cytotoxicity. Thus, this study demonstrates for the first time that lower dosage of genipin results in the induction of JNK/Nrf2/ARE signaling pathway and protection from cell death.

An alternate application of MgO film for the improvement of discharge characteristics and lifetime in an ac plasma display panel (PDP) is suggested. In the normal ramp reset, the accumulated wall charges on the phosphor surface just after the reset period are different as to their color (red, green, blue) of each phosphor, which reduces the common voltage margin of ac PDP. In general, MgO is used for the protective layer of dielectrics from ion sputtering and the improvement of secondary electron emission which affects the driving voltage in an ac PDP. In this research, we deposited MgO on the phosphor to get the same addressing characteristics irrespective to the color of each phosphor, high speed addressing, and prolonged lifetime of phosphor despite the vacuum ultraviolet irradiation and ion bombardment. An optimized MgO thickness was used to avoid the excessive luminance and luminous efficacy degradation by the MgO deposition on the phosphor. Results showed that PDP with MgO coated phosphor has more uniform formative delays in address discharges and improved degradation characteristics.

In general, the background light that is produced during the reset period deteriorates the dark room contrast ratio in AC PDP. In this paper, we propose a modified ramp reset pulse that can reduce the background light to an imperceptible level. In the new reset waveform, the discharges between the scan and sustain electrodes are minimized by applying a positive bias voltage to the sustain electrode and only the weak discharges between the scan and address electrodes are found to occur during the reset period. We also adopted the MgO coated phosphor layer to improve the address voltage margin that was reduced when the bias voltage in the modified ramp reset waveform was applied. As a result, the address voltage margin of 45 V which is the same level of the conventional method was ortained and the dark room contrast ratio was improved up to 7500 : 1.

The U-tubes installed inside the steam generator experience high temperature and pressure as a role of heat transfer. Specially, during the secondary side hydrostatic test which ensures the integrity of steam generator, the U-tube is subjected to high external pressure. The purpose of this paper is to investigate the allowable external pressure of the U-tube in steam generator. In the ASME B&PV Code Section III [1], the allowable external pressure is determined by the rules of NB-3133. Alternatively, NB-3228 analysis may be applied. In order to determining the allowable external pressure of steam generator tube, the buckling analysis is performed. The analysis consists of the collapse pressure and elastic instability pressure analyses. In this study, these pressures are determined by finite element analysis (FEA) using ANSYS computer program. The non-linear static analysis is performed with ideally elastic-plastic material properties for the collapse analysis. On the other hand, the elastic instability pressure is calculated by eigenvalue analysis in elastic range. These allowable pressures are found to be 24.1 ksi and 10.5 ksi. Therefore the lower pressure of 24.1 ksi is the allowable external pressure of tube. In addition, the results of analysis are compared with other research [6] and Det Norske Veritas (DNV) offshore standards [11]. In conclusion, the results of buckling analysis are well matched with other research [6] and standard [11]. For steam generator tubes, the collapse pressure is dominant factor in failure. Also, the collapse pressure is largely influenced by the ovality of tube.

Soluble receptor for advanced glycation end products RAGE (sRAGE), a secretory form of RAGE, plays an important role in suppressing RAGE signals that induce pro-inflammatory gene activation in a range of inflammatory diseases, such as Alzheimer's disease, complications of diabetes mellitus and atherosclerosis. Recent studies have suggested that fluid shear stress generated by laminar blood flow protects blood vessels from atherosclerosis, whereas low and oscillatory shear stress (OSS) generated by disturbed blood flow causes atherosclerosis. Although RAGE levels are increased in atherosclerotic plaque, the regulatory mechanisms of sRAGE in the occurrence of atherosclerotic plaque induced by disturbed blood flow remain largely unknown. This study aimed to determine the effects of sRAGE as a competitive inhibitor of RAGE in atherogenesis induced by disturbed blood flow. To determine the role of sRAGE in atherosclerosis induced by disturbed blood flow, we used a mouse model of partial carotid artery ligation using ApoE(-/-) and C57BL/6 mice. Our results revealed that the expression of RAGE was significantly increased in the region of atherosclerotic plaque and that treatment with sRAGE attenuated the development of plaque formation. We found that the expression levels of RAGE and high mobility group box 1 (HMGB1), the agonistic ligand of RAGE, were significantly increased in human umbilical vein endothelial cells (HUVECs) under shear stress conditions induced by disturbed blood flow and suppressed following treatment with sRAGE. We further observed that treatment with sRAGE decreased the expression of vascular cell adhesion molecule‑1 (VCAM-1) and markedly attenuated monocyte-endothelial cell adhesion. Taken together, our results reveal that sRAGE exerts anti-atherogenic effects by blocking the activation of the RAGE signaling pathway induced by disturbed blood flow and may thus be a potential therapeutic target for the prevention of atherosclerosis.

Rationale: The Rad-Gem/Kir-related family (RGKs) consists of small GTP-binding proteins that strongly inhibit the activity of voltage-gated calcium channels. Among RGKs, Rem1 is strongly and specifically expressed in cardiac tissue. However, the physiological role and regulation of RGKs, and Rem1 in particular, are largely unknown. Objective: To determine if Rem1 function is physiologically regulated by adrenergic signaling and thus impacts voltage-gated L-type calcium channel (VLCC) activity in the heart. Methods and Results: We found that activation of protein kinase D1, a protein kinase downstream of α 1 -adrenergic signaling, leads to direct phosphorylation of Rem1 at Ser18. This results in an increase of the channel activity and plasma membrane expression observed by using a combination of electrophysiology, live cell confocal microscopy, and immunohistochemistry in heterologous expression system and neonatal cardiomyocytes. In addition, we show that stimulation of α 1 -adrenergic receptor-protein kinase D1-Rem1 signaling increases transverse-tubule VLCC expression that results in increased L-type Ca 2+ current density in adult ventricular myocytes. Conclusion: The α 1 -adrenergic stimulation releases Rem1 inhibition of VLCCs through direct phosphorylation of Rem1 at Ser18 by protein kinase D1, resulting in an increase of the channel activity and transverse-tubule expression. Our results uncover a novel molecular regulatory mechanism of VLCC trafficking and function in the heart and provide the first demonstration of physiological regulation of RGK function.

Objective— Grb2-associated binder 1 (Gab1), a scaffolding adaptor protein, plays an important role in transmitting key signals that control cell growth, differentiation, and function from multiple tyrosine kinase receptors. The study was designed to investigate the role of endothelial Gab1 in angiogenesis and its underlying molecular mechanisms. Methods and Results— Using Cre-Lox recombination technology, we generated endothelial-specific Gab1 knockout (Gab1-ecKO) mice. Gab1-ecKO mice are viable and showed no obvious developmental defects in the vascular system. To analyze the role of Gab1 in postnatal angiogenesis, we used hindlimb ischemia and Matrigel plug models. We found that loss of endothelial Gab1 in mice dramatically impaired postnatal angiogenesis. Gab1-ecKO mice had impaired ischemia-initiated blood flow recovery, exhibited reduced angiogenesis, and were associated with marked limb necrosis. We further observed significant endothelial cell (EC) death in the ischemic hindlimb of Gab1-ecKO mice. Matrigel plug assay showed that hepatocyte growth factor (HGF)–mediated angiogenesis was inhibited in Gab1-ecKO mice. In vitro studies showed that Gab1 was required for HGF-induced EC migration, tube formation, and microvessel sprouting. Mechanistically, HGF stimulated Gab1 tyrosine phosphorylation in ECs, leading to activation of extracellular regulated MAP kinase 1/2 and Akt, which are angiogenic and survival signaling. Conclusion— Gab1 is essential for postnatal angiogenesis through mediating angiogenic and survival signaling.

Various kinds of nozzles are attached to a pressure vessel including Steam Generator (SG) in a pressurized water reactor plant. The downcomer feedwater nozzle on the upper vessel shell and the economizer feedwater nozzle in the lower vessel shell of the SG are representative nozzles which have a non axi-symmetric shape. In most cases, external loads composed with forces and moments are imposed on those nozzles during the plant operation. In order to evaluate structural integrity of junctures between the nozzles and vessels in compliance with the ASME Boiler and Pressure Vessel Code, Section III, it is essential to find the maximum stress intensity resulting from those loads. Welding Research Council (WRC) Bulletin 297 has been used to find the maximum stress intensity since it is not straightforward to calculate the stress intensity with a non axi-symmetric two dimensional finite element model. However, the compatibility of adopting WRC Bulletin 297 to nozzles which have a variety of geometries shall be considered. Moreover, the applicability of the stress intensity resulting from the bulletin should be into consideration when interested lines where stress intensity linearization is to be performed are not exactly consistent with the line defined in the Bulletin. In this study, the nozzles in cylindrical vessel shells are developed as three dimensional finite element models, which are loaded with unit forces and moments. The stress intensities from finite element models are investigated through a comparison of WRC Bulletin 297. In addition, a methodology to apply the stress intensity results from WRC 297 to different lines is proposed.Copyright © 2011 by ASME

Dynamic nucleocytoplasmic shuttling of class IIa histone deacetylases (HDACs) is a fundamental mechanism regulating gene transcription. Recent studies have identified several protein kinases that phosphorylate HDAC5, leading to its exportation from the nucleus. However, the negative regulatory mechanisms for HDAC5 nuclear exclusion remain largely unknown. Here we show that cAMP-activated protein kinase A (PKA) specifically phosphorylates HDAC5 and prevents its export from the nucleus, leading to suppression of gene transcription. PKA interacts directly with HDAC5 and phosphorylates HDAC5 at serine 280, an evolutionarily conserved site. Phosphorylation of HDAC5 by PKA interrupts the association of HDAC5 with protein chaperone 14-3-3 and hence inhibits stress signal-induced nuclear export of HDAC5. An HDAC5 mutant that mimics PKA-dependent phosphorylation localizes in the nucleus and acts as a dominant inhibitor for myocyte enhancer factor 2 transcriptional activity. Molecular manipulations of HDAC5 show that PKA-phosphorylated HDAC5 inhibits cardiac fetal gene expression and cardiomyocyte hypertrophy. Our findings identify HDAC5 as a substrate of PKA and reveal a cAMP/PKA-dependent pathway that controls HDAC5 nucleocytoplasmic shuttling and represses gene transcription. This pathway may represent a mechanism by which cAMP/PKA signaling modulates a wide range of biological functions and human diseases such as cardiomyopathy.

A tube support plate is one of the significant parts of a steam generator, which confines the rotational and translational motion of tubes caused by the hydraulic and seismic load. It also provides a flow path along the tubes. There are various types of tube support plates according to the component designer’s preference. In this investigation, ten types of trefoil Broached Tube Support Plate (BTSP) specimens made from ASME stainless steel were analyzed and tested to determine the appropriate shape of trefoil BTSP in the view of the elastic properties including elastic modulus and Poisson’s ratio. The types of trefoil BTSP specimens were designated as SI through S5 and L1 through L5 for S and L types, respectively. These specimens are categorized by the shape and dimension of broached hole. Ten specimens were investigated through finite element analysis, and compression and bending tests. The dimensions of the test specimens were decided through a previous research study done to examine appropriate shape for the compression and bending tests. The equivalent elastic properties of BTSP were obtained by the finite element analysis as per different loading orientation as well as the various specimen types. Autodesk® Inventor™ software was used to make the analytical model and ABAQUS® software was used for the analysis and post-processing. The equivalent elastic properties of BTSP specimens were also acquired by the compression and bending tests. From the results of the finite element analysis, and the compression and bending tests, the appropriate shapes of trefoil BTSP with regard to the equivalent elastic modulus, and Poisson’s ratio are suggested as L4, S3, and S4.Copyright © 2010 by ASME

Fluid shear stress generated by steady laminar blood flow protects vessels from atherosclerosis. Krüppel-like factor 2 (KLF2) and endothelial nitric oxide synthase (eNOS) are fluid shear stress–responsive genes and key mediators in flow anti-inflammatory and antiatherosclerotic actions. However, the molecular mechanisms underlying flow induction of KLF2 and eNOS remain largely unknown. Here, we show a novel role of histone deacetylase 5 (HDAC5) in flow-mediated KLF2 and eNOS expression. We found for the first time that fluid shear stress stimulated HDAC5 phosphorylation and nuclear export in endothelial cells through a calcium/calmodulin-dependent pathway. Consequently, flow induced the dissociation of HDAC5 and myocyte enhancer factor-2 (MEF2) and enhanced MEF2 transcriptional activity, which leads to expression of KLF2 and eNOS. Adenoviral overexpression of a HDAC5 phosphorylation–defective mutant (Ser259/Ser498 were replaced by Ala259/Ala498, HDAC5-S/A), which shows resistance to flow-induced nuclear export, suppressed flow-mediated MEF2 transcriptional activity and expression of KLF2 and eNOS. Importantly, HDAC5-S/A attenuated the flow-inhibitory effect on monocyte adhesion to endothelial cells. Taken together, our results reveal that phosphorylation-dependent derepression of HDAC5 mediates flow-induced KLF2 and eNOS expression as well as flow anti-inflammation, and suggest that HDAC5 could be a potential therapeutic target for the prevention of atherosclerosis.

There are various types of tube support plates installed in a steam generator according to the component designer’s preference. Most widely used types of tube support plates are BTSP (broached tube support plate), ATSG (advanced tube support grid), and the eggcrate. In this study, trefoil BTSP specimens made of ASME stainless steel are analyzed and tested. This study is to investigate the effect of specimen shape on an elastic behavior of trefoil BTSP through the compression and bending tests. Prior to the compression and bending tests of BTSP specimens, the equivalent elastic properties of BTSP unit cell are analyzed by the finite element analysis according to the different loading orientation as well as size of the model. Autodesk® Inventor™ software was used to make an analytical model and ANSYS® software was used for the finite element analysis and post-processing. Five and three different shapes of trefoil BTSP specimens are machined and utilized for the compression and bending (4-point and 3-point side bending) tests, respectively. Through the finite element analyses, compression, and bending tests, the equivalent elastic modulus of trefoil BTSP specimen is suggested to be 6,254MPa (907ksi) and the equivalent Poisson’s ratio as 0.64. Specifically the CS5 type specimen which has a ratio of one-fourth (= width/length) was revealed as an appropriate shape of specimen to show those elastic behavior.Copyright © 2009 by ASME

Objective— Histone acetylation/deacetylation plays an important role in the control of gene expression, tissue growth, and development. In particular, histone deacetylases 7 (HDAC7), a member of class IIa HDACs, is crucial in maintaining vascular integrity. However, whether HDAC7 is involved in the processes of vascular endothelial signaling and angiogenesis remains unclear. Here, we investigated the role of HDAC7 in vascular endothelial growth factor (VEGF) signaling and angiogenesis. Methods and Results— We show for the first time that VEGF stimulated phosphorylation of HDAC7 at the sites of Ser178, Ser344, and Ser479 in a dose- and time-dependent manner, which leads to the cytoplasmic accumulation of HDAC7. Using pharmacological inhibitors, siRNA, and adenoviruses carrying dominant-negative mutants, we found that phospholipase Cγ/protein kinase C/protein kinase D1 (PKD1)-dependent signal pathway mediated HDAC7 phosphorylation and cytoplasmic accumulation by VEGF. Infection of ECs with adenoviruses encoding a mutant of HDAC7 specifically deficient in PKD1-dependent phosphorylation inhibited VEGF-induced angiogenic gene expression, including matrix metalloproteinases MT1-matrix metalloproteinase (MMP) and MMP10. Moreover, HDAC7 and its targeting genes were involved in VEGF-stimulated endothelial cell migration, tube formation, and microvessel sprouting. Conclusions— Our results demonstrate that VEGF stimulates PKD1-dependent HDAC7 phosphorylation and cytoplasmic accumulation in endothelial cells modulating gene expression and angiogenesis.